Doses Of Misonidazole Research Articles

NEUROTOXICITY OF MISONIDAZOLE IN RATS FOLLOWING INTRAVENOUS ADMINISTRATION

Misonidazole is a hypoxic cell radiosensitizer that induces a peripheral neuropathy in humans after exceeding a schedule-dependent cumulative threshold dose. Clinical studies of misonidazole have been conducted using oral administration, whereas most other radiosensitizers have been administered intravenously. Since route of exposure can potentially influence the toxicity of xenobiotics, the objective of this study was to assess the neurotoxicity of misonidazole in rats following intravenous dosing using a battery of routine clinical, neurofunctional, biochemical, and histopathologic screening methods. Male Sprague–Dawley rats were administered intravenous doses of misonidazole at 0 (vehicle control), 100, 200, 300, or 400mgkg −1once per day, 5 days per week, for 2 weeks. Animals were evaluated for neurofunctional and pathological changes following termination of treatment (Days 15–17) and at the end of a 4 week observation period (Days 43–45). During the dosing phase, hypoactivity, salivation, rhinorrhea, chromodacryorrhea, rough pelage and ataxia were observed at 400mgkg −1, and body weight gain of the 300 and 400mgkg −1groups was significantly decreased relative to the vehicle controls by 24% and 49%, respectively. Corresponding reductions in food consumption were 8% and 23%, respectively. Although most 400mgkg −1animals appeared normal on Day 15 prior to the neurofunctional evaluations, rotorod testing precipitated a number of clinical signs including: ataxia, impaired righting reflex, excessive rearing, tremors, vocalization, circling, head jerking, excessive sniffing and hyperactivity. All of these animals recovered and appeared normal from Day 17 through study termination. There were no treatment-related effects on motor activity, acoustic startle response, rotorod performance, forelimb group strength, toe and tail pinch reflexes, tibial nerve β-glucuronidase activity or tail nerve conduction velocity. Although hindlimb grip strength of the 400mgkg −1group was significantly decreased by 17% relative to the vehicle controls on Day 15, this finding appeared related to the reduced food consumption and body weight gain in these animals. No microscopic changes were detected in peripheral nerves. Necrosis and proliferation of fibrillary astrocytes (gliosis) were seen in the cerebellum and medulla of the 400mgkg −1animals on Day 16. Gliosis in these same brain regions was observed in the 300 and 400mgkg −1groups on Day 44. The results show that intravenous administration of misonidazole to rats causes dose-limiting central nervous system toxicity without effects on peripheral nervous tissue. The lack of peripheral neurotoxicity was most likely due to a combination of several interrelated factors including route of administration, duration and intensity of the dosing regimen, and total cumulative dose.

Dose-dependent increase in the frequency of micronuclei and chromosomal aberrations by misonidazole in mouse bone marrow

Cytogenetic effects produced in bone marrow of mice by various doses of misonidazole (MISO, 100 to 1000 mg/kg bodyweight) were studied by assaying the induction of micronuclei (MN) and chromosomal aberrations. Misonidazole increased the frequency of both micronuclei and chromosomal aberrations over normal at 24 h after treatment, however, the values were statistically significant from normal only at drug doses above 250 mg/kg. The increase was proportional to the drug dose with a best fit to linear quadratic model for MN induction. For chromosomal aberrations the data fitted eqully well to linear as well as linear quadratic models.

Interaction of misonidazole, hyperthermia, and irradiation in a C3H mammary carcinoma and its surrounding skin in vivo

The interaction of irradiation, Misonidazole (MISO), and hyperthermia was studied in a C3H mouse mammary carcinoma and its surrounding skin in vivo. MISO (0.5–1.0 mg/g) was injected 30 min before irradiation. Hyperthermia (41.5°–43.5°C for 60 min) was given either simultaneously, 0.5 hr, or 4 hr after X rays. The results were evaluated as the radiation dose to achieve tumor control (TCD 50) or moist desquamation of the skin (DD 50) in half of the treated animals. A therapeutic gain was found when the enhancement in tumors were greater than that found in skin. The combination of simultaneous heat and irradiation caused great enhancement in radiation response, but with no therapeutic gain. A slightly lower enhancement of the damage in both tissues was found with a 30 min interval between irradiation and hyperthermia, whereas heat 4 hr after X rays gave a small, but significant therapeutic gain. MISO significantly enhanced the response in tumors but not in skin. Combined trimodality treatment with MISO, irradiation, and hyperthermia resulted in enhancement ratios up to 15, dependent on temperature, radiation-heat interval, and to a lesser extent the MISO dose. The enhancement was for all schedules most pronounced in the tumors, resulting in an improved therapeutic effect. The combination of MISO and hyperthermia may be a valuable addition to radiotherapy, especially if heat and irradiation can be applied with close interval and with one of the modalities given selectively to the tumor.

Structure-activity relationships for tumour radiosensitization by analogues of nicotinamide and benzamide.

Nicotinamide has been shown in our laboratory and those of other investigators to be an effective radiosensitizer of a variety of mouse tumours, while producing little or no radiosensitization of normal tissues. Its mechanism of action is different from classical electron-affinic compounds and appears to be the result of improved tumour oxygenation. In this study we have synthesized 29 analogues of nicotinamide and benzamide and characterized them for their tumour radiosensitization and acute toxicity in mice. The data show that a wide range of additions to the nicotinamide and benzamide ring produce tumour radiosensitization similar to that produced by equimolar doses of misonidazole, but that substitutions of the amide tend to reduce radiosensitization. Other structure-activity relationships are evident. Although some compounds produce similar tumour radiosensitization to nicotinamide at equimolar doses, and are comparably low in acute toxicity, none appears sufficiently superior to supplant nicotinamide itself as a candidate for clinical trials. Thus these data provide evidence that nicotinamide, because of the extensive experience with its use in man, is likely to be the best drug in the benzamide-nicotinamide series for development as a radiosensitizer of human tumours.

Enhancement of chemotherapy and nitroimidazole-induced chemopotentiation by the vasoactive agent hydralazine

Nitroimidazoles have been shown to be potent sensitisers of certain clinically active chemotherapeutic agents. This process of chemopotentiation has been shown to be hypoxia-mediated. The present studies evaluated whether increasing the level of hypoxia in the tumour tissue, by treatment with the vasoactive agent hydralazine, could modify the chemosensitising ability of nitroheterocyclics. Administering either misonidazole or RSU 1164 before, or hydralazine after, the chemotherapeutic agents melphalan, cyclophosphamide or the nitrosourea CCNU, increased the extent of cell kill in both the KHT sarcoma and RIF-1 tumour. However, even greater enhancements could be achieved when hydralazine was used in treatment protocols in which a nitroimidazole was combined with chemotherapy. For example, a 5.0 mg kg-1 dose of hydralazine given 30 min after melphalan, or a 2.5 mmol kg-1 dose of misonidazole administered 30 min before melphalan, increased, compared to melphalan alone, the resultant tumour cell kill by factors of approximately 1.9 and approximately 1.3, respectively. By comparison, when hydralazine was given after the melphalan plus misonidazole combination, treatment efficacy was enhanced approximately 3-fold compared to melphalan alone. Yet in contrast to the results of the tumour response studies, the inclusion of hydralazine did not increase the bone marrow toxicity associated with the chemotherapeutic agent when used alone or in conjunction with a nitroimidazole. The results, therefore, imply that the addition of hydralazine to the chemotherapy, or chemotherapy-sensitiser protocol, led to a therapeutic advantage.

Manipulation of oxygenation in a human tumour xenograft with BW12C or hydralazine: effects on responses to radiation and to the bioreductive cytotoxicity of misonidazole or RSU-1069.

The influence of altered tumour oxygenation on the responses to radiation and/or bioreductive 2-nitroimidazole compounds was studied in a well differentiated, human, colon adenocarcinoma (MAWI), grown as a subcutaneous xenograft in nude mice. Tumour growth delays were measured after local, single 5-18 Gy doses of X-rays. BW12C, which inhibits dissociation of oxyhaemoglobin, produced radioprotection similar to that resulting from clamping off the tumour blood supply during irradiation. Hydralazine, a vasoactive agent, also appeared to give radioprotection. BW12C or misonidazole (MISO) alone had no measurable inhibitory effect on xenograft growth. Hydralazine or RSU-1069 slightly increased the time for tumours to reach 6 times their original volumes. When hydralazine was given 40 min after a dose of 800 mg/kg of MISO, without X-rays, growth delays in excess of 5 tumour volume doubling times resulted and fewer tumour cells were present in histological sections. Lower doses of MISO combined with hydralazine were ineffective. Other combinations of bioreductive cytotoxic agents and methods of manipulation of tumour blood flow/oxygenation induced slight and inconsistent growth delays. Hydralazine was injected after irradiation of tumours in mice previously treated with various doses of MISO in an attempt to exploit the bioreductive cytotoxic potential of MISO in conjunction with its radiosensitizing properties; however, tumour growth delays were similar with or without hydralazine after irradiation. Thus, post-irradiation restriction of tumour blood flow appears to be an ineffective therapeutic strategy in this human xenograft tumour model.

Effect of thiol manipulation on chemopotentiation by nitroimidazoles

Highly electron affinic compounds such as the nitroimidazole misonidazole (MISO) have been shown both in vitro and in vivo to be effective potentiators of certain conventional chemotherapeutic agents. Mechanistically, the observation that nitroheterocyclics reduce intra-cellular thiols by enhancing the oxidation of glutathione (GSH), has suggested that thiol depletion by MISO may be a key factor in this enhancement. The present investigations were undertaken to determine whether the use of buthionine sulfoximine (BSO) to affect GSH metabolism may lead to more effective potentiation of chemotherapeutic agents by sensitizers. KHT/iv cells were treated in exponential phase under hypoxic conditions with variable doses of the activated form of cyclophosphamide (4-hydroxy-cyclophosphamide, 40H-CY) administered concominantly with or without MISO (2.5 mM) for an exposure time of 4 hr. Inclusion of the sensitizer in the treatment protocol resulted in a dose modifying factor of ∼2.4. Exposing cells to 1.0 mM BSO for 2 hr prior to treatment reduced intracellular GSH levels to 70–80% of control and increased the efficacy of 40H-CY ∼1.2-fold. If BSO was administered prior to the 40H-CY + MISO combination, severe tumor cell toxicity resulted. For example, when combined with 40H-CY, similar cell kill could be achieved with 5 to 6-fold lower MISO doses in the presence of BSO as in the absence of BSO. Ultrastructural evaluations revealed that in the three agent combination, membrane damage, as reflected by the formation of surface blebs, may play a key role in the mechanism of the observed enhanced cytotoxicity.

High-dose melphalan, misonidazole, and autologous bone marrow transplantation for the treatment of metastatic colorectal carcinoma. A phase I study.

To augment the antitumor effect of high-dose melphalan and determine pharmacokinetics we conducted a phase I trial of escalating doses of high-dose IV melphalan with the chemosensitizer misonidazole for patients with advanced colorectal carcinoma. Fourteen patients with modified Dukes D adenocarcinoma of the colorectum were treated with a single course of melphalan (40-60 mg/m2 i.v. bolus q.d. X 3 days) and misonidazole (1-3 g/m2 p.o. q.d. X 3 days) followed by autologous bone marrow transplantation. Toxicity consisted of severe myelosuppression, moderate nausea and vomiting, and mild mucositis and diarrhea. One patient developed unexplained renal tubular acidosis, and a diffuse encephalopathy occurred in another patient. Three patients died within the first 30 days after the start of treatment, two due to tumor progression and one due to sepsis and disseminated intravascular coagulation-induced intracerebral hemorrhage. Six of 14 patients achieved a partial response, and the median response duration was 4 months (range 3-10 months). Analysis of misonidazole serum concentrations showed similar pharmacokinetics to those previously reported, suggesting no significant drug interaction with intravenous melphalan. Mean peak serum concentrations ranged from 81.8 micrograms/ml to 115.2 micrograms/ml at the second and third misonidazole dose levels, which approximate those known to provide effective chemosensitization with melphalan in animal models. In this phase I study, we showed that maximally tolerated doses of intravenous melphalan can safely be combined with oral misonidazole. In view of the large volumes of oral misonidazole required at the highest dose level, subsequent studies to determine the maximally tolerated dose of misonidazole should employ the intravenous form.

Induction of Tumour Hypoxia Post-irradiation: A Method for Increasing the Sensitizing Efficiency of Misonidazole and RSU 1069in Vivo

It is known that hydralazine can decrease blood flow to experimental murine tumours. A consequence of this, in the KHT sarcoma, is the induction of close to 100 per cent radiobiological hypoxia, which lasts for nearly 2 h following i.v. injection of 5 mg/kg hydralazine to the mouse. This phenomenon is exploitable in order to increase the apparent sensitizing efficiency of the nitroheterocyclic radiosensitizers, misonidazole and RSU 1069, and is demonstrated using the treatment schedule: sensitizer----60 min----X-rays----1 min----hydralazine. Such a strategy will first take advantage of the radiosensitizing properties of the nitroimidazole, then after irradiation the hydralazine should allow expression of the differential toxicity towards hypoxic cells known to occur with misonidazole and RSU 1069. Misonidazole gives an enhancement ratio (ER) of 1.3 at 100 mg/kg, rising to 2.0 at 1000 mg/kg. Where hydralazine is given after irradiation, no additional cell kill is observed with 1000 mg/kg. In contrast, at lower doses of misonidazole, hydralazine induces a substantial increase in cell killing such that the ER obtained with 100 mg/kg is the same as that achieved with 1000 mg/kg misonidazole when used alone with radiation. Similarly, 20 mg/kg RSU 1069 with radiation followed by hydralazine is equivalent to the radiosensitizing effect of 80 mg/kg RSU 1069 without hydralazine. In addition, doses of RSU 1069 that normally give no radiosensitization (5 or 10 mg/kg) produce substantial increases in cell killing when combined with hydralazine.

Characterization of radiation resistant hypoxic cell subpopulations in KHT sarcomas. (II). Cell sorting.

Hypoxic cells in KHT sarcomas were characterized using fluorescence activated cell sorting based on the diffusion properties of the fluorochrome Hoechst 33342. Tumour-bearing female C3H/HeJ mice were injected i.v. with 10 micrograms g-1 Hoechst 33342 and the cells derived from the tumours sorted on the basis of their staining intensities. For each sorted fraction the DNA histogram was evaluated using FCM analysis. The results indicated that the bright and dim cells were not equally distributed about the cell cycle. For example, a greater proportion of S phase cells were in the bright subpopulations whereas the dim subpopulations contained an increased proportion of cells in G1. When the tumours were irradiated with a single dose of radiation prior to cell sorting, the dim cells survived preferentially. Dose response curves for the 20% most dim and 20% most bright cells, sorted on the basis of fluorescence intensity, then were determined. The survival curves of the dim and bright cells were found to have slopes similar to those of KHT cells irradiated in situ in dead animals or in vitro under fully oxic conditions, respectively. In addition, when KHT sarcoma-bearing mice were given a 2.5 mmol kg-1 dose of misonidazole (MISO) prior to irradiation and cell sorting, the dim subpopulation was sensitized whereas the bright subpopulation was not. These findings suggest that (i) compared to well-oxygenated areas, hypoxic regions of KHT tumours contain a smaller percentage of cells actively proliferating and (ii) Hoechst 33342 sorting may allow the detailed in situ evaluation of agents acting directly against hypoxic cells in solid tumours.

Misonidazole reduces blood flow in two experimental murine tumours

The effects of single doses of misonidazole (MISO) on blood flow and vascular volume in the SaFA and CaNT tumours and normal tissues of the mouse have been studied. MISO was administered in the dose range 250-1,000 mg kg-1 and blood flow measured at different times after MISO by the 86RbCl extraction technique. Vascular volume was assessed by the distribution of 51Cr-labelled red blood cells. MISO at doses of 500 mg kg-1 or greater decreased flow in both tumours by up to 60% within 2 h. Flow remained reduced for up to 24 h. Similar but less profound changes were seen in the skin, although flow had recovered by 24 h. Only slight changes were seen in muscle, and none in kidney. The apparent loss of flow in tumours seen after large single doses of MISO may have important implications for its use as a chemosensitizer.

Misonidazole in fractionated radiotheraphy of a murine mammary carcinoma: Comparison of tumor and normal tissue response

The potential therapeutic benefit of misonidazole was tested in radiotherapy with 1, 2, 5, and 10 equal fractions, using as endpoints local tumor control (TCD50) of murine mammary carcinoma MDAH-MCa-4 and leg contracture at the TCD50, measured 120 days after irradiation. In controls and misonidazole-treated mice, the TCD50 increased with the number of fractions, from 66.7 to 114.6 Gy in controls, and from 43.3 to 75.7 Gy with misonidazole. At doses of ≥0.1 mg/g body weight, misonidazole reduced the TCD50 in all fractionation schedules; however, because of toxicity, 1.0 and 0.6 mg/g could be given with only 1 or 2 fractions. Leg contracture at the TCD50 was greatest (14.5 mm) in control mice treated with a single dose of radiation, and was least (7.2 to 7.4 mm) in those treated with a single dose of radiation preceded by 1.0 or 0.6 mg misonidazole/g body weight. With 0.1 mg misonidazole/g, the leg contracture at the TCD50 was less (9.8 to 12.2 mm with the various schedules) than in controls (12.0 to 14.5 mm) for 1, 5, or 10 fractions. Therefore, a therapeutic gain could be obtained by using misonidazole with 1, 2, 5, or 10 fractions, but the greatest gain occurred with 1 fraction, with high doses of misonidazole, that is, 0.6 to 1.0 mg/g.

Influence of misonidazole, SR-2508, RSU-1069 and WR-2721 on spontaneous metastases in C57BL mice

The effect of treatments with the hypoxic cell radiosensitizers misonidazole (MISO), SR-2508, RSU-1069, and with the radioprotector WR-2721 on spontaneously disseminated Lewis lung carcinoma (LLC) and B16 melanoma (B16M) was investigated. Tumors were implanted into the tails of C57BL mice and were surgically removed after reaching volumes of approximately 40 mm 3. This technique results in the induction of metastatic disease in lungs and at other anatomical sites and allows the independent treatment of disseminated tumor cells. The radiosensitizers and the radioprotector were administered 24 hr after surgery and animals were killed 14 and 30 days after removal of LLC and B16M, respectively. The time to death from spontaneous metastases was also measured. Both single treatments with large doses of MISO (1.0 g/kg) and fractionated therapy with smaller doses (0.5 g/kg on 5 consecutive days) promoted the formation of metastases to the lungs, lymph nodes and other organs. The survival times of MISO treated animals did not differ from control animals but manifestation of metastatic disease in the lungs and other organs occurred at earlier times. Administration of equitoxic doses of SR-2508 (3.0 g/kg) and RSU-1069 (0.1 g/kg) also promoted metastases formation. Mice treated with these radiosensitizers developed more metastases in the lungs and at other sites. Treatment with a single dose of WR-2721 (0.4 g/kg) promoted lung metastases but exerted a suppressive effect on lymph node tumors. When the radioprotector was given in fractionated schedules in three different doses (0.05 g/kg, 0.1 g/kg and 0.2 g/kg for 10 consecutive days) a slight enhancement of lung metastases and suppression of extrapulmonary metastases was observed.

Failure of misonidazole-sensitized radiotherapy to impact upon outcome among stage III–IV squamous cancers of the head and neck

As part of the RTOG research effort in the treatment of advanced, inoperable squamous cancer of the head and neck region, the hypoxic cell sensitizer, misonidazole, was selected for investigation as an adjuvant to definitive irradiation. Based upon a pilot experience (78-02) showing a 67% complete response rate among 36 AJC Stage III–IV patients receiving full-dose irradiation and 6 weekly p.o. doses of misonidazole, a phase III trial was carried out from '79–'83. Three hundred and six patients were entered, 42% of whom had oropharyngeal primaries and with 78% of all cases representing T3 or T4 (inoperable) lesions. Only 16% of the entire series presented with N 0 necks. Fractionation was altered among the misonidazole-receiving patients, in contrast to “standard” 5 treatments per week among “control” patients, such that 2 separate treatments were given on each day of p.o. misonidazole administration (2.0 gm/m2/wk X 6 doses, 2.5 Gy in a.m., 2.1 Gy in p.m.). Total tumor doses were identical among the two treatment arms except that a limitation of 40.0 Gy to spinal cord was specified for sensitized radiotherapy vs., 45.0 Gy for “control” patients. Primary tumor clearance was observed to be 55–60%, with minor variations according to tumor stage and site. The local regional control rate among radiotherapy-alone patients was 26% at 2 years compared to 22% (2 years) within the misonidazole-receiving group. Analysis of survival revealed no advantage to the sensitized patients, with 55 ± 2% surviving 1 year and 22 ± 1 % living 3 years following treatment in both treatment categories. Distant metastases as first site of failure (12–13%) and the local failure among initial complete responders (46%) showed no advantage to the misonidazole group. Although a misonidazole dosage of 2.0 gm/m2/wk X 6 (12 gm/m2 total) is well tolerated, no clinical benefit was demonstrated in this randomized trial. Other nitromidazole analogs (e.g. SR-2508) are now being investigated.

Radiosensitizing and Cytocidal Effects of Misonidazole: Evidence for Separate Modes of Action

Hypoxic BP-8 murine sarcoma cells were exposed to misonidazole and/or radiation and the kinetics and extent of cell death were evaluated with the [125I]iododeoxyuridine-prelabeling assay. Cell death after treatment with lethal doses of misonidazole was rapid and essentially complete within 2 or 3 days after drug exposure. In contrast, radiation death became apparent only after a delay period of 4 days and was complete by Day 10 after irradiation. Radiosensitization by short exposures to sublethal doses of misonidazole affected only the delayed component of cell death, that is, the radiation component of death. In experiments involving sequential radiation and drug treatment, prior irradiation of cells did not enhance the direct cytocidal effects of misonidazole, as evidenced by the fact that the early component of cell death was equal in control and preirradiated cells. However, postirradiation treatment with misonidazole did enhance the delayed radiation component of cell death. These results suggest that radiosensitization and direct killing by misonidazole are two distinct phenomena mediated by different cellular mechanisms, and radiosensitization by misonidazole represents a two-component effect composed of true dose modification and dose additive damage interactions, but these additive effects must occur at a site different from the cellular structure responsible for direct drug-induced cell death.

Potentiation of CCNU activity by misonidazole in metastases.

The KHT sarcoma is a model system in which metastases can be studied in multiple organs without prior clonal selection. The present series of experiments were designed to evaluate and compare the extent of potentiation of chemotherapeutic agent activity by radiosensitizers when KHT sarcoma cells were grown in different anatomical sites. In these studies the effect of combining misonidazole (MISO) and 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea (CCNU) on KHT lung nodules and ovarian metastases was determined. Ovarian metastases were a consequence of the direct spread of tumor cells growing as lung nodules. Once established, KHT cells in the lungs and ovaries grew with a similar doubling time (1-2 days). Response of tumors at either site to chemotherapy in the presence or absence of a sensitizer was assessed using an in vivo to in vitro excision assay. MISO (1.0 mmol/kg) was administered simultaneously with a range of CCNU doses and survival of clonogenic tumor cells was measured 24 h after treatment. The results demonstrate that the addition of MISO to CCNU treatment potentiated the action of this chemotherapeutic agent at both tumor sites although greater cell kill enhancement occurred in the ovarian metastases. In the lung nodules, when combined with CCNU, a 1.0 mmol/kg dose of MISO was found to yield a dose modifying factor (DMF) of approximately 1.3. The same combination resulted in a DMF of approximately 1.8 in the ovarian metastases. This difference in DMF was not a result of an intrinsic difference in sensitivity to CCNU since cells grown at either site gave rise to the same dose response curve. Rather the difference in dose modification by MISO appears to be a consequence of the larger fraction of radiobiologically hypoxic cells in the ovarian tumors (approximately 50 per cent) than in the lung nodules (approximately 5 per cent) at the time of treatment. These findings suggest the use of drug-sensitizer combinations to treat disseminated disease and provide further evidence for the requirement of hypoxia in chemopotentiation by radiosensitizers.

The radiosensitizing effects of misonidazole (MISO) in combination with diethyl maleate (DEM) in mouse mammary tumors

Large radiosensitization of C3H/He mouse mammary tumors was obtained with the combination of a non-protein sulfhydryl (NPSH) depletor, diethyl maleate (DEM), and misonidazole (MISO), compared with MISO alone over a range of MISO dose. The difference in enhancement ratios (ER's) for these two treatments was especially prominent at small MISO doses. ER's of 2.06 and 1.44 were obtained, respectively, by combined treatment with DEM (760 mg/kg) and MISO (100 mg/kg) or treatment with MISO alone. Radiosensitization of tumors by DEM alone was observed for doses over 600 mg/kg. When DEM was combined with MISO (100 mg/kg), ER's of the combination were larger than that of MISO alone, for doses over 400 mg/kg of DEM. Similarly, in case of DEM plus MISO (300 mg/kg), the ER's became larger than MISO alone, for doses over 200 mg/kg of DEM. The NPSH content in untreated tumors was 1.08 mmole/kg on the average and no changes in NPSH content was observed after MISO treatment. DEM treatment markedly reduced the NPSH content of tumors as a function of DEM dose and this decrease in NPSH was not significantly affected by MISO treatment. Tumor NPSH was reduced to 24% or less of control by administration of 760 mg/kg of DEM with or without MISO. These results are consistent with competition theory of NPSH and electron affinic radiosensitizers.

Potentiation of in vitro cytotoxic effects of misonidazole on human colon tumor cells by the differentiation-inducing agent N-methylformamide

Human colon tumor cells (clone A) were studied in vitro with regard to modification of dose-dependent cytotoxicity to misonidazole (MISO) treatment by pre-exposure growth in medium containing the differentiation-inducing agent N-methylformamid(NMF). Cells were grown as exponential cultures and were exposed for 2 passages to 170 mM NMF before exposure to graded doses of MISO (0–100 mM, 3 hours at 37°C, oxic or hypoxic). Both oxic and hypoxic cells could be sensitized to MISO cell killing. Using the 10% level of survival for comparison, the calculated MISO doses (mM) were: 105, 37, 50, and 10 for oxic control cells, hypoxic control cells, oxic-NMF treated cells, and hypoxic-NMF treated cells, respectively. Therefore, for NMF treated oxic cells, cell killing was increased by a factor of about 2.1, while for NMF treated hypoxic cells, cell killing increased by a factor of about 3.7. These data indicate that NMF treatment, while potentiating effects on both oxic and hypoxic cells, appears to have selectivity towards hypoxic cells. NMF may therefore have use in combined modality radiation therapy of solid tumors with electron-affinic radiosensitizers.

Phase II trial of misonidazole (MISO) and cyclophosphamide (CYC) in metastatic renal cell carcinoma

In animal models pre-treatment with misonidazole, a hypoxic cell radiosensitizer, enhances the antineoplastic effects of alkylating agent chemotherapy. Laboratory data suggest that hypoxic tumor cells may be more resistant to chemotherapy because of suboptimal drug delivery, reduced rates of cell division, or because hypoxia confers relative drug resistance. The therapeutic potential depends on the tumor type, doses of radiosensitizer and alkylating agent, the time interval between drug administration, and the ratio of sensitization of normal and malignant tissues. A Phase II trial of misonidazole and cyclophosphamide was initiated by the Eastern Cooperative Oncology Group to determine the response rate and toxicity in patients with metastatic renal cell cancer. Patients received 5 gm/m 2 of misonidazole intravenously two hr before 1200 mg/m 2 of cyclophosphamide every 3 wk. Patients with prior chemotherapy or radiotherapy received 1000 mg/m 2 of cyclophosphamide. Misonidazole was discontinued after a total dose of 15 gm/m 2. The median total misonidazole dose was 23.5 gm (range 4.5–34.5 gm). The median number of cyclophosphamide cycles was 2 (range 1–12). Of the 30 patients evaluable for response, only one patient had an objective partial response. Twenty-nine patients had stable or progressive disease. One patient remains on cyclophosphamide after 9 mo. Estimated median survival is 4.8 mo. There have been no lethal toxicities; however, 9 patients (25%) experienced life-threatening leukopenia and an additional 42% experienced severe hematologic toxicity. Eight patients had WBC < 1000 on days 7–14 of cycle 1. Thrombocytopenia and grade 3 anemia occurred in 1 and 2 patients, respectively. Moderate or severe nausea and vomiting occurred in 47% and 19% of patients, respectively. Only 3 patients experienced severe neurotoxicity. Four additional patients had moderate neurotoxicity. In summary, misonidazole in this dosing schedule does not enhance the anititumor activity of cyclophosphamide in renal cell carcinoma.

Effect of misonidazole dose on survival in patients with stage IIIB-IVA squamous cell carcinoma of the uterine cervix: An RTOG randomized trial

Between August 1980 and November 1984, 120 patients with FIGO Stage IIIB or IVA squamous cell carcinoma of the uterine cervix were randomized to receive radiation therapy (RT) (46 Gy pelvis + 10 Gy parametrial boost) followed by intracavitary or external boost to the primary ± misonidazole (MISO) (400 Mg/M 2 2–4 hours prior to RT daily, maximum 12 gm/M 2). The median at 24–28 hr misonidazole plasma level was 20 μg/ml 2–6 hr and 3.5 μg/ml. Approximately 60% of the patients on RT + MISO received 100% of expected total Misonidazole dose; peripheral neurologic toxicity was reported for nine patients receiving misonidazole (8 with mild and 1 with moderate paresthesia or pain). Time-dependent regression analyses found that actual cumulative misonidazole dose was not related to duration of survival from start of treatment ( p = 0.5). MISO dose expressed as a percent of expected dose was marginally related to increased survival measured from 14 weeks on study ( p = 0.1). No improvement in survival was observed with the addition of misonidazole to RT (64% of the patients on RT alone were alive at 18 months versus 54% of those on RT + MISO).