Arsenic trioxide treatment decreases the oxygen consumption rate of tumor cells and radiosensitizes solid tumors.

Arsenic trioxide (As(2)O(3)) is an effective therapeutic against acute promyelocytic leukemia and certain solid tumors. Because As(2)O(3) inhibits mitochondrial respiration in leukemia cells, we hypothesized that As(2)O(3) might enhance the radiosensitivity of solid tumors by increasing tumor oxygenation [partial pressure of oxygen (pO(2))] via a decrease in oxygen consumption. Two murine models of radioresistant hypoxic cancer were used to study the effects of As(2)O(3). We measured pO(2) and the oxygen consumption rate in vivo by electron paramagnetic resonance oximetry and (19)fluorine-MRI relaxometry. Tumor perfusion was assessed by Patent blue staining. In both models, As(2)O(3) inhibited mitochondrial respiration, leading to a rapid increase in pO(2). The decrease in oxygen consumption could be explained by an observed decrease in glutathione in As(2)O(3)-treated cells, as this could increase intracellular reactive oxygen species that can disrupt mitochondrial membrane potential. When tumors were irradiated during periods of As(2)O(3)-induced augmented oxygenation, radiosensitivity increased by 2.2-fold compared with control mice. Notably, this effect was abolished when temporarily clamped tumors were irradiated. Together, our findings show that As(2)O(3) acutely increases oxygen consumption and radiosensitizes tumors, providing a new rationale for clinical investigations of As(2)O(3) in irradiation protocols to treat solid tumors.

In vivo mapping of tumor oxygen consumption using (19)F MRI relaxometry.

Recently, we have developed a new electron paramagnetic resonance (EPR) protocol in order to estimate tissue oxygen consumption in vivo. Because it is crucial to probe the heterogeneity of response in tumors, the aim of this study was to apply our protocol, together with (19)F MRI relaxometry, to the mapping of the oxygen consumption in tumors. The protocol includes the continuous measurement of tumor po(2) during the following respiratory challenge: (i) basal values during air breathing; (ii) increasing po(2) values during carbogen breathing until saturation of tissue with oxygen; (iii) switching back to air breathing. We have demonstrated previously using EPR oximetry that the kinetics of return to the basal value after oxygen saturation are mainly governed by tissue oxygen consumption. This challenge was applied in hyperthyroid mice (generated by chronic treatment with L-thyroxine) and control mice, as hyperthyroidism is known to dramatically affect the oxygen consumption rate of tumor cells. Our recently developed snapshot inversion recovery MRI fluorocarbon oximetry technique allowed the po(2) return kinetics to be measured with a high temporal resolution. The kinetic constants (i.e. oxygen consumption rates) were higher for tumors from hyperthyroid mice than from control mice, data that are consistent with our previous EPR study. The corresponding histograms of the (19)F MRI data showed that the kinetic constants displayed a shift to the right for the hyperthyroid group, indicating a higher oxygen consumption in these tumors. The color maps showed a large heterogeneity in terms of oxygen consumption rate within a tumor. In conclusion, (19)F MRI relaxometry allows the noninvasive mapping of the oxygen consumption in tumors. The ability to assess the heterogeneity of tumor response is critical in order to identify potential tumor regions that might be resistant to treatment and therefore produce a poor response to therapy.

Captopril and S-nitrosocaptopril as potent radiosensitizers: Comparative study and underlying mechanisms.

In an effort to improve the issue of radiotherapy treatments, we tested whether S-nitrosocaptopril, a molecule combining a NO donor and an angiotensin converting enzyme inhibitor (ACE inhibitor), could temporarily improve the hemodynamic status of experimental tumors. We monitored the effect of S-nitrosocaptopril in TLT tumors using non rinvasive magnetic resonance techniques. We identified a time window during which tumor oxygenation was improved, as a result of a combined effect on tumor blood flow and oxygen consumption. Consequently, the administration of S-nitrosocaptopril contributed to the increase in efficacy of radiation therapy, an effect that was not observed with captopril alone.

Comparison of methods for measuring oxygen consumption in tumor cells in vitro.

The oxygen consumption rate of tumor cells affects tumor oxygenation and response to therapies. Highly sensitive methods for determining cellular oxygen consumption are, therefore, needed to identify treatments that can modulate this parameter. We compared the performances of three different methods for measuring cellular oxygen consumption: electron paramagnetic resonance (EPR) oximetry, the Clark electrode, and the MitoXpress fluorescent assay. To compare the assays, we used K562 cells in the presence of rotenone and hydrocortisone, compounds that are known to inhibit the mitochondrial electron transport chain to different extents. The EPR method was the only one that could identify both rotenone and hydrocortisone as inhibitors of tumor cell oxygen consumption. The Clark electrode and the fluorescence assay demonstrated a significant decrease in cellular oxygen consumption after administration of the most potent inhibitor (rotenone) but failed to show any significant effect of hydrocortisone. EPR oximetry is, therefore, the most sensitive method for identifying inhibitors of oxygen consumption on cell assays, whereas the Clark electrode offers the unique opportunity to add external compounds during experiments and still shows great sensitivity in studying enzyme and chemical reactions that consume oxygen (non-cell assays). Finally, the MitoXpress fluorescent assay has the advantage of a high-sample throughput and low bulk requirements but at the cost of a lower sensitivity.

A New EPR oximetry protocol to estimate the tissue oxygen consumption in vivo.

Abstract The oxygen consumption rate in tumors affects tumor oxygenation and response to therapies. A new EPR method was developed to measure tissue oxygen consumption non-invasively. The protocol was based on the measurement of pO(2) during a carbogen challenge. The following sequence was used: (1) basal value during air breathing; (2) saturation of tissue with oxygen by carbogen breathing; (3) switch back to air breathing. The assumption was that the kinetics of the return to the basal value after oxygen saturation would be governed mainly by tissue oxygen consumption. This challenge was applied in hyperthyroid mice (generated by chronic treatment with l-thyroxine) and control mice because hyperthyroidism is known to dramatically affect the oxygen consumption rate of tumor and muscle cells. Muscle and tumor cells from the hyperthyroid mice consumed oxygen faster than muscle and tumor cells from the control mice, which is consistent with the results of in vitro studies. Tumor perfusion was not affected by the treatment with l-thyroxine. This method gives an index that may reasonably be ascribed to the local oxygen consumption and has the unique advantage of being adaptable to in vivo studies.

Targeting lactate-fueled respiration selectively kills hypoxic tumor cells in mice.

Tumors contain oxygenated and hypoxic regions, so the tumor cell population is heterogeneous. Hypoxic tumor cells primarily use glucose for glycolytic energy production and release lactic acid, creating a lactate gradient that mirrors the oxygen gradient in the tumor. By contrast, oxygenated tumor cells have been thought to primarily use glucose for oxidative energy production. Although lactate is generally considered a waste product, we now show that it is a prominent substrate that fuels the oxidative metabolism of oxygenated tumor cells. There is therefore a symbiosis in which glycolytic and oxidative tumor cells mutually regulate their access to energy metabolites. We identified monocarboxylate transporter 1 (MCT1) as the prominent path for lactate uptake by a human cervix squamous carcinoma cell line that preferentially utilized lactate for oxidative metabolism. Inhibiting MCT1 with alpha-cyano-4-hydroxycinnamate (CHC) or siRNA in these cells induced a switch from lactate-fueled respiration to glycolysis. A similar switch from lactate-fueled respiration to glycolysis by oxygenated tumor cells in both a mouse model of lung carcinoma and xenotransplanted human colorectal adenocarcinoma cells was observed after administration of CHC. This retarded tumor growth, as the hypoxic/glycolytic tumor cells died from glucose starvation, and rendered the remaining cells sensitive to irradiation. As MCT1 was found to be expressed by an array of primary human tumors, we suggest that MCT1 inhibition has clinical antitumor potential.