MOLECULAR & CELLULAR Biophysics and

BiochemistryGraduate Studies Program 2011-2012

Faculty Research

Robert E. Baier ...................................................................... 4

Interfacial Biophysics, addressing structure and function ofmacromolecules at boundaries of living tissues with prostheticdevices such as artificial heart valves and dental implants

Sathy V. Balu-Iyer (Sathyamangalam V. Balasubramanian) ............................. 6 Pharmaceutical Biotechnology; Formulation and Delivery ofProtein Therapeutics

David A. Bellnier..................................................................... 6Photodynamic Therapy: Basic and Translational Research

Mikhail V. Blagosklonny ........................................................ 7Intracellular signal transduction pathways in cancer and aging

William Cance ....................................................................... 9Biology of focal adhesion kinase (FAK); Role of FAK in preventing apoptosis; Novel drugs that target FAK pathway;Other survival mechanisms of cancer cells

Mikhail Chernov ................................................................... 10Bioactive Small Molecules and Functional Screening

Vita M. Golubovskaya.......................................................... 11Focal Adhesion Kinase Expression and Signaling in cancer

Andrei V. Gudkov.................................................................. 11Cancer treatment and normal tissue protection by targetingmajor stress response pathways; functional screeningapproaches to gene and drug discovery.

Katerina V. Gurova ............................................................... 14Anti-cancer drug discovery through modulation of transcriptional factors activity in tumor cells

Eugene S. Kandel ............................................................... 15Genetic dissection of signal transduction in mammalian cells

A. Latif Kazim ....................................................................... 16Biomolecular Resources: Proteomics, Mass Spectrometry, DNA Sequencing and NMR Spectroscopy; Research Interests:Proteomics, Metabolomics and Imaging Mass Spectrometry

Eleva V. Kurenova ................................................................ 17Protein complexes of Focal Adhesion Kinase as the targets in tumor growth, angiogenesis and metastasis

Asoke K. Mal ........................................................................ 18Epigenetics, Transcription factors and Childhood Cancer

Harish K. Malhotra............................................................... 19Applications of Medical Physics to cancer diagnosis andtreatment

Janet Morgan ....................................................................... 20Mechanisms and enhancement of Photodynamic Therapy incancer and cancer stem cells

Daryl P. Nazareth ................................................................. 21Applications of Medical Physics to cancer diagnosis andtreatment

Mikhail A. Nikiforov.............................................................. 21Molecular mechanisms of transformation and senescence

Ravindra K. Pandey ............................................................. 22Multifunctional agents for imaging and therapy

Matthew B. Podgorsak........................................................ 23Applications of Medical Physics to Cancer Diagnosis andTreatment

Arindam Sen ......................................................................... 24Nanoparticle drug formulation and tumor microenvironment

Mukund Seshadri ................................................................ 25Cancer Imaging/Targeted Therapies

Joseph Spernyak ................................................................. 25Magnetic resonance imaging in animal models of disease

Robert M. Straubinger ........................................................ 26Modulation of anticancer drug biodisposition andpharmacodynamics by drug carriers

Ulas Sunar ............................................................................ 27Novel optical imaging techniques for therapy monitoring

CONTENTSMolecular & Cellular Biophysics & Biochemistry Program . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

About Buffalo, New York . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

Map – Directions to Roswell Park Cancer Institute . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

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GRADUATE PROGRAM “BETTER CANCER THERAPIES THROUGH KNOWLEDGE AND INNOVATION.”

The current understanding of cancer as a disease and clinical approaches to treating cancer have made enormous gains in the pastcouple of decades. This was made possible by tremendous technological advances that delivered unparalleled capabilities and awealth of knowledge to the biomedical field. In our research, we use the insights into stress response mechanisms in normal andcancerous cells to improve the outcomes of cancer treatment, including the design of novel anti-cancer therapies, and ameliorationof the side effects and enhancement of efficacy of the available ones.

Our faculty members maintain a strong tradition of technological innovation, which leads to the establishment of new concepts intreatment and diagnosis of the disease, and to the discovery of prospective therapeutic targets and potent chemotherapeutic agents.Our students are exposed to a challenging curriculum delivered by a team of experts from RPCI and other institutions and conducthigh-impact research in several crucial and interlinked areas of modern oncology:

• Through the studies of signal transduction and gene regulation we expand the knowledge of the origins of cancer and thetherapeutic responses of the disease.

• Gene and drug discovery capitalizes on our expertise in biology of normal and tumor cells in order to identify andcharacterize therapeutic targets and chemotherapeutic agents.

• Biophysical therapies intensively studied by our scientists include radiation, thermal and photodynamic (PDT) therapies. PDT has been developed at Roswell Park Cancer Institute and our Institution remains the world leader in research and clinical applications of this technology.

• Our teams of biophysicists, chemists, and molecular biology and nanotechnology experts develop and optimize cancer targeting strategies for drug delivery, as well as for imaging and diagnostic purposes.

Successful integration of basic and clinical research remains the main strength of Roswell Park Cancer Institute. A network of closecollaborations with clinicians and industrial scientists expedites the transition of relevant findings “from bench to bedside” andprovides a comprehensive and versatile training experience for our students.

Molecular & Cellular Biophysics & Biochemistry

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About Buffalo, New YorkRoswell Park Cancer Institute, one of the oldest cancer research institutes in the world, is located on several blocks near otherhospitals within a mile of downtown Buffalo. Buffalo, the second largest city in New York, enjoys the cultural and social advantagesof many larger cities and offers a relaxed pace of life and exceptionally easy access to the surrounding countryside and lakefronts.Located at the eastern end of Lake Erie, buffalo is 15 miles from Niagara Falls and across Lake Ontario from Toronto. Lake Eriemoderates winter and summer temperatures and provides outstanding recreational opportunities in boating, swimming, fishing, and diving. The surrounding hills, fields, and forests in western New York and southern Ontario provide excellent downhill and cross-country skiing, hiking, and camping. Accessible, inexpensive, and convenient flights offer year-round access to New York and other major East Coast, Midwestern and Southern cities.

Directions to Roswell Park Cancer Institute

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Interfacial Biophysics,addressing structure andfunction of macromoleculesat boundaries of livingtissues with prostheticdevices such as artificial

heart valves and dental implants.Robert E. Baier, PhD, PEProfessor and Director, Biomaterials Graduate ProgramState University of New York at Buffalo

Beginning as a surgical technician operating heart-lung machineand dialysis equipment at the Buffalo General Hospital in 1959,he progressed through Bachelor of Engineering Science(Physics, Cleveland State University) and Ph.D. (Biophysics)degrees to post-doctoral training as a National Academy ofSciences fellow (Surface Chemistry) in Washington D.C. (1966-68). Dr. Baier spent sixteen years on the professional researchstaff of Calspan Advanced Technology Center prior to joiningSUNYAB full time. He was Executive Director of the New YorkState Center for Advanced Technology in Health-careInstruments and Devices (1984-1989), and now is ExecutiveDirector of the Industry/University Center for Biosurfacessponsored by the U.S. National Science Foundation. He isextensively published in many areas of biosurface physics,particularly involving dental and medical implant technology.

Dr. Baier is available for consulting assignments on a limitedbasis due to his teaching obligations in the Schools of Medicine,Dentistry, and Engineering.

EDUCATION/TRAINING

Cleveland State University (Cleveland, OH), B.E.S. 1962,Engineering Sci/Physics

State University of New York at Buffalo, Ph.D. 1966, Biophysics

National Academy of Sciences/National Research Council(Washington, DC), Postdoctoral Associate 1966-1968, SurfaceChemistry [U.S. Naval Research Lab]

RESEARCH AND PROFESSIONAL EXPERIENCE

Employment1959-1961 Surgical Research Laboratory Technician, Buffalo

General Hospital, Buffalo, NY

1961-1962 Nucleonics Engineer, Republic Steel Corporation,Cleveland, OH

1968-1984 Staff Scientist (Principal Physicist 1976-1980,Research Physicist 1968-1976), AdvancedTechnology Center Arvin/Calspan (formerly CornellAeronautical Laboratory of Cornell University),Buffalo, NY

1985-1989 Director, NY State Center for Advanced Tech.,Health-care Instruments & Devices Institute (SUNYBuffalo)

1972-1996 Professor (Assoc. Prof. 1990-1993, Res. Asst. Prof.1976-1990 Dept. Biomaterials, SUNY Buffalo

1996-pres Professor, Department of Oral Diagnostic Sciences,SUNY Buffalo

1998-pres Director, Biomaterials Graduate Program, SUNYBuffalo

Additional Experience1968-pres Registered Professional Engineer, State of Ohio

License #E-033541

1970-1995 Faculty Associate in the University Seminar onBiomaterials, Columbia University in the City of NewYork

1971-1976 Adjunct Associate Professor, School of ChemicalEngineering, Cornell University, Ithaca, NY

1971-pres Registered Professional Engineer, State of New YorkLicense #047547

1972-pres Associate Research Professor (Asst Res Prof 1972-1990), Dept. Molecular and Cellular Biophysics,Roswell Park Division, Graduate School of SUNYBuffalo

1977-pres Editorial Board, Journal of Biomedical MaterialsResearch

1978-1980 NIH/NHLBI Working Group on PhysicochemicalCharacterization of Biomaterials

1982 Expert Panel, NIH Consensus DevelopmentConference, “Clinical Applications of Biomaterials”

1983-pres Research Professor, Dept. Physiology andBiophysics, SUNY Buffalo

Faculty Research

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1988 Expert Panel, NIH Consensus DevelopmentConference on “Dental Implants”

1988-pres Executive Director (Co-Director 1988-1992), NSFIndustry/University Center for Biosurfaces, SUNYBuffalo

1988-2004 Editorial Consulting Staff, The International Journalof Oral & Maxillofacial Implants

1991-pres Adjunct Professor, Center for Bioengineering &Dept. Mechanical and Aerospace Eng., SUNYBuffalo

1992 Expert Panel, DHHS Breast Implant Task Force

1992 Member, U.S. Army Research Laboratory (ARL)Biotechnology Assessment Committee

1992-1993 President, Society For Biomaterials

1994-1996 U.S. Chairman, Implant Data:Record/Report/Review (3-year internationalconference series) Hyannis, MA, 1994; Melbourne,Australia, 1995; Buffalo, NY, 1996

1995-1999 Editorial Board, Journal of Dental Research

2000 Expert Panel, NIH Technology AssessmentConference on Implant Retrieval

2000-pres Editorial Board, The Journal of Adhesion

2004-pres Member, U.S. Army Biotechnology Working Group

2009-pres AAMI Blood/Gas Exchange Device Committee, U.S.Sub-TAG for ISO/TC 150, Blood-contact coatings

Memberships: Nat’l Soc Prof. Engineers (1962-pres); Amer Chem Soc (1965-pres); Amer Soc Cell Biol (1965-pres); Biophys Soc (1965-pres);Amer Soc Artif Internal Organs (1970-pres); Sigma Xi, The SciRes Soc (1972-pres); Soc For Biomaterials (Founding Member,1974-pres); Int’l Assoc Colloid and Surface Chem (1980-pres);Canadian Biomaterials Soc (1981-pres); Int’l Assoc Dent Res(1984-pres); The Adhesion Soc (1984-pres); Acad Surg Res(1989-pres); Amer Conf Gov Industrial Hygienists (1990-pres);Int’ Soc for Artif Cells & Immobilization Biotech (1992-pres);Amer Acad Cardiovasc Perfusion(1993-pres); Amer Inst Med BiolEng (1993-pres); Amer Ceramic Soc (1999-pres)

Honors1966-1968 NRC Post-Doctoral Associateship [1962-1965 NIH

Pre-Doctoral Traineeship]

1971 Union Carbide Chemicals Prize, awarded byAmerican Chemical Society

1983 Clemson Award for Basic Research, awarded bySociety For Biomaterials, 1983

1987 Award for Innovation in Medical Devices, AmericanSociety for Artificial Internal Organs

1989 Chairman, Gordon Research Conference onBiocompatibility and Biomaterials

1993 Founding Fellow, American Institute of Medical andBiomedical Engineering

1994 Fellow, Biomaterials Science and Engineering, Int’lUnion of Societies for Biomaterials Science &Engineering

1994 D.Odont., honoris causa, Lund University, Sweden

2000 Engineer of the Year Award, Fenn College ofEngineering, Cleveland State University

2001 Humanitarian Award, Health Care IndustriesAssociation of the Niagara Frontier

2002 Founders Award, Society For Biomaterials

2005 Sharma Award, International, Society forBiomaterials and Artificial Organs, India

Select Publications, including Relevant Abstracts andProceedings (1995 - present)

1. Baier RE, Zobel CR (1966) Structure in Protein Films: Myosin Monolayers,Nature, 212:351-353.

2. Baier RE, Shafrin EG, Zisman WA (1968) Adhesion: Mechanisms that Assist orImpede It, Science 162:1360-1368.

3. Baier RE, Dutton RC (1969) Initial Events in Interactions of Blood with ForeignSurfaces, J Biomed Mater Res 3:191-206.

4. Baier RE, Gott VL, Dutton RC (1972) Thromboresistance of Stellite 21:Adventitious Wax, J Biomed Mater Res, 6:465-470.

5. Boretos JW, Pierce WS, Baier RE, Leroy AF, Donachy HJ (1975) Surface andBulk Characteristics of a Polyether Urethane for Artificial Hearts, J BiomedMater Res 9:237-340.

6. Baier RE, Glantz P-O (1978) Characterization of Oral In Vivo Films Formed onSolid Surfaces, Acta Odontol Scan 36:289-301.

7. Baier RE (1978) Noninvasive, Rapid Characterization of Human Skin ChemistryIn Situ, J Soc Cosmet Chem, 29:283-306.

8. Pierce WS, Donachy JH, Rosenberg G, Baier RE (1980) Calcification InsideArtificial Hearts: Inhibition by Warfarin-Sodium, Science, 208:601-603.

9. Baier RE, Akers CK, Natiella, Jr, Meenaghan MA, Wirth J (1980) PhysiochemicalProperties of Stabilized Umbilical Vein, Vascular Surgery 14(3):145-157.

10. Baier RE (1982) Conditioning Surfaces to Suit the Biomedical Environment:Recent Progress, J Biomech Eng 104:257-271

11. Baier RE, Meyer AE, Akers CK, Natiella JR, Meenaghan MA, Carter JM (1982)Degradation effects of conventional steam sterilization on biomaterial surfaces,Biomaterials 3:241-245.

12. Baier RE, Meyer AE, Natiella JR, Carter JM (1984) Surface Properties DetermineBioadhesive Outcomes: Methods and Results, J Biomed Mater Res, 18:337-355.

13. Baier RE (1985) Cell Seeding: Biomaterial Surface Preparation, ASAIO Journal8:104-108.

14. Baier RE, DePalma VA, Goupil DW, Cohen E (1985) Human platelet spreadingon substrata of known surface chemistry, J Biomed Mater Res, 19:1157-1167.

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15. Baier RE, Meyer AE (1988) Implant Surface Preparation, Int J Oral MaxillofacImplants 3:9-20.

16. Baier RE, Meenaghan MA, Hartman LC, Flynn HE, Meyer AE, Natiella Jr, CarterJM (1988) Implant Surface Characteristics and Tissue Interaction, J OralImplant XIII:594-606.

17. Rittle KH, Helmstetter CE, Meyer AE, Baier RE (1990) Escherichia coliRetention on Solid Surfaces as Functions of Substratum Surface Energy andCell Growth Phase, Biofouling 2:121-130.

18. Muller-Mai CM, Voigt C, Baier RE Gross U (1992) The Incorporation of Glass-Ceramic Implants in Bone After Surface Conditioning Glow DischargeTreatment, Cells and Materials 2: 309-327.

19. Baier RE (1999) Physical and biomechanical issues in graft design. SemVascular Surg 12(1):8-17.

20. Baier R, Meyer A, Glaves-Rapp D, Axelson E, Forsberg R, Kozak M, NickersonP (2000) The Body's Response to Inadvertent Implants: Respirable Particles inLung Tissues. Journal of Adhesion 74:103-124.

21. Baier R, Axelson E, Meyer A, Carter L, Kaplan D, Picciolo G, Jahan S (2000)The Body's Response to Deliberate Implants: Phagocytic Cell Responses toLarge Substrata vs. Small Particles. Journal of Adhesion 74:79-102.

22. Edsberg LE, Natiella JR, Baier RE, Earle J (2001) Microstructural characteristicsof human skin subjected to static versus cyclic pressures. Journal ofRehabilitation Research and Development 38(5):477-486.

23. Baier RE (2002) A Challenging Anomaly – Glass that Does NOT Clot Blood!.The Glass Researcher 12(1&2):23-24.

24. Kukulka, DJ, Baier, RE, Mollendorf, JC (2004) Factors Associated with Foulingin the Process Industry. Heat Transfer Engineering 25: 23-29.

25. Drake, LA, Meyer, AE, Forsberg, RL, Baier RE, Doblin MA, Heinemann S,Johnson WP, Kock, M, Rublee, PA, Dobbs, FC (2005) Potential Invasion ofMicroorganisms and Pathogens via “Interior Hull Fouling”:Biofilms InsideBallast Water Tanks. Biological Invasions 7:969-982.

26. Baier RE (2006) Surface Behaviour of biomaterials: The theta surface forbiocompatibility. J Mater Sci: Mater Med 17:1057-1062.

27. Meyer AE, Baier RE, Chen H, Chowhan, M (2007) Differential Tissue-on-TissueLubrication by Ophthalmic Formulations. J Biomed Mater Res, Part B: AppliedBiomaterials 82B:74-88

28. Rangwala HS, Ionita C, Baier RE, Rudin S (2009) Partially PolyurethaneCovered Stents (PCCS) for Cerebral Aneurysm Treatment, J Biomed Mater ResB Appl Biomater 89B(@):415-429.

Professional Organizations

- Society for Biomaterials (Past President)

- American Institute for Medical and Biological Engineering (Vice President)

- Health Care Industries Association of the Niagara Frontier(Director)

- National Society of Professional Engineers

- American Chemical Society

PharmaceuticalBiotechnology; Formulationand Delivery of ProteinTherapeutics

Dr. Sathy V. Balu-Iyer (Sathyamangalam. V. Balasubramanian)Associate Professor, Pharmaceutical Sciences

Dr. Sathy V. Balu-Iyer (Sathyamangalam V. Balasubramanian) isan Associate Professor of Pharmaceutical Sciences. His researchinterest is in the area of Pharmaceutical Biotechnology, inparticular, formulation and delivery of protein therapeutics usingan interdisciplinary approach of Biophysics (Bioengineering),Immunology and Pharmaco Kinetics and Pharmacodynamics. Dr. Balu-Iyer is the Associate Director of the PharmaceuticalSciences Instrumentation Facility. He was a post doctoral fellowin the Department of Pharmaceutical Sciences, State Universityof New York at Buffalo. He received his Ph.D. in MolecularBiophysics from the Indian Institute of Science in Bangalore, India.

Photodynamic Therapy: Basic andTranslational Research

David A. Bellnier, PhDAssistant Professor of Oncology

The objective of this program is to devise and implement moreeffective approaches to Photodynamic Therapy (PDT). PDTinvolves the administration of a photodynamically-active drug(photosensitizer) or pro-drug followed by drug-activating visiblelight. This therapy has been successfully applied to bothneoplastic and non-neoplastic diseases.

This research program takes place in the multidisciplinary, highlyinteractive environments of the Photodynamic Therapy Centerand the Biophysical Therapies Program. As such, numerous linesof attack are being taken to improve both the efficacy andselectivity of PDT, including (i) the rational design, synthesis andtesting of new photosensitizers 1, 2 [directed by Dr. Pandey inour group], (ii) the design and application of optimal therapeuticregimens, e.g., drug dosage and schedule based onphamacokinetic/pharmacodynamic studies3 [directed by Dr.Bellnier] and light dosage and schedule based on fluence/fluencerate studies4 [directed by Dr. Henderson in our group] and (iii)the study of multimodal approaches 5-7.

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We have had a long-standing program to study the interactionbetween vascular disrupting agents (VDAs) and PDT. Much ofour past and current work has focused on the antivascular agentvadimezan (DMXAA; 5,6-dimethylxanthenone-4-acetic acid) 4.Although vadimezan does not appear to be directly cytotoxic totumor cells, it induces biological responses that include early-onset endothelial cell apoptosis, the production of cytokines byboth host and tumor cells, as well as the induction of vasoactivecompounds like NO and serotonin. The synthesis of the cytokineTNF-alpha is largely responsible for vadimezan’s dramaticantitumor-antivascular effects in murine tumors and xenografts.We have shown that neoadjuvant DMXAA dramatically increasesthe antitumor activity of Photofrin-sensitized PDT in rodent tumormodels, and have observed a similar interaction betweenvadimezan both HPPH and delta-aminolevulinic acid (ALA)-protoporphyrin IX-sensitized PDT. Recent experiments, inconjunction with Dr. Gollnick, suggest that vadimezan mayinitiate an antitumor immune response. We have recently foundthat vadimezan also may enhance direct photodynamic effectson the vasculature by increasing photosensitizer levels in tumorendothelial cells (with Dr. Morgan:http://classic.roswellpark.org/Site/Research/Research_Staff/Morgan_Janet_PhD). This appears to be due to vadimezan-mediatedinhibition of the ABCG2 transporter that is expressed onendothelial cells and which pumps out known substrates such asthe photosensitizer HPPH. In addition, we plan to study theinteraction between PDT and tubulin-binding VDAs, in particulardinitrogen-substituted stilbene analogues structurally similar tocombretastatin.

Select Publications1. Henderson BW, DA Bellnier, WR Greco, A Sharma, RK Pandey, LA Vaughan, KRWeishaupt and TJ Dougherty. An in vivo quantitative structure-activityrelationship for a congeneric series of pyropheophorbide derivatives asphotosensitizers for photodynamic therapy. Cancer Res 57:4000-4007, 1997.

2. Zheng G, WR Potter, SH Camacho, JR Missert, G Wang, DA Bellnier, BWHenderson, MAJ Rodgers, TJ Dougherty and RK Pandey. Synthesis,photophysical properties, tumor uptake, and preliminary in vivo photosensitizingefficacy of a homologous series of 3-(1’-alkyoxy)ethyl-3-devinylpurpurin-18-N-alkylimides with variable lipophilicity. J Med Chem 44:1540-1559, 2001.

3. Bellnier DA, WR Greco, H Nava, AR Oseroff, GM Loewen, T Tsuchida, RKPandey and TJ Dougherty. Population pharmacokinetics of the photodynamictherapy agent 2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide-a (HPPH) incancer patients. Cancer Res 63:1806-1813, 2003.

4. Henderson BW, TM Busch, LA Vaughan, NP Frawley, D Babich, TA Sosa, JDZollo, AS Dee, MT Cooper, DA Bellnier, WR Greco and AR Oseroff. Photofrinphotodynamic therapy can significantly deplete or preserve oxygenation inhuman basal cell carcinomas during treatment, depending on fluence rate.Cancer Res 60:525-529, 2000.

5. Snyder JW, WR Greco, DA Bellnier, L Vaughan and BW Henderson.Photodynamic therapy: A means to enhanced drug delivery to tumors. CancerRes 63:8126-8131, 2003.

6. Bellnier DB, SO Gollnick, SH Camacho, WR Greco and RT Cheney. Treatmentwith the tumor necrosis factor-alpha-inducing drug 5,6-dimethylxantheone-4-acetic acid enhances the antitumor activity of the photodynamic therapy of RIF-1 tumors. Cancer Res 63:7584-7590, 2003.

7. Bellnier DA. Potentiation of photodynamic therapy in mice with recombinanthuman tumor necrosis factor-alpha. J Photochem Photobiol 8:203-210, 1991.

8. Zheng X, J Morgan, SK Pandey, Y Chen, E Tracy, H Baumann, J Missert, C Batt,J Jackson, D Bellnier, BW Henderson and RK Pandey. Conjugation of HPPH tocarbohydrates changes its subcellular distribution and enhances photodynamicactivity in vivo. J Med Chem 52: 4306–4318, 2009.

9. Seshadri M and DA Bellnier. The vascular disrupting agent 5,6-dimethylxanthenone-4-acetic acid improves the antitumor efficacy and shortenstreatment time associated with Photochlor-sensitized photodynamic therapy invivo. Photochem Photobiol 85:50-56, 2009.

10. Seshadri M, R Mazurchuk, JA Spernyak, A Bhattacharya, YM Rustum and DABellnier. Activity of the vascular-disrupting agent 5,6-dimethylxanthenone-4-aceticacid against human head and neck carcinoma xenografts. Neoplasia 8:534-542,2009.

Intracellular signaltransduction pathways in cancer and aging

Mikhail V. Blagosklonny, MD, PhDProfessor of Oncology

Dr. Blagosklonny is the author of over 170 research articles,reviews and book chapters. He is the Founding Editor andEditor-in-Chief of Cell Cycle and also Co-editor and co-founderof Aging and also serves as an Associate Editor for CancerBiology & Therapy, Autophagy, Cancer Research, Cell Death andDifferentiation, International Journal of Cancer, The AmericanJournal of Pathology and PLOS ONE.

His research interests range from molecular and cellular biologyto clinical investigations and specifically include oncogenes andtumor suppressors, signal transduction, cell cycle, mitosis,apoptosis, anticancer therapeutics with emphasis on translationof basic science into new anticancer strategies such asexploiting cancer cell cycling and drug resistance for selectiveprotection of normal cells. He has extended this approach toother age-related diseases and aging itself, thus revealing ananti-aging drug to be used today (Cell Cycle, 2006, 5 : 2087-2102).

Area of General Research Interest- Cancer biology and therapy

- Selective targeting cancer cells using drug combinations

- Mechanisms of aging and anti-aging drugs

Current Program- Targeted combinations of anti-cancer agents and protection ofnormal cells

- Pharmacologic suppression of cellular and organismal aging

We have suggested that aging is not caused by moleculardamage (nor by free radicals) but instead is a purposeless quasi-program driven in part by TOR (Target of Rapamycin). Theoreticalanalysis of cellular senescence, organismal aging, diseases ofaging and effects of rapamycin reveals that rapamycin is an anti-aging drug that could be used today to slow down aging inhumans and to prevent age-related diseases including cancer.

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In 2009-2010, we have demonstrated that:a. mTOR is required for cellular senescence and that rapamycinsuppresses cellular senescence, by transforming it intoquiescence.

b. Inhibitors of MEK and PI-3K, at non-toxic concentrationsinhibit mTOR and also suppresses senescence. Therefore,certain agents, currently viewed as anti-cancer agents, couldbe potentially used, at low doses, as anti-aging agents.

c. Resveratrol, at concentrations that inhibit mTOR, alsosuppreses cellular aging.

d. Cellular aging is accompanied by pseudo-DNA damageresponse, which is inhibited by rapamycin.

e. p53 suppresses cellular senescence by inhibiting mTOR

f. Rapamycin prevents age-related weight gain, decreases rateof aging, increases life span and decreases carcinogenesis intransgenic HER-2/neu cancer-prone mice. We suggest that,by slowing down organismal aging, rapamycin delays cancer.

Select Publications

1. Demidenko ZN, Blagosklonny MV. Flavopiridol induces p53 via initial inhibition ofMdm2 and p21 and, independently of p53, sensitizes apoptosis-reluctant cellsto tumor necrosis factor. Cancer Res. 64: 3653-60, 2004.

2. Demidenko ZN, Halicka D, Kunicki J, McCubrey JA, Darzynkiewicz Z,Blagosklonny MV. Selective killing of adriamycin-resistant (G2 checkpoint-deficient and MRP1-expressing) cancer cells by docetaxel. Cancer Res. 65:4401-7, 2005.

3. Demidenko ZN, Rapisarda A, Garayoa M, Giannakakou P, Melillo G,Blagosklonny MV. Accumulation of hypoxia-inducible factor-1alpha is limited bytranscription-dependent depletion. Oncogene 24:4829-38, 2005.

4. Demidenko ZN, Vivo C, Halicka HD, Li CJ, Bhalla K, Broude EV, BlagosklonnyMV. Pharmacological induction of Hsp70 protects apoptosis-prone cells fromdoxorubicin: comparison with caspase-inhibitor- and cycle-arrest-mediatedcytoprotection. Cell Death Differ. 13:1434-41, 2006.

5. Demidenko ZN, Kalurupalle S, HankoC, Lim CU, Broude EV, Blagosklonny MV.Mechanism of G1-like arrest by low concentration of paclitaxel: next cell cyclep53- Dependent arrest with sub G1 DNA content mediated by prolongedmitosis. Oncogene. 27: 4402-10, 2008.

6. Demidenko ZN, Zubova SG, Bukreeva EI, Pospelov VA, Pospelova TV,Blagosklonny MV. Rapamycin decelerates cellular senescence. Cell Cycle8:1888-95.

7. Pospelova TV, Demidenko ZN, Bukreeva EI, Pospelov VA, Gudkov AV,Blagosklonny MV. Pseudo-DNA damage response in senescent cells. Cell Cycle.2009;8:4112-8.

8. Anisimov VN, Zabezhinski MA, Popovich IG, Piskunova TS, Semenchenko AV,Tyndyk ML, Yurova MN, Antoch MP, Blagosklonny MV. Rapamycin ExtendsMaximal Lifespan in Cancer-Prone Mice. Am J Pathol. 2010 Apr 2. [Epub aheadof print]

9. Demidenko ZN, Korotchkina LG, Gudkov AV, Blagosklonny MV. Paradoxicalsuppression of cellular senescence by p53. Proc Natl Acad Sci USA, 2010, May

Conceptual ReviewsBlagosklonny MV, Pardee AB. Exploiting cancer cell cycling forselective protection of normal cells. Cancer Res. 61: 4301-4305,2001.

Blagosklonny MV. Oncogenic resistance to growth-limitingconditions. Nature Reviews Cancer 2, 221-225, 2002.

Blagosklonny MV. Cell senescence and hypermitogenic arrest.EMBO Rep. 4:358-62, 2003.

Blagosklonny MV. Targeting cancer cells by exploiting theirresistance. Trends Mol Med. 9:307-12, 2003.

Blagosklonny MV. Matching targets for selective cancer therapy.Drug Discov Today 8:1104-7, 2003.

Blagosklonny MV. Antiangiogenic therapy and tumorprogression. Cancer Cell 5:13-7, 2004.

Blagosklonny MV. Prospective strategies to enforce selectivelycell death in cancer cells. Oncogene 23:2967-75, 2004.

Blagosklonny MV. How cancer could be cured by 2015. CellCycle 4:269-78, 2005.

Blagosklonny MV. Overcoming limitations of natural anticancerdrugs by combining with artificial agents. Trends Pharmacol Sci.26:77 81, 2005.

Blagosklonny MV. Carcinogenesis, cancer therapy andchemoprevention. Cell Death Differ. 12:592-602, 2005.

Blagosklonny MV. An anti-aging drug today: from senescence-promoting genes to anti-aging pill. Drug Discov Today 12:218-24, 2007.

Blagosklonny MV. "Targeting the absence" and therapeuticengineering for cancer therapy.Cell Cycle 7:1307-12, 2008.

Blagosklonny MV. Prevention of cancer by inhibiting aging.Cancer Biol Ther. 2008;7: 1520-1524, 2008.

Blagosklonny MV. Aging: ROS or TOR. Cell Cycle 7:3344-54,2008.

Blagosklonny MV and Hall MN. Growth and Aging: a commonmolecular mechanism. Aging 1: 357-362, 2009.

Blagosklonny MV. Validation of anti-aging drugs by treating age-related diseases. Aging. 2009;1:281-8.

Blagosklonny MV. Calorie restriction: decelerating mTOR-drivenaging from cells to organisms (including humans). Cell Cycle.2010; 9: 683-8.

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Biology of focal adhesionkinase (FAK); Role of FAKin preventing apoptosis;Novel drugs that target FAKpathway; Other survivalmechanisms of cancer cells

William Cance, MD Surgeon-in-ChiefChair, Department of Surgical OncologyProfessor of Oncology

Our team is focused on developing inhibitors of Focal AdhesionKinase (FAK), a non-receptor tyrosine kinase that plays animportant role in survival signaling. Our laboratory was the firstgroup to isolate human FAK cDNA from sarcoma tissues and toshow that this protein was upregulated in a wide variety ofhuman solid tumors. Focal adhesion kinase has also beenshown to be overexpressed in breast cancer tumors at earlystages of tumorigenesis. Our laboratory cloned the FAKpromoter and found transcription factors binding this regulatoryregion, including p53 and NF-ΚB. Our research work is currentlyfunded by the National Cancer Institute and the Susan G. KomenBreast Cancer Foundation.

Our group is focused on isolation of novel FAK-binding proteinsthat interact with the N-terminal domain of FAK, such asreceptor-interacting protein (RIP), p53 and NF-1. Receptorinteracting protein was the first of a few FAK-interacting pro-apoptotic proteins and analysis of this interaction led to thesuggestion of a possible mechanism of sequestering suchproteins by FAK and providing additional survival function in thecancer cells. We used the defined FAK site of RIP binding for insilico molecular docking of small molecules from the NCI DrugDiscovery Program database with the purpose to find somewhich disrupt the FAK-RIP complex and lead to apoptosis ofcancer cells, in collaboration with Dr. David Ostrov, University ofFlorida.

Another binding partner in the C-terminal domain of FAK isvascular endothelial growth factor receptor, VEGFR-3. Dr.Cance’s lab has demonstrated that FAK and VEGFR-3 areimportant protein tyrosine kinases that physically interact and areinvolved in the process of tumor progression. We demonstratedthat the FAK-VEGFR3 interaction provided essential survivalsignals for different types of cancer (breast, pancreatic andmelanoma). For the first time, we have demonstrated thatVEGFR-3 overexpression significantly promotes breast cancercell proliferation, motility, survival, anchorage-independentgrowth and tumorigenicity in the absence of ligand expression.By computer modeling and screening of NCI small moleculedatabase in collaboration with Dr. Ostrov (University of Florida)and biological function approaches the novel small moleculeinhibitor, C4 that specifically targeted VEGFR-3- FAK interactionhas been identified. The inhibitor blocked cellular viability in vitroand inhibited tumor growth of breast, melanoma and pancreatic

cancers on mouse xenograft models in vivo. Dr. Elena Kurenovaleads the VEGFR-3 project and collaborative study onmelanoma.

Our group isolated that p53 protein as a binding partner of FAK.Targeting this interaction with peptides and small moleculeinhibitors will be developed for future therapies. Computermodeling and NCI screening database isolated small moleculeinhibitors targeting the FAK-p53 and FAK-Mdm-2 interactions.The model that involves targeting these protein interactions inaddition to or independently of FAK-kinase function wererecently discussed in a review. In addition, Dr. VitaGolubovskaya conducted a study on a novel interaction of FAKand p53, funded by Susan G/ Komen Breast Cancer Foundation.We found in collaboration with University of North Carolina ahigh correlation of FAK overexpression with p53 mutations in 596breast cancer tumors that provide a basis for future targetedFAK-p53 therapy.

Finally, our group isolated a novel inhibitor of FAK, called Y15(compound 14) that targets the main autophosphorylation site ofFAK and caused breast tumor regression. The development ofFAK inhibitors will be the aim of future studies for an efficientcancer therapy. Dr. Golubovskaya directs both the p53/Mdm-2and the Y15 projects.

Select Publications1. Golubovskaya V, Kaur A, Cance WG. Cloning and characterization of thepromoter region of human focal adhesion kinase gene: nuclear factor kappa Band p53 binding sites. Biochim Biophys Acta 2004; 25;1678(2-3):111-125.

2. Kurenova E, Xu L, Yang X, Baldwin A, Craven R, Hanks S, Liu Z, Cance WG.Focal adhesion kinase suppresses apoptosis by binding to the death domain ofreceptor interacting protein. Mol Cell Biol 2004; 24(10):4361-4371.

3. Golubovskaya VM, Finch R, Cance WG. Direct interaction of the N-terminaldomain of focal adhesion kinase with the N-terminal transactivation domain ofp53. J Biol Chem 2005; 280(26):25008-25021.

4. Garces CA, Kurenova EV, Golubovskaya VM, Cance WG. Vascular endothelialgrowth factor receptor-3 and focal adhesion kinase bind and suppressapoptosis in breast cancer cells. Cancer Res 2006; 66(3):1446-1454.

5. Cance WG and Golubovskaya VM. Focal Adhesion Kinase (FAK) versus p53:apoptosis or survival? Sci Signal 2008; 1(20):pe22.

6. Golubovskaya VM, Nyberg C, Zheng M, Kweh F, Magis A, Ostrov D, Cance WG.A small molecule inhibitor, 1,2,4,5-benzenetetraamine tetrahydrochloride,targeting the y397 site of focal adhesion kinase decreases tumor growth. J MedChem 2008; 51(23):7405-7416.

7. Kweh F, Zheng M, Kurenova E, Wallace M, Golubovskaya V, Cance WG.Neurofibromin physically interacts with the N-terminal domain of focal adhesionkinase. Mol Carcinog, 2009;48(11): 1005-1017.

8. Kurenova EV, Hunt DL, He DH, Fu AD, Massoll NA, Golubovskaya VM, GarcesAG, Cance WG. Vascular endothelial growth factor receptor-3 promotes breastcancer cell proliferation, motility, and survival in vitro and tumor formation invivo. Cell Cycle, 2009;8(14): 2266-2280.

9. Kurenova, EV, Hunt DL, He D, Magis AT, Ostrov DA, Cance WG. Small moleculechloropyramine Hydrocholoride (C4) targets the binding site of focal adhesionkinase and vascular endothelial growth factor receptor 3 and suppresses breastcancer growth in vivo. J Med Chem 2009;52(15):4716-4724.

10. Golubovskaya, VM, Zheng M, Zhang L, Li, JL, Cance WG. The direct effect offocal adhesion kinase (FAK), dominant-negative FAK, FAK-CD and FAK siRNAon gene expression and human MCF-7 breast cancer cell tumorigenesis. BMCCancer 2009: 9:280.

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11. Golubovskaya VM, Conway -Dorsey K, Edmiston SN, Tse C-K, Lark AA, LivasyCA, Moore D, Millikan RC, Cance WG. FAK overexpression and p53 mutationsare highly correlated in human breast cancer. Int J Cancer 2009;125(7):1735-1738.

12. Golubovskaya VM, Cance W. focal adhesion kinase and p53 signaltransduction pathways in cancer. Front Biosci 2010:15-901-912.

Bioactive Small Moleculesand Functional Screening

Mikhail Chernov, PhDDirector, Small Molecule Screening Core FacilityAssociate Professor of Oncology

Our research focuses on identification of small organic moleculeswith distinctive effects on biological systems. This type ofresearch was always in the center of drug development in thepharmaceutical industry, but over the last two decades thecontribution of academic and medical institutes in this fieldincreased dramatically.

Two main approaches to identification of bioactive molecules arerational design and screening of chemical libraries usingbiological readout systems. Chemical libraries are collections ofdifferent chemical compounds arranged in multi-well plates forfast and convenient high-throughput screening (HTS).

The Small Molecule Screening Core at RPCI is a state of the art,high throughput screening facility where investigators can screenthe chemical libraries of more than 50,000 compounds in avariety of readout systems. Our libraries consist of over 50,000diverse uncharacterized compounds which can be used for drugdiscovery and over 3,000 pharmacological and naturalcompounds with known activities, which can be used as ascientific research tool.

The Core provides support in all stages of screening projectstarting from assay optimization for HTS format, screening ofcompounds, and data analysis.

We also engage in collaborative efforts to develop new assaysystems. One example of such collaboration is development ofreadout system for monitoring p53 activity in live cellsundergoing stress. This assay was used in a collaborative projectwith Dr. Andrei Gudkov as a cell-based read-out for screening ofchemical library of small molecules in a search for inhibitors ofp53 activation. This work resulted in identification of the first p53inhibitory small molecule pifithrin alpha, the first reportedchemical screening study done in an academic lab. We continuestudying p53 regulation and functions and participate in thedevelopment of new assays to monitor various aspects of p53activity in normal and transformed cells.

The other examples of current or recent projects in the labinclude the search for the small molecules modulators of

circadian clock regulation, modulators of muscle celldifferentiation and inhibitors of metastasis development.

Select publications:1. Smith KA, Agarwal ML, Chernov MV, Chernova OB, Deguchi Y, Ishizaka Y,Patterson TE, Poupon M, Stark GR. (1995) Regulation and mechanisms of geneamplification. Phil. Trans. R. Soc. Lond. B 347, 49-56

2. Komarova EA, Chernov MV, Franks R, Wang K, Armin G, Zelnick CR, Chin DM,Bacus SS, Stark GR, Gudkov AV. (1997) Transgenic mice with p53-resonsivelacZ: p53 activity varies dramatically during normal development and determinesradiation and drug sensitivity in vivo. EMBO Journal, 16, 1391-1400.

3. Chernova OB, Chernov MV, Ishizaka Y, Agarwal M, Stark GR. (1998) MYCabrogates p53-mediated cell cycle arrest in PALA-treated REF52 cells,permitting CAD gene amplification. Mol. Cell. Biol. 18(1): 536-545.

4. Garkavtsev I, Grigorian IA, Ossovskaya VS, Chernov MV, Chumakov PM,Gudkov AV. (1998) p33ing1, a candidate tumor supressor gene, cooperates withp53 in cell growth control. Nature; 391, 295-298.

5. Chernov MV, Ramana CV, Adler VV, Stark GR. (1998) Stabilization and activationof p53 are regulated independently by different phosphorylation events. Proc.Natl. Acad. Sci. U S A, 95 2284-2289.

6. Komarov PG, Komarova EA, Kondratov RV, Christov-Tselkov K, Coon JS,Chernov MV, Gudkov AV. (1999) A chemical inhibitor of p53 that protects micefrom the side effects of cancer therapy Science (United States), Sep 10 1999,285 (5434) p1733-7

7. Chernov MV, Bean LJ; Lerner N, Stark GR. (2001) Regulation of ubiquitinationand degradation of p53 in unstressed cells through C-terminal phosphorylation.J Biol Chem (United States), 276(34) p31819-24.

8. Kondratov RV, Chernov MV, Kondratova AA, Gorbacheva VY, Gudkov AV, AntochMP. (2003) BMAL1-dependent circadian oscillation of nuclear CLOCK:posttranslational events induced by dimerization of transcriptional activators ofthe mammalian clock system. Genes Dev. 17(15):1921-32.

9. Komarova EA, Krivokrysenko V, Wang K, Neznanov N, Chernov MV, KomarovPG, Brennan ML, Golovkina TV, Rokhlin OW, Kuprash DV, Nedospasov SA,Hazen SL, Feinstein E, Gudkov AV. (2005) p53 is a suppressor of inflammatoryresponse in mice FASEB J., 19, 1030-2.

10. Kondratov RV, Kondratova AA, Lee C, Gorbacheva VY, Chernov MV, AntochMP. (2006) Post-translational regulation of circadian transcriptionalCLOCK(NPAS2)/BMAL1 complex by CRYPTOCHROMES Cell Cycle ;5(8), 890-5.

11. Antoch MP, Chernov MV. Pharmacological modulators of the circadian clock aspotential therapeutic drugs. Mutat Res. 2009 Nov-Dec;680(1-2):109-15. PMID:20336820

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Focal Adhesion KinaseExpression and Signalingin cancer

Vita M. Golubovskaya, PhD Associate Professor of Oncology

The research focus is to understand the role and function ofFocal Adhesion Kinase in survival pathways duringtumorigenesis. Focal adhesion Kinase is overexpressed in manytypes of tumors and is involved in many intracellular processes:adhesion, motility, invasion, proliferation, angiogenesis andmetastasis. To understand regulation of Focal Adhesion Kinaseexpression we have cloned promoter of Focal Adhesion Kinaseand found p53 and NF-kappaB transcription factors in theregulatory sequence of promoter. One of the projects is tounderstand the mechanism of up-regulation of Focal AdhesionKinase in different types of tumors. We found that p53 inhibitedFAK expression through repression of FAK promoter, andanalysis of 600 breast tumors with mutant p53 demonstratedhigh correlation between p53 mutations and FAK overexpression.In addition, we demonstrated direct interaction of FAK and p53proteins and that FAK inhibit p53-transcriptional activity. We arestudying interaction of FAK and p53 pathways. One of theprojects is to target this interaction with small molecule inhibitorsto decrease survival of cancer cells.

Another direction is to target Focal Adhesion Kinaseautophosphorylation activity with novel small molecule inhibitorstargeting autophosphorylation Y397 site. Recently, we developednovel inhibitor of FAK autophosphorylation by computermodeling, virtual screening of small molecule compounds andfunctional studies. This strategy has been applied to breast,pancreatic, neuroblastoma and colon cancer and we were ableto decrease tumorigenesis in mice xenograft models with theseFAK inhibitors. Inhibition of FAK autophosphorylation and itsdown-regulation with FAKsiRNA are used to reveal the functionof Focal Adhesion Kinase in survival signaling, interaction withother signaling pathways, involving Src and PI3-Kinase, invasionand metastasis.

Select Publications 1. Beierle EA, Ma X, Trujillo A, Stewart J, Nyberg C, Trujillo A, Cance WG,Golubovskaya VM. Inhibition of Focal Adhesion Kinase decreases tumor growthin human neuroblastoma. Cell Cycle 2010; 9 (5)

2. Beierle EA, Ma X, Trujillo A, Kurenova EV, Cance WG, Golubovskaya VM.Inhibition of focal adhesion kinase and src increases detachment and apoptosisin human pancreatic cancer Mol Carcinog 2010, 49:224-234

3. Hochwald SN, Nyberg C, Zheng M, Zheng D, Wood C, Massoll NA, Magis A,Ostrov D, Cance WG, Golubovskaya V. A novel small molecule inhibitor of FAKdecreases growth of human pancreatic cancer. Cell Cycle 2009;8(15):2435-2443.

4. Golubovskaya VM, Nyberg C, Zheng M, Kweh F, Magis A, Ostrov D, Cance WG.A small molecule inhibitor, 1,2,4,5-benzenetetraamine tetrahydrochloride,targeting the y397 site of focal adhesion kinase decreases tumor growth. J MedChem. 2008; 51(23):7405-7416.

5. Cance WG, Golubovskaya VM. FAK vs p53, Survival or Apoptosis? ScienceSign, 1/20/22,2008

6. Bieierle EA, Trujillo A, Nagaram A, Cance WG, Kurenova E, Golubovskaya V.2007. N-Myc regulates Focal Adhesion Kinase (FAK) expression. J Biol Chem,282, 12503-12516.

7. Golubovskaya VM, Cance WG. 2007. Focal Adhesion Kinase and p53 signalingin cancer cells. Internl Review of Cytology, 263, 103-153.

8. Garces CA, Kurenova EV, Golubovskaya VM, Cance WG. 2006. VascularEndothelial Growth Factor Receptor-3 (VEGFR-3) and Focal Adhesion Kinase(FAK) Bind and Suppress Apoptosis in Breast Cancer Cells. Cancer Res., 66,1446-1454.

9. Golubovskaya VM, Finch R, Cance WG. 2005. Direct Interaction of the N-terminal domain of focal adhesion kinase with the N-terminal transactivationdomain of p53. J Biol Chem., 280, 25008-25021.

10. Golubovskaya V, Kaur A., and Cance W. 2004. Cloning and characterization ofthe promoter region of human Focal Adhesion Kinase gene: nuclear factorkappa B and p53 binding sites. BBA, 25, 1678, 111-125.

11. Vita Golubovskaya, Steven Gross, Aparna S. Kaur, Richard Wilson, Li-Hui Xu,William G. Cance. 2003. Simultaneous Inhibition of Focal Adhesion Kinase(FAK) and Src Enhances Detachment and Apoptosis in Colon Cancer CellLines. Mol. Cancer Research, 1, 755-764.

12. Golubovskaya V, Beviglia L, Xu LH, Earp HS 3rd, Craven R, Cance W. 2002.Dual inhibition of focal adhesion kinase (FAK) and epidermal growth factorreceptor (EGFR) pathways cooperatively induces death receptor-mediatedapoptosis in human breast cancer cells. J Biol Chem. 277(41): 38978-38987.

13. Watson J, Hurding T, Golubovskaya VM, Hunter D, Li X, Earp HS, Haskill JS.2001. CADTK is critical for monocyte spreading and motility. J Biol Chem.276(5): 3536-3542.

Cancer treatment andnormal tissue protection by targeting major stressresponse pathways;functional screeningapproaches to gene anddrug discovery.

Andrei V. Gudkov, PhD, DSci Sr. Vice President of Basic ScienceProfessor of Oncology Garman Family Chair in Cell Stress Biology

Andrei V. Gudkov, PhD, DSci, a pre-eminent cancer researcherwas appointed Senior Vice President for Basic Research; Chairof the Department of Cell Stress Biology, and a member of thesenior leadership team for National Cancer Institute (NCI) CancerCenter Support Grant at Roswell Park Cancer Institute (RPCI) in2007. He is responsible for building on the basic andtranslational research strengths of the Cell Stress Biology andDrug Discovery programs. As Senior Vice President, he assiststhe President & CEO in developing and implementing strategicplans for new scientific programs and enhance collaborations in

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research programs with regional and national academic centersas well as with industry.

Dr. Gudkov comes to Roswell Park from the Lerner ResearchInstitute, Cleveland Clinic Foundation where he served as Chairof the Department of Molecular Genetics and professor ofbiochemistry at Case Western University. He earned his doctoraldegree in Experimental Oncology at the Cancer Research Center,USSR and a Doctorate of Science (D.Sci) in Molecular Biology atthe Moscow State University, USSR. He has authored or co-authored >160 scientific articles and holds 27 patents. He is afounder of biotech companies Cleveland BioLabs (NASDAQ,CBLI; www.cbiolabs.com), Tartis and Incuron –that developanticancer drugs based on his discoveries.

Area of general research interest: Discovery of novel anticancer drugs and molecular targets,development of new principles of cancer treatment and tissueprotection

RESEARCH Gudkov’s laboratory is running a broad research programinvolving several distinct but highly integrated branches of study.This includes identification of new disease-associated genes anddeciphering molecular mechanisms of activity of their productsas potential targets for therapeutic modulation by smallmolecules or biologics. The lab’s major focus is on developingand applying new technologies for functional gene discovery,which will lead to designing new therapeutic approaches tocancer treatment and protection of healthy tissues from cancertreatment side effects and other stresses.

Novel Gene Discovery Approaches Gudkov’s lab pioneered functional gene discovery field bydeveloping in mid 90s, in collaboration with Igor Ronionson, oneof the first gene discovery methods enabling identification ofgenes’ function based on gene repression, named GeneticSuppressor Element (GSE) methodology. GSE technique hasresulted in discovery and/or functional characterization of anumber of cancer-associated genes, including ING1, BTG2 andothers (Ossovskaya et al., 1996; Garkavstsev et al., 1996, 1998;Boiko et al., 2004; Komarov et al., 2008).

Selection-Subtraction Approach (SSA) is another functionalgenetic methodology developed in Gudkov’s lab, which allowedeffective isolation of growth suppressive and killing genes andgenetic elements based on negative selection (Singhi et al.,2006).

Main principles of GSE and SSA methodologies were combinedto create the most powerful version of gene discovery approachnamed DECIPHER technique. DECIPHER, developed incollaboration with Cellecta, Inc (Mountain View, California;www.cellecta.com/products-services/pooled-shRNA-libraries/),involves generation and screening of diverse bar-coded shRNAlibraries covering the whole transcriptome of humans and miceand enabling finding genes that control major cellular functions.

Another functional genomics approach recently developed byGudkov’s team in collaboration with George Stark’s lab

(Cleveland Clinic) is named Validation-Based InsertionalMutagenesis (VBIM). VBIM is a novel highly efficient version ofretrovirus-based promoter-insertion approach that has alreadyresulted in discovery of several new regulators of NF-kappaBsignaling pathway (Lu et al., 2009, 2010).

Currently Gudkov’s lab is working on a new functional genomics-based screening methodology, which, when developed, will allowhigh throughput identification of biologically active secretedpeptides (BASP) as a new class of prospective pharmaceuticals.

Role of p53 in Cancer and Tissue Damage p53 studies conducted by Gudkov’s team are focused on themechanism and role of this tumor suppressor in how normaltissues respond to genotoxic stresses associated with cancertreatment and other types of acute stresses (Garkavstev et al.,1998; Gudkov and Komarova, 2003, 2005). Observations madein his lab in late 90s demonstrated tissue specificity of p53-mediated apoptosis and its major role in determining theradiation sensitivity of mammals (Komarova et al., 1997). Thisresulted in development of a paradigm-shifting strategy ofpharmacological inhibition of p53 for tissue protection. It wasproven by isolating a small molecule p53 inhibitor Pifithrin-a thatrescues mice from lethal doses of gamma irradiation (Komarov etal., 1999). This approach, covered by a series of patents, formedthe foundation for development of p53-inhibitory drugs that arecurrently in human trials (http://www.quarkpharma.com/qbi-en/products/QPI-1002/).

Targeting various branches of p53 signaling pathway by smallmolecules and biologics remains one of the major aspects ofGudkov’s research. For example, a small molecule, Pifithin-m,was isolated that blocks p53-mediated apoptosis by preventingits binding to mitochondria which acts as radioprotectant havingno effect on the majority of p53 functions as transcription factor(Strom et al., 2006).

An important aspect of p53 biology discovered in Gudkov’s labis its interaction with NF-kappaB, a signal transduction pathway,which controls inflammation and immune responses. It wasfound that the two major stress response mechanisms areinvolved in mutual negative regulation which makes p53 ageneral suppressor of inflammation and NF-kappaB – anoncogene (Komarova et al., 2005; Gurova et al., 2005). Thisfinding opened a new opportunity for developing drugs capableof simultaneously targeting p53 and NF-kB; an old anti-malariadrug quinacrine was shown to belong to this category and,therefore, to act as an anticancer agent (Gurova et al., 2005).

Tissue Protecting Drugs for Cancer Treatment andBiodefense Applications Healthy tissue protecting strategy invented and developed inGudkov’s lab involves the use of pharmacological agentsinhibiting p53 and activating NF-kappaB, thereby mimickingmechanisms acquired by tumors to escape apoptosis (Gudkovand Komarova, 2010). NF-kappaB activating approach resultedin development of a new class of drugs, named Protectans, thatare derivatives of natural NF-kappaB activators produced bynatural human microflora. For example, NF-kappaB-activatingagent Protectan CBLB502 is a pharmacologically optimized

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derivative of bacterial protein flagellin, natural agonist of Toll-likereceptor pathway, which mediates mobilization of endogenousmechanisms of resistance of mammalian organisms to a varietyof stresses, including lethal doses of ionizing radiation (Burdelyaet al., 2008). CBLB502 is currently at advanced stages of clinicaldevelopment as radioprotective antidote suitable for biodefenseand radiotherapy applications.

Other Directions of Anticancer Drug Discovery: Targeting MYC,MRP1, etc.

Gudkov’s team is focused on targeting a number of other cancertreatment targets, including MYC, an oncoprotein that isessential for growth of any type of cancer, MRP1, a multidrugtransporter responsible for resistance to chemotherapy of largeproportion of tumors (Burkhart et al., 2008), several targets forantiviral therapies (Thakur et al., 2007; Mao et al., 2008) andothers. In all of these projects, drug development is based onhigh throughput screening of small molecules using cell-basedreadout systems accompanied by comprehensive validation ofspecificity and followed by hit-to-lead optimization. Gudkov’s labis especially interested in exploring opportunities for safe cancertreatment approaches which could lead to the development ofnew drugs for treatment childhood cancers.

Select Publications 1. Ossovskaya, V.S., Mazo, I.A., Chernov, M.V., Chernova, O.B., Strezoska, Z.,Kondratov, R., Stark, G.R., Chumakov, P.M. and Gudkov, A.V. (1996) Dissectionof p53 functions by genetic suppressor elements: distinct biological effects ofseparate p53 domains. Proc Natl. Acad Sci. USA., 93, 10309-10314.

2. Garkavtsev, I.A., Kazarov, A.R., Gudkov, A.V. and Riabowol, K. (1996)Suppression of the novel growth inhibitor p33ING1 promotes neoplastictransformation. Nature Genetics 14, 415-420.

3. Komarova, E.A., Franks, R., Chernov, M.V., Armin, G., Chin, D.M., Zelnick, C.R.,Bacus, S.S. and Gudkov, A.V. (1997) Transgenic mice with p53-responsive lacZ:new insights into p53 function in normal development and in response to DNAdamage in vivo. EMBO J., 16, 1391-1400.

4. Garkavtsev, I.A., Grigorian, I.A., Chernov, M.V., Ossovskaya, V.S., Chumakov,P.M. and Gudkov, A.V. (1998) A candidate tumor suppressor p33ING1cooperates with p53 in cell growth control. Nature 391, 295-298.

5. Komarov, P.G. Komarova, E.A., Kondratov, R.V. Christov-Tselkov, K., Coon, J.C.,Chernov, M.V., and Gudkov, A.V. (1999) A Chemical Inhibitor of p53 ThatProtects Mice from the Side Effects of Cancer Therapy. Science 285, 1733-1737.

6. Gudkov, A.V. and Komarova, E.A. (2003) Role of p53 in determining sensitivity toradiotherapy. Nature Rev. Cancer 3, 117-129.

7. Singhi, A., Kondratov, R., Neznanov, N. and Gudkov, A.V. (2004) Selection-subtraction approach (SSA): a universal genetic screening technique thatenables negative selection. PNAS 101, 9327-9332.

8. Komarova, E.A., Krivokrysenko, V.I., Wang, K., Neznanov, N., Chernov, M.V.,Komarov, P.G., Brennan, M-L., Golovkina, T.V., Rokhlin, O.W., Kuprash, D.V.,Nedospasov, S.A., Hazen, S., Feinstein, E. and Gudkov, A.V. (2005) p53 is asuppressor of inflammatory response in mice. FASEB J 19, 1030-1032.

9. Gudkov, A.V. and Komarova E.A. (2005) Prospective therapeutic applications ofp53 inhibitors. Biochem. Biophys. Res. Commun. 331, 726-736.

10. Gurova K.V., Hill, J.E., Prokvolit, A., Burdelya, L.G., Samoylova, E.,Khodyakova, A.V., Ganapathi, R., Ganapathi, M., Tararova, N.D., Lvovsky, D.,Webb, T.R. and Gudkov, A.V. (2005) Small molecules, reactivating p53 in renalcell carcinoma, reveal a new NF-κB-dependent mechanism of p53 suppressionin tumors. PNAS 102, 17448-17453.

11. Boiko, A.D., Porteous, S., Razorenova, O.V., Krivokrysenko, V.I., Williams, B.R.and Gudkov, A.V. (2006) A systematic search for downstream mediators oftumor suppressor function of p53 reveals a major role of BTG2 in suppressionof Ras-induced transformation. Genes Dev, 20, 236-52.

12. Strom, E., Sathe, S., Komarov, P.G., Chernova, O.B., Pavlovska, I.,Shyshynova, I., Bosykh, D.A., Burdelya, L.G., Macklis, R.M., Skaliter, R.,Komarova, E.A. and Gudkov, A.V. (2006) Inhibition of p53 binding tomitochondria by small molecule is sufficient for radioprotection in vivo. NatureChem. Biol., 2, 474-9.

13. Thakur CS, Jha BK, Dong B, Das Gupta J, Silverman KM, Mao H, Sawai H,Nakamura AO, Banerjee AK, Gudkov A, Silverman RH. (2007). Small-moleculeactivators of RNase L with broad-spectrum antiviral activity. PNAS 104: 9585-90.

14. Mao H, Thakur CS, Chattopadhyay S, Silverman RH, Gudkov AV, Banerjee AK.(2008). Inhibition of human parainfluenza virus type 3 infection by novel smallmolecules. Antiviral Res. 77: 83-94.

15. Burdelya LG, Krivokrysenko VI, Tallant TC, Strom E, Gleiberman AS, Gupta D,Kurnasov OV, Fort FL, Osterman AL, Didonato JA, Feinstein E, Gudkov AV.(2008). An agonist of toll-like receptor 5 has radioprotective activity in mouseand primate models. Science 320: 226-30.

16. Logunov DY, Scheblyakov DV, Zubkova OV, Shmarov MM, Rakovskaya IV,Gurova KV, Tararova ND, Burdelya LG, Naroditsky BS, Ginzburg AL, GudkovAV. (2008). Mycoplasma infection suppresses p53, activates NF-kappaB andcooperates with oncogenic Ras in rodent fibroblast transformation. Oncogene27: 4521-31.

17. Kravchenko JE, Ilyinskaya GV, Komarov PG, Agapova LS, Kochetkov DV,Strom E, Frolova EI, Kovriga I, Gudkov AV, Feinstein E, Chumakov PM. (2008).Small-molecule RETRA suppresses mutant p53-bearing cancer cells through ap73-dependent salvage pathway. PNAS 105(17): 6302-7.

18. Komarov AP, Rokhlin OW, Yu C-A, Gudkov AV. (2008). Functional geneticscreening reveals the role of mitochondrial cytochrome b as amediator of FAS-induced apoptosis. PNAS 105(38):14453-8.

19. Burkhart CA, Watt F, Murray J, Pajic M, Prokvolit A, Xue C, Flemming C, SmithJ, Purmal A, Isachenko N, Komarov P, Gurova KV, Sartorelli AC, Marshall GM,Norris MD, Gudkov AV and Haber M. (2009). Reversan, a Novel MRP1 InhibitorThat Increases the Therapeutic Index of Chemotherapy in Mice. Cancer Res69:6573-80.

20. Lu T, Jackson MW, Singhi AD, Kandel E, Yang M, Zhang Y, Gudkov AV, andStark GR. (2009). Validation-based insertional mutagenesis identifies lysinedemethylase FBXL11 as a negative regulator of NFκB. PNAS 106:16339-44.

21. Gudkov AV and Komarova EA (2010) Pathologies associated with the p53response. Cold Spring Harbor Perspectives in Biology, Arnold Levine, DavidLane, eds., published online April 7, 2010.

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Anti-cancer drug discoverythrough modulation oftranscriptional factorsactivity in tumor cellsKaterina V. Gurova, MD, PhDAssistant Professor of Oncology

Major goal of our lab is the discovery of new anti-cancer agentsthrough different approaches, their testing and early stagedevelopment as well as understanding of the mechanisms oftheir activity. The preferred way of discovery of new compoundsis through identification and genetic validation of pathways,critical for survival of different types of tumor cells, generation ofcell based readout monitoring the state of a critical pathway intumor cells and screening of small molecule libraries forcompounds capable of modulation of a critical signaling pathwayin a desired direction. Right now we have two parallel studies inthe lab which are based on this approach. Both projects alloweddiscovery of active in vivo anti-tumor compounds with lowtoxicity profile and good perspectives of drug development.Besides this they also helped us to uncover new mechanisms ofregulation of two very important pathways in tumor cells.

One project was initially focused on p53 activation by non-genotoxic stress in tumor cells with wild type but inactive p53(1). We have isolated several compounds which activate p53, butsimultaneously inhibit NF-kB in tumor cells (2). NF-kB was alsofound to be responsible for p53 inhibition in many different tumortypes (2). These compounds bind DNA and RNA and causeprofound effect on some types of transcription and translation,not being general inhibitors of transcription or translation. Theyalso do not cause any mutagenic effect or structuralmodifications of DNA. Most probably they interfere with theactivity of transcription elongation complexes responsible fornucleosome assembly in cells. As we believe this leads in thefirst turn to the inhibition of stress-related transcription andtherefore hit predominantly tumor cells, which are dependent onstress signaling permanently in contrast to normal cells. Weobserve that NF-kB and unfolded protein response relatedtranscription in inhibited in tumor cells treated with thesecompounds. We believe that this class of compounds maybecame a new effective and safe type of tumor therapy. Severallines of research in the lab is devoted to the testing of thesecompounds in the most difficult to cure cancer types, likepancreatic cancer, understanding of the compounds effect onnucleosome chaperones (FACT, HMG domain proteins) andtranscription of different genes, mechanism of p53 activation andNF-kB and heat shock factor 1 repression as well as effect onother signaling pathways.

Another project was aimed on the inhibition of androgen receptor(AR) activity in androgen insensitive advanced prostate cancer(PC). We have shown that AR controls death and proliferation ofPC cells on different stages, including androgen-refractorydisease (3). Therefore targeting of AR may be approach for thetreatment of even androgen –insensitive disease. AR on thisstage does not respond anymore to androgen stimulation, but isstill active. It is usually mutated in ligand binding domain to

accommodate stimulation by other ligands in the absence ofandrogens, which were depleted in the course of initial anti-androgen therapy. We generated a readout system which allowsselection of compounds acting downstream of AR – ligandinteractions. Several groups of compounds were isolated andtested. Some of them are very specific and potent AR inhibitorsin androgen sensitive and insensitive PC in vitro and in vivo.Compounds we isolated in this screening have differentmechanism of activity, one acting only against AR-dependenttransactivation and others disturbing stability of AR mRNA.Experiment with these two groups of compounds allowedproposing some transcription independent role of AR in thecontrol of PC cells survival, as well as new mechanisms of ARregulation. More detailed investigation of these questions, as wellas development of anti-cancer agents based on thesecandidates, are the major focuses of the research in thisdirection.

Select Publications 1. Gurova, K.V. and Gudkov, A.V. (2003). Paradoxical role of apoptosis in tumorprogression. (Review) J. Cell. Biochem. 88, 128-137.

2. Gurova KV, Hill JE, Guo C, Prokvolit A, Burdelya LG, Samoylova E, KhodyakovaAV, Ganapathi R, Ganapathi M, Tararova ND, Bosykh D, Lvovskiy D, Webb TR,Stark GR, Gudkov AV. (2005) Small molecules that reactivate p53 in renal cellcarcinoma reveal a NF-kappaB-dependent mechanism of p53 suppression intumors. Proc Natl Acad Sci U S A, 102:17448-53.

3. Guo C, Gasparian AV, Zhuang G, Bosykh DA, Komar AA, Gudkov AV, Gurova KV.9-Aminoacridine-based anticancer drugs target the PI3K/AKT/mTOR, NF-kappaB and p53 pathways. Oncogene. 2009 Feb 26;28(8):1151-61. Epub 2009Jan 12.

4. Tararova ND, Narizhneva N, Krivokrisenko V, Gudkov AV, Gurova KV. Prostatecancer cells tolerate a narrow range of androgen receptor expression andactivity. Prostate. 2007 Dec 1;67(16):1801-15.

5. Jung KJ, Dasgupta A, Huang K, Jeong SJ, Pise-Masison C, Gurova KV, BradyJN. Small molecule inhibitor which reactivates p53 in HTLV-1 transformed cells JVirol. 2008 Sep;82(17):8537-47.

6. Narizhneva N, Tararova ND, Ryabokon P, Shyshynova I, Prokvolit A, KomarovPG, Purmal AP, Andrei V. Gudkov, Katerina V. Gurova. Small molecule screeningreveals a transcription-independent pro-survival function of androgen receptor incastration-resistant prostate cancer Cell Cycle, 8, 24, 2009

7. Gurova KV. New hope from an old drug: revisiting DNA-binding small moleculesas anti-cancer agents. Future Oncology. 2009

15

Genetic dissection of signal transduction in mammalian cells

Eugene S. Kandel, PhDAssistant Professor of OncologyDirector of Graduate Studies

Mammalian cells posses an intricate network of signaltransduction pathways, which module cell behavior in responseto a variety of external and internal stimuli. These cellularmechanisms normally ensure the survival of cells in specificorganismal niches, as well as in conditions of temporaryenvironmental stress. Excessive activation of these cellularmechanisms during oncogenesis contributes to enhancedsurvival and growth of cancer cells, modulates tumor responsesto therapy and offsets the otherwise deleterious consequencesof other oncogenic mutations. A better understanding of thesesignaling networks in health and disease are needed to identifythe changes that could serve to diagnose the disease, as well asto predict its clinical properties, such as the likelihood ofmetastasis and the responsiveness to various therapeuticregimens. It is also needed to identify the molecules and specificmolecular interactions, which could be targeted with selectivetoxicity against cancerous, but not normal cells. Our research inthis area relies on a broad spectrum of traditional techniques ofcell and molecular biology, as well as on the development ofnovel tools and approaches for gene discovery andcharacterization.

The specific signaling pathways that we have investigatedinclude those of protein kinase B (a.k.a. Akt), NFkB, and heatshock.

Akt is frequently hyperactivated in cancers. The work of othersand us has shown that this abnormality contributes to enhancedgrowth, reduced cell death, restructured micro-environment, andhigher likelihood of additional mutations in cancer cells (Kennedyet al. 1999; Gottlob et al. 2001; Kandel et al. 2002; Somanath etal. 2007). However, non-transformed cells often respond tohyperactivation of Akt by undergoing growth arrest ordifferentiation, rather than oncogenic transformation. This meansthat cancer cells harbor additional alterations, which allow themto utilize Akt as an oncogene without arresting or differentiating,and reversal of such alterations is likely to have an anti-cancereffect. We have already identified some factors that are neededfor oncogenic function of Akt (Somanath et al. 2009) and arecontinuing the work in this direction.

Transcription factors from NFkB family control expression ofvarious genes that contribute to cell survival and immuneresponse. Abnormal function of these proteins is an importantfactor in cancer, inflammatory and autoimmune disorders. Weconducted several genetic screens for the genes whoseproducts can control the function of NFkB, identifying bothpositive and negative regulators (Kandel et al. 2005; Dasgupta etal. 2008; Lu et al. 2009). We are interested in identification ofadditional genes with such properties, as well as in betterunderstanding the mechanism of action of the ones that have

been discovered. Our functional genetic screens utilize a newgene discovery methodology, which is based on improvedinsertional mutagenesis(Kandel and Stark 2003; Kandel et al.2005). We are working on enhancing the technical aspects of thisapproach, including increasing the throughput of identificationand validation of relevant genes and extending this approach toin vivo gene discovery applications.

We were part of the team that had identified a novel RNAmolecule, which modulates the response of mammalian cell tothermal stress (Shamovsky et al. 2006). This was one of the firstexamples of a non-coding RNA that is a structural element ofmammalian signal transduction. Our interest in the role of non-coding RNAs in signal transduction continues in the ongoingwork on microRNAs, which have emerged as the major class ofregulators of various cellular processes (reviewed in (Gartel andKandel 2006; Gartel and Kandel 2008)).

We heavily rely on retroviruses as tools of gene delivery andgene discovery. This fuels our interest the in properties of theseviruses and led to the finding of a new mechanism of emergenceof recombinant retroviruses (Kandel and Nudler 2002) and theinteraction between the viral vectors and a newly discoveredhuman virus (Dong et al. 2008).

Select Publications1. Kennedy SG, Kandel ES, Cross TK, Hay N (1999). "Akt/Protein kinase B inhibitscell death by preventing the release of cytochrome c from mitochondria." MolCell Biol 19(8): 5800-10.

2. Gottlob K, Majewski N, Kennedy S, Kandel E, Robey RB, Hay N (2001)."Inhibition of early apoptotic events by Akt/PKB is dependent on the firstcommitted step of glycolysis and mitochondrial hexokinase." Genes Dev 15(11):1406-18.

3. Kandel ES, Nudler E (2002). "Template switching by RNA polymerase II in vivo.Evidence and implications from a retroviral system." Mol Cell 10(6): 1495-502.

4. Kandel ES, Skeen J, Majewski N, Di Cristofano A, Pandolfi PP, Feliciano CS,Gartel A, Hay N (2002). "Activation of Akt/protein kinase B overcomes a G(2)/mcell cycle checkpoint induced by DNA damage." Mol Cell Biol 22(22): 7831-41.

5. Kandel ES, Stark GR (2003). Forward genetics in mammalian cells. SignalTransducers and Activators of Transcription (STATs): Activation and Biology. P. B.Sehgal, D. E. Levy and T. Hirano. The Netherlands, Kluwer Academic Publishers.

6. Kandel ES, Lu T, Wan Y, Agarwal MK, Jackson MW, Stark GR (2005)."Mutagenesis by reversible promoter insertion to study the activation of NF-kappaB." Proc Natl Acad Sci U S A 102(18): 6425-30.

7. Shamovsky I, Ivannikov M, Kandel ES, Gershon D, Nudler E (2006). "RNA-mediated response to heat shock in mammalian cells." Nature 440(7083): 556-60.

8. Gartel AL, Kandel ES (2006). "RNA interference in cancer." Biomol Eng 23(1): 17-34.

9. Somanath PR, Kandel ES, Hay N, Byzova TV (2007). "Akt1 signaling regulatesintegrin activation, matrix recognition, and fibronectin assembly." J Biol Chem282(31): 22964-76.

10. Dasgupta M, Agarwal MK, Varley P, Lu T, Stark GR, Kandel ES (2008)."Transposon-based mutagenesis identifies short RIP1 as an activator of NF-kB." Cell Cycle 7(14 ): 2249-56.

11. Dong B, Silverman RH, Kandel ES (2008). "A natural human retrovirusefficiently complements vectors based on murine leukemia virus." PLoS ONE3(9): e3144.

12. Gartel AL, Kandel ES (2008). "miRNAs: Little known mediators ofoncogenesis." Semin Cancer Biol 18(2): 103-10.

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13. Lu T, Jackson MW, Singhi AD, Kandel ES, Yang M, Zhang Y, Gudkov AV, StarkGR (2009). "Validation-based insertional mutagenesis identifies lysinedemethylase FBXL11 as a negative regulator of NFkB." Proceedings of theNational Academy of Sciences 106(38): 16339-44.

14. Somanath PR, Vijai J, Kichina JV, Byzova T, Kandel ES (2009). "The role ofPAK-1 in activation of MAP kinase cascade and oncogenic transformation byAkt." Oncogene 28(25): 2365-9.

Biomolecular Resources:Proteomics, MassSpectrometry, DNASequencing and NMRSpectroscopy; ResearchInterests: Proteomics,

Metabolomics and Imaging MassSpectrometryA. Latif Kazim, PhDDirector, Biomolecular Resource FacilityAssociate Professor of Oncology

EDUCATION

Boston University, Boston, MA, BA, 1971, Chemistry/Biology

University of Minnesota/Mayo Medical School, Rochester, MN,PhD, 1979, Biochemistry/Immunology

Mayo Medical School, Rochester, MN, Postdoctoral Fellowship,1979-1981, Immunology

Biomolecular Resource Facility and General Research Interests

The Biomolecular Resource Facility is a core laboratory thatprovides advice and technical assistance to institute staff in DNAsequencing, genotyping, mass spectrometry, proteomics, andnuclear magnetic resonance spectroscopy. The main objectivesof the facility are to assess the technical needs of the staff in thecontext of these analytical methods, and develop resources toassist investigators in accomplishing their research goals.

Research and developmental interests in this laboratory currentlyfocus on structure/activity relationships in proteins and peptides,and the use of mass spectrometry-based methods in studyingthe effects of oxidative stress on DNA and proteins,metabolomics and tissue imaging by mass spectrometry.

Heat Shock Proteins are known to play essential roles in normalcellular functions and are involved in numerous pathwaysinvolving protein processing and interactions. In collaborationwith Dr. John Subjeck, we employed a grp170-secreting tumor

cell system to determine whether tumor cells engineered tosecrete grp170 generate an anti-tumor specific immuneresponse. Further, we examined the possibility that secretedgrp170 can bind to and co-transport out of tumor cells full-length tumor antigens that may play a role in the anti-tumorimmune response. Immunization of animals with grp170-secreting tumor cells results in rejection of the tumor byinduction of antigen-specific, CD8+dependent immuneresponses. The secreted grp170 was able to deliver full-lengthtumor antigens to the tumor microenvironment, thus makingthem available for uptake by antigen presenting cells (APCs) toinitiate tumor-specific immune responses. These data paralleledearlier studies showing that hsp110 or grp170 are able tochaperone full-length proteins, and when complexed with proteinantigens and used as vaccines, these complexes elicit immuneresponses in vivo against the protein antigens. This cell-basedapproach has the potential to be utilized as a tumor-specificvaccine in tumors of various histological origins.

A recent research interest of the laboratory is in the area ofimaging mass spectrometry (IMS) – the use of massspectrometric methods to “image” analytes such as drugs andmetabolites in tissue sections. Tissue imaging is a promisingtechnique as it allows the determination of the distributionpatterns derived from drug treatment (precursor drug andmetabolites) in specific tissues, and at a lateral resolutionranging from 100 μ (MALDI) to ~3 μ or less (TOF-SIMS). Incollaboration with Dr. Mari Prieto at Thermo-Fisher Scientific,using a MALDI source and an ion-trap mass spectrometer forIMS we recently identified and confirmed most of the knownmetabolites of irinotecan in liver and tumor tissues obtained frommice treated with this drug. Additional experiments are underwayto determine whether the distribution of the drug andmetabolites are related to the vascularity of the tumor. In relatedstudies and in collaboration with Dr. Youcef Rustum of RPCI andDr. Joseph Gardella at the University at Buffalo we have usedtime-of-flight secondary ion mass spectrometry (TOF-SIMS) todetermine the spatial distribution of metabolites ofmethylselenocysteine (MSC) in liver and in xenografts of humantumor tissues, demonstrating that the metabolites were in directproximity to the vasculature. MSC and other seleniumcompounds have been utilized by the Rustum laboratory incancer chemotherapy regimens to enhance the efficacy ofchemotherapeutic drugs. The mechanism by which thisenhanced efficacy occurs is unknown. The results from IMSshow that MSC metabolites are localized to the vasculature oftumor and liver tissue, which may result in a more effectivedelivery of chemotherapeutic agents into the tumor.

Select Publications1. Arnouk H, Zynda ER, Wang X-Y, Hylander, BL, Manjili MH, Repasky EA, SubjeckJR, Kazim AL. Tumour secreted grp170 chaperones full-length proteinsubstrates and induces an adaptive anti-tumour immune response in vivo.International Journal of Hyperthermia. 2010 26(4): 366-375.

2. Prieto Conaway MC, Cao S, Durrani F, Rustum YM, Wang P, Marlar K, KazimAL. The role of MALDI-enabled linear ion trap mass spectrometry as a sensitivetool in tissue imaging. Spectroscopy Europe 2008; 20:15-17.

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3. Burns SA, Ventura RS, Marlar K, Montecinos VP, Cao S, Durrani FA, Rustum YM,Kazim AL, Gardella JA, Jr. ToF-SIMS Imaging of Selenium Metabolites ofChemotherapeutic Agents within the Vascular Network of Tumor and LiverTissues. Submitted, 2010.

4. Protein complexes of Focal Adhesion Kinase as the targets in tumor growth,angiogenesis and metastasis.

Protein complexes of Focal Adhesion Kinase as the targets in tumorgrowth, angiogenesis and metastasis.

Elena V. Kurenova, PhDAssociate Professor of Oncology

My research is focused on Focal Adhesion Kinase (FAK) -tyrosine kinase that functions as a key orchestrator of signalsleading to survival of cancer cells, invasion and metastasis. FAKinteracts with a number of critical proteins involved in survivalsignaling in cancer cells, some of which were discovered byphage display approach. We hypothesized that targeting aprotein-protein interface with drug-like small molecules is afeasible strategy for inhibiting tumor growth. We aim to elucidatethe mechanisms of these FAK-related survival signals anddevelop novel therapeutic to inhibit this signaling in humantumors. We selected small molecules that disrupt some protein-protein interaction of FAK and cause cancer cell death in vitroand in vivo.

We have shown that serine/threonine kinase RIP - a majorcomponent of the death receptor complex, binds to FAK. Wehypothesize that this interaction is a critical component forsuppressing apoptosis in tumor cells. We are focused onunderstanding the mechanism of FAK-RIP interaction andsignaling which may provide a molecular basis for thedevelopment of new cancer therapeutics.

FAK and VEGFR-3 are tyrosine kinases that have been identifiedas critical signaling molecules not only for survival of cancercells, but also important in vascular development. Previously wehave shown that VEGFR-3 and FAK physically interact and areoverexpressed in cancer cells to provide a significant survivaladvantage for the tumor cells. We subsequently identified anovel small molecule inhibitor that targeted VEGFR-3-FAK site ofinteraction and disrupted the survival function of these twoproteins. We utilized the crystal structure of the FAK focaladhesion targeting (FAT) domain for molecular docking of smallmolecules that targeted the VEGFR-3 binding site on FAK. Weidentified a small molecule C4 that disrupted VEGFR-3-FAKbinding. In vitro testing of this compound resulted in theselective growth inhibition, reduction in motility and invasion, andinduction of apoptosis in a time- and dose-dependent manner inmany cancer cell lines, especially those that overexpressed

VEGFR-3. In vivo, C4 showed a marked reduction of tumorgrowth and was synergistic with doxorubicin chemotherapy inbreast cancer xenograft models, with dacarbazine in melanomaxenograft models, and with gemcitabine in pancreatic cancerxenograft models. These results demonstrate that targeting theFAK-VEGFR-3 interaction with small molecules inhibited thesurvival function of these two tyrosine kinases, representing aunique approach for molecular-targeted highly-specific cancertherapeutics. Now we aim to elucidate the effect of this inhibitionon blood and lymphatic vasculature of the tumor and tumormetastasis.

Select Publications1. Kurenova E., Xu L-H., Yang X., Baldwin A.S. Jr., Craven R.J., Hanks S.K., Liu Z-G., and Cance W.G. 2004 The Focal Adhesion Kinase Suppresses Apoptosis byBinding to the Death Domain of Receptor Interacting Protein. Mol Cell Biol.24:4361-4371.

2. C. A. Garces, E. V. Kurenova, V. M. Golubovskaya, and W. G. Cance. 2006Vascular Endothelial Growth Factor Recptor-3 (VEGFR-3) and Focal AdhesionKinase (FAK) Bind and Suppress Apoptosis in Breast Cancer Cells. Cancer Res.66:1446-54.

3. E. Kurenova, V. Golubovskaya, W.G. Cance. 2006 The Tumor Biology of FocalAdhesion Kinase. (Review) Cellscience Reviews, 3 (1):1742-8130

4. Liu W, Bloom DA, Cance WG, Kurenova EV, Golubovskaya VM, Hochwald SN.2008 FAK and IGF-IR interact to provide survival signals in human pancreaticadenocarcinoma cells. Carcinogenesis. 29(6):1096-107

5. Golubovskaya V, Finch R, Zheng M, Kurenova EV, Cance WG. 2008 The 7amino-acid site in the proline-rich region of the N-terminal domain of p53 isinvolved in interaction with FAK and is critical for p53 functioning. Biochem J.,411(1):151-60

6. Magis AT, Bailey KM, Kurenova EV, Hernández Prada JA, Cance WG, Ostrov DA.2008 Crystallization of the focal adhesion kinase targeting (FAT) domain in aprimitive orthorhombic space group. Acta Crystallogr Sect F Struct Biol CrystCommun. 64(Pt 6):564-6. Epub 2008 May 30.

7. Kweh F, Zheng M, Kurenova E, Wallace M, Golubovskaya V, Cance WG. 2009Neurofibromin physically interacts with the N-terminal domain of focal adhesionkinase. Mol Carcinog; 48(11):1005-1017.

8. Kurenova EV, Hunt DL, He D, Fu AD, Massoll NA, Golubovskaya VM, GarcesCA, Cance WG. 2009 Vascular endothelial growth factor receptor-3 promotesbreast cancer cell proliferation, motility and survival in vitro and tumor formationin vivo. Cell Cycle; 8(14):2266-2280.

9. Kurenova EV, Hunt DL, He D, Magis AT, Ostrov DA, Cance WG. 2009 Smallmolecule chloropyramine hydrochloride (C4) targets the binding site of focaladhesion kinase and vascular endothelial growth factor receptor 3 andsuppresses breast cancer growth in vivo. J Med Chem; 52(15):4716-4724.

10. Zheng D, Kurenova E, Ucar D, Golubovskaya V, Magis A, Ostrov D, Cance WG,Hochwald SN. Targeting of the protein interaction site between FAK and IGF-1R. Biochem Biophys Res Commun 2009; 388(2):301-305.

11. Zheng D, Golubovskaya V, Kurenova E, Wood C, Massoll NA, Ostrov D, CanceWG, Hochwald SN. A novel strategy to inhibit FAK and IGF-1R decreasesgrowth of pancreatic cancer xenografts. Mol Carcinog 2010; 49(2):200-209.

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Epigenetics, Transcriptionfactors and ChildhoodCancer

Asoke K. Mal, PhDAssistant Professor of Oncology

Malignant tumors of childhood represent an uncommonheterogeneous group of neoplasms originating from virtually any anatomical structure. Despite major improvement throughthe integrated, multimodality treatment approach includingcombination chemotherapy, childhood cancer remains theleading cause of disease-related death in children. Moreover,recently approved chemotherapeutic agents that have significantantitumor activity in adult cancers appear to be inactive inconventional therapy performed in children with aggressive and recurrent cancers. The prevailing explanation for thedevelopment of cancer over the past 50 years has raised thenotion that apparently cancers represent the failure of the tumorcells to differentiate along a pathway that would be expected fortheir tissue of origin. Hence, investigation of the mechanismabout the underlying molecular defect in cancer cells todifferentiate would be important to develop new biologicallybased treatment approach in making bad cells go good.Aberrant gene function and alteration in epigenetic regulation of gene expression are now increasingly recognized to causedifferentiation dysregulation in cancer development andprogression.

Our research program is focused on basic and translationalresearch that connects the normal skeletal muscle differentiationprogram and its dysregulation in the development of childhoodrhabdomyosarcoma (RMS). Our laboratory pursues two majorresearch programs: Epigenetic gene regulation and Drugdiscovery.

The epigenetic gene regulation program involves deciphering theepigenetic mechanism as potential target strategy for therapy.This program is aimed to identify the epigenetic modifiersregulating normal and perturb skeletal muscle differentiation.Particularly, the research focuses on epigenetic modulation ofgene expression regulating skeletal muscle transcription inmodel system and in rhabdomyosarcoma (RMS).

The drug discovery program involves searching small moleculepharmaceuticals capable of activating muscle differentiationprogram in rhabdomyosarcoma for the treatment of childhoodrhabdomyosarcoma.

The ultimate goal of both research programs is aimed to developbiologically based approach for the invention of differentiatingagents as new anti-rhabdomyosarcoma pharmaceuticals.

ONGOING RESEARCH PROJECTS:

Epigenetic mechanism regulating skeletal muscle differentiation transcriptionOur laboratory has discovered that epigenetic modifier histoneH3-lysine 9 methytransferase Suv39h represses muscle geneexpression in executing muscle differentiation. We are studyingthe mechanism of Suv39h mediated epigenetic events inarresting muscle cell differentiation and the balance betweenhistone H3-lysine 9 methylation by Suv39h and demethylation ofthis epigenetic repressive mark in regulating myogenictranscription during muscle differentiation.

Epigenetic mechanism regulating rhabdomyosarcoma developmentRhabdomyosarcoma is a childhood malignant tumor comprising5-8% of all cases of cancer in children and thought to arise dueto arrest of muscle differentiation program despite theexpression of MyoD that acts as a key myogenic transcriptionalregulator of the muscle differentiation. We are investigating theepigenetic mechanism that is encountered in the failure ofMyoD-mediated differentiation in rhabdomyosarcoma. Inaddition, we are investigating the potential contribution ofoncogenic fusion protein Pax3-Fkhr, a genetic signature inaggressive alveolar rhabdomyosarcoma (ARMS) development,mediating epigenetic alteration in the defect of muscledifferentiation program in these tumor cells.

Identification of small molecule modulators by screeningchemical library

This research program involves the identification of smallmolecule modulators in biological readout systems targetingmyogenic regulator MyoD, epigenetic modifier Suv39h andoncogenic fusion protein Pax3-Fkhr capable of inducing muscledifferentiation program in rhabdomyosarcoma.

Select Publications1. Mal A: Histone Methyltransferase Suv39h1 Represses MyoD-stimulatedMyogenic Differentiation. EMBO J., 25: 3323-3334, 2006.

1. Mal A, Harter ML: MyoD is functionally linked to silencing of a muscle-specificregulatory gene prior to skeletal myogenesis. Proc. Natl. Acad. Sci., 100: 1735-1739, 2003.

1. Mal A, Chattopadhyay D, Ghosh MK, Poon RY, Hunter T, Harter ML: p21cip1and retinoblastoma proteins control the absence of DNA replication in terminallydifferentiated muscle cells. J. Cell. Biol., 149: 281-292, 2000.

1. Mal A, Sturniolo M, Schiltz RL, Ghosh MK, Harter ML: A role of histonedeacetylase HDAC1 in modulating the transcriptional activity of MyoD: Inhibitionof myogenic program. EMBO J., 20: 1739-1753, 2001.

Applications of MedicalPhysics to cancerdiagnosis and treatment

Harish K. Malhotra, PhD, DABRAssistant Professor of Oncology

Surgery, chemotherapy and radiation medicine are threemodalities which are used, often together, in the management ofcancer. Therapeutic Medical Physics is an applied branch ofphysics which is associated with the application of physics toradiation medicine so that delivery of ionizing radiation to apatient is safely and accurately targeted to the tumor.Accordingly, the research in this field is very rewarding personallyas the results and the benefits to society are usually immediate.Besides, providing clinical physics care to all patients undergoingtreatment at the institute, half a dozen medical physicists in thedepartment of Radiation Medicine of Roswell Park CancerInstitute, also pool together their knowledge and expertise inadvancing scientific research in this field often even collaboratingwith professionals from other fields like physics, computerscience, etc. not to mention our clinical colleagues. At any givenmoment of time, there are various research projects in variousstages of completion in the department.

Brachytherapy plays an important role in the treatment of femalegynecological cancers. A good part of the century long clinicalexperience has come from use of a tandem and ovoid applicatorset which has tungsten shields in it to minimize the radiationdose and hence radiation toxicity to the nearby criticalstructures, rectum and bladder. Unfortunately due to practicallimitations, the dose reduction due to their presence was ignoredand hence the clinical experience gained relies on incorrectrecorded dosage to these two vital organs. Thus the publishedtolerance values in literature for these structures can not be usedfor new set of CT/MR compatible applicators which can not haveany shielding due to imaging artifacts. We are presentlydeveloping experimental methods to determine the difference inthe recorded and actual radiation dosage to these organs usingvarious detectors. This study will provide guidance to properlyapply corrected constraints for rectum and bladder using CT/MRapplicators.

Project 2 is based on collaboration with researchers inCarestream Health, Rochester, NY. We have successfullydemonstrated a novel algorithm to use Electronic Portal ImagingDetector (EPID) to track the lung tumor in real time withoutrequiring any external surrogate. It is very promising field whichone day may help us to gate the treatments of lung patientsusing this technique which will be simple and yet free of lessaccurate external surrogates. This will also result in the reductionin tumor margin commonly employed for lung cases resulting inlower radiation toxicity to these patients.

In radiation therapy, it is very important to accurately reproducethe patient position during treatment as was used during

treatment simulation. This is presently carried out using thetechnical information recorded manually during simulation. Inanother project, we are seeing the feasibility of using digitalcameras along with information extracted from the patientstreatment plan to verify that the patients is not only in its rightposition before the treatment is started but also to continuouslytrack and monitor the patients’ position during his/her entiretreatment. Our initial results are very promising in this study.

In yet another project, we are trying to find optimal beamarrangements and their mathematical characteristics to achievean optimal treatment plan which best satisfies clinical constraintsusing a super computer at CCR, UB. We have alreadydemonstrated the proof of concept in which patient specificimages along with appropriate image segmentations and desiredclinical constraints are fed to a cluster of computers at CCRwhere hundreds of combinations are tried using a process calledGenetic Algorithm to come to an optimal solution. The processhas been successfully demonstrated for 3D conformal therapyand we wish to extend it for intensity modulated radiotherapy(IMRT) in near future as well.

Select Publications:1. Mukhraj Hira, Matthew B Podgorsak, Wainwright Jaggernauth, Harish KMalhotra. Measurement of dose perturbation around shielded ovoids in high-dose-rate brachytherapy. Brachytherapy [in press].

2. J. S. Schildkraut, N. Prosser, A. Savakis, J. Gomez, D. Nazareth, A. K. Singh, H.K. Malhotra. Level Set Segmentation of Pulmonary Nodules in MegavoltElectronic Portal Images Using a CT Prior. Med. Phys. 37(11), 5703-5710, 2010.

3. Harish K Malhotra, J S Avadhani, Steven F deBoer, W Jaggernauth, Michael R.Kuettel, and M.B. Podgorsak; Duplicating a tandem and ovoid distribution withIMRT: A feasibility study. Journal of Applied Clinical Medical Physics Journal ofClinical Medical Physics. Vol. 8, Issue 3, 91-98, 2007.

4. Tran T, Stanley T, Malhotra HK, deBoer SF, Prasad D, Podgorsak MB. Target andperipheral dose during patient repositioning with the Gamma Knife automaticpositioning system (APS) device. J. Appl. Clin. Med. Phys. 11(3): 88-98, 2010.

5. Daryl P. Nazareth, Stephen Brunner, Matthew D. Jones, Harish K. Malhotra,Mohammad Bakhtiari. Optimization of beam angles for intensity modulatedradiation therapy treatment planning using genetic algorithm on a distributedcomputing platform. Journal of Medical Physics, Vol. 34, No. 3, (ICMP 2008Special Issue) 129-132, 2009.

6. Malhotra H.K., Raina S., Avadhani J.S., deBoer S., and Podgorsak M.B.Technical and dosimetric considerations in IMRT treatment planning for largetarget volumes, Journal of Clinical Medical Physics. Vol. 6, Issue 4, 77-87, 2005.

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Mechanisms andenhancement ofPhotodynamic Therapy in cancer and cancer stem cells

Janet Morgan, PhD Clinical Assistant Professor of Oncology

My research interests lie primarily in the field of photobiology andphotodynamic therapy (PDT) a treatment for cancer and otherdiseases in which photosensitive drugs are activated by light todestroy malignant or other target tissue. Currently approvedphotosensitizing drugs for PDT are not optimally selective andhave drawbacks such low absorbance at wavelengths which donot readily penetrate tissue. In addition, heterogeneity of tumorsand of photosensitizer distribution, may affect the response. Tobe most effective, a selective tumor response is required withminimal side effects.

To achieve this goal, we use several in vitro and in vivo models,and examine the effects of photosensitizers, their distributionand phototoxic mechanisms by different approaches includingphotoreactivity, biochemistry, molecular biology and fluorescenceimaging.

In particular we are interested in targets and mechanisms of PDTtoxicity a) to understand the mechanisms by which the PDTresponse is effected from the whole animal response to thetissue, cellular, subcellular and molecular levels b) to aid indesigning and synthesizing more efficacious drugs and deliverysystems for photodynamic therapy and c) to help to optimizePDT protocols, and their effectiveness.

Our current focus is using small drug molecules which interferewith or inhibit mechanisms and pathways which control cancergrowth, survival and proliferation and which may enable us toenhance the PDT response by using combination therapies. Inparticular stem cell-like cancer cells PDT have properties whichmay help them to evade PDT toxicity including the expression ofpumps which efflux photosensitizers, and innate resistance tooxidative stresses. Also, targets at or within the mitochondriadefine critical sites of photosensitizer uptake, synthesis from pro-drugs, binding and damage. Some of the binding sites we areexamining include the peripheral (mitochondrial) benzodiazepinereceptor (PBR), which is expressed at high level in tumors andwhich may help to concentrate photosensitizers in mitochondria.The PBR is a potential sensitive site for PDT and the role of itsnatural ligands in enhancing photosensitizer uptake andenhancement of reactive oxidative species is under investigation.

Select publications

1. J. Morgan, J.E. Whitaker and A. R. Oseroff, (1998). GRP78 induction by calciumionophore potentiates photodynamic therapy using the mitochondrial targetingdye Victoria Blue-BO. Photochem. Photobiol. 61:155-164.

2. I.J. MacDonald, J. Morgan, D.A. Bellnier, G. Paszkiewicz, J.E. Whitaker, D. J.Litchfield and T.J. Dougherty. (1999) Subcellular localization patterns and theirrelationship to photodynamic activity of pyropheophorbide-a derivatives.Photochem. Photobiol. 70 789-797.

3. J. Morgan, W.R. Potter and A.R. Oseroff. (2000) Comparison of photodynamictargets in a carcinoma cell line and its mitochondrial DNA negative derivative.Photochem. Photobiol. 71(6) 747-757.

4. J. Morgan and A.R. Oseroff. (2001) Mitochondria-based anti-cancerphotodynamic therapy. Advanced Drug Delivery Reviews - Drug and DNAdelivery to Mitochondria. Elsevier Press. Eds.V. Weissig and Torchilin.Vol. 49(1-2), 71-86.

5. Janet Morgan, Allan R. Oseroff and Richard T. Cheney (2004). The peripheralbenzodiazepine receptor expression is decreased in skin cancers compared tonormal skin. Br. J Dermatol. 151:846-856.

6. Barbara W. Henderson, Sandra O. Gollnick, John W. Snyder, Theresa M. Busch,Philaretos Kousis, Richard T. Cheney, Janet Morgan (2004). Fluence RateDetermines Tumor Oxygenation and Inflammatory Responses to PhotodynamicTherapy. Cancer Res. 64: 2120-2126.

7. Ingegerd Eggen Furre, Susan Shahzidi, Zivile Luksiene, Michael T.N. Møller, ElinBorgen, Janet Morgan, Kinga Tcazc-Stachowska, Jahn M. Nesland, and QianPeng (2005). Targeting PBR by hexaminolevulinate-mediated photodynamictherapy induces apoptosis through translocation of apoptosis-inducing factor inhuman leukaemia cells. Cancer Res. 65: 11051-11060.

8. Andrew Rosenfeld, Janet Morgan, Lalit N.Goswami, Tymish Ohulchansky, XiangZheng, Paras N. Prasad, Allan Oseroff, and Ravindra K. Pandey (2006).Photosensitizers derived from 132-oxo-methyl pyropheophorbide-a: enhancedeffect of indium (III) as a central metal in vitro and in vivo photosensitizingefficacy. Photochem. Photobiol. 82: 626–634.

9. Weiguo Liu, Maria R. Baer, Mary Jo Bowman, Paula Pera, Xiang Zheng, JanetMorgan, Ravindra A. Pandey and Allan R. Oseroff (2007) The tyrosine kinaseinhibitor Imatinib Mesylate enhances the efficacy of photodynamic therapy byinhibiting ABCG2. Clin Cancer Res. 13(8): 2463-70.

10. Xiang Zheng, Janet Morgan, Suresh K. Pandey, Yihui Chen, Erin Tracy, HeinzBaumann, Joseph Missert, Carrie Batt, Jennifer Jackson, David A. Bellnier,Barbara W. Henderson and Ravindra K. Pandey (2009). Conjugation of HPPHto carbohydrates changes its subcellular distribution and enhancesphotodynamic activity in vivo. J. Med. Chem. 52, 4306–4318 PubMed CentralPMCID: PMC2913405

11. Janet Morgan and Cara M. Petrucci. (2009). The effect of ALA/PpIX-PDT onputative cancer stem cells in tumor side populations.. Photodynamic Therapy:Back to the Future, edited by David H. Kessel, Proc. of SPIE Vol. 7380,738011-1-9.

12. Janet Morgan, Jennifer D. Jackson, Xiang Zheng, Suresh K. Pandey andRavindra K. Pandey. (2010) Substrate affinity of photosensitizers derived fromchlorophyll-a: The ABCG2 transporter affects the phototoxic response of sidepopulation stem-cell like cancer cells to photodynamic therapy. MolecularPharmaceutics 5, 1789–1804 NIHMSID # 229924

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Applications of MedicalPhysics to cancerdiagnosis and treatment

Daryl P. Nazareth, PhDAssistant Professor of Oncology

Computational methods are now playing a larger role than everin all aspects of science, including the diagnosis and treatmentof disease. No other technology has seen such tremendousadvances in the last half century as computer technology. Theexponential advances in computer power make it criticallyimportant to understand its potential, and leverage itscapabilities in our fight against cancer. As a medical physicist, Iam interested in applying computational techniques to problemsin radiation medicine.

A large and promising branch of computation is optimization, ortechniques for selecting values of independent variables thatmaximize or minimize a particular parameter, or goal. In radiationoncology, there is the goal of maximizing the radiation dosedelivered to the tumor region, while at the same time minimizingthe dose absorbed by healthy tissue. This is accomplished byvarying the independent parameters (such as radiation beamenergy, intensity, shape, and delivery geometry) of a treatmentplan.

One exciting clinical treatment modality that incorporates theseideas is intensity modulated radiation therapy, or IMRT. Ourrecent work has focused on applying principles of computationto the optimization of patient-specific parameters in IMRT. Wehave explored promising optimization techniques, such as thegenetic algorithm, which is based on ideas from biologicalevolution. These methods become even more powerful whenimplemented in a distributed-computing framework. To this end,we have formed a collaboration with the Center forComputational Research (CCR), an academic supercomputingcenter that forms part of UB’s Center for Excellence. Byemploying the CCR’s computational resources, we have appliedthe genetic algorithm to improve IMRT treatment plans.

Another project involving optimization addresses the patientsetup problem: how do we best position a patient for a complexradiation treatment, ensuring that he or she is in the correct 3Dlocation and orientation? We are investigating a novel approachto this problem, by using optical information obtained from aregular digital camera, and applying optimization techniques todetermine the 3D geometric parameters accurately. This methodwill allow us to improve patient setup without subjecting thepatient to increased ionizing radiation exposure.

Select Publications:

1. J.S. Schildkraut, N. Prosser, A. Savakis, J. Gomez, D. Nazareth, A.K. Singh, H.K.Malhotra. “Level Set Segmentation of Pulmonary Nodules in Megavolt ElectronicPortal Images Using a CT Prior,” Medical Physics, 37(11): p. 5703-5710 (2010).

2. M Bakhtiari, H Malhotra, MD Jones, V Chaudhary, JP Walters, D Nazareth,“Applying graphics processor units to Monte Carlo dose calculation in radiationtherapy,” Journal of Medical Physics, 35(2), 120-122 (2010).

3. D. Nazareth, S. Brunner, M. Jones, H. Malhotra, M. Bakhtiari, ”Optimization ofbeam angles for intensity modulated radiation therapy treatment planning usinggenetic algorithm on a distributed computing platform,” Journal of MedicalPhysics, 34(3), 129-132 (2009).

4. H. Zhang, L. Shi, R. Meyer, D. Nazareth, W. D’Souza, ”Solving Beam-AngleSelection and Dose Optimization Simultaneously via High-ThroughputComputing,” INFORMS Journal on Computing, 21(3), 427-444 (2009).

5. W. D’Souza, H. Zhang, D. Nazareth, L. Shi, R. Meyer, ”A nested partitionsframework for beam angle optimization in intensity-modulated radiationtherapy,” Physics in Medicine and Biology, 53, 3293-3307 (2008).

6. R. Meyer, H. Zhang, L. Goadrich, D. Nazareth, L. Shi, W. D’Souza “A MultiplanTreatment-Planning Framework: A Paradigm Shift for Intensity-ModulatedRadiotherapy.” International Journal of Radiation Oncology Biol Phys, Volume68, Issue 4, Pages 1178-1189.

Molecular mechanisms of transformation and senescence

Mikhail A. Nikiforov, PhD Associate Professor of Oncology

Dr. Nikiforov joined the staff of Roswell Park Cancer Institute in2007. He completed his PhD in Genetics at the University ofIllinois at Chicago, IL after receiving his masters and bachelor'sdegree in Biochemistry from Moscow State University, Moscow,Russia.

Dr. Nikiforov completed his postdoctoral fellowships at theUniversity of Rochester and Princeton University and since 2004worked as an Assistant Professor in the Department ofDermatology at the University of Michigan. He is a member ofthe American Association for Cancer Research and Society forInvestigative Dermatology.

Research InterestsMolecular mechanisms of oncogene-mediated transformationand senescence of melanocytic cells. Role of nucleotidemetabolism in oncogenic transformation, genomic instability andsenescence. Mechanisms regulating stability of the oncoproteinC-MYC in melanocytic cells.

Select Publications 1. Denoyelle C, Abou-Rjaily G, Bengtson AL, Miller TP, Tang WH, Nikiforov MA,Kaufman RJ, Soengas M. Anti-oncogenic role of the endoplasmic reticulumdifferentially activated by mutations in the MAPK pathway. Nature Cell Biology8(10):1053-63, 2006.

2. Nikiforov MA, Riblett M, Tang WH, Gratchouck V, Hari M, Fernandez Y,Verhaegen M, Jakubowiak AJ, Soengas MS. Tumor cell-selective regulation ofNOXA by c-MYC in response to proteasome inhibition. Proc. Natl. Acad. Sci.104(49):19488-19493, 2007.

3. Wang H, Mannava S, Grachtchouk V, Zhuang D, Soengas MS, Gudkov AV,Prochownik EV, Nikiforov MA. c-Myc depletion inhibits proliferation of humantumor cells at various stages of the cell cycle. Oncogene 27(13):1905-1915,2008.

4. Flanagan SA, Krokosky CM, Mannava S, Nikiforov MA, Shewach DS. MLH1Deficiency Enhances Radiosensitization with 5-Fluorodeoxyuridine by IncreasingDNA Mismatches. Mol Pharmacol 2008;74:863-871. PMCID: PMC2615187

5. Zhuang D, Mannava S, Grachtchouk V, Tang W-H, Patil S, Wawrzyniak JA,Berman AE, Giordano TJ, Prochownik EV, Soenas MS, Nikiforov MA. C-MYCoverexpression is required for continuous suppression of oncogene-inducedsenescence in melanoma cells. Oncogene 27:6623-6634 (featured on the cover)2008.

6. Mannava S, Grachtchouk V, Wheeler LJ, Im M, Zhuang D, Slavina EG, MathewsCK, Schewach DS, Nikiforov MA. Direct role of nucleotide metabolism in C-MYC-dependent proliferation of melanoma cells. Cell Cycle 7:2392-2400(featured on the cover) 2008.

Multifunctional Agents forImaging and Therapy

Ravindra K. Pandey, PhD Distinguished Professor of Oncology

Dr. Pandey was awarded PhD in medicinal chemistry from theUniversity of Rajasthan, Jaipur, India. He then worked as apostdoctoral fellow and research associate in the laboratories ofProfessor Kevin M. Smith (University of California, Davis, 1980-1983 and 1984-1986) and the late Professor A. H. Jackson(University College Cardiff, Wales, 1983-1984) on the chemistryand biochemistry aspects of porphyrin-based compounds. Hejoined Oncologic Foundation of Buffalo in 1986 and was involvedin a photodynamic therapy (PDT) project related to Photofrin®funded by Johnson & Johnson. In 1990 he left the Foundationand joined the Photodynamic Therapy Center, Roswell ParkCancer Institute, Buffalo, where he is currently associated asDistinguished Professor and Director of PharmaceuticalChemistry, Department of Cell Stress Biology. He also has anappointment as Professor with the Institute of Lasers, Photonicsand Biophotonics, SUNY at Buffalo.

Description of Research:Photosensitizers for Photodynamic Therapy: For the last severalyears one of the objectives of Dr. Pandey’s laboratory has beento synthesize and evaluate tumor-avid porphyrin-basedphotosensitizers for photodynamic therapy (PDT) exhibiting thelong wavelength absorption in the range of 660-800 nm. Suchcompounds on exposing to light at appropriate wavelengthsconvert the molecular oxygen present in tumors to singletoxygen, a cytotoxic agent responsible for the destruction oftumors. Starting from chlorophyll-a and bacteriochlorophyll-a, hisgroup synthesized and evaluated a series of photosensitizers. Onthe basis of SAR and QSAR studies conducted in a highlystimulating collaboration with other members of the PDT groupof our department, Dr. Pandey’s group has been able to selectthe best candidate from each series, and these photosensitizersare currently at various stages of clinical (Phase I/II) andpreclinical trials.

Molecular Modeling-Based Target-Specific Photosensitizers: Themajor challenge of cancer therapy is the selective destruction ofmalignant cells while sparing normal tissue. While certainphotosensitizers show a degree of selectivity, the parameterschosen for treatment in patients are limited by reactions of thenormal tissue within the light field. Therefore, one of theobjectives of Dr. Pandey's research program has been toimprove and optimize PDT by targeting photosensitizers totumors. His current approach is to target the photosensitizers tointegrins (avb3), folate receptors and certain cellularcarbohydrate receptors (e.g. galectins) over-expressed in avariety of malignant tumors. Dr. Pandey's group is usingmolecular modeling-based approach for designing target-specific photosensitizers in collaboration.

Multifunctional Tumor-Imaging (MRI, PET and Fluorescence) andTherapeutic Agents: In recent years, cellular and molecularbiology have ushered in a new era for the characterization of thetumor tissue at the molecular level. The non-invasive molecularimaging of the specific type of tumors in vivo followed by tailoredmedical intervention is becoming the new frontier of cancertreatment. Therefore, for investigating the utility of tumor-avidphotosensitizers as vehicles to deliver the imaging agents (MRI,Nuclear Imaging, Optical Imaging) to tumors, HPPH (achlorophyll-a analog developed in our laboratory currently inPhase I/II human clinical trials) was conjugated with a variety ofimaging agents. In preliminary studies, the correspondingconjugates e.g., HPPH-DTPA, was found to be a promising dualfunction (tumor MR imaging and therapy) agent. This approach isnow being extended for developing target-specific agents forimproved efficacy.

PDT and Nanotechnology: In collaboration with the Biophotonicsgroup at SUNY Buffalo, and Dr. Kopelman, University ofMichigan, Dr. Pandey’s group has initiated several researchproject focused on investigating the utility of nanotechnologyplatforms (Ormosil, gold and polyacrylamide) in developingmultifunctional nana-platforms for tumor imaging andphototherapy. These nanoconstructs also provide an opportunityto introduce tumor-targeting moieties at the peripheral positionof the nanoparticles.

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Select Publications (from a list of 250 publications) 1. Liu C, Dobhal MP, Missert JR, Pandey RK. Highly Selective Synthesis of B-ringReduced Chlorins by Ferric Chloride Mediated Oxidation of Bacteriochlorins: Aremarkable Effect of the Fused Imide Ring in the Electronic AbsorptionCharacteristics of New Chromophores. J Am Chem Soc 2008, 130, 14311-14323

2. Pandey RK. Lighting Up the Lives of Cancer Patients by Developing Drugs forTumor Imaging and Photodynamic Therapy, Oncology Issues, March/April 2008,22-23.

3. Pandey SK, Sajjad M, Chen Y, Yao R, Missert JR, Batt C, Nabi HA, Oseroff AR,Pandey RK. Comparative PET Imaging and Phototherapeutic Potential of 124I- Labeled methyl-3-(1’-iodobenzyloxyethyl) pyropheo phorbide-a vs. thecorresponding glucose and galactose conjugate, J. Med. Chem. 2009, 52, 445-455. PMCID:PMC 2699564

4. Zheng X, Morgan J, Pandey RK, Chen Y, Baumann H, Missert JR, Batt C,Jackson J, Bellnier DA, Henderson BW, Pandey RK. Conjugation of HPPH tocarbohydrates changes its subcellular distribution and enhances photodynamicactivity in vivo. J. Med. Chem. 2009, 52, 4306-4318.

5. Goswami LN, White WH, Spernyak JA, Ethirajan M, Chen Y, Missert JR, MorganJ, Mazurchuk R, Pandey RK. Synthesis of tumor avid photosensitizer-Gd(III)DTPA conjugates: Impact of the number of gadolinium units in T1/T2relaxivity, intracellular localization and photosensitizing efficacy. BioconjugateChem. 21, 816-827, 2010.

6. Spernyak JA, White WH, Ethirajan E, Patel NJ, Goswami L, Chen Y, Turowski S,Missert JR, Batt C, Mazurchuk R, Pandey RK. Hexyl ether derivative ofpyropheophorbide-a (HPPH) on conjugating with eGd(III)ADTPA shows potentialfor in vivo imaging (MR, fluorescence) and PDT. Bioconjugate Chem. 21, 828-835, 2010.

Applications of MedicalPhysics to CancerDiagnosis and TreatmentMatthew B. Podgorsak, PhD, DABMP, FAAPMAssociate Professor of Oncology

Medical Physics is an applied branch of physics that isconcerned with the application of physical concepts andmethods to the diagnosis and treatment of disease. Medicalphysicists provide clinical services and are involved in researchin any of the four subfields of medical physics: therapeuticradiological physics, diagnostic radiological physics, medicalnuclear physics, and medical health physics. At Roswell ParkCancer Institute, the medical physics division is primarilyinterested in research aimed at improving the delivery ofradiation therapy to patients, although we do collaborate withmedical physicists at other facilities that are involved in the otherbranches of medical physics. We also collaborate with scientistsin other fields, such as physics, engineering, chemistry,computer science, and biology, with whom we share a commoninterest.

Research in a typical medical physics laboratory is either appliedin nature or it is initially theoretical but is quickly translated toclinical practice. Currently, research under my direction isfocused on three main themes: delivery of radiation therapyusing state-of-the-art techniques, development of models topredict tumor motion during a patient’s respiratory cycle, and

enhancement of quality assurance techniques for ourdepartment’s clinical services. Implicit in all research is anoverarching interest in process development in order to ensurepatient safety during state-of-the-art clinical treatment delivery.

The Department of Radiation Medicine has recently acquired twonew clinical linear accelerators equipped with image guidanceenabling very precise and accurate dose delivery. One deliverytechnique is known as volumetric modulated arc therapy (VMAT),and we have been involved in research aimed at identifying theanatomies that are best treated with this new approach. Earlyindications suggest that VMAT is a very powerful tool thatprovides clinical benefit to many patients, however, there isnonetheless a subset of patients for whom VMAT is notappropriate. We are currently in the process of identifying thissub-group of patients through treatment plan simulations andenhanced dose calculations coupled with empiricalmeasurement.

The second theme is based on collaboration with colleagues inthe Department of Mechanical and Aerospace Engineering atUB. We are developing models that will hopefully successfullypredict the excursion of tumors within a patient’s thoracic cavityduring respiration. Applications of these models during radiationtherapy treatment planning for lung tumors may enable us tominimize the size of radiation beams with an anticipateddecrease in treatment related side effects.

The research project satisfying the third theme has been on-going for several years and is based on optimization of qualityassurance techniques used to ensure safe and accurate dosedelivery. We have recently developed a mathematical algorithmthat has been applied to patient-specific quality assurancemeasurements of intensity modulated radiation beams, and weare in the process of expanding the model to VMAT delivery. It isexpected that research along the quality assurance theme willcontinue indefinitely. The need for safety in clinical care willalways be present, and as new treatment techniques andparadigms are introduced it will be necessary to develop newquality assurance tests, equipment, and programs to ensure safedelivery of care.

Select Publications:1. Van Benthuysen L, Hales L Podgorsak MB. Volumetric modulated arc therapycompared to IMRT for the treatment of distal esophageal cancer. Med.Dosimetry (in press).

2. Bailey DW, Kumaraswamy L and Podgorsak MB. A fully electronic intensity-modulated radiation therapy quality assurance (IMRT QA) process implementedin a network comprised of independent treatment planning, record and verify,and delivery systems. Radiol. Oncol. 44(2): 124-130, 2010.

3. Park CC, Yom SS, Podgorsak MB, Harris E, Price RA, Bevan A., Pouliot J,Konski AA and Wallner PE. American Society for Therapeutic Radiology andOncology (ASTRO) Emerging Technology Committee Report on ElectronicBrachytherapy. Int. J. Radiat. Oncol. Biol. Phys. 76(4): 963-972, 2010.

4. Tran T, Stanley T, Malhotra HK, deBoer SF, Prasad D, Podgorsak MB. Target andperipheral dose during patient repositioning with the Gamma Knife automaticpositioning system (APS) device. J. Appl. Clin. Med. Phys. 11(3): 88-98, 2010.

5. Bailey DW, Kumaraswamy LK and Podgorsak MB. An effective correctionalgorithm for off-axis portal dosimetry errors. Med. Phys. 36: 4089-4094, 2009.

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Nanoparticle drugformulation and tumormicroenvironment

Arindam Sen, PhDAssistant Professor of Oncology

EDUCATION

BSc, Physics, Delhi University, Delhi, India

MSc, Physics, Delhi University, Delhi, India

PhD, Life Sciences, J. Nehru University, N. Delhi, India

Postdoctoral, Biophysics, Chelsea College, University of London,London, UK

Research Interest

My general research interests include structure-functionrelationship of biological membranes, model membrane systemsand biomolecules using biophysical techniques includingmicroscopy, spectroscopy, x-ray and electron diffraction. Myspecialization is in the area of electron, optical and atomic forcemicroscopy and, uv/vis and infrared spectroscopic techniques.Specific interests include developing strategies for parenteraldrug delivery including transdermal, lipidic nanoparticles andmacromolecular assemblies as drug delivery systems. Further,we are investigating the physical barriers posed by the tumormicroenvironment on effective chemo and radiation therapy, andpossible approaches for overcoming those barriers.

CURRENT RESEARCH

Transdermal Drug Delivery and Analyte MonitoringThe broad aim of this program is to develop technologies thatcan automate transdermal drug delivery and analyte monitoringwith minimal discomfort and inconvenience. Recent transdermalelectroporation work in this laboratory has resulted in a novelmethod using electric pulses and lipid formulations to achieve atransient increase in skin permeability, enabling sampling ofsystemic glucose and delivery of insulin. We have received anUS patent for transdermal monitoring of analytes and have apatent pending for transdermal delivery.

Lipidic Nanoparticles for Drug DeliveryThe aim of this program is to design and test novel lipidicnanoparticles for drug delivery to tumors with the aim oftranslating this program to the clinic. Amongst the lipidicnanoparticles that we have tested include those that aretemperature sensitive for which we have received an US patent.Research is aimed at tumor targeting of nanoparticles by usinghyperthermia and other adjuvant therapies to enhance thetherapeutic efficacy in difficult to treat solid tumors includingmetastatic cancers.

Tumor MicroenvironmentThe aim of this program is to examine changes in the tumorvasculature and microenvironment in response to hyperthermiaor other therapies that can influence tumor drug delivery. Ouraim is to establish role of different adjuvant treatments likehyperthermia in increasing tumor uptake of drugs possibly byremodeling tumor vasculature and/or by inducing changes intumor hydrostatic pressure.

Patents1. Method for increasing the efficiency of transfection. Patent

No. US 6,187,588 B1 Inventors: S.W. Hui, S.P. Murphy, L. H.Li and A. Sen

2. Method for transdermal sampling of analytes. Patent No. US6,383,138 B1 Inventors: A. Sen, S.W. Hui and Y.L. Zhao

3. Temperature controlled content release from liposomes.Patent No. US 6,964,778 B1. Inventors: S.W. Hui and A. Sen.

4. Temperature-sensitive control of liposomes-cell adhesion.Patent No. US 6,991,805B1. Inventors: S.W. Hui and A. Sen.

5. Method for transdermal or intradermal delivery of molecules.Patent pending. Serial No. 60/184,918. Inventors: A. Sen,S.W. Hui, Y.L. Zhao and L. Zhang.

Select Publications1. Murthy SN, Zhao Y, Hui SW, Sen A. (2005) Electroporation and transcutaneousextraction (ETE) for pharmacokinetics studies of drugs. J Controlled Rel 105:132-141.

2. Murthy SN, Zhao Y, Hui SW, Sen A. (2006) Lipid and electroosmosis enhancedtransdermal delivery of insulin by electroporation. J Pharmaceutical Sci 95,2041-4050.

3. Murthy SN, Zhao Y, Hui SW, Sen A. (2006) Synergistic effect of lipid treatmentand electroosmosis for transdermal delivery of insulin. Intl J Pharm 326: 1-6.

4. Burgess SE, Zhao Y, Sen A, Hui SW. Resealing of electroporation of porcineepidermis using phospholipids and poloxamers. Intl J Pharm 2007; 336: 269-275.

5. Xu Y, Choi J, Sen A, Hylander BL, Evans SS, Kraybill WG, Repasky EA. Fever-range whole body hyperthermia enhances the delivery and therapeutic efficacyof liposomally encapsulated doxorubicin by specific functional alterations intumor vasculature. Int J Hyperthermia 2007; 23: 513-527.

6. Sckolnick M, Hui SW, Sen A. Influence of DMPS on the water retention capacityof porcine skin. Int J Pharm 2008; 350: 138-144.

7. Bhattacharya Arup, Tóth K, Sen A, Seshadri M, Cao S, Durrani FA, Faber E,Repasky Subjeck EA, Rustum YM. Histomorphological structure andmicrovessel distribution of colorectal cancers influences antiangiogenicchemomodulation effects of methylselenocysteine. Clin Colorectal Can 2009 8:155-162.

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Cancer Imaging/TargetedTherapies

Mukund Seshadri, DDS, PhDCo-Director, Preclinical Imaging FacilityAssistant Professor of Oncology

Research programMy research program is focused on the use of advanced imagingtechniques such as magnetic resonance imaging (MRI) andoptical molecular imaging techniques in preclinical and clinicalstudies to assess the biological characteristics of tumors andtheir response to traditional and novel anticancer treatments.

The program is also focused on the development of novelmultifunctional and multimodal (MRI and optical imaging)nanoparticles for targeted imaging and therapy through activeinter- and intradepartmental collaborations at Roswell Park andthe University at Buffalo.

We have previously demonstrated the potential of targetingangiogenesis for therapeutic benefit in head and neck squamouscell carcinomas (HNSCC), gliomas, colorectal cancers. UsingMRI, we have successfully characterized the vascular responseof tumors to antiangiogenic, antivascular and chemopreventiveagents such as selenium and vitamin D in head and neck, colon,lung and prostate cancers. We are currently focused ondeveloping non-invasive imaging based early responsebiomarkers that can reliably predict therapeutic outcome inHNSCC xenografts derived from primary human tumors. Studiesare also focused on the development of MR contrast agents andimaging techniques for the detection of metastatic disease inanimal models of head and neck and colon cancer.

Select Publications1. Seshadri M, Spernyak JA, Mazurchuk R, Camacho SH, Oseroff AR, Cheney RTand Bellnier DA (2005). Tumor vascular response to photodynamic therapy andthe antivascular agent, 5,6- dimethylxanthenone-4-acetic acid (DMXAA):Implications for combination therapy. Clin Can Res. 11(11):4241-50. (Coverfeature).

2. Seshadri M, Mazurchuk R, Spernyak JA, Bhattacharya A, Rustum YM andBellnier DA (2006). Activity of the vascular disrupting agent 5,6-dimethylxanthenone-4-acetic acid (DMXAA) against human head and neckcarcinoma xenografts. Neoplasia 8(7): 534-542.

3. Seshadri M, Spernyak JA, Maier PG, Cheney RT, Mazurchuk R and Bellnier DA(2007). Visualizing the acute effects of vascular-targeted therapy in vivo usingintravital microscopy and magnetic resonance imaging: Correlation withendothelial apoptosis, cytokine induction and treatment outcome. Neoplasia9(2):128-135.

4. Bhattacharya A*, Seshadri M, Oven SD, Tóth K, Vaughan M and Rustum YM(2008). Tumor Vascular Maturation and Improved Drug Delivery Induced byMethylSelenocysteine Leads to Therapeutic Synergy with Anticancer Drugs. ClinCancer Res. 14(12):3926-3932.

5. Seshadri M*, Bellnier DA and Cheney RT (2008). Assessment of the Early Effectsof 5,6-dimethylxanthenone-4-acetic acid using Macromolecular Contrast MediaEnhanced Magnetic Resonance Imaging: Ectopic versus orthotopic tumors. Int.J. Radiation Oncology Biol. Phys. 72(4):1198–1207.

6. Chung I, Guangzhou H, Seshadri M, Gillard B, Wei-dong Y, Trump DL andJohnson CS* (2009). Aberrant tumor angiogenesis in vitamin D receptorknockout mice leads to enhanced tumor growth. Cancer Res, 69(3):967-975.

7. Seshadri M, Ciesielski MJ (2009). Magnetic resonance imaging-basedcharacterization of vascular disruption by 5,6-dimethylxanthenone-4-acetic acid(DMXAA) in gliomas. J Cer Blood Flow Meta 2009 Aug 29(8):1373-82 (coverfeature)

8. Seshadri M*, Merzianu M, Tang H, Rigual NR, Sullivan-Nasca M, Loree TR,Popat SR, Repasky EA and Hylander BL (2009). Establishment andcharacterization of patient tumor-derived head and neck squamous cellcarcinoma xenografts. Cancer Biol Therapy. 8(23): 2261-2269.

Magnetic resonance imagingin animal models of disease

Joseph Spernyak, PhDCo-Director, Preclinical Imaging FacilityImaging Research Scientist

Dr. Spernyak has been involved in a broad range of researchprojects incorporating the use of magnetic resonance imaging(MRI) for studying small animal model of disease. His recentinterests have focused primarily in three areas, a) developmentand assessment of novel MRI/PDT contrast agents for thepurpose of combining imaging and therapy within a singlecompound, b) use of MRI to non-invasively characterize changesin tumor microvasculature arising from anti-cancer therapies, and c) application of balanced, steady-state free precessionimaging for efficient and accurate measurement of tissue MRrelaxation rates.

Collaborating with Dr. Ravindra Pandey (Cell Stress Biology), ourlab has been developing and characterizing novel, multi-functional imaging agents for use in both imaging andphotodynamic therapy (PDT). Combining imaging capabilitieswith a tumor-specific therapeutic has multiple advantages,including improved tumor boundary delineation and non-invasivetracking of tumor uptake and pharmacokinetics. Thesecapabilities may lead to improved prognostic indicators on theefficacy of PDT treatment based upon imaging biomarkers.

Dynamic-contrast enhanced MR imaging is being utilized toprobe changes in vascular function as a result of chemotherapy(e.g. doxorubicin) or biophysical therapies such as PDT or wholebody hyperthermia. The microvasculature architecture withintumors differs from normal vasculature in a number of ways.Tumor vasculature is chaotic and hyper-permeable, oftenexhibiting reduced blood flow, which in turn can reduce theefficacy of anti-cancer treatments. Using MRI, changes in tumorblood flow and vascular function can be studied in situ, leadingto greater understanding of the vascular effects of suchtreatments and potentially to better outcomes.

Critical to this work is the ability to perform rapid measurementof tissue relaxation rates in vivo with a high degree of accuracyand sensitivity. Balanced, steady-state free precession (bSSFP)

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imaging is being explored in a preclinical environment as ameans of rapid relaxometry, as it offers many advantages overalternative methods, namely a) high signal to noise ratios, b)improved spatial homogeneity over a large volume, c) shortacquisition times, and d) simultaneous extraction of both T1 andT2 relaxation rates within a single MRI dataset. This methodologyis currently being applied for use in novel contrast agentdevelopment, microvascular assessment, and high-resolution invivo microscopy.

Select Publications:

1. Spernyak JA, White WH, Ethirajan M, Patel NJ, Goswami L, Chen Y, Turowski S,Missert JR, Batt C, Mazurchuk R, Pandey RK. “Hexylether Derivative ofPyropheophorbide-a (HPPH) on Conjugating with 3Gadolinium(III)Aminobenzyldiethylenetriaminepentaacetic Acid Shows Potential for In vivoTumor-Imaging (MR, Fluorescence) and Photodynamic Therapy” BioconjugateChem. In Press

2. Chintala S, Novak EK, Spernyak JA, Mazurchuk R, Torres G, Patel S, Busch K,Meeder BA, Horowitz JM, Vaughan MM, Swank RT. “The Vps33a gene regulatesbehavior and cerebellar Purkinje cell number.” Brain Res. 2009 Apr 17;1266:18-28.

3. Paul AK, Lobarinas E, Simmons R, Wack D, Luisi JC, Spernyak J, Mazurchuk R,Abdel-Nabi H, Salvi R. Metabolic imaging of rat brain during pharmacologically-induced tinnitus. Neuroimage. 2009 Jan 15.

1. Seshadri M, Bellnier DA, Vaughan LA, Spernyak JA, Mazurchuk R, Foster TH,Henderson BW. Light delivery over extended time periods enhances theeffectiveness of photodynamic therapy. Clin Cancer Res. 2008 May1;14(9):2796-805.

4. Torres G, Hallas B, Gross KW, Spernyak JA, and Horowitz JM. “Magneticresonance imaging and spectroscopy in a mouse model of schizophrenia.” BrainResearch Bulletin 2008; Mar 28;75(5):556-61.

5. Seshadri M, Spernyak JA, Maier PG, Cheney RT, Mazurchuk R and Bellnier DA.“Visualizing the Acute Effects of Vascular-Targeted Therapy In Vivo Using IntravitalMicroscopy and Magnetic Resonance Imaging: Correlation with EndothelialApoptosis, Cytokine Induction and Treatment Outcome.” Neoplasia 2007; 9(2):128-135.

6. Pliss L, Mazurchuk RV, Spernyak JA, Patel MS. MR Imaging and Proton MRSpectroscopy in Female Mice with Pyruvate Dehydrogenase ComplexDeficiency. Special Issue: Neurochemical Research 2007; 32:645-54.

7. Jell J, Merali S, Hensen ML, Mazurchuk R, Spernyak JA, Diegelman P, Kisiel ND,Barrero C, Deeb KK, Alhonen L, Patel MS, Porter CW. “Genetically AlteredExpression of Spermidine/Spermine N1-Acetyltransferase Profoundly Affects FatMetabolism in Mice via Acetyl-CoA.” J Biol Chem. 2007 Mar 16; 11:8404-8413

8. Bouvet M, Spernyak J, Katz MH, Mazurchuk RV, Takimoto S, Bernacki R,Rustum YM, Moossa AR, and Hoffman RM. “High Correlation Of Whole-BodyRed Fluorescent Protein Imaging And MRI On An Orthotopic Model OfPancreatic Cancer”. Cancer Research, 2005 Nov 1; 65(21):9829-33.

9. Seshadri M, Spernyak JA, Mazurchuk R, Camacho SH, Oseroff AR, Cheney RT,Bellnier DA. “Tumor vascular response to photodynamic therapy and theantivascular agent 5,6-dimethylxanthenone-4-acetic acid: implications forcombination therapy.” Clin Cancer Res. 2005 Jun 1; 11(11):4241-50.

10. Mazurchuk R, Spernyak JA. Magnetic resonance imaging of tumor response tochemotherapy. Methods Mol Med. 2005;111:381-415.

11. Li G, Slansky A, Dobhal MP, Goswami LN, Graham A, Chen Y, Kanter P,Alberico RA, Spernyak J, Morgan J, Mazurchuk R, Oseroff A, Grossman Z,Pandey RK Chlorophyll-a analogues conjugated with aminobenzyl-DTPA aspotential bifunctional agents for magnetic resonance imaging andphotodynamic therapy. Bioconjug Chem. 2005 Jan-Feb;16(1):32-42.

Modulation of anticancerdrug biodisposition and pharmacodynamics by drug carriersRobert M. Straubinger, PhDProfessor, State University of NewYork at Buffalo

Oncology drugs as a therapeutic class tend to have the lowesttherapeutic index of any drugs in widespread clinical use, basedon their antitumor efficacy vs. toxicity to the patient. Our longterm objective is to employ drug carrier technology to mitigatedrug toxicity, increase antitumor potency, or overcome drugphysicochemical, pharmaceutical, or biodispositionalshortcomings. The three main foci of our lab are to (i) developnew carrier-based formulations that have specific properties thatcould improve the therapeutic utility of specific drugs, (ii)investigate the mechanisms by which carrier incorporation ofdrugs can change the ‘apparent pharmacology’ and confer uponthe drug/carrier complexes new mechanisms of antitumor effectthat would not be predicted from the mechanism of action of thedrug itself, and (iii) develop a rational, mechanistic, andquantitative basis upon which to combine drug carrier -basedformulations with conventional cytotoxics or novel target-selective agents.

In terms of the development of new carrier-based anticancerdrug formulations, our group was the first to elucidate thephysicochemical basis for designing stable, liposome-basedformulations containing taxanes such as paclitaxel, the activeagent in Taxol®. These formulations eliminated the toxic co-solvent in which this poorly-soluble drug was administered topatients, and in addition showed significantly reduced toxicity tocritical normal tissues. Our most recent publication on taxane-containing liposomes showed that in a drug-resistant,intracranial rat brain tumor model, paclitaxel showed activitysuperior to the clinical standard (1). Mechanistic studies thatemployed quantitative, systems-level modeling of antitumorpharmacodynamic effects that were observed by using magneticresonance imaging, which provided repeated, non-invasivemeasurement of tumor volume progression, demonstrated thatthe enhanced antitumor efficacy of the liposome formulationresulted not only from its reduced toxicity, but also from agreater propensity to exert a tumor ‘priming’ effect. Other groupshave shown that an initial administration of taxanes creates adefined temporal window in which a subsequent dose undergoesgreater deposition. Our group demonstrated that in this drugresistant rat brain tumor model, the taxane clinical standardshowed no priming effect, whereas with the liposome-basedformulation, a pharmacodynamic model that hypothesized apriming effect best captured the effect of varying taxaneliposome dose and schedule of administration (1).

Because of the utility of quantitative, systems-levelpharmacodynamic models in creating experimentally-testablehypotheses to explain how carrier-based formulations canchange the apparent pharmacology of anticancer drugs, our lab

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continues to explore the strengths and limitations of differenttypes of semi-mechanistic models in terms of describinganticancer drug action quantitatively (2,3).

The necessity to acquire experimental data that enablesdevelopment and testing of theses novel pharmacokinetic/pharmacodynamic models often challenges existing analyticaltechnology. A long-standing effort of our group is to develop ultra-sensitive approaches for drug quantification in the small samplesthat are obtainable in animal model systems (4).

The laboratory also investigates other types of carrier-basedanticancer drug formulations that have distinct properties. Ourgroup was the first to demonstrate that an FDA-approvednanoparticulate formulation, consisting of doxorubicinencapsulated in highly-stable, long-circulating liposomes, couldexert a profound ‘anti-vascular’ effect in the intracranial rat braintumor model. When administered on a specific schedule, thedoxorubicin liposome formulation compromised tumor vascularpermeability, which permitted greater deposition of subsequentdoses. A recent publication demonstrated that the amount ofdrug deposited in tumor doubled on the second administration,as a result of the vascular compromise (5). This effect is notobserved with the free drug. An upcoming publication providesevidence that the nanoparticulate doxorubicin formulation exertsthese novel antivascular effects by extravasation andsequestration near the tumor blood vessel wall. This creates anintra-tumor depot that kills tumor vascular endothelium overtime, resulting in higher tumor perfusion and permeability of thetumor vasculature. These studies also suggest that theantivascular effect of nanoparticulate doxorubicin formulationscould be exploited to increase tumor deposition of other,conventional anticancer drugs, thus overcoming the barrier ofthe tumor vasculature to achieving drug levels within tumors thatare sufficient to reverse tumor progression.

Select Publications:1. Zhou R, Mazurchuk RV, Tamburlin JH, Harrold JM, Mager DE, Straubinger RM.Differential pharmacodynamic effects of paclitaxel formulations in an intracranialrat brain tumor model. J Pharmacol Exp Ther 332: 479-88 (2010).

2. Yang J, Mager DE, Straubinger RM. Comparison of two pharmacodynamictransduction models for the analysis of tumor therapeutic responses in modelsystems. AAPS J 12: 1-10 (2010).

3. Straubinger RM, Krzyzanski W, Francoforte CM, Qu J. Applications ofquantitative pharmacodynamic effect markers in drug target identification andtherapy development. Anticancer Res 27: 1237-46 (2007).

4. Yu H, Straubinger RM, Cao J, Wang HS, Qu J. Ultra-sensitive quantification ofpaclitaxel using selective solid-phase extraction in conjunction with reversed-phase capillary liquid chromatography/tandem mass spectrometry. J ChromatogA 1210: 160-7 (2008).

5. Arnold RD, Mager DE, Slack JE, Straubinger RM. Effect of repetitiveadministration of doxorubicin-containing liposomes on plasma pharmacokineticsand drug biodistribution in a rat brain tumor model. Clin Cancer Res11: 8856-65 (2005).

Novel optical imagingtechniques for therapymonitoring

Ulas Sunar, PhD Assistant Professor of Oncology

CURRENT PROGRAM– Project 1: Fluorescence molecular tomography

– Project 2: Noninvasive tumor therapy monitoring with optical imaging

– Project 3: High resolution photoacoustic imaging

Profile:Our research focuses on the development of novel optical imagingtechniques such as fluorescence molecular imaging, optical bloodflow, volume and oxygenation imaging for noninvasive tumortherapy monitoring and high resolution photoacoustic imaging. We utilize state of the art instrumentation techniques and developmathematical (forward/inverse modeling of photon diffusion)modeling of tissue, image reconstruction algorithms. Thesetechniques are being applied in preclinical models and beingtranslated into clinical studies of therapy monitoring, particularlyon head and neck cancer and skin cancer patients duringphotodynamic therapy.

Optical imaging can utilize intrinsic functional contrasts such as hemoglobin in blood to extract highly valuable functionalcontrast of tissue blood oxygenation and blood volume.Furthermore, optical imaging is also sensitive to motion of bloodcells and can quantify blood flow without contrast agentadministration. Fluorescence molecular tomography (FMT), arelatively new optical imaging modality, allows quantitative imagingof tissue noninvasively, in cases of exogenous contrasts agentsadministered to tissue for improved contrast. FMT has beensuccessfully utilized for breast and brain imaging in humans, andquantitative, comparative studies performed with other imagingmodalities. Since FMT is fast and noninvasive technique, it hasbeen also utilized for repetitive and longitudinal studies such astherapy monitoring. With clinical relevance in mind, weconstructed a whole-body fluorescence tomography instrument to monitor therapeutic drugs in deeply seated tumors in smallanimals. The instrument allows dense source and detectorsampling with a fast galvo scanner and a CCD detector forimproved resolution and sensitivity.

Our recent research area focuses on photoacoustic imaging. Oneof the main issues with optical-alone imaging technique is that itsuffers from the inherent high scattering of photons by biologicaltissue resulting in low spatial resolution. To address this issue welike several other research groups are combining optics andultrasound to provide high functional optical contrast at superiorultrasound resolution. Photoacoustic imaging can achieve spatialresolution of a single capillary level (~5µm) at ~700µm tissuepenetration depth, and ~50µm resolution up to ~3mm depth.Since it inherits functional optical contrast, it can assess bloodvessel vasculature, oxygenation, and flow at high resolution.

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Select Publications:

Sunar U, Makonnen S, Zhou C, Durduran T, Yu G, Wang HW, Lee W, Yodh AG.Hemodynamic responses to antivascular therapy and ionizing radiation assessedby diffuse optical spectroscopies. Opt. Express 15:15507-15516 (2007).

Sunar U, Quon H, Durduran T, Zhang J, Du J, Zhou C, Yu G, Choe R, Kilger A,Lustig R, Loevner L, Nioka S, Chance B, Yodh AG. Noninvasive diffuse opticalmeasurement of blood flow and blood oxygenation for monitoring radiation therapyin patients with head and neck tumors: A pilot study. J. Biomed. Opt., 11(6):p.064021 (2006).

Hall DJ, Sunar U, Heydari S F and Han S H. In vivo simultaneous monitoring of twofluorophores with lifetime contrast using a full-field time domain system. Appl. Opt.48:10, D74-D78 (2009).

Hall DJ, Sunar U, Heydari SF. Quadrature detection of ultrasound-modulatedphotons with a gain-modulated, image-intensified, CCD camera. The Open OpticsJournal 2:75-78 (2008).

Song L, Li H, Sunar U, Cao W, Chen J, Yodh AG, Zheng G. Naphthalocyanine-Reconstituted LDL Nanoparticles for in vivo Cancer Imaging and Treatment. Intl. J.of Nanomedicine, 2(4):1-8 (2007).

Lee I, Kim DH, Sunar U, Magnitsky S, Shogen K. The physiological mechanisms ofranpirnase-induced enhancement of radiation response on A549 human NSCLCxenografts in nude mice. In Vivo, 21:5-12 (2007).

Nioka S, Kime R, Sunar U, Im J, Izzetoglu M, Zhang J, Alacam B, Chance B. Anovel method to measure muscle blood flow continuously using NIRS kineticsinformation. Dynamic Medicine, 5:5 (2006).

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