Targeting Tumor Perfusion and Oxygenation to Improve the Outcome of Anticancer Therapy1

Radiotherapy and chemotherapy are widespread clinical modalities for cancer treatment. Among other biological influences, hypoxia is a main factor limiting the efficacy of radiotherapy, primarily because oxygen is involved in the stabilization of the DNA damage caused by ionizing radiations. Radiobiological hypoxia is found in regions of rodent and human tumors with a tissue oxygenation level below 10 mmHg at which tumor cells become increasingly resistant to radiation damage. Since hypoxic tumor cells remain clonogenic, their resistance to the treatment strongly influences the therapeutic outcome of radiotherapy. There is therefore an urgent need to identify adjuvant treatment modalities aimed to increase tumor pO2 at the time of radiotherapy. Since tumor hypoxia fundamentally results from an imbalance between oxygen delivery by poorly efficient blood vessels and oxygen consumption by tumor cells with high metabolic activities, two promising approaches are those targeting vascular reactivity and tumor cell respiration. This review summarizes the current knowledge about the development and use of tumor-selective vasodilators, inhibitors of tumor cell respiration, and drugs and treatments combining both activities in the context of tumor sensitization to X-ray radiotherapy. Tumor-selective vasodilation may also be used to improve the delivery of circulating anticancer agents to tumors. Imaging tumor perfusion and oxygenation is of importance not only for the development and validation of such combination treatments, but also to determine which patients could benefit from the therapy. Numerous techniques have been developed in the preclinical setting. Hence, this review also briefly describes both magnetic resonance and non-magnetic resonance in vivo methods and compares them in terms of sensitivity, quantitative or semi-quantitative properties, temporal, and spatial resolutions, as well as translational aspects

Chapter 13 Targeting Tumor Perfusion and Oxygenation Modulates Hypoxia and Cancer Sensitivity to Radiotherapy and Systemic Therapies

Hypoxia, a partial pressure of oxygen (pO2) below physiological needs, is a limiting factor affecting the efficiency of radiotherapy. Indeed, the reaction of reactive oxygen species (ROS, produced by water radiolysis) with DNA is readily reversible unless oxygen stabilizes the DNA lesion. While normal tissue oxygenation is around 40 mm Hg, both rodent and human tumors possess regions of tissue oxygenation below 10 mm Hg, at which tumor cells become increasingly resistant to radiation damage (radiobiological hypoxia) (Gray, 1953). Because of this so-called “oxygen enhancement effect”, the radiation dose required to achieve the same biologic effect is about three times higher in the absence of oxygen than in the presence of normal levels of oxygen (Gray et al., 1953; Horsman & van der Kogel, 2009). Hypoxic tumor cells, which are therefore more resistant to radiotherapy than well oxygenated ones, remain clonogenic and contribute to the therapeutic outcome of fractionated radiotherapy (Rojas et al., 1992)

Metabolite changes in HT-29 xenograft tumors following HIF-1 alpha inhibition with PX-478 as studied by MR spectroscopy in vivo and ex vivo

The hypoxia-inducible transcription factor (HIF-1 alpha) plays a central role in tumor development. PX-478 is an experimental anti-cancer drug known to inhibit HIF-1 alpha in experimental tumors. The purpose of this study was to identify MRS-visible metabolic biomarkers for PX-478 response prior to phase I/II clinical trials. Single-voxel in vivo localized H-1 spectra were obtained from HT-29 tumor xenografts prior and up to 24 h after treatment with a single dose of PX-478. Profiles of water-soluble and lipophilic metabolites were also examined ex vivo with both H-1 and P-31 spectroscopy for peak identification and to interrogate the underlying biochemistry of the response. The total choline (tCho) resonance was significantly decreased in vivo 12 and 24 h following treatment with PX-478 and this was confirmed with high-resolution 1 H and P-31 MRS. In non-aqueous extracts, significant reductions in cardiolipin, PtdEtn (phosphatidylethanolamine) and PtdI (phosphatidylinositol) were seen in response to PX-478. Although there were trends to a decrease in lactate (and lipid) resonances in vivo and ex vivo, these changes were not significant. This is in contrast to inhibition of in vitro glucose consumption and lactate production by PX-478 in HT-29 cells. The significant and robust change in tCho has identified this as a potential 1H MRS-visible biomarker for drug response in vivo while high-resolution spectroscopy indicated that GPC, PC, myol, PE, GPE, CL, PtdEtn and Ptdl are potential ex vivo response biomarkers. Copyright (c) 2005 John Wiley & Sons, Ltd

The acidic tumor microenvironment promotes the reconversion of nitrite into nitric oxide: towards a new and safe radiosensitizing strategy

PURPOSE: The biological status of nitrite recently evolved from an inactive end product of nitric oxide catabolism to the largest intravascular and tissue storage of nitric oxide (NO). Although low partial O(2) pressure favors enzymatic reconversion of nitrite into NO, low pH supports a nonenzymatic pathway. Because hypoxia and acidity are characteristics of the tumor microenvironment, we examined whether nitrite injection could preferentially lead to NO production in tumors and influence response to treatments. EXPERIMENTAL DESIGN: The effects of nitrite were evaluated on arteriole vasorelaxation, tumor cell respiration and tumor blood flow, oxygenation, and response to radiotherapy. RESULTS: We first showed that a small drop in pH (-0.6 pH unit) favored the production of bioactive NO from nitrite by documenting a higher cyclic guanosine 3',5'-monophosphate-dependent arteriole vasorelaxation. We then documented that an i.v. bolus injection of nitrite to tumor-bearing mice led to a transient increase in partial O(2) pressure in tumor but not in healthy tissues. Blood flow measurements failed to reveal an effect of nitrite on tumor perfusion, but we found that O(2) consumption by nitrite-exposed tumor cells was decreased at acidic pH. Finally, we showed that low dose of nitrite could sensitize tumors to radiotherapy, leading to a significant growth delay and an increase in mouse survival (versus irradiation alone). CONCLUSIONS: This study identified low pH condition (encountered in many tumors) as an exquisite environment that favors tumor-selective production of NO in response to nitrite systemic injection. This work opens new perspectives for the use of nitrite as a safe and clinically applicable radiosensitizing modality

The thioredoxin-1 inhibitor 1-methylpropyl 2-imidazolyl disulfide (PX-12) decreases vascular permeability in tumor xenografts monitored by dynamic contrast enhanced magnetic resonance Imaging

Purpose: The purpose of this study was to use dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) to measure changes in tumor xenograft permeability produced by the antitumor thioredoxin-1 (Trx-1) inhibitor 1-methylpropyl 2-imidazolyl disulfide (PX-12) and to assess the relationship to Trx-1 and vascular endothelial growth factor (VEGF) levels. Experimental Design: DCE-MRI was used to monitor the dynamics of gadolinium-diethylenetriaminepentaacetic acid coupled bovine serum albumin as a macromolecular contrast reagent to measure hemodynamic changes in HT-29 human colon xenografts in immunodeficient mice treated with PX-12. Blood vessel permeability was estimated from the slope of the enhancement curves, and tumor vascular volume fraction from the ordinate. Tumor Trx-1 and VEGF was also measured. Results: PX-12 caused a rapid 63% decrease in the average tumor blood vessel permeability within 2 hours of administration. The decrease lasted 24 hours and had returned to pretreatment values by 48 hours. The changes in vascular permeability were not accompanied by alterations in average tumor vascular volume fraction. There was a decrease in tumor and tumor-derived VEGF in plasma at 24 hours after treatment with PX-12, but not at earlier time points. However, tumor redox active Trx-1 showed a rapid decline within 2 hours following PX-12 administration that was maintained for 24 hours. Conclusion: The rapid decrease in tumor vascular permeability caused by PX-12 administration coincided with a decrease in tumor redox active Trx-1 and preceded a decrease in VEGF. DCE-MRI responses to PX-12 in patients of Trx-1 inhibition at early time points and decreased VEGF at later times. may be useful to follow tumor response and even therapeutic benefit

Impact of myo-inositol trispyrophosphate (ITPP) on tumour oxygenation and response to irradiation in rodent tumour models.

Tumour hypoxia is a well-established factor of resistance in radiation therapy (RT). Myo-inositol trispyrophosphate (ITPP) is an allosteric effector that reduces the oxygen-binding affinity of haemoglobin and facilitates the release of oxygen by red blood cells. We investigated herein the oxygenation effect of ITPP in six tumour models and its radiosensitizing effect in two of these models. The evolution of tumour pO upon ITPP administration was monitored on six models using 1.2 GHz Electron Paramagnetic Resonance (EPR) oximetry. The effect of ITPP on tumour perfusion was assessed by Hoechst staining and the oxygen consumption rate (OCR) in vitro was measured using 9.5 GHz EPR. The therapeutic effect of ITPP with and without RT was evaluated on rhabdomyosarcoma and 9L-glioma rat models. ITPP enhanced tumour oxygenation in six models. The administration of 2 g/kg ITPP once daily for 2 days led to a tumour reoxygenation for at least 4 days. ITPP reduced the OCR in six cell lines but had no effect on tumour perfusion when tested on 9L-gliomas. ITPP plus RT did not improve the outcome in rhabdomyosarcomas. In 9L-gliomas, some of tumours receiving the combined treatment were cured while other tumours did not benefit from the treatment. ITPP increased oxygenation in six tumour models. A decrease in OCR could contribute to the decrease in tumour hypoxia. The association of RT with ITPP was beneficial for a few 9L-gliomas but was absent in the rhabdomyosarcomas

Dynamic Contrast-Enhanced and Diffusion MRI Show Rapid and Dramatic Changes in Tumor Microenvironment in Response to Inhibition of HIF-1α Using PX-478

PX-478 is a new agent known to inhibit the hypoxia-responsive transcription factor, HIF-1α, in experimental tumors. The current study was undertaken in preparation for clinical trials to determine which noninvasive imaging endpoint(s) is sensitive to this drug's actions. Dynamic contrast-enhanced (DCE) and diffusion-weighted (DW) magnetic resonance imaging (MRI) were used to monitor acute effects on tumor hemodynamics and cellularity, respectively. Mice bearing human xenografts were treated either with PX-478 or vehicle, and imaged over time. DW imaging was performed at three b values to generate apparent diffusion coefficient of water (ADCw) maps. For DCE-MRI, a macromolecular contrast reagent, BSA-Gd-DTPA, was used to determine vascular permeability and vascular volume fractions. PX-478 induced a dramatic reduction in tumor blood vessel permeability within 2 hours after treatment, which returned to baseline by 48 hours. The anti-VEGF antibody, Avastin, reduced both the permeability and vascular volume. PX-478 had no effect on the perfusion behavior of a drug-resistant tumor system, A-549. Tumor cellularity, estimated from ADCw, was significantly decreased 24 and 36 hours after treatment. This is the earliest significant response of ADC to therapy yet reported. Based on these preclinical findings, both of these imaging endpoints will be included in the clinical trial of PX-478

A Fast Hydrogen Sulfide-Releasing Donor Increases the Tumor Response to Radiotherapy

Hydrogen sulfide (H2S) is the last gaseous transmitter identified in mammals, and previous studies have reported disparate conclusions regarding the implication of H2S in cancer progression. In the present study, we hypothesized that sodium hydrosulfide (NaHS), a fast H2S-releasing donor, might interfere with the mitochondrial respiratory chain of tumor cells, increase tumor oxygenation, and potentiate the response to irradiation. Using electron paramagnetic resonance (EPR) oximetry, we found a rapid increase in tumor pO2 after NaHS administration (0.1 mmol/kg) in two human tumor models (breast MDA-MB-231 and cervix SiHa), an effect that was due to a decreased oxygen consumption and an increased tumor perfusion. Tumors irradiated 15 minutes after a single NaHS administration were more sensitive to irradiation compared with those that received irradiation alone (increase in growth delay by 50%). This radiosensitization was due to the oxygen effect, as the increased growth delay was abolished when temporarily clamped tumors were irradiated. In contrast, daily NaHS injection (0.1 mmol/kg/day for 14 days) did not provide any effect on tumor growth in vivo. To understand these paradoxical data, we analyzed the impact of external factors on the cellular response to NaHS. We found that extracellular pH had a dramatic effect on the cell response to NaHS, as the proliferation rate (measured in vitro by BrdU incorporation) was increased at pH = 7.4, but decreased at pH = 6.5. Overall, our study highlights the complex role of environmental components in the response of cancer cells to H2S and suggests a new approach for the use of H2S donors in combination with radiotherapy

Mechanism of reoxygenation after antiangiogenic therapy using SU5416 and its importance for guiding combined antitumor therapy. Cancer Res

Abstract Emerging preclinical studies support the concept of a transient ''normalization'' of tumor vasculature during the early stage of antiangiogenic treatment, with possible beneficial effects on associated radiotherapy or chemotherapy. One key issue in this area of research is to determine whether this feature is common to all antiangiogenic drugs and whether the phenomenon occurs in all types of tumors. In the present study, we characterized the evolution of the tumor oxygenation (in transplantable liver tumor and FSAII tumor models) after administration of SU5416, an antagonist of the vascular endothelial growth factor receptor. SU5416 induced an early increase in tumor oxygenation [measured by electronic paramagnetic resonance (EPR)], which did not correlate with remodeling of the tumor vasculature (assessed by CD31 labeling using immunohistochemistry) or with tumor perfusion (measured by dynamic contrast enhanced-magnetic resonance imaging). Inhibition of mitochondrial respiration (measured by EPR) was responsible for this early reoxygenation. Consistent with these unique findings in the tumor microenvironment, we found that SU5416 potentiated tumor response to radiotherapy but not to chemotherapy. In addition to the fact that the characterization of the tumor oxygenation is essential to enable correct application of combined therapies, our results show that the long-term inhibition of oxygen consumption is a potential novel target in this class of compounds