oxygen effect and hypoxia

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and Hypoxia DR SAILENDRA SENIOR RESIDENT DEPT OF RADIOTHERAPY MAULANA AZAD MEDICAL COLLEGE Oxyge n effec

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Page 1: Oxygen effect and hypoxia

and Hypoxia

DR SAILENDRASENIOR RESIDENT

DEPT OF RADIOTHERAPYMAULANA AZAD MEDICAL COLLEGE

Oxygen effect

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Road map• What is oxygen effect ?• What is the mechanism ?• What is the timing of action of oxygen ?• What is the concentration of oxygen ?• Different types of hypoxia ?• What is reoxygenation ?• Biology of tumour hypoxia.• Modes to overcome hypoxia effect.

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history• The oxygen effect was observed as early as 1912 in Germany by Swartz, who noted that the skin reaction produced on his forearm by a radium applicator was reduced if the applicator was pressed hard onto the skin.• 1921,it had been noted by Holthusen that Ascaris eggs were relatively resistant to radiation in the absence of oxygen• In England in the 1930s, Mottram explored the question of oxygen in detail.

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Oxygen effect

OER IS LOW AT LOWER DOSES AND A BIT HIGH AT HIGHER DOSES

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THE NATURE OF THE OXYGEN EFFECToxygen enhancement ratio(OER)The ratio of doses administered under hypoxic to aerated conditions needed to achieve the same biologic effect.

Oxygen enhancement ratio = For sparsely ionizing radiations, such as x-rays and γ-rays, the OER at high doses has a value of between 2.5 and 3.5.

Radiation dose in HypoxiaRadiation doseIn Air

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SPARSELY IONIZING

HIGHLY IONIZING

INTERMEDIATE IONIZING

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summary

The oxygen effect is large and important in the case of sparsely ionizing radiations, such as x-rays.

Absent for densely ionizing radiations, such as α-particles.

Intermediate value for fast neutrons.

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THE TIME AT WHICH OXYGEN ACTS AND THE MECHANISM OF THE OXYGEN EFFECT

For the oxygen effect to be observed, oxygen must be present during the radiation exposure or, to be precise, during or within microseconds after the radiation exposure. Howard-Flanders and Moore, 1958; Michael et al., 1973

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The DNA radical are deactivated to its reduced form through reaction with a sulfhydryl (SH) group.

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• The damage produced by free radicals in DNA can be repaired under hypoxia but may be “fixed” (made permanent and irreparable) if molecular oxygen is available.This is known as the oxygen fixation hypothesis.

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THE CONCENTRATION OFOXYGEN REQUIRED

• If the radiosensitivity under extremely anoxic conditions is assigned a value of 1, the relative radiosensitivity is about 3 under well-oxygenated conditions.• Maximum change of sensitivity occurs at the oxygen tension 30 mm Hg.• A further increase of oxygen content has little further effect.

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CHRONIC AND ACUTE HYPOXIA

Chronic hypoxiaLimited diffusion distance of oxygen through tissue

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Chronic Hypoxia• Chronic hypoxia was first described by Thomlinson and Gray in 1955.• By viewing histological sections of fresh specimens of human bronchial carcinomas.

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• Thickness of the sheath of viable tumor cells remains essentially constant.From Thomlinson RH, Gray LH

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• They calculate the distance to which oxygen could diffuse in respiring tissue and came up with a distance of about 150μm which was close enough to the thickness of viable tumor cords on their histologic sections.

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From Thomlinson RH, Gray LH, Br J Cancer. 1955;9:539–549

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Acute Hypoxia• Postulated in the early 1980s by Martin Brown.• Temporary closing or blockage of a particular blood vessel.• Tumor blood vessels open and close in a random fashion, so that different regions of the tumor become hypoxic intermittently.• That is why fractionated radiotherapy can overcome acute hypoxia.

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EXPERIMENTAL DEMONSTRATION OF HYPOXIC CELLS IN A TUMOR

• If the shallow component of the curve is extrapolated backward to cut the surviving-fraction axis, it does so at a survival level of about 1%.• From this, it may be inferred that about 1% of the clonogenic cells in the tumor were deficient in oxygen.Powers WE, Tolmach LJ.survival curve for mouse lymphosarcoma cells irradiated in vivo.Nature. 1963;197:710–711

D0 =1.1 Gy

D0 =2.6 Gy

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TECHNIQUES TO MEASURETUMOR OXYGENATION

•Gold standard•InvasiveOxygen Probe Measurements•Pimonidazole•Noninvasive •IHC from biopsy specimen for carbonic anhydrase IX (CA9) and HIF-1•Differentiate between viable and necrotic tissue

Markers of Hypoxia

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pretreatment frozen biopsy from a patient withcarcinoma of the cervix

• Green – pimonidazole (hypoxic cells)• Red - nuclei that express the HIF-1 (regions with low oxygen tension) • Blue - blood vessels Cancer Res. 2005;65:7259–7266

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REOXYGENATION• Van Putten and Kallman determined the proportion of hypoxic cells in a transplantable sarcoma in the mouse.• Hypoxic cells in the untreated tumor was about 14%.• When groups of tumors were exposed to five daily doses of 1.9 Gy delivered Monday through Friday, the proportion of hypoxic cells was determined on the following Monday to be 18%.• In another experiment, four daily fractions were given Monday through Thursday, and the proportion of hypoxic cells measured the following day, Friday, was found to be 14%.

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summary• Proportion of hypoxic cells in the tumor is about the same at the end of a fractionated radiotherapy regimen.• This phenomenon, by which hypoxic cells become oxygenated after a dose of radiation, is termed Reoxygenation.• If reoxygenation is efficient between dose fractions, the presence of hypoxic cells does not have a significant effect on the outcome of a multifraction regimen.

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MECHANISM OF REOXYGENATION

• As the tumor shrinks in size, surviving cells that previously were beyond the range of oxygen diffusion are closer to a blood supply and so reoxygenate.• This takes several days.• It overcomes chronic hypoxia• Other mechanism is reopening of the blood vessels those were temporarily closed causing acute hypoxia.It occurs within hours of irradiation.

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HYPOXIA AND CHEMORESISTANCE

• Decreased free-radical generation.• Associated with a low pH that can also diminish the activity of some chemotherapy agents.

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HYPOXIA AND TUMOR PROGRESSION

• A clinical study in Germany in the 1990s showed a correlation between local control in advanced carcinoma of the cervix treated by radiotherapy and oxygen-probe measurements.• Specifically, patients in whom the probe measurements indicated pO2s greater than 10 mm Hg did better than those with pO2s less than 10 mm Hg.• This suggested that the presence of hypoxic cells limited the success of radiotherapy.

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• Studies carried out in the United States on patients receiving radiotherapy for soft tissue sarcoma highlighted the correlation between tumor oxygenation and the frequency of distant metastases.• 70% metastasis pO2s < 10 mm Hg • 35% metastasis pO2s > 10 mm Hg.

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RADIORESISTANCECHEMORESISTANCEAGGRESSIVE BEHAVIOUR

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BIOLOGY OF TUMOUR HYPOXIAHYPOXIA-INDUCIBLE FACTOR• Hypoxia-inducible factors (HIFs) are transcription factors that facilitate both oxygen delivery and adaptation to oxygen deprivation by regulating the expression of genes that are involved in many cellular processes including glucose uptake and metabolism, angiogenesis,erythropoiesis, cell proliferation, and apoptosis.• It has an α-sub unit and a β-sub unit.• Three HIFs (HIF-1,-2, and -3) have been identified.

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OXYGEN PRESENT

OXYGEN ABSENT

AngiogenesisErythropoiesisTissue remodelingGlycolysis

MECHANISM OF ACTION OF HIF

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Cancer Mutations that ActivateHypoxia-Inducible Factor

• VHL and PTEN mutation can lead to activation of HIF leading to multiple tumour syndromes like MEN,renal cell carcinoma,retinal hemangioblastoma,CNS neoplasms,pancreatic tumours etcs.

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Roles of Hypoxia Inducible Factor in Tumors

Angiogenesis By activating VEGF-A Tumor Metabolism shift glucose metabolism from an oxidative to glycolytic pathwayTumor Metastasis

promotes metastasis through the transcriptional regulation of key factors such as E-cadherin, lysyl oxidase, and CXCR4 that govern cell adhesion,extracellular matrix formation, and cell migration.Radiotherapy HIF-1 deficient tumors are more sensitive to radiation compared to wildtype tumors.

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UNFOLDED PROTEIN RESPONSE

• Stress -----------accumulation of unfolded proteins-----------cell death• IRE 1--------activate XBP1(transcription factor)----------induce chaperon synthesis--------folding of proteins----------cell survive----------tumourigenesis• PERK(translation inhibitor)--------activated due to hypoxic stress----------inhibit translation or protein synthesis---------misfolded protein synthesis stops---------cell survive----------tumourigenesis

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IRE INDUCE CHAPERON SYNTHESIS CELL SURVIVES

PERK MISFOLDED PROTEIN SYNTHESIS STOPSTUMOURIGENESIS

UNFOLDED PROTEIN RESPONSE

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RADIOSENSITIZING HYPOXIC CELLS

• Hyperbaric oxygen• Chemical radiosensitizers• Hypoxic cytotoxins.• Blood transfusion

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HYPERBARIC OXYGEN • Patients were sealed in chambers filled with pure oxygen raised to a pressure of 3 atmospheres• Patient compliance was a problem• The clinical trials involved small numbers of patients and unconventional fractionation schemes. (mostly large fractions and short duration)• Risk of fire

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• The largest multicenter trials performed by the Medical Research Council in the United Kingdom • Showed a significant benefit both in local control and in survival for patients with carcinoma of the uterine cervix and advanced head and neck cancer.• The trials showed a 6.6% improvement in local control• Increase in late normal tissue damage.

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• Hyperbaric oxygen lost its popularity• Because smoking can decrease tumor oxygenation, it is clearly advisable for patients to give up smoking,at least during radiotherapy

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Properties essential for hypoxic cell sensitizer

Selectively sensitize hypoxic cellsChemically stableHighly soluble in water or lipidsShould be effective at the relatively low daily doses.

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MISONIDAZOLE• Misonidazole has a dramatic effect on tumors in experimental animals.• If x-rays(single dose) are used alone, the dose required to control half of the mouse mammary tumors is 43.8 Gy. This falls to 24.1 Gy if the radiation is delivered 30 minutes after the administration of misonidazole (1 mg/g body weight).This corresponds to an enhancement ratio of 1.8.

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Sheldon PW, Foster JL, Fowler JF. Radiosensitization of C3Hmouse mammary tumours by a 2-nitroimidazole drug. Br J Cancer. 1974;30:560–565

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• This dramatic effect is only seen in single-dose treatments,in contrast to the multifraction regimens common in conventional radiotherapy.• More than 20 trials conducted by RTOG,but none yielded a statistically significant advantage for misonidazole.

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• Danish head and neck cancer trial of misonidazole. • Significant improvement of tumor control by radiotherapy only for males with tumors of the pharynx and depended on hemoglobin status.

Data from Dr. Jens Overgaard

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• Dose limiting toxicity of misonidazole was found to be peripheral neuropathy that progressed to central nervous system toxicity on continuous use.

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Metronidazole↓ Misonidazole: more active, toxicbenefit in subgroups↓Etanidazole: less toxic, no benefit↓Nimorazole: less active, much less toxicbenefit in head and neck cancer

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Overgaard’s Meta-analysis

• Overgaard and colleagues performed a meta-analysis.• They identified 10,602 patients treated in 82 randomized clinical trials involving hyperbaric oxygen, chemical sensitizers, carbogen breathing, or blood transfusions.• Overall, local tumor control was improved by 4.6%, survival by 2.8%, and the complication rate increased by only 0.6%, which was not statistically significant.

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• Head and neck tumors showed the greatest benefit.• It was also concluded that the problem of hypoxia may be marginal in most adenocarcinomas and most important in squamous cell carcinomas.

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HYPOXIC CYTOTOXINS1. Quinone antibiotics e.g Mitomycin C2. Nitroaromatic compounds....high toxicity3. Benzotriazine di-N-oxides...tirapazamine4. Dinitrobenzamide modified nitrogen mustard5. 2-nitroimidazole attached to dibromo isophosphoramide

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TIRAPAZAMINE

Tumor volume as a function of time after various treatments of an SCCVII transplantable mouse carcinomaInt J Radiat Oncol Biol Phys. 1991;20:457–461

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There were no significant differences in Overall survival, failure-free survival, time to locoregional failure, or quality of life.

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toxicitY• Nausea and vomiting• Diarrhea• Weight loss• Skin rash • Muscle cramps• Tinnitus ,acute reversible hearing loss• Visual disturbances• Cardiac ischemia and transient loss of consciousness.• Grade 3 and 4 neutropenia was reported in 4 of 39 patients treated with TPZ at 159 mg/m 2 three times a week for 12 doses with radiotherapy for head and neck cancers.

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TO REMEMBER…..• Hypoxia not only makes tumours radioresistant but aggressive too.• Misonidazole and Tirapazamine are two drugs that are effective against hypoxic cells.• OER is more marked in sparsely ionising elements like x-rays and gamma rays.• Chronic hypoxia is due to decreased Oxygen diffusion.• Acute hypoxia is due to temporary vasospasm.

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Thank

you