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 Dartmouth
Clinical
problem
Parameter to be
measured
Status of measurements in human
subjects
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
Radiationinduced 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
Rationale for using in vivo EPR
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:




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
(a) Clinical EPR spectrometer
(b) Subject positioned for
oximetry using India ink as
reporter
Unique or Exceptional Capabilities of In Vivo EPR
In vivo EPR is sensitive
to a wide array of
physiologic parameters.
• Partial pressure of oxygen (pO2; using oxygensensitive paramagnetic materials)
• Free radicals observed directly or by spin
trapping, including oxygen-, carbon-, and sulfurcentered radicals
• Nitric Oxide (spin trapping)
This information is
observed through
simple changes in the
shapes of the detected
spectra.
• 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 : Oximetry
2500
2000
(nitrogen)
pO , mmHg
1500
Signal amplitude, au
Collisions between the spin
probe and molecular O2
promote relaxation and broaden
the linewidth of the observed
spectrum.
a
1000
500
(air)
0
-500
-1000
412.0
412.5
413.0
Magnetic field, Gauss
LiPc
413.5
414.0
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.
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
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.
Overall goal
 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
Optimize Delivery
Timing of dose
Application of pO2 modifiers
Combination therapies
Initiation of
intervention
*
Design
Select patients
Tumor profiling
pO2 heterogeneity
*
*
Points of pO2
measurement
Improved
Treatment
Survival
Outcome
Therapeutic
Cycle
*
Assessment
pO2 redistribution
Vascular remodeling
Monitor normal tissues
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
30
F = y0+a*x/(b+x)
y0 = 1.1229
25
a = 51.2401
b = 120.4903
LW (G)
20
15
10
5
regression
PO mmHg vs LW
2
0
0
20
40
60
PO2 mmHg
Ink injection and calibrated pO2 response
80
100
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.
V7 - pO2 Measurments during RT
pO2 Variability across Serial Measurements
12
pO2 [mmHg]
10
8
Scalp Pre-treatment
Scalp Post-treatment
6
Neck Pre-treatment
Neck Post-treatment
4
2
0
3/3/2008
3/5/2008
3/7/2008
3/9/2008
3/11/2008
3/13/2008
3/15/2008
Measurement Day
Tumor pO2 was monitored in melanoma metastases at two sites, in the
scalp and neck, during the course of radiation treatment (66 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 DpO2 due to radiation.
Oximetry with Implantable Resonators
The coupling loop of the
implantable resonator is
placed subcutaneously and
the detection loop(s) are
inserted into the tissue of
interest.
The development of implantable
resonators will extend
applicability of EPR oximetry
• Implantable resonators extend
the range of EPR oximetry to
deeper tissues.
The implantable
resonators can be
fabricated with great
flexibility in configuration.
‒ 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
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 sublobar 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
PPG Layout: Projects & Cores
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 deeptissue oxygen sensors (PI: Swartz)
CORE A:
Administrative Core
CORE B:
Instrument/Resource Core
CORE C:
Biostatistics Core
PROJECT
1
Carbon
Primary:
CORE
S
India ink
OxyChip
ImOS
PROJECT A,B,C PROJECT
2
3
OxyChip
Primary:
India ink
OxyChip
ImOS
Deep-tissue Sensor
Synopsis of Project 1
Oxygen measurements in human tumors using EPR oximetry with carbonbased 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 measurements
in 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 2
Monitoring of pO2 in human tumors using EPR oximetry with an implantable oxygensensing probe (OxyChip) (PI: Kuppusamy; Sites: Dartmouth, Emory)
•
Will use OxyChip – a newly developed, high-sensitive, PDMScoated implantable/retrievable sensor for EPR oximetry
•
•
Awaiting FDA approval for Investigational Device Exemption
•
Repeated pO2 measurements will be made in superficial
tumors (up to 10-mm depth)
•
Studies will be performed to establish:
The sensor has been tested and validated in preclinical animal
models
 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 3
Oxygen measurements in human tumors using EPR oximetry with implantable deeptissue oxygen sensors (PI: Swartz; Sites: Dartmouth, Emory)
• Will use implantable deep-tissue oxygen sensors for repeated measurement of
pO2 in deep-sited tumors
•
•
•
•
External surface-loop resonator of the EPR
spectrometer
Seek FDA approval for Investigational Device Exemption
Skin
Biological tissue
Transmission lines
The sensor has been tested and validated in animal models
Coupling loops of the
implantable oxygen
sensor
Sensory tips
pO2 measurements will be made in tumors deeper than 1 cm
Studies will be performed to establish:
Tissue of interest
B
 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
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.
Acknowledgements
NIH, Norris Cotton Cancer Center, ACS Institutional Research Grants,
Dartmouth Center for Clinical and Translational Science (DCCTS)
Department of Radiology, Section of Radiation Oncology
Investigators
Harold Swartz
Lesley Jarvis
Periannan Kuppusamy
Phil Schaner
Nadeem Khan
Eunice Chen
Huagang Hou
Alan Eastman
Ann Flood
Eugene Demidenko
Bernard Gallez (UCL, Brussels, Belguim)