effects of oxygen on radiation therapy
TRANSCRIPT
Effects of Oxygen on Radiation Therapy
February 19, 2009
Zhong Yun, PhD
Department of Therapeutic Radiology, HRT-313Yale University School of MedicineTel: 737-2183; [email protected]
Outline
1. A mechanism of hypoxia signal transduction
2. Oxygen effects on Radiotherapy---OER and factors affecting OER---Reoxygenation---Overcoming tumor hypoxia
---Oxygenation enhancement---Hypoxia sensitizers---Hypoxia toxins---Hypoxia-activated gene therapy
The Hypoxia-Inducible Factor Family
Denko, N.C., Nature Reviews Cancer 8, 705-713 (September 2008)
Mechanisms of hypoxia-inducible factor 1 (HIF1) stabilization
pOpO22
PHD1,2,3
Anaerobic Metabolism
degradation
Angiogenesis
VE
GF
Glut-1
ROS
mt
Metastasis LOX
; E-cadherin
Muscle tissue survival
Cul2
Elong.
HIF1,2
Notch
Embryogastrulation
Oct
-4
Lung Morphogenesis
EG
FR
Ten
asci
n-C
Erythropoesis
EPO
FIH
VHL
DNA Repair
Stem Cell Differentiation
Tu
mor
igen
esis
Tu
mor
igen
esis
Dif
fere
nti
atio
n-D
evel
opm
ent
Dif
fere
nti
atio
n-D
evel
opm
ent
msh2/msh6
Mitoch. O2 consumption PDK1 Proteasome
OH
OH
HIF
O2
Maxwell & Coumenis, 2006
Semenza GL, Drug Discovery Today, 12:853-9, 2007
Semenza GL, Drug Discovery Today, 12:853-9, 2007
Biology of Tumor Hypoxia
HypoxicRegion
Blood Vessel
O2 / DrugConcentration
Gene/Protein Regulation
Increased Glycolysis
Increased Angiogenesis
Increased Genomic Instability
Selection of Apoptosis Resistance
Chemo/Radio-therapy Resistance
Evans and Koch, 01
Hypoxia (EF5, Red) and Proliferation (Ki67, Green)
Fei P. et al. Cancer Cell, 6:597, 2004
Proapoptotic BNIP3L is Induced by p53 under Hypoxia
C.V. Dang, et al. Nature Reviews Cancer 8, 51-56, 2008
The HIF1, HIF2 and MYC protein–protein interaction transcriptional network.
Kinzler KW and Vogelstein B Nature, 379, 1996Graeber TG et al. Nature 379, 1996
Hypoxia Selects for Apoptosis Resistant Clones
Graeber TG, et al. Nature, 379:88, 1996
Hypoxia Selects for Apoptosis Resistant Clones
• Point mutations• Oxidative base damage• DSBs/SSBs• Gene amplification• DNA over-replication
Hypoxia Increases Genome Instability
Genetic instability induced by the tumor microenvironment. Reynolds, T.Y., S. Rockwell, and P.M. Glazer. Cancer Res, 1996. 56(24): p. 5754-7.
Diminished DNA repair and elevated mutagenesis in mammalian cells exposed to hypoxia and low pH. Yuan, J., Narayanan, L., Rockwell, S. and P.M Glazer. Cancer Res, 2000. 60(16): p. 4372-6.
Decreased expression of the DNA mismatch repair gene Mlh1 under hypoxic stress in mammalian cells. Mihaylova, V.T., Bindra, R.S., Yuan, J., Campisi, D, Narayanan, L., Jensen, R., Giordano, F., Johnson, R.S., Rockwell S, and P.M. Glazer. Mol Cell Biol, 2003. 23(9): p. 3265-73.
Mihaylova VT, et al. Mol. Cell. Biol. 24:8504, 2004
Hypoxia Suppresses Mismatch Repair (MMR)
Bindra RS, et al. Mol. Cell. Biol. 24:8504, 2004
Hypoxia/Reoxygenation Represses Homologous Recombination (HR)
Hypoxia/Reoxygenation Represses Homologous Recombination (HR)
Bindra RS, et al. Mol. Cell. Biol. 24:8504, 2004
Oxygen Influences Biological Effects of Radiation
1. Less damage to tightly pressed skin, Swartz 1912,
2. Radiation inhibited germination of vegetable seeds only in the presence of O2, Petry 1923
3. Oxygen effect on tumor radiosensitivity championed by Mottram 1930s
4. 1st quantitative study on O2 effects on radiation-inducedgrowth inhibition of broad bean, Gray 1953
5. O2 levels decrease in respiring tumor cells located away from blood vessels, Tomlinson and Gray 1955
Brown JM & Wilson WR, 2004
Ability of ionizing radiation to kill cells is highly dependent on pO2
---Tomlinson and Gray , 1950s
Oxygen Enhancement Ratio (OER)
The ratio of HYPOXIC to AEROBIC IR doses needed to achieve the SAME biological effects.
Dose (Gy)
0 10 20 30
Su
rviv
ing
Fra
ctio
n
0.01
0.1
1
AerobicHypoxic
1/e1/e
OER =D0 (hypoxic)
D0 (aerobic)= 3.22
D0D0
OER =D0 (hypoxic)
D0 (aerobic)
Radiosensitivity and pO2
1.0
2.0
3.0
0 20 40 60 80155 760 2280
Air 100%O2
HBO(3 ATA)
VenousBlood
Rel
ati
ve
Rad
iose
nsi
tivi
ty
Oxygen tension (mmHg)
(Rockwell et al.)
Half maximum sensitivity = 3 mmHg or 0.5% O2
Kirkpatrick JP, et al. Intl. J. Rad. Oncol. Biol. Phys. 59:822, 2004
CHO Cells
Re
lati
ve
Ra
dio
se
ns
itiv
ity
The Oxygen Fixation Hypothesis
X-rays + H2O H + H3O+ + OH
HO2
H2O2
O2-
H2O+ + e-H2O OH
R
RHRO2
O2
H+
O2
O2
t1/2: 10-10 sec for ion pairs; 10-5 sec for free radicals
Non-repairable
Parameters Affecting OER
1. Nature of cellular sensitivity
2. Nature of radiation (x-ray, neutron, alpha particle, or LET)
3. pO2
5. Cell Cycle ( cells in S-phase are more resistant.)
4. Time of oxygen presence
1. CellularSensitivity
Resistant Cells
High OER
Sensitive Cells
Low OER
2. Nature of Radiation X-rays
neutrons -particles
LET = 2.0
LET and OER
60
4. Oxygen effect is time-dependent
1. O2 needs not to be present DURING radiation
2. O2 has to be present WITHIN milliseconds after radiation
Radiation Ion pairs Free Radicals
T = 10-10 sec T = 10-5 sec (0.01 msec)
Time delay is determined by lifetime of free radicals
Sinclair W and Morton R, Radiat. Res. 29:450, 1966Sinclair W, Radiat. Res. 33:620, 1968
5. Cell Cycle and Radiation Sensitivity
Most Sensitive Phase:G2/M
Most Resistant Phase:Late SEarly G1
OER varies slightly during cell cycle
S phase(OER = 2.8-2.9)
G2/M phase(OER = 2.3-2.4)
G1 phase >>
OER remains unchanged for ALL dose levels in a given phase of cell cycle.
ReoxygenationNormoxic
Hypoxic
Necrotic
X-ray
Recovery
X-ray
X-ray
X-ray
Hypoxic Fraction
No change
Decreased
Increased
Factors Affecting Reoxygenation
2. Clearance of dead cells by macrophages, etc.
1. Rate of angiogenesis.
3. Inflammatory response
4. Tissue remodeling.
Mouse mammary Carcinoma
Rat sarcoma
Mouse fibrosarcoma
Mouse osteosarcoma
Mouse fibrosarcoma
Variable Rates of Reoxygenation
Strategies to Overcome Tumor Hypoxia
1. Radiation Dose Fractionation- Limiting damage to normal tissue- Reoxygenation of hypoxic tumor cells
2. Improve Tumor Oxygenation- Hyperbaric oxygen breathing- Compounds to improve oxygenation (Motexafin Gadolinium or MGd)- Allosteric modifiers of hemoglobin-oxygen binding such as RSR13
(to reduce O2 affinity for efficient O2 discharge)
3. Hypoxic Cell Sensitizers (Nitroimidazoles with high electron affinity)- Misonidazole: more active, but toxic (peripheral neuropathy)- Etanidazole: less toxic, but no benefit- Nimorazole: less active, much less toxic (benefit in H&N cancer--Danish study)
4. Hypoxia Cytotoxins (Bioreductive agents that selectively kill hypoxic cells)- Tirapazamine: high Hypoxic toxicity ratio- Mitomycin C: low Hypoxic toxicity ratio
MGd Improves Tumor Oxygenation
0 10 20 30 40 50
Rel
ativ
e F
req
uen
cy (
%)
0
10
20
30
40
50
60
70
0 10 20 30 40 500
10
20
30
40
50
60
70
Oxygen Partial Pressure (mmHg)
0 10 20 30 40 50
Rel
ativ
e F
req
uen
cy (
%)
0
10
20
30
40
50
60
70
Oxygen Partial Pressure (mmHg)
0 10 20 30 40 500
10
20
30
40
50
60
70
(A) Control
(C) 40 mol/kg MGd - 6 Hours
(B) Mannitol Control - 6 Hours
(D) 40 mol/kg MGd - 8 Hours *
*
* p < 0.01
**
**
E. Donnelly & S. Rockwell, 2005
MGd: Clinical Tumor Oxygenation Observations
• Effects of MGd on tumor oxygenation were measured during Phase I head and neck cancer clinical trials– Median Tumor pO2
• Before treatment: 6.8 mmHg• 1 mg/kg MGd, one week: 27.0 mmHg
– Improved Tumor Response• 8 of 9 patients experienced complete tumor response• 7 patients still in remission after 9 months
D.M. Brizel, Duke University
Brown JM & Wilson WR, 2004
Activation of Hypoxia-Selective Prodrugs
JM Brown, Stanford Univ.
Topo II
TPZ Preferentially Kills Hypoxic Cells
Brown JM & Wilson WR, 2004
A Strategy for Hypoxia-Activated Gene Therapy
Semenza GL, Drug Discovery Today, 12:853-9, 2007