Allosteric Modifier Of Hemoglobin Research Articles

Modulation of red blood cell oxygen affinity with a novel allosteric modifier of hemoglobin is additive to the Bohr effect

PurposeThe Bohr effect describes hemoglobin's affinity for oxygen dependent on solution pH. Within pH range 6.0–8.5, hemoglobin's oxygen affinity decreases with decreasing pH. This results in increased oxygen delivery to metabolically active, acidic tissues and improved oxygen uptake in basic regions including lung tissue. Myo-Inositol tripyrophosphate (ITPP) translocates the erythrocyte membrane and allosterically modifies hemoglobin (Hb). We tested the hypothesis that ITPP does not abrogate the Bohr effect. MethodsExperiments were conducted to determine the effect of increasing concentrations of ITPP on P50 with varying pH. We incubated 10 mL red blood cells at 37 °C for 1 h with ITPP concentrations from 0 to 240 mM. The Clark oxygen electrode (Hemox-Analyzer; TCS Scientific, New Hope, PA) determined oxygen affinity of each sample, in triplicate, using buffers pH 6.8, 7.4, and 7.6. A mixed linear regression model with fixed effects for ITPP concentration and pH was used. ResultsIncreasing ITPP concentration and decreasing pH increased P50 (p < 0.0001 for ITPP concentration, p < 0.0001 for pH). ITPP modulated increased P50 in normal pH (7.4) and acidic condition pH (6.8); with no effect at alkaline pH (7.6). ConclusionThe Bohr effect is conserved, with ITPP augmenting the decreased oxygen affinity seen with tissue acidosis, while not affecting oxygen affinity in conditions similar to a pulmonary microenvironment.

Abstract 4371: RRx-001 modulates intratumor blood flow in SCCVII and U87 tumors

Abstract The tumor-specific provascular effects of the anti-cancer agent RRx-001, a novel nitrogen oxide donor and allosteric hemoglobin modifier in Phase 1 clinical trials were investigated using contrast enhanced ultrasound (CEUS) and whole tissue section quantitative immunohistological staining. Changes in tumor blood flow by CEUS in SCCVII tumor bearing mice following RRx-001 (3, 6, 12 mg/kg) were dose and time dependent, reaching maximum perfusion 6 h after treatment, returning to baseline within 72 h. Microregional effects of RRx-001 (15 mg/kg) on tumor blood flow and oxygenation by immunohistological staining were studied in mouse SCCVII and human U87 tumors. SCCVII tumors exhibited regions of intermittent perfusion exemplified by co-localization of vessels with the hypoxia marker pimonidazole commonly occurring throughout the tissue. Only 21% of vessels showed perfusion after a bolus dose of the perivascular stain DiOC7. This increased to 28% 12 h post RRx-001. Paradoxically, an increase in tissue oxygenation was not seen while modestly increased pimonidazole binding was observed. However, preliminary radiobiological assessment suggested tumors were radiosensitized when irradiated 10 min after 12 mg/kg RRx-001. In the U87 tumor a decrease in vessel perfusion at 90 min post RRx-001 administration was observed. Blood flow over large areas of treated tumors was completely shut down. These areas stained positive for CD31 and tissue did not appear necrotic. Analysis indicated 25% reduction in the fraction of perfused vessels and un-perfused tissue by area at 90 min returning to near normal levels at 12 h. There was no significant increase in the fraction of necrosis at 12 h suggesting that the large un-perfused regions seen at 90 min had recovered rather than become necrotic. No significant change in tumor hypoxia was seen at 90 min or 12 h. For both tumor types, RRx-001 appeared to cause the loss of perfusion in large regions of the tumor however, at the 12 h time point, both tumor types showed increase in vessel perfusion but no significant decrease in hypoxia. These data suggest a redistribution of blood flow in both tumor types while differences between the tumors were related to tumor architecture and distribution of α-SMA. RRx-001 shows promise for short-term blood flow redistribution in tumors with a pericyte-rich vasculature with contractile proteins such as α-SMA. Expression of α-SMA in tumor vasculature could therefore be useful for predicting tumor response to RRx-001. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4371. doi:1538-7445.AM2012-4371

Reduction in Hemoglobin–Oxygen Affinity Results in the Improvement of Exercise Capacity in Mice With Chronic Heart Failure

This study examined whether a reduction in hemoglobin-oxygen affinity improves exercise capacity in mice with heart failure. Exercise intolerance is a major determinant of quality of life in patients with chronic heart failure. One of the major goals of the treatment for chronic heart failure is to improve quality of life. Four weeks after left coronary ligation, we transplanted bone marrow cells isolated from the transgenic mice expressing a hemoglobin variant with low oxygen affinity, Presbyterian, into the lethally irradiated mice with heart failure or administered a synthetic allosteric modifier of hemoglobin. The mice were then exercised on a treadmill. Four weeks after the left coronary artery ligation, mice showed cardiac dysfunction and chamber dilation, which were characteristics of heart failure. The transplantation led to a reduction in hemoglobin-oxygen affinity and an increase in oxygen supply for skeletal muscle without changes in muscle properties. The transplanted mice showed improved running performance on a treadmill despite impaired cardiac contractility. Furthermore, administration of the synthetic allosteric modifier of hemoglobin showed a similar effect. Allosteric modification of hemoglobin represents a therapeutic option for improving exercise capacity in patients with chronic heart failure. One mechanism of improvement in exercise capacity is enhanced oxygen delivery in the skeletal muscle.

Improved Survival, Quality of Life, and Quality-Adjusted Survival in Breast Cancer Patients Treated With Efaproxiral (Efaproxyn) Plus Whole-Brain Radiation Therapy for Brain Metastases

To determine whether efaproxiral, an allosteric modifier of hemoglobin, improves quality of life and quality of survival in patients with primary breast cancer and brain metastases when used as an adjunct to whole-brain radiation therapy (WBRT). Patients with brain metastases from breast cancer were randomly assigned to receive WBRT and either efaproxiral or no efaproxiral. The primary endpoint for this analysis was quality of life and quality-adjusted survival. Quality of life was assessed prior to initiation of WBRT and periodically in follow-up using the Spitzer Quality of Life Index (SQLI). A subgroup of 106 eligible breast cancer patients with baseline SQLI were randomized into this study and represent the target population discussed in this report. Treatment, age, and SQLI were significant predictors of survival. The addition of efaproxiral to WBRT reduced the death rate by 46% (P = 0.0086). Quality of life was improved in the WBRT + efaproxiral arm compared with the WBRT alone arm (P = 0.019). Quality-adjusted survival was statistically significantly improved by the addition of efaproxiral to WBRT (P = 0.001). Survival, quality of life, and quality-adjusted survival were all improved in breast cancer patients with brain metastases receiving efaproxiral and WBRT compared with those receiving WBRT alone.

The Effects of Efaproxyn™ (Efaproxiral) on Subcutaneous RIF-1 Tumor Oxygenation and Enhancement of Radiotherapy-Mediated Inhibition of Tumor Growth in Mice

Efaproxiral, an allosteric modifier of hemoglobin, reduces hemoglobin-oxygen binding affinity, facilitating oxygen release from hemoglobin, which is likely to increase tissue pO(2). The purpose of this study was to determine the effect of efaproxiral on tumor oxygenation and growth inhibition of RIF-1 tumors that received X radiation (4 Gy) plus oxygen breathing compared to radiation plus oxygen plus efaproxiral daily for 5 days. Two lithium phthalocyanine (LiPc) deposits were implanted in RIF-1 tumors in C3H mice for tumor pO(2) measurements using EPR oximetry. Efaproxiral significantly increased tumor oxygenation by 8.4 to 43.4 mmHg within 5 days, with maximum increases at 22-31 min after treatment. Oxygen breathing alone did not affect tumor pO(2). Radiation plus oxygen plus efaproxiral produced tumor growth inhibition throughout the treatment duration, and inhibition was significantly different from radiation plus oxygen from day 3 to day 5. The results of this study provide unambiguous quantitative information on the effectiveness of efaproxiral to consistently and reproducibly increase tumor oxygenation over the course of 5 days of treatment, modeling the clinical use of efaproxiral. Also, based on the tumor growth inhibition, the study shows the efaproxiral-enhanced tumor oxygenation was radiobiologically significant. This is the first study to demonstrate the ability of efaproxiral to increase tumor oxygenation and to increase the tumor growth inhibition of radiotherapy over 5 days of treatment.

The influence of efaproxiral on breast cancer xenograft hypoxia and HIF-1-alpha expression

10102 Background: Efaproxiral (E), a synthetic allosteric modifier of hemoglobin, has demonstrated clinical safety and efficacy as a radiosensitizer in patients with brain metastases from breast cancer. Although shown to enhance oxygenation in a murine mammary tumor, efaproxiral has not previously been tested in a human breast cancer xenograft. We studied changes in tumor oxygenation in a human xenograft and evaluated whether efaproxiral induces cell signaling events of potential therapeutic value. Methods: MDA-MB-468 breast cancer cells in matrigel were injected into the flank of nude mice. After tumors grew to 1–1.5 cm, animals were subjected to 1 of 3 treatments: ip saline +room air breathing (RA), ip saline + 50% oxygen breathing (O2), or ip efaproxiral (300 mg/kg) + 50% oxygen breathing (E+O2). Twenty minutes later the hypoxia marker, pimonidazole (pimo), was given, and 70 minutes later tumors were harvested for immunohistochemical study of hypoxia and hypoxia-inducible factor 1-alpha (HIF1-α) and RNA extraction to identify early changes in gene expression. Image analysis software was used to quantify observations. Results: Tumor hypoxia and HIF1-α staining were significantly decreased by efaproxiral (Table). HIF1-α staining did not entirely colocalize with pimo, implying different oxygen tension levels for HIF1-α ubiquitination and pimo reduction. Gene arrays indicated that after E+O2, expression of the hypoxia-induced DR1 transcription repressor was reduced compared with O2. Conclusions: The combination of E+O2 reduced hypoxia and HIF1-α expression in MDA-MB-468 human breast tumors in vivo, and an early effect on gene expression was reduced DR1. The results demonstrate an efaproxiral-mediated enhanced oxygenation of human hypoxic breast cancer. Furthermore, the efaproxiral-mediated down-regulation of HIF1-α suggests possible new opportunities in the clinical application of efaproxiral, notably as an adjuvant to systemic agents for which HIF1-α-mediated resistance limits efficacy. [Table: see text] [Table: see text]

Whole-brain radiotherapy with or without efaproxiral for the treatment of brain metastases: Determinants of response and its prognostic value for subsequent survival

To determine the prognostic factors for radiographic response and its prognostic value for subsequent survival in patients undergoing whole-brain radiotherapy (WBRT) for brain metastases. Five hundred fifteen eligible patients were randomized in a phase III trial evaluating WBRT and supplemental oxygen with or without efaproxiral, an allosteric modifier of hemoglobin that reduces hemoglobin oxygen-binding affinity and enhances tumor oxygenation, potentially increasing tumor radiosensitivity. Brain images were obtained at baseline and at scheduled follow-up visits after WBRT. Landmark analysis was used to assess the ability of response at selected time points to predict subsequent survival. Logistic regression was used to assess determinants of response at 3 months. Treatment arm, Karnofsky Performance Status, presence or absence of liver metastases, and primary site were all determinants of response at the 3-month follow-up visit, with patients in the efaproxiral arm experiencing a 67% greater odds of response at this visit (p = 0.02). Response at 3 and 6 months was a significant prognostic factor for longer subsequent survival. The 3-month scan is a valuable prognostic factor for subsequent survival in patients with brain metastases treated with WBRT. Patients in the efaproxiral arm had a higher response rate at 3 and 6 months than those in the control arm.

Efaproxiral: A Radiation Enhancer Used in Brain Metastases from Breast Cancer

To review the mechanism of action and clinical data of efaproxiral use in brain metastases of breast cancer. Articles were identified through MEDLINE (1966-June 2005) and EMBASE (1980-May 2005) searches using the key words efaproxiral and RSR13. Published abstracts over the previous 10 years from various scientific meetings, including American Society of Clinical Oncology and San Antonio Breast Cancer Symposium, were also searched for investigations of efaproxiral. Data on efaproxiral were also provided by Allos Therapeutics. All published clinical data in humans regarding efaproxiral use in brain metastases from breast cancer were selected for this review. In addition, published studies in humans that discussed the pharmacokinetics, pharmacodynamics, and safety of efaproxiral were evaluated. Efaproxiral is a synthetic allosteric modifier of hemoglobin that results in a shift of the hemoglobin oxygen dissociation curve to the right. Therefore, oxygen is more readily released from hemoglobin into tissues. Efaproxiral demonstrated a significant survival benefit when used as a radiation enhancer in patients with brain metastases originating from breast cancer. The safety profile of efaproxiral and improved survival rates make this agent advantageous over radiation alone. Further investigation and results from the ongoing clinical trials will help to define the role of efaproxiral in clinical practice. Efaproxiral is the first synthetic allosteric modifier to demonstrate significant improvement in survival in patients undergoing radiation therapy for brain metastases of breast cancer. Validation of this effect in ongoing clinical trials will be important in determining the role of efaproxiral in brain metastases from breast cancer.

Phase III Study of Efaproxiral As an Adjunct to Whole-Brain Radiation Therapy for Brain Metastases

To determine whether efaproxiral, an allosteric modifier of hemoglobin, improves survival in patients with brain metastases when used as an adjunct to whole-brain radiation therapy (WBRT). Patients with brain metastases from solid tumors and a Karnofsky performance score of > or = 70 were randomly assigned to receive WBRT with supplemental oxygen and either efaproxiral at 75 or 100 mg/kg (efaproxiral arm) or no efaproxiral (control arm). The primary end point was survival. The study consisted of 515 eligible patients (efaproxiral arm, n = 265; control arm, n = 250). The median survival time (MST) was 5.4 months for the efaproxiral arm versus 4.4 months for the control arm (hazard ratio [HR] = 0.87; P = .16). For the subgroup of patients with non-small-cell lung cancer (NSCLC) or breast cancer, the MST was 6.0 and 4.4 months, respectively (HR = 0.82; P = .07). Cox multiple regression analysis demonstrated a significant reduction in the risk of death for the efaproxiral arm in both primary populations. Further analysis indicated that the benefit may be restricted to the subgroup of patients with breast cancer. Response rates (radiographic complete response plus partial response) improved by 7% (P = .10) and 13% (P = .01) for all patients and for NSCLC and breast cancer patients in the efaproxiral arm, respectively. The most common severe adverse event in patients treated with efaproxiral was hypoxemia, which was reversible and effectively managed with supplemental oxygen in most patients. The addition of efaproxiral, a noncytotoxic radiation sensitizer, to WBRT may improve response rates and survival in patients with brain metastases, particularly metastases from breast cancer. A confirmatory trial for breast cancer patients has been initiated.

Effects of a Pharmacologically-Induced Shift of Hemoglobin-Oxygen Dissociation on Myocardial Energetics During Ischemia in Patients with Coronary Artery Disease

Conventional strategies to treat myocardial ischemia include interventions that reduce oxygen demand and/or increase myocardial blood flow. Animal experiments suggest that right-shifting the hemoglobin-oxygen dissociation curve may also attenuate the metabolic consequences of myocardial ischemia. We evaluated whether exercise-induced myocardial ischemia can be alleviated in subjects with coronary artery disease (CAD) by enhancing oxygen release with an allosteric modifier of hemoglobin's affinity for oxygen (RSR13). Seven subjects with CAD underwent a randomized, double-blind, cross-over study of the metabolic consequences of RSR13 administration on myocardial ischemia. Myocardial high-energy phosphates were quantified with 31P nuclear magnetic resonance (NMR) spectroscopy before, during, and after isometric handgrip-exercise. Subjects underwent NMR studies at baseline and on two separate occasions following the infusion of RSR13 (100 mg/kg) or placebo. RSR13 infusion significantly increased mean p50 by 8.1 +/- 2.7 mmHg at the end of the infusion, and it was still elevated by 4.9 +/- 3.3 mmHg after the completion of the treadmill tests while placebo had no effect. The myocardial creatine-phosphate (PCr) to adenosine-triphosphate (ATP) ratio decreased during handgrip-exercise in the baseline studies (from 1.39 +/- 0.23 before exercise to 0.95 +/- 0.21 during handgrip-exercise, p = 0.0001) and in the placebo studies (from 1.29 +/- 0.16 to 0.98 +/- 0.37, p = 0.06) but not during administration of RSR13 (from 1.28 +/- 0.18 to 1.02 +/- 0.24, p = 0.12). However, the mean values of cardiac PCr/ATP during handgrip-exercise did not differ significantly among the three measurements (baseline, placebo, RSR13). A single infusion of RSR13 to subjects with CAD increased mean p50 by 4.9-8.1 mmHg but did not significantly alter myocardial PCr/ATP during exercise. This is the largest right-shift in hemoglobin-oxygen binding affinity achieved in CAD subjects, and it did not provide clear evidence of protection from cardiac ischemia.

Increased oxygenation of intracranial tumors by efaproxyn (efaproxiral), an allosteric hemoglobin modifier: In vivo EPR oximetry study

To determine quantitatively the changes in oxygenation of intracranial tumors induced by efaproxiral, an allosteric hemoglobin modifier. Efaproxiral reduces hemoglobin-oxygen binding affinity, which facilitates oxygen release from hemoglobin into surrounding tissues and potentially increases the pO(2) of the tumors. The study was performed on 10 male Fisher 344 rats with 9L intracranial tumors. Electron paramagnetic resonance (EPR) oximetry was used to measure quantitatively the changes in the pO(2) in the tumors. Lithium phthalocyanine (LiPc) crystals were implanted in the tumors and in the normal brain tissue in the opposite hemispheres. We monitored the cerebral pO(2) starting 7 to 10 days after the tumor cells were implanted. NMR imaging determined the position and size of tumor in the brain. After an initial baseline EPR measurement, efaproxiral (150 mg/kg) was injected intravenously over 15 minutes, and measurements of tumor and normal brain oxygen tension were made alternately at 10-minute intervals for the next 60 minutes; the procedure was repeated for 6 consecutive days. Efaproxiral significantly increased the pO(2) of both the intracranial tumors and the normal brain tissue on all days. The maximum increase was reached at 52.9 to 59.7 minutes and 54.1 to 63.2 minutes after injection, respectively. The pO(2) returned to baseline values at 106 to 126.5 minutes after treatment. The maximum tumor and normal tissue pO(2) values achieved after efaproxiral treatment from Day 1 through Day 6 ranged from 139.7 to 197.7 mm Hg and 103.0 to 135.9 mm Hg, respectively. The maximum increase in tumor tissue pO(2) values from Day 2 to Day 5 was greater than the maximum increase in normal tissue pO(2). We obtained quantitative data on the timing and extent of efaproxiral-induced changes in the pO(2) of intracerebral 9L tumors. These results illustrate a unique and useful capability of in vivo EPR oximetry to obtain repeated noninvasive measurements of tumor oxygenation over a number of days. The information on the dynamics of tumor pO(2) after efaproxiral administration illustrates the ability of efaproxiral to increase intracranial tumor oxygenation.

Population pharmacokinetic-pharmacodynamic modeling of efaproxiral in cancer patients receiving radiation therapy

Aims Efaproxiral (EFP), a synthetic allosteric modifier of hemoglobin (Hb), reduces O2-binding affinity in blood (p50) and is investigated as a radiation therapy sensitizer. The goal of this work was to describe the pharmacokinetics (PK) and pharmacodynamics (PD) of EFP in cancer (CA) patients. Methods Pooled data from 6 phase I -III trials included: 451 patients, 2582 plasma and 2881 RBC concentrations, and 2483 p50 values. Covariates were age, weight, height, sex, concomitant medications, CA type, Hb, albumin, creatinine clearance, body surface area (BSA) and dose. Data were analyzed using NONMEM. Results A linear, 2-compartment model with RBC:plasma proportionality constant and a linear RBC-p50 model described the PK-PD relationship. Parameters were consistent with previously reported values (%SE): CL=1.88 L/hr (8%), V1=10.5 L (2%), Q=2.58 L/hr (12%), V2=18.1 L (10%), SLPRBC=0.982 (1%), INTp50=26.9 mmHg (~0%), SLPp50=0.0193 mmHg/(μg/mL) (5%). The inclusion of covariates resulted in improvements in goodness of fit and decreases in the estimated inter-individual variances. The final model adequately described the central tendency and population variability for the observed data. Conclusions Differences in PK-PD response due to CA type were small, may have been study-related and were probably not clinically relevant. EFP exposure increased with age, but the clinical relevance of this effect was unknown. BSA was the most important predictor of EFP disposition. Clinical Pharmacology & Therapeutics (2005) 77, P89–P89; doi: 10.1016/j.clpt.2004.12.233

Safety profile of efaproxiral (RSR13), a novel radiation sensitizer, in patients undergoing radiation therapy

3090 Background: Tumor hypoxia is known to decrease radiation sensitivity of solid tumors. RSR13, a novel radiation sensitizer, is an allosteric modifier of hemoglobin. RSR13 reduces the oxygen-binding affinity of hemoglobin to facilitate the release of oxygen, leading to an increase in tumor oxygenation. Methods: A total of 538 patients across phase 1 through phase 3 clinical trials have received at least 1 dose of RSR13 as sole adjunct to radiation therapy (RT). Patients were dosed daily up to 100 mg/kg/d RSR13, 5 days a week for up to 32 doses, immediately prior to RT. Due to the pharmacodynamic effect of RSR13, and to maximize oxygen saturation, all patients received supplemental oxygen during RSR13 and RT treatments. Results: Eight treatment-emergent adverse events have been identified as components of the RSR13 safety profile: nausea/vomiting, headache, hypoxemia (hypoxia), hypotension/dizziness, infusion symptoms, rash/allergic reaction, anemia, and renal dysfunction. The majority of the hypoxemia events were asymptomatic, consisted of low pulse oximeter (SpO2) measurements alone, and were treated with increased supplemental oxygen (increased duration or flow) only. Non-hypoxemia events were typically treated with supportive care (eg, hydration, antihistamines, or antiemetics). Overall 11% of the patients treated in these studies developed a Serious Adverse Event related to RSR13. Conclusions: The use of RSR13 was well tolerated in patients with various solid tumors receiving RT. Events of the RSR13 safety profile were typically reversible and managed with supportive care alone. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Allos Therapeutics, Inc. Allos Therapeutics, Inc. Allos Therapeutics, Inc. Allos Therapeutics, Inc.

Evaluation of safety and tolerance of escalating doses of RSR13 administered with a fixed dose of BCNU every six weeks in patients with recurrent malignant glioma: Results of the phase I NABTT 9806 clinical trial

1533 Background: RSR13 is an allosteric modifier of hemoglobin which has been shown to increase tumor oxygen delivery, with potential to improve cytotoxic therapy. Previous studies in glioblastoma have demonstrated that the agent is tolerable at 100 mg/kg when delivered daily prior to radiotherapy, and survival was reported in excess of one year. Preclinical studies have shown enhancement of the response to certain chemotherapeuticagents as well. This protocol is assessing the safety and tolerance of escalating doses of RSR13 when administered prior to BCNU in patients with recurrent high grade glioma, as well as maximum tolerated dose and pharmacokinetics of RSR13 when administered with BCNU. Methods: RSR13, in escalating doses of 25 mg/kg, to a maximum of 100 mg/ kg, in cohorts of six patients each, was administered over 30 minutes prior to infusion of BCNU 200 mg., every six weeks, for a maximum of six cycles. Only nonhematologic toxicities are assessed in determining the dose limiting toxicity (DLT) of the combination of agents. Results: A total of 33 patients have been accrued, 14 females, 19 males. Two grade 3/4 hematologic toxicities occurred in the first cohort of six patients at 25 mg/kg, therefore this cohort was expanded to 12. One non-hematologic DLT was observed in this cohort (generalized weakness) and in the 50 mg/kg cohort, a DLT of pulmonary edema was observed. There were no DLTs in the 75 mg/kg cohort. Symptomatic hypoxia was observed as a significant adverse event in two patients in the 100 mg/kg cohort which is still enrolling. Conclusions: Hematologic toxicity is manageable in this drug combination. The maximum tolerated dose of RSR13 with BCNU is projected to be 75 or 100 mg/kg. No significant financial relationships to disclose.

Evaluation of safety and tolerance of escalating doses of RSR13 administered with a fixed dose of BCNU every six weeks in patients with recurrent malignant glioma: Results of the phase I NABTT 9806 clinical trial

1533 Background: RSR13 is an allosteric modifier of hemoglobin which has been shown to increase tumor oxygen delivery, with potential to improve cytotoxic therapy. Previous studies in glioblastoma have demonstrated that the agent is tolerable at 100 mg/kg when delivered daily prior to radiotherapy, and survival was reported in excess of one year. Preclinical studies have shown enhancement of the response to certain chemotherapeuticagents as well. This protocol is assessing the safety and tolerance of escalating doses of RSR13 when administered prior to BCNU in patients with recurrent high grade glioma, as well as maximum tolerated dose and pharmacokinetics of RSR13 when administered with BCNU. Methods: RSR13, in escalating doses of 25 mg/kg, to a maximum of 100 mg/ kg, in cohorts of six patients each, was administered over 30 minutes prior to infusion of BCNU 200 mg., every six weeks, for a maximum of six cycles. Only nonhematologic toxicities are assessed in determining the dose limiting toxicity (DLT) o...

Standard whole brain radiation therapy (WBRT) with supplemental oxygen (O2), with or without RSR13 (efaproxiral)in patients with brain metastases: Results of the randomized REACH (RT-009) study

1534 Background: Brain metastases represent a common cause of morbidity and mortality among cancer pts. The effectiveness of WBRT as primary therapy for brain metastases is limited by tissue hypoxia, which has been shown to cause radioresistance in solid tumors. RSR13 (efaproxiral) is a novel radiation sensitizer that acts as an allosteric modifier of hemoglobin, facilitates O2 release and decreases tissue hypoxia. Methods: A randomized, open-label Phase 3 study was conducted comparing RSR13 and WBRT to WBRT alone in pts. with newly diagnosed brain metastases from various solid tumors. In the RSR13 arm (n=271), pts. received RSR13 (75–100 mg/kg/d, IV) plus O2 followed by WBRT (30 Gy, 3 Gy/d x 10 days). In the Control arm (n=267), pts. received WBRT plus O2. The primary endpoint was survival. Secondary endpoints were RR in the brain, TTP, cause of death and QoL. Results: A total of 538 pts. were enrolled over 29 months at 82 sites in 12 countries. In the overall study population (n=538), the RSR13 arm demonstrated a 17.6% improvement in median survival time (MST) over the Control arm (5.26 mo vs. 4.47 mo; p=0.17). The survival benefit for RSR13 was statistically significant in a Cox multiple regression model (Hazard Ratio 0.78 [95% CI 0.64, 0.94], p=0.010). Pts. with breast cancer or NSCLC (n=414) receiving RSR13 had a 32.4% increase in MST vs. Control (5.91 vs. 4.47 mo; p=0.12). In pts. with breast cancer (n=115) the MST almost doubled in those who received RSR13 vs. Control (8.67 vs. 4.57 mo; p=0.006). The percentage of breast cancer pts. with stable/improved KPS at 3 months was significantly higher in the RSR13 arm (35% vs 18%, p=0.001). Related SAEs that are part of the RSR13 safety profile occurred in 7% of pts. The most common RSR13-related SAE was hypoxemia (3.4 %), which is dose-dependent and effectively managed with increased supplemental O2. Conclusions: The addition of RSR13 to WBRT was well tolerated and led to a 22% reduction in the risk of death for all pts. A near doubling in MST and significant improvement in QoL were observed in pts. with brain metastases from breast cancer. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Allos Therapeutics, Inc. Allos Therapeutics Allos Therapeutics, Inc. Allos Therapeutics

Standard whole brain radiation therapy (WBRT) with supplemental oxygen (O2), with or without RSR13 (efaproxiral)in patients with brain metastases: Results of the randomized REACH (RT-009) study

1534 Background: Brain metastases represent a common cause of morbidity and mortality among cancer pts. The effectiveness of WBRT as primary therapy for brain metastases is limited by tissue hypoxia, which has been shown to cause radioresistance in solid tumors. RSR13 (efaproxiral) is a novel radiation sensitizer that acts as an allosteric modifier of hemoglobin, facilitates O2 release and decreases tissue hypoxia. Methods: A randomized, open-label Phase 3 study was conducted comparing RSR13 and WBRT to WBRT alone in pts. with newly diagnosed brain metastases from various solid tumors. In the RSR13 arm (n=271), pts. received RSR13 (75–100 mg/kg/d, IV) plus O2 followed by WBRT (30 Gy, 3 Gy/d x 10 days). In the Control arm (n=267), pts. received WBRT plus O2. The primary endpoint was survival. Secondary endpoints were RR in the brain, TTP, cause of death and QoL. Results: A total of 538 pts. were enrolled over 29 months at 82 sites in 12 countries. In the overall study population (n=538), the RSR13 arm demo...

Effect of RSR13, an allosteric hemoglobin modifier, on oxygenation in murine tumors: an in vivo electron paramagnetic resonance oximetry and bold MRI study

Purpose RSR13, an allosteric modifier of hemoglobin, reduces hemoglobin-oxygen binding affinity facilitating oxygen release from hemoglobin, resulting in increases in tissue pO 2. The purpose of this study was noninvasively to monitor the time course and effect of RSR13 on tumor oxygenation, directly using in vivo electron paramagnetic resonance (EPR oximetry), and indirectly using blood oxygen level dependent magnetic resonance imaging (BOLD MRI). Methods and materials The study was performed in transplanted radiation-induced fibrosarcoma tumors (RIF-1) in 18 female C3H/HEJ mice, which had two lithium phthalocyanine (LiPc) deposits implanted in the tumor when the tumors reached about 200–600 mm 3. Baseline EPR measurements were made daily for 3 days. Then, for 6 consecutive days and after an initial baseline EPR measurement, RSR13 (150 mg/kg) or vehicle (same volume) was injected intraperitoneally, and measurements of intratumoral oxygen were made at 10-min intervals for the next 60 min. In each mouse, every third day, instead of EPR oximetry, BOLD MRI measurements were made for 60 min after administration of the RSR13. Results Based on EPR measurements, RSR13 produced statistically significant temporal increases in tumor pO 2 over the 60-min time course, which reached a maximum at 35–43 min postdose. The average time required to return to the baseline pO 2 was 70–85 min. The maximum increase in tumor tissue pO 2 values after RSR13 treatment from Day 1 to Day 5 (8.3–12.4 mm Hg) was greater than the maximum tumor tissue pO 2 value for Day 6 (4.7 mm Hg, p < 0.01). The maximum increase in pO 2 occurred on Day 2 (12.4 mm Hg) after RSR13 treatment. There was little change in R 2*, indicating that the RSR13 had minimal detectable effects on total deoxyhemoglobin and hemoglobin-oxygen saturation. Conclusion The extent of the increase in tumor pO 2 achieved by RSR13 would be expected to lead to a significant increase in the effectiveness of tumor radiotherapy. The lack of a change in the BOLD MRI signal suggests that the tumor physiology was largely unchanged by RSR13. These results illustrate a unique and useful capability of in vivo EPR oximetry and BOLD MRI to obtain repeated measurements of tumor oxygenation and physiology. The dynamics of tumor pO 2 after RSR13 administration may be useful for the design of clinical protocols using allosteric hemoglobin effectors.

Detection in urine of efaproxiral (RSR13), a potential doping agent, by a routine screening procedure based on methylation followed by gas chromatography/mass spectrometry

Efaproxiral (also known as RSR13) is a synthetic allosteric modifier of hemoglobin. The ability to increase maximal muscle oxygen uptake makes RSR13 a potential performance-enhancing agent for athletes and, therefore, a new agent to be taken into account by the antidoping control laboratories. In this work, a new method for the detection of efaproxiral in urine by gas chromatography/mass spectrometry is described.

RSR13 modulates tumor hypoxia and HIF-1-alpha expression in H226 xenografts in vivo

Purpose/Objective: Interstitial tumor oxygen measurements and radiosensitivity assays have established that RSR13 (efaproxiral), a synthetic allosteric modifier of hemoglobin that decreases hemoglobin-oxygen binding affinity, improves radiosensitivity in vivo by reducing the hypoxic fraction in solid tumors. To characterize the micro-regional distribution of RSR13-mediated oxygenation and evaluate its effect upon hypoxia-induced gene expression, a xenograft model was analyzed for the geographic pattern of changes in hypoxia and the resulting impact upon hypoxia-inducible factor 1-alpha (HIF1-α) expression.