Lung Cancer: The Breathtaking Battle of TKIs and EGFR Mutants

Lung Cancer: The Breathtaking Battle

of TKIs and EGFR MutantsBobby Hodges, Elizabeth Ross, Natalie Simak, Keith Solvang and

Eliot Vildaver

DiseasedGenomeHospital

http://blogs.usask.ca/medical_education/archive/ElksRehabFront%2520Door.jpg

http://1.bp.blogspot.com/_HI2mCx5P3H0/SLlEQoHj8AI/AAAAAAAAA8Q/My7tf78TNc8/s320/hospital-bed-786282.jpg

http://img.quamut.com/chart/9471/02_healthy_sick_x-ray.jpg

http://www.cardiothoracicsurgery.org/content/2/1/16/figure/F1?highres=y

Where Are We Headed?What Is Lung Cancer?

• Types

• Risks

Predisposition

Mutations

• EGFR

• Other proteins

http://www.freefoto.com/images/21/16/21_16_2---Single-Carriageway-Road_web.jpg

Cancer

http://www.odec.ca/projects/2005/thog5n0/public_html/Tumour.gif

History of Lung Cancer

http://www.icowman.com/uploadfiles/2007072409225175.jpg http://www.icowman.com/uploadfiles/2007072409225175.jpg

Pathology

NON SMALL CELL

Squamous cell carcinoma

Adenocarcinoma Large-cell carcinoma

SMALL CELL

Bronchi Grows rapidly Spreads quickly

http://images.google.com/imgres?imgurl=http://www.dkimages.com/discover/previews/852/20223038.JPG

http://images.google.com/imgres?imgurl=http://upload.wikimedia.org/Lynch et al., (2004)

Risk Factors

http://www.taconichills.k12.ny.us/webquests/noncomdisease/lungcancerpic.jpg

You Are Not Alone

This year approximately 200,000 people will be diagnosed with lung cancer

NSCLC accounts for 80% of all lung cancers

http://www.cdc.gov/cancer/lung/statistics/http://seo2.0.onreact.com/wp-content/uploads/2008/09/you-re-not-alone.jpg

Epidemiology

http://medicineworld.org/cancer/lung/ http://images.google.com/

What Next?

While there is no known cure as of right now, there ARE options!

http://dspace.mit.edu

ImatinibGefinitib

Cisplatin

http://dspace.mit.edu

Cisplatin

Imatinib

Gefitinib

EGF ER

BB2

EGFR

Nucleus

P

Protein Cascade

?

?

P

P

EGF ER

BB2

EGFR

PI3K

AKT

P

P

MET

P

VEG

F-A

Cisplatin

Imatinib Gefiti

nib

Nucleus

Protein Cascade

PDGFR

Protein Cascade

Implications of Chromosomal Loci in the Development of Lung Cancer

http://rosenblumtv.files.wordpress.com

http://www.pharmgkb.org

http://academic.evergreen.edu

Chromosomes

Compact structures of DNA

Contains thousands of genes

Centromere

Chromatid

Genes

http://staff.jccc.net/PDECELL/celldivision/chromosome1.gif

Single Nucleotide Polymorphisms (SNPs)

Single variation in nucleotide base pairs

Alleles are a result of SNPs within gene sequence

SNP’s are the basis for lung cancer susceptibility

http://www.dnalandmarks.com/english/pictures/what_are_snps.jpg

Genome-wide Association Studies

Variation across genomes

Genotype cases along with controls

Find roles that genes play in disease

Large sample sizes

http://www.blog.speculist.com/archives/dna2two

Identifying Regions On Chromosomes

Centromere

p-arm

q-arm

6 Regions Away

Region 5q6

5http://images.google.com/

Three Regions of Association

http://upload.wikimedia.org/ http://upload.wikimedia.org/

Nicotinic Acetylcholine Receptor Genes Found on 15q24 and 15q25

Nicotine binds and stimulates acetylcholine ion channel-linked in

neurons

http://www.jyi.org/articleimages/88/originals/img0.jpg

How were these regions discovered?

(Hung et al., 2008)

15q24 highly linked with smoking behavior

(Thorgeirson et al., 2008)

TERT gene found on 5p15.33

Cancerous Growth

http://images.google.com/i

5p15.33 shows similar results

(McKay et al., 2008)

P

P

EGF ER

BB2

EGFR

PI3K

AKT

P

P

MET

P

VEG

F-A

Cisplatin

Imatinib Gefiti

nib

Nucleus

Protein Cascade

PDGFR

Protein Cascade

How Do Cells Proliferate?

EGFR/ERBB2 Forms a Heterodimer

EG

FR

P

EG

FR

EGF

ProliferationSurvival

Apoptosis

ERBB2

Nucleus

EG

FR

P

Protein Cascade

ERBB2

How Can This Lead to Cancer?

Mutated EGFR Leads to Cancer

ERBB2EG

FR

Nucleus

Proliferation

P

Protein Cascade

Mutated EGFRs have Distinct Patterns of Tyr Phosphorylation

(Chen et al., 2006)

Tyrosine Kinase Inhibitors (TKIs)Gefitinib

EG

FR

ATP TKI

ATP binding cleft on the kinase

TyrosineKinasedomain

Gefitinib Suppresses Phosphorylation of Most EGFR

Mutants

(Chen et al., 2006)

What About the Proteins in the

Signaling Pathway?

Effectiveness of Gefitinib is Dependent on Concentration

(Pedersen et al., 2005)

High Doses of Gefitinib Decreases Proliferation of Mutated EGFR Cells

(Pedersen et al., 2005)

Gefitinib has Variable Growth-Inhibitory Effects

(Chen et al., 2006)

P

P

EGF ER

BB2

EGFR

PI3K

AKT

P

P

MET

P

VEG

F-A

Cisplatin

Imatinib Gefiti

nib

Nucleus

Protein Cascade

PDGFR

Protein Cascade

What Are the Roles of Proteins Associated with Lung Cancer?

http://www.taconichills.k12.ny.us/webquests/noncomdisease/lungcancerpic.jpg

Recap

Nucleus

N-terminus

C-terminus

P

Serine

P

Threonine

Tyrosine

P

Leucine

Lysine

Alanine

How Does The Pathway Work?

Domain Attraction

ERBB2 P

PI3KSH2

Nucleus

P

Protein Cascade

Change in Gene

Expression

Why are EGFR and ERBB2 Special?

Main Contributors to NSCLC

(Rikova et al. 2007)

Link between Concentration and Interaction

Green and Blue circles = Proteins involved in cascade

Red lines = Interactions

Red circles = EGFR Receptor

(Jones et al. 2006)

Other ERBB2 Associations

(Jones et al. 2006)

No

. p

rote

in r

ece

pto

rs

No

. p

rote

in r

ece

pto

rs

No

. p

rote

in r

ece

pto

rs

Affinity Threshold (nM) Affinity Threshold (nM) Affinity Threshold (nM)

Are there other types of Proteins?

http://en.wikipedia.org/wiki/Proteins

Other Contributors to NSCLC

(Rikova et al. 2007)

ROS ALK MET PDGFRα ErbB2 EGFR VEGFR-10

5

10

15

20

25

30

35

40

45

50

Tumor Samples

Cell Lines

Chart configured from Table1 of Rikova et al. 2007

Specifically, some proteins more than others

Am

ount

of

over

phos

phor

ylat

ed p

rote

in in

sam

ples

Previous Research, Amplification of MET

(Engelman et al., 2007)

P

P

EGF ER

BB2

EGFR

PI3K

AKT

P

P

MET

P

VEG

F-A

Cisplatin

Imatinib Gefiti

nib

Nucleus

Protein Cascade

PDGFR

Protein Cascade

What is the Role of Angiogenesis in Lung Cancer?

http://www.taconichills.k12.ny.us/webquests/noncomdisease/lungcancerpic.jpg

Angiogenesis and VEGF

http://www.researchvegf.com/researchvegf/multimedia/index.m#type-slides

Tumor Angiogenesis Is Unregulated

VEGF-A

VEGFR

Angiogenesis

Tumors

PDGF-β

PDGFR-β

PI3K / Akt Pathway in Lung Cancer

P85-α PIP2

P

P

PTEN

P

PIP3P

P

P

P110-α

PI3KAKT

SH2

PP

Endothelial Cell

Akt is a Downstream Regulator

AKT

P

P

P

BAD

MDM2

mTORGrowth,

Translation

Apoptosis

p53

P

Nucleus

FLJ10540 Is An Oncogene

Cancer Cell

Proto-Oncogene Oncogene

VEGF-A Upregulates FLJ10540

(Chen et al., 2009)

FLJ10540 Mediates Cell Migration and Invasion Through PI3K/Akt

Giemsa-Stained Cells

(Chen et al., 2009)

Recap: FLJ10540 Missing Link in PI3K/Akt

P85-α PIP2

P

P

PTEN

P

PIP3P

P

P

P110-α

PI3K

AKT

SH2

PP

VEGF-A

VEGFR

FLJ10540

Endothelial Cell

Imatinib is a TKI

http://www.medicinescomplete.com/mc/martindale/2007/images/MRT9229C001.gif

Binds to ATP binding site of many enzymes:

p-PDGFR-β Inhibitor Reduced IFP, Hypoxia

Decreased VEGF Expression

Imatinib Targets IFP and Hypoxia

http://www.medscape.com/pi/editorial/clinupdates/2000/583/art-tu02.fig04.jpghttp://www.nature.com/nm/journal/v7/n9/images/nm0901-987-F1.gif

Normal

Tumor

Imatinib Reduces p-PDGFR-β and VEGF Expression

(Vlahovic et al. 2007)

Imatinib Reduces Hypoxia and MVD

Vlahovic et al. 2007

Imatinib Treatment

http://onctalk.com/wp-content/uploads/2008/01/angiogenesis-summary.jpg

Imatinib

P

P

EGF ER

BB2

EGFR

PI3K

AKT

P

P

MET

P

VEG

F-A

Cisplatin

Imatinib Gefiti

nib

Protein Cascade

PDGFR

Protein Cascade

Nucleus

Treatments

Dr. Vildaver

• ImatinibDr. Ross

• Gefinitib

Discovery of Cisplatin

(Rosenberg et al., 1969)

Old Model of Cisplatin

Cisplatin

NucleusSimple Diffusion

Pt

NH3NH3

GGP73

C-Abl

Cell Cycle Arrest

&Apoptosis

P53

HMG1?HMG1

?

How Pt Drugs Enter Cells

nucleus

Pt

Pt

Pt

Pt

Pt

Transporter

protein

Transporter

protein

Pt

(Hall et al., 2007)

Crosslinks

crosslink

nucleus

Pt

NH3

NH3

Cell Cycle Arrest

&Apoptosis

GG

1,2-intrastrand d(GpG)adduct

(Ohndorf et al., 1999)

Pt-DNA adduct

http://www.unc.edu/~shantanu/images/bkstcg_aacr07.png

High Mobility Group Proteins

Pt-DNA Adduct

Apoptosis

HMGB

HMGB

P73

P53

(Stross et al., 2002)

(Ohndorf et al., 1999)

*Nucleotides

crosslinked

Bold- nucleotides contacted by the

protein

Phenylalanine Mutations

(Ohndorf et al., 1999)

Old Model of Cisplatin

Cisplatin

Nucleus

?Simple Diffusion

Pt

NH3NH3

GGP73

C-Abl

Cell Cycle Arrest

&Apoptosis

P53

HMG1?HMG1

?

2 Distinct Regulating Mechanisms

tyrosine kinase

mismatch repair protein

cisplatin

homologue of p53

Tumor suppresor

(Gong et al., 1999)

Role of p73 in Cell Death

P53 knockout= less sensitive to CDDP, survived better than wild type in lower concentrations of CDDPMLH1-/- & Abl= less sensitive to CDDP , survived better than wild type in lower concentrations of CDDP

(Gong et al., 1999)

New Model of Cisplatin

CisplatinNucle

usSimple Diffusion

Pt

NH3NH3

GG

P73

Cell Cycle Arrest

&Apoptos

is

P53

HMG1

HMG1

ATM

TransporterProteins

C-AblP

Cisplatin

Imatinib

Gefitinib

EGF ER

BB2

EGFR

Nucleus

P

Protein Cascade

??

P

P

EGF ER

BB2

EGFR

PI3K

AKT

P

P

MET

P

VEG

F-A

Cisplatin

Imatinib Gefiti

nib

Nucleus

Protein Cascade

PDGFR

Protein Cascade

Cocktail of Drugs

http://immunodefence.com/ii/gefitinib.gif

http://www.themesotheliomalibrary.com/cisplatin.jpg

http://www.specialityformulations.com/pcat-gifs/products-small/imatinib.jpg

?

Current & Future Hypotheses

New plasma membrane proteins have unique mechanisms and effects on NSCLC

Gefitinib effectively inhibits particular EGFR mutants

SiRNA could be used as a possible treatment for various mutated proteins

Future Studies

How does cisplatin enter the nucleus?

Roles and mechanisms behind the various mutated proteins involved in lung cancer

New drugs , secondary mutations

Acknowledgements

Professor Sleiter

?Schmooing yeast!!!

Acknowledgements

Dr. DebBurman

Acknowledgements

Mike Fiske

Abid MR, Guo S, Minami T, Spokes KC, Ueki K, et al. (2004) Vascular endothelial growth factor activates PI3K/Akt/forkhead signaling in endothelial cells. Arterioscler Thromb Vasc Biol 24: 294-300.

Agami, R., Blandino, G., Oren, M., and Shaul, Y. (1999). Interaction of c-Abl and p73alpha and their collaboration to induce apoptosis. Nature 399, 809-813.

Alberg AJ, Ford JG, Samet JM; American College of Chest Physicins. Epidemiology of lung cancer: ACCP evidence-based clinical practice guidelines (2nd edition). Chest. 2007; 132:29S-55S.

Alberts, B., Bray, D., Hopkin, K., Johnson, A., Lewis, J., Raff, M., Roberts, K., and Walter, P. 2004. Essential Cell Biology. Garland Science. New York, pp740.

Chanock, Stephen J., Hunter, David J. 2008. When the smoke clears. Nature. 452: 537-538.

Chen, C.H., Lai, J.M., Chou, T.Y., Chen, C.Y., Su, L.J., Lee, Y.C., Cheng, T.S., Hong, Y.R., Chou, C.K., Whang-Peng, J., Wu, Y.C., and Huang, C.Y. (2009). VEGFA upregulates FLJ10540 and modulates migration and invasion of lung cancer via PI3K/AKT pathway. PLoS ONE 4, e5052.

Chen, Y.R., Fu, Y.N., Lin, C.H., Yang, S.T., Hu, S.F., Chen, Y.T., Tsai, S.F., and Huang, S.F. (2006). Distinctive activation patterns in constitutively active and gefitinib-sensitive EGFR mutants. Oncogene 25, 1205-1215.

Engelman, J.A., Janne, P.A., Mermel, C., Pearlberg, J., Mukohara, T., Fleet, C., Cichowski, K., Johnson, B.E., and Cantley, L.C. (2005). ErbB-3 mediates phosphoinositide 3-kinase activity in gefitinib-sensitive non-small cell lung cancer cell lines. Proc. Natl. Acad. Sci. U. S. A. 102, 3788-3793.

Ferrara, N. and Kerbel, R.S. (2005). Angiogenesis as a therapeutic target. Nature 438, 967.

Franks, L.M. and Teich, N. (1986). Introduction to the Cellular and Molecular Biology of Cancer (New York:Oxford University Press).

Gao, B., Lee, S.M., and Fang, D. (2006). The tyrosine kinase c-Abl protects c-Jun from ubiquitination-mediated degradation in T cells. J. Biol. Chem. 281, 29711-29718.

Hall, M.D., Okabe, M., Shen, D.W., Liang, X.J., and Gottesman, M.M. (2008). The role of cellular accumulation in determining sensitivity to platinum-based chemotherapy. Annu. Rev. Pharmacol. Toxicol. 48, 495-535.

Helotera, H. and Alitalo, K. (2007). The VEGF Family, the Inside Story. Cell 130:591-592

Hung, Rayjean J. et al. 2008. A susceptibility locus for lung cancer maps to nicotinic acetylcholine receptor subunit genes on 15q25. Nature. 452: 633-637.

Huang, J.C., Zamble, D.B., Reardon, J.T., Lippard, S.J., and Sancar, A. (1994). HMG-domain proteins specifically inhibit the repair of the major DNA adduct of the anticancer drug cisplatin by human excision nuclease. Proc. Natl. Acad. Sci. U. S. A. 91, 10394-10398.

Jemal A, Siegel R, Ward E, Hao Y, Xu J, et al. (2008) Cancer statistics, 2008. CA Cancer J Clin 58: 71-96.

References

ReferencesJohnson DH, Blot WJ, Carbone DP, et al. Cancer of the lung_Non-small cell lung cancer and small cell lung cancer. In: Abeloff MD, Armitage JO, Niederhuber JE, Kastan MB, McKena WG. Clinical Oncology. 4th ed.

Philadelphia, Pa: Churchill Livingstone Elsevier: 2008: chap 76.

Jones, R.B., Gordus, A., Krall, J.A., and MacBeath, G. (2006). A quantitative protein interaction network for the ErbB receptors using protein microarrays. Nature 439, 168-174.

Kwak, E.L., Sordella, R., Bell, D.W., Godin-Heymann, N., Okimoto, R.A., Brannigan, B.W., Harris, P.L., Driscoll, D.R., Fidias, P., Lynch, T.J. et al. (2005). Irreversible inhibitors of the EGF receptor may circumvent acquired resistance to gefitinib. Proc. Natl. Acad. Sci. U. S. A. 102, 7665-7670.

McKay, James D. et al. 2008. Lung cancer susceptibility locus at 5p15.33. Nature Genetics. 40(12): 1404-1406.

Moyer JD, Barbacci EG, Iwata KK, Arnold L, Boman B, et al. (2006) Induction of apoptosis and cell cycle arrest by CP-358,774, and inhibitor of epidermal growth factor receptor tyrosine kinase. Cancer Res 57: 4838-4848.

Ohndorf, U., Rould, M., He, Q., Pabo, C., Lippard (1999). Basis for recognition of cisplatin-modified DNA by high-mobility-group proteins. Nature. 399: 708-712.

Pedersen, M.W., Pedersen, N., Ottesen, L.H., and Poulsen, H.S. (2005). Differential response to gefitinib of cells expressing normal EGFR and the mutant EGFRvIII. Br. J. Cancer 93, 915-923.

Pietras K, Ostman A, Sjoquist M, Buchdunger E, Reed RK, Heldin CH, Rubin K (2001) Inhibition of platelet-derived growth factor receptors reduces interstitial hypertension and increases transcapillary transport in tumors. Cancer Res 61: 2929-2934.

Poklar, N., Pilch, D.S., Lippard, S.J., Redding, E.A., Dunham, S.U., and Breslauer, K.J. (1996). Influence of cisplatin intrastrand crosslinking on the conformation, thermal stability, and energetics of a 20-mer DNA duplex. Proc.Natl. Acad. Sci. U. S. A. 93, 7606-7611.

Rikova, K., Guo, A., Zeng, Q., Possemato, A., Yu, J., Haack, H., Nardone, J., Lee, K., Reeves, C., Li, Y. et al. (2007). Global Survey of Phosphotyrosine Signaling Identifies Oncogenic Kinases in Lung Cancer. Cell 131, 1190-1203.

Ruddon, R.W. (2007). Cancer Biology: Fourth Edition (New York: Oxford University Press).

Schiller JH, Harrington D, Belani CP, Langer C, Sandler A, Krook J, Zhu J, Johnson DH (2002) Comparison of four chemotherapy regimens for advanced non-small-cell lung cancer. N Engl J Med 346: 92-98.

Shilo, B., Yarden, Y. (2007). Snap Shot: EGFR Signaling Pathway. Cell.131, 1018e1-1018e2. Stros, M., Ozaki, T., Bacikova, A., Kageyama, H., and Nakagawara, A. (2002). HMGB1 and HMGB2 cell-specifically down-regulate the p53- and p73-dependent sequence-specific transactivation from the human Bax gene

promoter. J. Biol.Chem. 277, 7157-7164.

Thorgeirson, Thorgeir E. et al. 2008. A variant associated with nicotine dependence, lung cancer and peripheral arterial disease. Nature. 452: 638-642.

Vlahovic G, Rabbani ZN, Herndon II JE, Dewhirst MW, Vujaskovic Z (2006) Treatment with Imatinib in NSCLC is associated with decrease of phosphorylated PDGFR-beta and VEGF expression, decrease in interstitial fluid pressure and improvement of oxygenation. Br J Cancer 95: 1013– 1019.

Yarden, Y., and Sliwkowski, M.X. (2001). Untangling the ErbB signalling network. Nat. Rev. Mol. Cell Biol. 2, 127-137.

Yuan, Z.M., Shioya, H., Ishiko, T., Sun, X., Gu, J., Huang, Y.Y., Lu, H., Kharbanda, S., Weichselbaum, R., and Kufe, D. (1999). p73 is regulated by tyrosine kinase c-Abl in the apoptotic response to DNA damage. Nature 399, 814-817.

Yun, C.H., Mengwasser, K.E., Toms, A.V., Woo, M.S., Greulich, H., Wong, K.K., Meyerson, M., and Eck, M.J. (2008). The T790M mutation in EGFR kinase causes drug resistance by increasing the affinity for ATP. Proc. Natl. Acad. Sci. U. S. A. 105, 2070-2075.