Direct and Repeated Clinical Measurement of pO2 for Enhancing Cancer Therapy

2014 Norris Cotton Cancer Center Comprehensive Thoracic Oncology Program (CTOP)

Retreat

Benjamin B. Williams, Ph.D.

The Geisel School of Medicine at Dartmouth, Departments of Medicine (Radiation Oncology) and Radiology

May 22, 2014

Clinical EPR at DartmouthClinical problem

Parameter to be measured

Status of measurements in human subjects

Rationale for using in vivo EPR

Cancer pO2 in tumors

• Numerous measurements performed in patients with superficial tumors.

• FDA IDE submitted for application of implantable oxygen sensors for application at increased depth and with increased sensitivity

• NCI PPG proposal in review • Established collaborations with

external academic institutions

The response of tumors to cytotoxic therapy, especially ionizing radiation, is critically dependent on pO2. Anti-tumor therapies are given repeatedly and often change pO2. Knowledge of the changes in individual patients would significantly optimize the timing of the therapy

Potential exposure to

clinically significant doses of radiation

Radiation-induced EPR signals in teeth (magnitude is proportional to dose)

• Underway in unirradiated volunteers and patients receiving significant doses to teeth from radiation therapy.

• Measurements at remote locations possible using transportable device.

• Established collaborations with external academic institutions

Exposures may occur from terrorism, war, or accident. In vivo EPR is the only physical method capable of making measurements ‘after-the-fact

Peripheral vascular disease

Oxygen at sites of likely pathologies

• Measurements underway in normal volunteers.

The pO2 in the tissues is the most significant pathophysiological variable; no other method available to make such direct measurements.

Radiation Induced Fibrosis

pO2 in irradiated tumor beds and peripheral normal tissue

• Recruitment and measurements underway

Radiation-induced hypoxia may play a critical role in the signaling of pro-inflammatory, pro-fibrotic, and pro-angiogenic growth factors and cytokines that lead to tissue fibrosis.

Wound healing

pO2 near wounds and in transplanted tissues

• Conceptual stages, with established pre-clinical investigations

The pO2 is a critical variable for successful healing of wounds. Direct measurements would identify patients likely to have poor healing and follow responses to therapy

EPR: Background Fundamentals

• EPR is a form of magnetic resonance spectroscopy that measures the absorption of RF energy by unpaired electrons in a magnetic field.

• The physics is similar to that of MRI, but unpaired -e are detected.

• Features and considerations:

(a) Clinical EPR spectrometer

(b) Subject positioned for oximetry using India ink as reporter

- Non-ionizing modality- Spectroscopic data available- Sensitive to the magnetic

environment- In most cases exogenous spin probes

must be introduced.- Lower magnetic fields- Higher RF frequencies, shallower

penetration- Rapid relaxation of electron spins

affects detection strategies

Unique or Exceptional Capabilities of In Vivo EPR

In vivo EPR is sensitive to a wide array of physiologic parameters.

This information is observed through simple changes in the shapes of the detected spectra.

• Partial pressure of oxygen (pO2; using oxygen-sensitive paramagnetic materials)

• Free radicals observed directly or by spin trapping, including oxygen-, carbon-, and sulfur-centered radicals

• Nitric Oxide (spin trapping)• Redox status (using metabolism of

nitroxides)• Thiol groups (using specific nitroxides)• pH (using specific nitroxides)• Perfusion (using washout of paramagnetic

tracers)• Metal ions (in paramagnetic states such as

chromium, manganese)• Absorbed dose of radiation (in teeth, nails,

or bones)

EPR Sensitivities : OximetryCollisions between the spin probe and molecular O2 promote relaxation and broaden the linewidth of the observed spectrum.

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Magnetic field, Gauss

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1000

1500

2000

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(air)

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Significance of Hypoxia for Cancer Therapy

• Hypoxia plays crucial roles in tumor development and treatment.– The response of tumors to cytotoxic therapies, especially

ionizing radiation, is critically dependent on pO2.– Hypoxia may promote metastatic growth and tumor

phenotypes that are more resistant to therapy.– Hypoxia is dynamic: diffusion, perfusion (incl. cycling),

anemic

Tatum JL, Kelloff GJ, Gillies RJ, et al. Hypoxia: Importance in tumor Biology, noninvasive measurement by imaging, and value of its measurement in the management of cancer therapy. Int J Radiat Biol. 2006 Oct;82(10):699-757.

Eric J. Hall, Amato J. Giaccia. Radiobiology for the radiologist. Lippincott Williams & Wilkins, 2006; 546p.

Hockel M, Schelnger K, Aral B, Mitze M, Schaffer U, Vaupel P. Association between Tumor Hypoxia and malignant Progression in Advanced Cancer of the Uterine Cervix. Can Res. 1996 Oct;56:4509-4515.

Significance of Hypoxia for Cancer Therapy

• Hypoxia plays crucial roles in tumor development and treatment.– The response of tumors to cytotoxic therapies, especially

ionizing radiation, is critically dependent on pO2.– Hypoxia may promote metastatic growth and tumor

phenotypes that are more resistant to therapy.– Hypoxia is dynamic: diffusion, perfusion (incl. cycling),

anemic

Therapeutic Cycle

Optimize Delivery Timing of dose Application of pO2 modifiers Combination therapies

*Assessment pO2 redistribution Vascular remodeling Monitor normal tissues

*Design Select patients Tumor profiling pO2 heterogeneity

*

*Points of pO2 measurement

Initiation ofintervention

ImprovedSurvival

Treatment Outcome

To establish EPR oximetry as a clinical tool for noninvasive monitoring of tumor oxygen in human subjects, under conditions that are fully compatible with clinical practice

Overall goal

Capabilities of EPR Oximetry

EPR oximetry can provide repeated and direct measurements of absolute pO2 in tumors and other tissues. – Following introduction of the reporters (injection or implantation), the

measurements are non-invasive– Measurements are made in the tissue at the site of interest– Measurements can be made continuously or repeatedly as needed, often

indefinitely, at the same site– Sub-mm resolution can be achieved – The measurements are especially sensitive and accurate at the low levels

of pO2 (sensitivity to D < 1 mmHg)– Compatibility with complementary techniques for imaging and

measurement– Immediate clinical applicability using India ink, advanced techniques in

stages of regulatory approval

EPR Oximetry with India Ink

• Patient preparation - Injection of India ink– India ink formulated with Printex-U carbon black

(200mg/ml) in 0.9% NaCl and 1.25% CMC– Sterilized via autoclave prior to injection– 20-50mL injected into tissue of interest using 22 gauge

needle– 1-10mm depth

• Measurements– Clinical whole-body spectrometer with temperature

control– EPR data collection (10s/scan, 1-30 min/set) – Measurements repeated over the course of treatment as

desired

PU ink new calibration 200701

PO 2 mmHg

0 20 40 60 80 100

LW

(G)

0

5

10

15

20

25

30

regression PO

2 mmHg vs LW

F = y0+a*x/(b+x) y0 = 1.1229 a = 51.2401 b = 120.4903

Ink injection and calibrated pO2 response

Preliminary Measurements in Humans

• Tumor pO2 has been measured in 14 volunteers.• Various tumor types of superficial tumors• Measurement sites have ranged from the feet to the scalp.• Tumor pO2 varied among the patients studied and over the course

of treatment.• Tumors were tested for response to changes of inhaled oxygen

gas.• Large differences among tumors were found showing potentially

very important implications for improving personalized therapy based on the oxygen levels in that person’s tumor.

pO2 Variability across Serial Measurements

V7 - pO2 Measurments during RT

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3/3/2008 3/5/2008 3/7/2008 3/9/2008 3/11/2008 3/13/2008 3/15/2008

Measurement Day

pO

2 [m

mH

g]

Scalp Pre-treatment

Scalp Post-treatment

Neck Pre-treatment

Neck Post-treatment

Tumor pO2 was monitored in melanoma metastases at two sites, in the scalp and neck, during the course of radiation treatment (66 Gy). Spectra were recorded immediately before and after each fraction while the patient inspired room air. These results indicate hypoxic environments that vary on a day-to-day basis, but show little acute response in pO2 due to radiation.

Oximetry with Implantable Resonators

The development of implantable resonators will extend applicability of EPR oximetry

• Implantable resonators extend the range of EPR oximetry to deeper tissues.

‒ Improved SNR at all depths‒ Improved spatial localization

• Ability to use optimal materials for added sensitivity

‒ Biocompatible coating contains paramagnetic oximetric material

‒ Ability to remove implants following use

The implantable resonators can be fabricated with great flexibility in configuration.

The coupling loop of the implantable resonator is placed subcutaneously and the detection loop(s) are inserted into the tissue of interest.

Installation for deep tissue measurements of pO2 in the flank of a pig

pO2 Assessment following Sub-lobar Resection with Adjuvant SABR

• NCCC-ACS-IRG funded effort – PI: Philip Schaner (Rad. Onc), Co-I: Erkmen, Williams, Hou, Black

• Proposes that local control of early-stage NSCLC following sub-lobar resection could be improved with adjuvant SABR– Efficacy of radiation is related to tissue pO2, which may be diminished

following surgical disruption. – Tissue pO2 will allow a more complete understanding of the potential of

adjuvant SABR in the post-operative setting.

• Primary Objective: to evaluate the technical feasibility of adjuvant stereotactic ablative radiotherapy (SABR) after sub-lobar resection, and obtain preliminary safety data, in a large animal model (pig)

• Secondary Objective: to evaluate oxygenation using a novel application of EPR in the pulmonary parenchyma, both after sub-lobar resection and post-operative SABR.

• Status: Entering in vivo stage, following procedural evaluation and refinements with cadaveric animals

PROJECT 1: Oxygen measurements in human tumors using EPR oximetry with carbon-based sensors (PI: Gallez)

PROJECT 2: Monitoring of pO2 in human tumors using EPR oximetry with implantable oxygen-sensing probe, OxyChip (PI: Kuppusamy)

PROJECT 3: Oxygen measurements in deep-seated human tumors using EPR oximetry with implantable deep-tissue oxygen sensors (PI: Swartz)

CORE A: Administrative Core

CORE B: Instrument/Resource Core

CORE C: Biostatistics Core

Primary: India inkOxyChipImOS

Primary: India inkOxyChip

ImOS

PROJECT 2

PROJECT 3

Deep-tissue Sensor

PROJECT 1

OxyChip

Carbon

CORES

A,B,C

PPG Layout: Projects & Cores

Oxygen measurements in human tumors using EPR oximetry with carbon-based sensors (PI: Gallez; Sites: Dartmouth, Brussels, Emory, UPenn)

• Make use of already approved materials (India ink in USA and charcoal in Europe) for human measurements

• pO2 measurements in superficial tumors, up to 10 mm depth

• Dartmouth has performed initial clinical measurementsin human tumors

• Studies will be performed to establish:

Intra-tumor and inter-tumor variations of pO2 levels using repeated measurements over a period of weeks in the same tumor

Tumor response to hyperoxygenation treatments

Comparison of EPR pO2 data with those obtained using indirect methods such as PET, MRI, and hypoxia gene signatures

Monitor pO2 dynamics in tumors and normal tissue following radiation therapy

Role of oxygen in radiation-induced fibrosis

Tumors: Head & Neck (oral), Cervical, Breast, Mycosis Fungoides

Synopsis of Project 1

Synopsis of Project 2Monitoring of pO2 in human tumors using EPR oximetry with an implantable oxygen-sensing probe (OxyChip) (PI: Kuppusamy; Sites: Dartmouth, Emory)

• Will use OxyChip – a newly developed, high-sensitive, PDMS-coated implantable/retrievable sensor for EPR oximetry

• Awaiting FDA approval for Investigational Device Exemption

• The sensor has been tested and validated in preclinical animal models

• Repeated pO2 measurements will be made in superficial tumors (up to 10-mm depth)

• Studies will be performed to establish:

Safety and efficacy for repeated measurements of pO2 in human subjects

Monitor temporal changes in tumor pO2 over a period of weeks to months

Tumor response to hyperoxygenation treatments

Comparison of EPR pO2 data with hypoxia gene signatures

Monitor pO2 dynamics in tumor and normal tissue following radiation therapy

Role of oxygen in neo-adjuvant trastuzumab therapy for HER2+ breast cancer

Tumors: Head & Neck (oral), Cervical, Breast (HER2+), Mycosis Fungoides

Synopsis of Project 3Oxygen measurements in human tumors using EPR oximetry with implantable deep-tissue oxygen sensors (PI: Swartz; Sites: Dartmouth, Emory)• Will use implantable deep-tissue oxygen sensors for repeated measurement of

pO2 in deep-sited tumors

• Seek FDA approval for Investigational Device Exemption

• The sensor has been tested and validated in animal models

• pO2 measurements will be made in tumors deeper than 1 cm

• Studies will be performed to establish:

Safety and efficacy for repeated measurements of pO2 in human subjects

Monitor changes in tumor pO2 over a period of weeks to months

Tumor response to hyperoxygenation treatments

Comparison of EPR pO2 data with hypoxia gene signatures

Monitor pO2 dynamics during concurrent treatment with radiation and oxygen-modifying interventions

Skin

External surface-loop resonator of the EPR spectrometer

Biological tissue

Sensory tips

Tissue of interest

Coupling loops of the implantable oxygen sensor

Transmission lines

B

Tumors: Head & Neck (lymph node), high-grade sarcoma, glioblastoma

Clinical EPR Oximetry Summary

• In vivo EPR oximetry has been used successfully in the clinical setting to make repeated non-invasive direct measurements of tissue pO2.– pO2 in peripheral tissues can be measured, including responses

to changes in FIO2 and perfusion

– In most tumors, baseline pO2 values were observed to be quite low (>10mmHg) and the application of inhaled oxygen led to dramatic increases in tumor pO2.

– Tumor pO2 values have varied among the patients studied and over the courses of treatment

– Different responses of tumor pO2 to increased fractions of inhaled oxygen are observed.

• Implantable probes and resonators and are being developed to extend the clinical applicability of EPR oximetry to include deep tumor sites.

• Based on the measurements to date, we believe that it is feasible that in vivo EPR oximetry could be used to monitor tumor pO2 in the clinical setting and guide the optimal application of therapies.

NIH, Norris Cotton Cancer Center, ACS Institutional Research Grants,Dartmouth Center for Clinical and Translational Science (DCCTS)

Department of Radiology, Section of Radiation Oncology

Acknowledgements

InvestigatorsHarold Swartz Lesley JarvisPeriannan Kuppusamy Phil SchanerNadeem Khan Eunice ChenHuagang Hou Alan EastmanAnn Flood Eugene DemidenkoBernard Gallez (UCL, Brussels, Belguim)