Inl. 1. Radiation Oncology Biol. Phys., Vol. 5, pp. 775-786 0 Pergamon Press Ltd., 1979. Printed in the U.S.A.

0

Original Contribution INITIAL UNITED STATES CLINICAL AND PHARMACOLOGIC EVALUATION OF MISONIDAZOLE (Ro-07-0582), AN HYPOXIC CELL RADIOSENSITIZER? TODD H.

WASSERMAN,

M.D.,S

RICHARD J. JOHNSON, M.B.,§

THEODORE L.

PHILLIPS, M.D.,+

CHARLES J. GOMER,

M.S.,tt

GILBERT A. LAWRENCE, M.B.,O WOLFGANG SADEE, Ph.D.,TJ ROBERTO A. MARQUES, B.S.,7 VICTOR A. LEVIN, M.D.H and GRETCHEN VANRAALTE, R.N., M.S.$ The hypoxic cell radiosensitixer, misonidaxole, is in a Phase I clinical pharmacology and toxicology triai. Forty wasgiven on the dose schedule of a singk oral patients wereevafuated on a weekly dose schedule. Misonidazole

dose weekly for 3 or 6 weeks in separate patient groups. Radiation therapy, of a palliative nature, was given 46 hr after drug administration to aflow for maximal tumor levels. The initial dose level was 1 gmlm’ and escalated to 3 gm/m’ weekly x 6 weeks and 5 gm/m2 weekly x 3 weeks. Toxicologic evaluation included serial clinical and laboratory evaluations. The principle toxicities observed were nausea and vomiting and neurotoxicity. The nausea and vomiting were acute, and dose limiting at 5 gm/m’. The neurotoxicity was primarily a peripheral sensory polyueuropathy, either seen on objective exam only or with mild to moderate pares&Gas. Three patients manifested motor neuropathy with moderate weakness. Some ototoxicity and encephalopathy was observed. All patients had reversible, non-progressive neuropathy. The development of neuropathy was not related to the pharmacologic pprruaeters of serum level or ha&life. The incidence of neuropathy was related to total drug dose. Patients receiving less than or equal to 10 gms/m’ had an incidence of 4 of 19 (21%) compared to patients receiving more than 10 gms/m’ who had an incidence of 15 of 19 (79%). (HPLC) Pharmacologic evaluation included ultraviolet (UV) or high pressure liquid chromatography measurements of blood, urine, stool, and tumor biopsies. The HPLC identified Ro-059963, the desmethylated compound, as the principal metabolite in both blood and urine. Comparison of sera levels at the time of radiation (4-6 hr) with oral drug dose showed a good correlation, with maximum oral drug absorption not reached. Clfnically significant blood level of 100 &ml could be achkved by a dose of about 2.5 gmlm’ or 65-80 n&g. The pharmacology yielded peak sera levels at 2-4 hr with a mean half life of 15 hr and medfan of 14 hr; the haff lffe was not related to drug dose. Urinary excretion was principally of the active metabolite (9963); total per cent excretion of both compounds in 24 hr ranged from 1265% with a mean of 2ft% and a medhn of 23%. Tumor levels in 2 patients were 80 and 90% of peak sera levels. No increased toxicity of normal tissues in the radiation field was observed. Some interesting patient responses were observed. Further toxicologic, pharmacofogic, and efficacy studies are underway. Misonidazole,

Ro-07-0582, Hypoxic cell sensitizers

INTRODUCTION

tance

cells are known to be more resistant to the effects of ionizing radiation than aerated cells. Hypoxic cells are known to be present in solid tumors in man and animal

models.

Whether

of such

cells

is a limiting

factor

in the local

control of solid tumors (radioresponsiveness) that are treated with fractionated radiotherapy is a subject of current study. Attempts to improve the cell kill of hypoxic cells involve the use of hyperbaric oxygen,

Hypoxic

the radioresis-

Inst., Buffalo, NY 14263. l/Clinical Pharmokinetics Lab., Univ. of California, San Francisco, CA 94143. “Dept of Neurosurgery, Univ. of California, San Francisco, CA 94143. VMammalian Biology Group, Los Alamos Scientific Lab., Los Alamos, NM 87545. Reprint requests to: Todd H. Wasserman, MD,, Department of Radiation Oncology, M-330, University of California, San Francisco, CA 94143, U.S.A. Accepted for publication 15 January 1979.

tsponsored by the National Cancer Institute (NCI), Division of Cancer Treatment (DCT) through the Radiation Therapy Oncology Group (Grant No. CA21439). Misonidazole supplied by the Drug Development Program, DCTNCI. Some patients were admitted to the General Clinical Research Center, University of California, San Francisco. Presented in part at the annual meeting of the American Society of Therapeutic Radiologists, Los Angeles, 2 November 1978. $Dept of Radiation Oncology, Univ. of California, San Francisco, CA 94143. $Dept of Radiation Medicine, Roswell Park Memorial 775

776

Radiation

Oncology

0 Biology 0 Physics

high linear energy transfer radiation, and hypoxic cell sensitizers.‘.’ The nitroimidazoles appear to have the greatest potential of the compounds tested to date as hypoxic cell sensitizers, because of their superior pharmacologic properties and greater effectiveness in mammalian cells.2-4~”Among these, metronidazole (Flagyl) and misonidazole (Ro-07-0582) have proven to be active in animal model systems2~6~“~13~‘6~‘ and7 are now in early clinical evaluation in several countries.‘9~‘2*14*21-24 These compounds act by mimicking oxygen in fixing the DNA damage caused by radiation. They thus improve the radiosensitivity or cell kill of hypoxic cells, as tested both in vitro and in vivo. They apparently do not sensitize normal oxic cells to radiation. Thus there is a selective effect between tumors and normal tissues. New data suggests that misonidazole may be directly cytotoxic to hypoxic cells without radiation. Preclinically, misonidazole was felt to possibly be a better radiosensitizer than metronidazole. Misonidazole is a 2-nitroimidazole compared to metronidazole which is a 5-nitroimidazole. Initial clinical pharmacologic and toxicologic evaluation of misonidazole by Dische et al. in England8~9*‘2and by Urtasun et al. in Canada**,” have provided some qualitative and quantitative data on these drug properties for single doses and some multiple dose schedules. Pharmacologic studies using ultra violet (UV) and gas liquid chromatography (GLC) assays established that: peak serum levels of nitroimidazoles occur at l-4 hr; serum half-life values ranged between 9-17 hr; peak serum levels in pg/ml were linearly proportional to the drug dose administered in mglkg; good percentages (SO-70%) of serum levels were detectable in tumor biopsies (both in the tumor center and periphery) and in the cerebrospinal fluid (CSF); and that acute toxicity of the oral formulation was bad taste and occasional nausea and vomiting. Multiple dose schedules revealed the dose limiting toxicity to be sensory peripheral neuropathy; the incidence of this toxicity apparently was related to total drug dose given, dose frequency, and size of each drug dose.9 Studies are ongoing in both these centers to further delineate the properties of this neurotoxicity as it relates to drug schedule. Initial clinical efficacy trials have also just begun in both centers. The National Cancer Institute, Division of Cancer Treatment (NCI-DCT) in conjunction with the Radiation Therapy Oncology Group (RTOG), has embarked upon a program to develop and apply hypoxic cell sensitizers in the United States according to the Committee on Radiation Oncology Studies (CROS) Research Plan.’ This program involves: an NCI-DCT supported Hypoxic Cell Sensitizer Coordinating and

June 1979, Volume

5, Number

6

Review Committee; direct NCI-DCT supported contracts in drug synthesis; in vitro and in vivo tumor model evaluation; small and large animal toxicology: dose formulation; and clinical evaluation in Phases I-III clinical trials. This entire mechanism is set up to work in conjunction with the existing chemotherapy drug development program.“,” As a first clinical step in this program, misonidazole was obtained by the NCI-DCT and an application for initial clinical trial was filed with the U.S. Food and Drug Administration (FDA) and approved for activation in July 1977, via the RTOG. This paper will describe the early clinical results of this initial Phase I study, which is still ongoing. The study has involved a team of investigators in 2 clinical centers. The objectives of this trial were: to determine the maximum tolerated dose of misonidazole, administered orally, on a once per week dose schedule for up to 6 weeks; to determine the qualitative and quantitative toxicities of the drug; to determine the pharmacologic properties of the drug as it related to serum peak levels, half-life, renal excretion, metabolism, and tumor tissue levels. When possible, objective parameters of tumor effect of the combined drug and radiation were studied, although this was not a primary objective of this trial. METHODS AND MATERIALS Patient population The patient population of this study was typical for a Phase I study except that no patient received the drug alone; all patients received some form of palliative radiotherapy in conjunction with the drug. The patients were all adults with advanced solid tumors of mixed primary sites and histologies. In most patients, prior surgery, radiation and chemotherapy had failed to cure their disease. Some patients were already at an advanced state and entered directly into the study. They had normal blood counts, renal function and liver function, (some patients had mild liver enzyme elevations). Careful pre-drug neurologic testing established that some patients had subclinical (objective signs) peripheral neuropathy secondary to their cancer or nutritional state. No patient was entered with a pre-existing major neurologic dysfunction. All patients gave signed informed consent using forms which had received multiple committee approval. TREATMENT SCHEMA The misonidazole was prepared as a scored 0.5 gm enteric coated tablet. All doses were oral and prescribed to the nearest 0.25gm/m2. The dose schedule prescribed single oral weekly doses for either 3 or 6 weeks alternately in separate groups of 3 or more patients each. The initial dose was 1.0 gm/m*; the dose was escalated by 1.0 gm/m* in successive groups

771

Misonidazole study 0 T. H. WASSERMANet al.

of 3 or more patients for each dose schedule. Radiation therapy was given at a dose of 600-800 rad, at an interval of 4-6 hr after each drug dose once per week, to allow for anticipated maximum tumor tissue drug concentration. There was no dose escalation in a given patient. The rationale for the weekly dose schedule was early clinical information from England concerning peripheral neuropathy on more frequent dose schedules. We desired the greatest frequency of drug administration with the highest degree of initial safety. Table 1 shows the schema in dose, dose frequency, total dose and number of patients. A total of 40 patients (Roswell Park Memorial Institute (RPMI)15: University of California, San Francisco (UCSF)25) were entered representing 149 drug doses. Treatment was listed as incomplete if a patient received one or more doses but did not complete the planned dose schedule. The reasons for the incomplete treatment patients are given in the table. The initial dose was escalated to 5 gm/m* x 3 weeks and 3 gm/m* x 6 weeks. Individual drug doses ranged from 22-143 mg/kg. Because of toxicity (described below) and because frequency of administration rather than higher dose levels will be important in scheduling the misonidazole with radiation further exploration of higher dose levels was not deemed desirable. We are currently evaluating more frequent dose schedules of twice and thrice per week.

measured by either an ultraviolet spectrophotometry (UV) method or a high pressure liquid chromatography (HPLC) method developed at UCSF.” The HPLC method is more sensitive than the UV because it separates metabolites from the parent compound. Comparisons of the 2 methods were made initially. No interfering endogenous compounds were found in baseline measurements. The sensitivity of the HPLC method was < lFg/ml. Both methods allowed for measurements of serial blood levels, which allowed for drawing of serum clearance curves and calculation of crude half-lives. Half life calculations were based on the observable peak serum level. A computer modeling of selected serum clearance curves has provided other data. We obtained serial urine samples and calculated urinary excretion of both the parent compound and its principal metabolite. Other blood levels were obtained at the time of radiation (4-6 hr after the drug dose) as this was felt to be useful for larger clinical trials where only limited pharmacology will be practical. Where patients had multiple such levels for multiple drug doses, a mean 4-6 hr serum level was calculated. RESULTS Pharmacology results Figure 1 shows typical UV serum concentration curves. Serum concentrations are expressed in pg/ml on a linear scale. Drug doses were given on a gm/m*

Table 1. Misonidazole Phase I study. Schema of patient entry by dose level and number of doses Incomplete treatment* Dose

gin/m’

Weekly doses Total dose gm/m* No. patients

D

(No. of patients) W ND

1.0

3

3

3

0

0

0

2.0 3.0

3 3

6 9

4 4

0

0

0

4.0 5.0 :::

3 3 6

12 15 12 6

3 4 93

1 1 0 1

0 0 2 1

2.5 3.0

6 6 149 Drug doses

15 18

5 5 40

0 0 4

2 5

1 0 0 0 2 0 0 5

ii

NO dose escalation in a given patient. Radiation given 4-6 hr after drug.

tIncomplete treatment because of premature death (‘W”), patient withdrawal (“W”), lack of drug (“ND”). TOXICOLOGIC

AND PHARMACOLOGIC EVALUATION The toxicologic evaluation consisted of pre-drug and serial clinical histories, exams, neurologic testing, and blood chemistries and counts. The pharmacologic evaluation consisted of blood and other levels

basis but calculations were made on a mg/kg basis also. Half-life calculations were performed as described above. Figure 2 shows a typical HPLC serum concentration curve. The HPLC allowed for measurement of serum concentrations of a metabolite Ro-059963.

Radiation Oncology 0 Biology ?? Physics

778

120

June 1979, Volume 5, Number 6

PATIENT 9 (gPMI )2 gmlm2.67

r

mg/kg

= 12.5 hrs

T%D@WOl

2 = 12.S hrs

00

3=

12hrs

60

HOURS AFTER MlSONlDAZOLE

Fig. 1. Serum concentration curves of a patient receiving 2 gm/m’ of misonidazole or 3 gm total. Each serum curve is for a dose. The calculated half-lives based on the peak serum concentration are shown. MISONIDAZOLE - PHASE I

00

J.L.415 2gm/d

70

O(2)

4.5gm 2.2ml

220 7

sj

0.

Neuropahy

200

o

180

0 No

N=19

neuropathy

??

N=17

f !!i

0 0 /’

140 /'

s

f

2 :

100

g

80

2

2 $

4020_

,.sr"' .

0

10

I

I

I

I

30

50

70

90

DOSE

mg/kg

I

110

I

I

130

150

Fig. 3. Relationship of mean 4-6 hr serum levels to oral dose given in mg/kg. The dashed line represents a one to one correlation of the two parameters. Also, the relationship of neurotoxicity to each parameter is shown. HOURS

Fig. 2. Serum concentration curve of a patient receiving 2 gm/m* or 45 mg/kg or 4.5 gm total of misonidazole. The serum curve is for the patient’s fifth drug dose with the drug levels for the other drug doses shown as points with the number of the drug dose in parentheses. The calculated half life of 14 hr is shown. This is based on the observed peak serum concentration of 64 pg/ml at 5 hr. A serum concentration curve is also shown for Ro-O5-9!%3.

This metabolite is the product of a terminal desmethylation of the parent compound. Low levels of Ro-05-9963 were seen in serum. Figure 3 shows the relationship between mean 46 hr serum levels (of misonidazole only) and drug dose expressed in mg/kg. The points show that a plateau of maximum oral absorption of drug was not reached as higher doses continued to give higher sera

levels. Blood levels are more than proportional to drug dose, expressed in n&kg. As shown in Figs. 1 and 2, peak sera levels were usually at 2-4 hr and were slightly higher than the levels of 4-6 hr. No prolonged (after 48-72 hr) sera levels were observed. All baseline levels before the next weekly dose of drug were less than 1% of administered drug. Table 2 gives the 6 different dose levels in gram/m* and the corresponding mean 4-6 hr blood levels f 1 standard deviation. These mean values are somewhat (lO-25%) lower than the peak values, and represent either UV values or the sum of the HPLC values of both parent compound and metabolite (Ro-059963). The metabolite accounted for 5-W% of the total HPLC value. Both Table 2 and Fig. 3 demonstrate that a blood level of about 100 pg/ml can be obtained by a drug dose of about 2.5 gm/m* or 65-M mglkg.

Misonidazole study 0 T.

Table 2. Misonidazole Phase I study. Relationship to mean 4-6 hr serum level

Dose gm/m’

No. of doses (determinations)

WASSERMAN

779

et al.

MISONIDAZOLE

of dose

- PHASE I o Newopothy 0 No nwropathy

0

Mean 4-6 hr serum level pglmlt

k Standard deviation of:

30 74 106 116 175 183

+9 k 15 4 16 226 +-56 k60

17 52 22 25 8 7

1.0 2.0 2.5 3.0 4.0 5.0

H.

N=O N=6

0

??

0

0

00

??

t(0582 + 9963). Half-life determinations were done on 24 patients (as not all patients were suitable for pharmacologic evaluation which required hospitalization); these evaluations yielded 33 results with a range of 3-25 hr

and a mean of 15 hr (standard deviation 2 5) and a median of 14 hr. Figure 4 shows that no apparent relationship was observed between half-life and drug dose in mg/kg.t A computer analysis of 17 serum curves by the HPLC method, yielded a somewhat shorter mean half-life of 10 hr (& 1 hr standard deviation; median = 10.2 hr). The HPLC method allowed for analysis of urinary This was done in 16 patients. Both excretion. misonidazole and the principal metabolite (Ro-05 9963) were identified in the urine. About two-thirds of the urine excretion was of Ro-05-9963. The total per cent excretion of both species over 24 hr in 14 patients (16 determinations) ranged from 1265% of the oral dose with a mean of 28% (standard deviation 2 15) and a median of 23%. Figure 5 shows that no MISONIDAZOLE 26

??

r

0

- PHASE I

10

30

50

70

90

DOSE

mg/kg

110

130

150

Fig. 4. Relationship of half-life to drug dose and the relationship of neurotoxicity to each parameter. tseparate calculations (not shown in the figure) also demonstrated no apparent relationship between half-life

I 0

I IO

I 30

I 50

I 70

I 90

I 110

I 130

I 150

DOSE mg/kg

Fig. 5. Relationship of per cent total urinary excretion in 24 hr to drug dose and relationship of neurotoxicity to each parameter.

apparent relationship was observed between the per cent urinary excretion and oral drug dose in mg/kg. There was also no apparent relationship between per cent urinary excretion and drug dose in grams/m’. Vomitus levels of drug were measured in several patients and were high if the patient vomited acutely (within 1 hr) after the drug dose. Blood levels in such cases were usually lower. Stool levels were measured in 3 patients and were less than 1% over 24 hr including 1 patient with liver disease. Perhaps longer stood collections would have reflected higher levels. Bacterial decomposition of misonidazole is known to occur with coliform bacteria preclinically, but probably into chemical species not detected by our assays. Tumor levels in excised skin nodules were measured in 2 patients and found to be 80 and 90% of peak sera levels. Excision time was at the time of radiation (4-6 hr), and the samples were placed in a sealed specimen container and frozen within 1 hr. Toxicity results The toxicity observed from the dose schedules in this study were gastrointestinal and neural. No apparent renal, liver, or bone marrow toxicity was found. There was also no enhanced radiation reaction of the normal tissues in the radiation field, for the short period of follow-up. The gastrointestinal toxicity consisted of acute nausea and vomiting. About one-fourth of patients experienced nausea alone and about one-fourth had associated vomiting. The vomiting usually occurred within 1-12 hr after the drug dose. It often occurred and drug dose in grams/m*.

780

Radiation Oncology 0 Biology 0 Physics

at the time of or after the radiation, and was related to gastrointestinal effects and the often additive radiation nausea. The nausea and vomiting was dose related in that patients at the higher dose levels had a higher incidence. This may be related to the number of tablets required to swallow (15-20 at higher doses). and another dose formulation might decrease this toxicity. One of 3 patients at 5 gm/m* had severe nausea and vomiting. Five to six gram/m* would seem to be the highest tolerable individual dose. The nausea and vomiting was ameliorated somewhat by pre-treatment of the patients with antiemetics (compazine suppository 25 mg. 1 hr before drug dose). Diarrhea was observed in 2 patients at high total doses, and it is unclear whether this represents a drug toxicity. The neurotoxicity was primarily a peripheral, sensory, polyneuropathy of hands and feet. This was manifested either by changes on objective serial neurological exams measuring vibratory sense or with paresthesias. We were very liberal in calling any neurologic change a drug-related neuropathy although many of the patients had advancing cancer, and a worsening nutritional state. Some patients were also suspected of developing central nervous system metastases. The incidence of neuropathy is clearly related to total drug dose as expressed in total grams or total grams/m* (Table 3). Patients receiving less than or equal to lOgrams/m* total dose had an incidence of neuropathy of 4 of 19 (21%), compared to patients receiving more than lOgrams/m* who had an incidence of 15 of 19 (7%). (Two patients were nonevaluable for neurotoxicity because of premature death after only 1 dose of drug.) More important, the severity of the neuropathy seemed to increase with increasing total doses. There is no apparent threshold dose for developing or not developing neuropathy. Most of the neuropathies occurred after the patient

June 1979, Volume

5,

Number 6

approached lOgm/m*, usually within 1 week of that total dose. The incidence of neuropathy below lOgm/m’ would appear to be partially related to the advanced state of the patients and the very careful neurologic screening. At its worst, the neuropathy was associated with weakness and inability to ambulate, for a period of about 1 week. This occurred in several patients at the 3 gm/m*x 6 dose level. None of the neuropathies progressed when the drug was stopped. Instead the neuropathy improved with time, lasting several weeks to months. A patient who had had a prior vincristine neuropathy, developed rapid, severe misonidazole neuropathy suggesting an additive toxic effect. No seizures were observed except for 1 patient who had probable brain metastases from an eye melanoma. Obtundation and mental confusion was observed in 1 patient each. (Two other patients on the recent twice per week dose schedule have developed symptoms of such encephalopathy; this was related to dehydration.) Ototoxicity manifested as decreased hearing of short term duration was observed in 3 patients at 3 gm/m* and 1 patient at 2.5 gm/m*. We are now in the process of obtaining serial audiograms on selected patients. As is shown in Figs. 3-5 the development of neuropathy was apparently unrelated to the pharmacologic parameters of blood level, half-life, or per cent urinary excretion. These parameters would not distinguish those patients who would be expected to develop neuropathy. Total dose was the only predictive factor. A computer analysis of serum concentration times time (c x t; gm-min/ml) established a was relationship of c x t to drug dose. Neurotoxicity related to c x t but only as an indirect measure of total drug dose (Fig. 6). Those patients who had objective signs of pre-drug neuropathy secondary to

Table 3. Misonidazole Phase I study. Incidence of neurotoxicity as related to total patient dose either expressed as total gm or gm/m* Total dose in gm o-5 > 5-10 > lo-15

15-20 > 20-25 > 25-30 > 30-35 > 35-40 Total >

Incidence of neuropathy o/4 I/6 4111 414 I 719 213 0

5/21 (24%)

14117(82%)

Total dose gmlm’

Incidence of neuropathy

O-3 4-6 7-9 IO-12 13-15 16-18

015 218 216 9/12 617 0

Total

19138

4/19 (21%)

15119(79%)

1/1J 19/38(50%)

Two patients not applicable for analysis of neuropathy because of premature death, after only 1 dose.

Misonidazole

study 0 T. H.

WASSERMAN

et

al.

781

The development of the hypoxic cell radiosensitizers opens the door for new avenues of radiotherapy research-pre-clinically and clinically. Misonidazole has been shown to be an active hypoxic cell radiosensitizer in experimental systems.6 The expectation is that this will allow for a selective increase in the cell kill of radiation against cancer cells, with no increased normal tissue damage.” The further expectation is that hypoxic cells which are still clonogenic are perhaps resistant to the cell killing of radiation and lead to persistence of local regional tumors and clinical failures. The adequate control of local regional cancer is known to be a major problem despite the attention paid recently to the parallel problem of occult distant metastases.3 It is not within the context of this paper to review the pre-clinical data and rationale of misonidazole and those interested may read several selected reviews.“.” The initial clinical trials of misonidazole by Dische

et a1.22v24in et a1.8,9s’2 in England and by Urtasun Canada established some basic clinical knowledge regarding the pharmacology and toxicology of this drug. This trial to some extent was repetitious of their work for three reasons. First, the amount of patient information available was not so large, that further confirmation of their results would be wasteful. Second, the regulations of the U.S. Food and Drug Administration required an initial Phase I trial in the U.S. before other larger clinical trials could be undertaken. Third, they did not explore a limited number of doses on a weekly schedule. We feel our data both adds to and supplements that of Dische et al. and Urtasun et al. The findings of a dose limiting peripheral neurotoxicity at multiple dose schedules has been confirmed and our data have also added to the knowledge of this toxicity being related to total dose. We have established no apparent relationship of this toxicity to pharmacologic variables of blood level, halflife or urinary excretion. The neurotoxicity may be related to the concentration x time (c times t) exposure of the peripheral nerves to the drug. Since the time course of drug action is very short (seconds or less at the time of radiation, once it reaches the tumor in good concentration) some selective alteration of the drug excretion or metabolism after the radiation might decrease neural exposure and decrease neurotoxicity. It is of interest to note that Ro-05-9963 the principal metabolite of misonidazole, which we have identified both in blood and urine is also an active radiosensitizer experimentally6 and may be more useful clinically because of its lower lipid solubility and higher urinary excretion. We plan to continue to apply the HPLC pharmacology method we developed in order to better understand the pharmacology of misonidazole and Ro-05-9963 and possibly alter the neurotoxicity. Indeed, it is not known which of these two compounds or if another as yet unknown metabolite is the neurotoxic species. An animal model for the neurotoxicity is needed to further study the questions raised. Our pharmacology data confirms that of Dische et al. and Urtasun et al. in showing good absorption of the oral misonidazole with blood levels achieved proportional to or above the drug dose given on a mg/kg basis. We have not observed lower blood levels with subsequent drug doses in a given patient as seen by Johnson et al.” in stumptail monkeys and thus cannot confirm an enzyme induction in man. Perhaps this is because of the infrequent, weekly dose schedule. t

tWe also observed no apparent relationship serum half-life and oral drug dose (expressed

mg/kg or gram/m*). Other studies have demonstrated apparent increase in half-life with larger oral doses.’

360

O=Neuropothy

320 _

.=No

N=9

Neuropothy

N=8

c=.a99

280 E .G

240 -

E 200 f

160-

ci

12080 40 -

,/I 0

15

30

I 45

I 60

I 75 DOSE

I 90

I 105

I 120

I 135

1 150

mg/kg

Fig. 6. Relationship of area under serum drug curve in gm-min per ml. to dose in mglkg, and relationship of neurotoxicity to each parameter. The solid line is a least square fit of the data points with a confidence limit of 0.899. their cancer or nutritional state did not seem to be at The one a higher risk for drug-related neuropathy.

exception was the patient with the vincristine neuropathy discussed above. Overall toxicity seemed to be worse in patients who were older, in those with poorer performance status, poorer nutritional status and those in whom a well hydrated state was difficult to maintain. Evaluation of effect of the misonidazole was not an objective of this study. However, our observations have yielded some good responses, particularly in patients with squamous cell carcinomas. Tables 4a-f provide all the individual patient data. DISCUSSION

between either as

an

1.5 1.5 1.3 2.0

1.25

1.8

UCSF UCSF UCSF RPM1

RPM1

UCSF

V.C. L.M. E.J. P.F.

V.F.

B.F.

1 2 3 4

5

6

24

22

1.25

1 10.5

26 29 30 24

4.5 4.5 3.9 4.0

3 3 3 2

6

mglkg

31

10

3 14 11 6

28 25 26 31(1 only) 38

Serum T; in hr

Mean 4-6 hr sera level pdml (0582 only)

RPM1

E.G.

17

1.25

5 6

1.6

RPM1

E.C.

12

3

1.7

RPM1

H.L.

11

3 5

1.4 1.6

RPM1 RPM1

E.J. A.R.

9 10

3

2.1

RPM1

C.L.

8

3

1.7

MZ

RPM1

C.G.

7

Instit.

Initials

Pt no.

2 gm/m’ x no. doses

46 62.5

52

10.5 15.0 15.0

67 50

42

60

mg/kg

9.0 15.0

12.0

10.5

Total dose in gm

77(4 only) 104(3 only)

78

82 87(1 only)

63

82

23

:f

Yes Gr I

NA

No No No

No

11 24.5 Tumor biopsy = 74 pg/gm Blood = 75 pg/ml 21

i2.5,

Yes

Yes

No

No No

No 12.5

20,

Neuropathy and grade

No

% urine excretion 24 hr 0582 9963 Total

weekly dose schedule

12

12 22

i8.5, 19,215 22

20

Serum T$ in hr

Phase I study (R.T.O.G.)

Mean 4-6 hr sera level pglml (0582 only)

Table 4b. Misonidazole-Ro-07-0582.

11

6 7

Neuropathy and grade

weekly dose schedule

% urine excretion 24 hr 0582 9963 Total

Phase I study (R.T.O.G.)

Weekly dose schedule = 40 pts. RPM1 values are UV and tend to be slightly higher because they include metabolites. tRPM1 = Roswell Park Memorial Inst. UCSF = University of California, San Francisco.

M*

1nstit.t

Initials

Pt no.

Total dose in gm

1 gm/m’ X no. doses

Table 4a. Misonidazole-Ro-07-0582.

Expired Did not Lack of Expired of dose Did not Lack of

complete dose schedule drug before completion schedule complete dose schedule drug

Comments and status post dl drug as of 9/l/78

Expired-7 weeks Expired-9 weeks Expired4 weeks Did not complete dose schedule Refused Expired before completion of dose schedule at 1 week Alive 12.5 months

Comments and status post dl drug as of 9/l/78

Initials

J.L.

R.S.

S.M.

N.W.

K.D.

A.G.

Initials

L.R.

H.F.

F.F.

S.A.

R.F.

Pt no.

15

16

18

19

20

21

Pt no.

27

31

32

33

35

UCSF

RPM1

UCSF

UCSF

UCSF

1.5

2.0

2.42

1.64

1.62

Instit.

24.0

21.0

17.5

14.0

21.0

27.0

Total dose in gm

48

56

56

54

58

45

mg/kg

58

66

61

60(3 only)

80

58

5

3

6

6

6

18.75

10

36

24

24

Total dose in gm

85

62

54

67

71

mg/kg

119(4 only)

115( 1 only)

98(5 only)

94

9 12

32

15

6 13 22 37 (Hydration) Yes Gr II

Yes Gr II No

No

Neuropathy and grade

8

26

Neuropathy and grade

Yes Gr IV

Yes Gr II 0% in stool-Tumor at 6.5 hr 0582 = 162 &gm 45 20 65 Yes Gr II Yes Gr II No

18

vomitus 3 hr after drug 0582 = 3700 pg/ml 9963=0

13

10

Serum T; in hr

Yes Gr I

weekly dose schedule

% urine excretion 24 hr 0582 9963 Total

Phase I study (R.T.O.G.)

Mean 4-6 hr sera level pLg/ml (0582 only) 90

7 15

3

8 7 0% in stool 27 15

% urine excretion 24 hr 0582 9963 Total

weekly dose schedule

Tumor at 7 hr 0582 = 50 pglgrn 88% peak sera level 9963 = 8 pg/gm 200% peak sera level 18 No

11.75 14.25

7.5

16

14

Serum Tf in hr

Phase I study (R.T.O.G.)

Mean 4-6 hr sera level pglml (0582 only)

Table 4d. Misonidazole-Ro-07-0582.

2.5 gm/m* X no. M* doses

6

6

5

4

6

6

2.02

1.73

1.68

1.75

1.7

2.2

MZ

UCSF

UCSF

UCSF

UCSF

UCSF

UCSF

Instit.

2 gmlm’ X no. doses

Table 4c. Misonidazole-Ro-07-0582.

CA

Incomplete-patient became jaundiced due to liver mets Patient refused last dose and last Rx not given-expired 3 days later-Had prior Vincristine

Dead 2 months

Alive 7.5 months

Dead 9 months

Comments and status post dl drug as of 9/l/78

Dead 9 months

Dead 4.5 months Good response-tongue

Expired 4 weeks, did not complete dose schedule, liver dysfunction-mets Pt. refused last dose of drug because of persistent N & V Dead 5 months

Died 6 months

Alive 10 months

Comments and status post dl drug as of 911178

Z

E B s

2 B

3:

! R s 2 Y 0 3

$ 9’ 2.

RPM1 UCSF

UCSF UCSF

UCSF

RPM1 UCSF

UCSF

Instit.

UCSF

RPM1

UCSF

UCSF

UCSF

RPM1 RPM1

M.K. R.P.

B.S. L.S.

C.B.

F.M. R.G.

I.W.

S.C.

Initials

V.H.

H.H.

A.J.

G.S.

F.G.

G.B. J.S.

13 14

24 26

36

37 39

46

52

Pt no.

25

28

29

30

41

38 38b

UCSF

Instit.

Initials

Pt No.

1.6 1.9

1.61

1.88

1.68

1.78

1.72

M*

2.2

1.48

2 2.24

l.%

1.62 1.4

1.7 1.7

M*

x

19.5

68

91

61 76

69

89 89

80 85

mg/kg

72(3)

122(4)

130(l) 109(4)

120(5)

112 100

14 20.5

2 3

24 8 9.5

3 1 1

3

25

21

3

5 gm/m* no. doses

Total dose in gm

4grnlm’ X no. doses

143 116

138

130

120

103

97

mglkg

Mean 4-6 hr s&a level pglml (0582 only)

157( 1 only) -

173

187

217

193(2)

106

11 13.75

10 9.75

17 16.5,17.5 pills)

12 5

15 18

17.5

22

Serum Tt in hr

31 17

23 53

12

(;!4% in424 hr

36

s.:tol)

% urine excretion 24 hr 0582 9963 Total

24

No Yes Gr II Yes Gr IV No Yes Gr III Yes Gr IV Yes Gr III

Yes Gr IV No No

No

Yes Gr IV

Yes Gr II No

Neuropathy and grade

weekly dose schedule

19 12

8 35

Yes NA

Neuropathy and grade

weekly dose schedule

% urine excretion 24 hr 0582 9963 Total

Phase I study (R.T.O.G.)

112(2 only) 26(vomited

Serum Ti in hr

Phase I study (R.T.O.G.)

Mean 4-6 hr sera level *g/ml (0582 only)

Table 4f. Misonidazole-Ro-05-0582.

3

18

6 27

1 4 4

28.75

14.25 12.75

3 3 5

10.0 5.0

Total dose in gm

2 1

3gm/m* x no. doses

Table 4e. Misonidazole-Ro-O7-0582.

refusal

refusal

refusal refusal refusal

Patient Patient

withdrew withdrew

after 1 dose after 1 dose

Alive 8 months Took only 6gm for 3rd dose Died-2.5 months

Expired before 3rd dose at 3.5 weeks Severe liver dis. Died 7 weeks

Died at 3 months

Comments and status post dl drug as of 9/l/78

Alive-6 months Incomplete-patient Incomplete-patient Incomplete-patient Alive-6.5 months Incomplete-patient Died-3.5 months Incomplete-patient Dead

Drug not available for last dose Expired before completion of dose schedule Expired 7.5 weeks Dead 4 months

Comments and status post dl drug as of 9/l/78

Misonidazole study @T H. WASSERMANet al.

It is uncertain from the pre-clinical data’what blood level is necessary to achieve specific radiation enhancement ratios in man. (Enhancement ratio expresses the improved cell kill of a given dose of radiation). One problem in interpreting the mouse data is the discrepancy or difference in drug half-life in the mouse vs man (0.5 hr vs 15 hr). Blood levels of lOO-2OOpg/ml may be needed to establish enhancement ratios of 1.75-2.0. At 50 pg/ml blood level, an enhancement ratio of 1.25-1.5 may be expected. As can be seen in Table 2, such blood levels (50200 pg/ml) (at 4-6 hr, and not at peak which would be higher) are achievable with doses of 1.5-5.0 gm/m2. The question is how frequently can such doses be given if neurotoxicity becomes dose limiting with total doses greater than 10 gm/m*. It is unlikely that misonidazole will be the ideal radiosensitizer since it will not be possible to administer it in frequent dose schedules (more than twice per week) at doses which are going to yield good blood levels (more than 50-100 pglml). We are participating in an ongoing project of the National Cancer Institute that attempts to develop other better drugs in the same class.20 The clinical lag time between good serum levels and good tumor levels has not been well established.

Some data by Urtasun suggests a lag time of l2 hr.22,24However more serial tumor levels are needed but these are often logistically difficult clinically. We gave our radiation 4-6 hr after the drug dose by

785

adding the known time of peak blood level of 2-4 hr to the proposed lag time between blood and tumor of l-2 hr. We feel we established some knowledge of the safety and pharmacology of misonidazole at the weekly dose schedules, so that further clinical trials seem justified. Our study was not designed to establish any efficacy data; nevertheless we have seen several patients who have had encouraging clinical responses. We have joined with other members of the RTOG in writing a series of disease specific Pilot Phase II trials using weekly schedules of both radiation and misonidazole (at doses of 2-4gm/m2; total doses 12gm/m2 over 3 weeks or 15 gm/m2 over 6 weeks). Several reports suggest that weekly radiation (by itself) might be useful in some solid tumors by either more practical advantages or by better radioThe RTOG pilot studies are responsiveness.“.” designed to be logical steps along this avenue of clinical research. We are continuing to do toxicologic and pharmacologic evaluation on patients receiving twice or thrice per week dose schedules. The future clinical efficacy of misonidazole and other hypoxic cell sensitizers is unknown. We are encouraged to continue to pursue their clinical usefulness and understanding of their limitations. Misonidazole is sufficiently tolerable to continue clinical research in conjunction with our colleagues from the United States and other countries.

REFERENCES 1. Adams, G.E.: Chemical radiosensitization of hypoxic cells. Br. Med. Bull. 29: 48-53, 1973. 2. Adams, G.E., Fowler, J.F.: Nitroimidazoles as hypoxic cell sensitizers in vitro and in uiuo. In Modification of Radiosensitivity of Biological Systems, I.A.E.A. Advisory Group Meeting, Vienna, 1976, pp. 103-l 17. 3. Chapman, J.D., Reuvers, A.P., Borsa, J., Henderson, J.D., Migliore, R.D.: Nitro-heterocyclic drugs as selective radiosensitizers of hypoxic mammalian cells. Cancer Chemo. Repts 58: 559-570,

1974.

4. Chapman, J.D., Urtasun, R.C.: The application in radiotherapy of substances which modify cellular radioactive response. Cancer 40: 484-488, 1977. 5. Committee on Radiation Oncology Studies Research Plan: Radiation Sensitizers. Cancer 37: 2062-2070, 1976. of solid 6. Denekamp, J., Fowler, J.F.: Radiosensitization tumors by nitroimidazoles. ht. J. Radiat. Oncol. Biol. Phys. 4: 143-151, 1978.

7. Dische, S., Gray, A.J., Zanelli, G.D.: Clinical testing of the radiosensitizer Ro-07-0582-11. Radiosensitization of normal and hypoxic skin. Clinical Radiology 27: 159-167, 1976.

8. Dische, S., Saunders, M.I., Lee, M.E., Adams, G.E., Flockhart, I.R.: Clinical testing of the radiosensitizer Ro-07-0582: Experience with multiple doses. Br. J. Cancer 35: 567-579,

1977.

9. Dische, S.: Hypoxic cell sensitizers in radiotherapy. ht. J. Radiat. Oncol. Biol. Phys. 4: 157-160, 1978. 10. Ellis, F., Goldson, A.L.: Once-a-week treatments. ht. J. Radiat. Oncol. Biol. Phys. 2: 537-548, 1977. 11. Foster, J.L.: Differential cytotoxic effects of metronidazole and other nitro-heterocyclic drugs against hypoxic tumor cells. ht. J. Radiat. Oncol. Biol. Phys. 4: 153-156, 1978. 12. Foster, J.L., Flockhart, I.R., Dische, S., Gray, A., Lenox-Smith, I., Smithen, C.E.: Serum concentration measurements in man of the radiosensitizer Ro-07-0582: Some preliminary results. Br. J. Cancer 31: 679-683, 1975. 13. Foster, J.L., Willson, R.L.: Radiosensitization of anoxic cells by metronidazole. Br. J. Radiol. 46: 234-235, 1973. 14. Gray, A.J., Dische, S., Adams, G.E., Flockhart, I.R., Foster, J.L.: Clinical testing of the radiosensitizer Ro07-0582-I. Dose tolerance, serum and tumor concentrations. Clin. Radiol. 27: 151-157, 1976. 15. Habermalz, H.J., Fischer, J.J.: Radiation therapy of malignant melanoma--experience with high individual treatment doses. Cancer 38: 2258-2262, 1976. 16. Hall, E.J., Bigalow, J.: Ro-O7-0582 as a radiosensitizer and cytotoxic agent. ht. J. Radiat. Oncol. Biol. Phys. 2: 521-530, 1977. 17. Johnson, R., Gomer, C., Pearce, J.: An investigation of radiosensitizing effects of Ro-07-0582 on hypoxic skin

786

18.

19.

20.

21.

Radiation

Oncology

0 Biology ?? Physics

in primates. Int. J. Radiat. Oncol. Biol. Phys. 1: 593599,1976. Marques, R.A., Stafford, B., Flynn, N., Sadee, W.: Determination of metronidazole and misonidazole and their metabolites in plasma and urine by high performance liquid chromatography. J. Chromatography 146: 163-166, 1978. National Cancer Institute: Division of Cancer Treatment (NCI-DCT); Program Linear Array-New Drug Development Flow-Radiosensitizers. July 1977. Phillips, T.L., Wasserman, T.H., Johnson, R.J., Gomer, C.J., Lawrence, G.A., Levine, M.L., Sadee, W., Penta, J.S., Rubin, O.J.: The hypoxic cell sensitizer programme in the United States. Br. J. Cancer 37: Suppl. III, 276280, 1978. Urtasun, R.C., Band, D. Chapman, J.D., Feldstein,

June 1979, Volume

5, Number

6

M.L., Mielka, B., Fryer, C.: Radiation and high dose metronidazole (Flagyl) in supratentorial glioblastomas. N.E.J.M. 294: 1364-1367, 1976. 22. Urtasun, R., Band, P., Chapman, J.D., Rabin, H.R., Wilson, A.F., Fryer, C.R.: Clinical phase I study of the hypoxic cell radiosensitizer RoM-QS82, a 2-nitroimidazole derivative. Radiology 122: 801-8@, 1977. 23. Urtasun, R.C., Chapman, J.D., Band, D., Rabin, H.R., Fryer, C.G., Sturmwind, J.: Phase I study of high dose metronidazole: a specific in vivo and in vim radiosensitizer of hypoxic cells. Radiology 117: 129-133, 1975. 24. Urtasun, R.C., Chapman, J.D., Feldstein, M.L., Band, R.P., Rabin, H.R., Wilson, A.F., Marynowski, B., Starrveld, E., Shnitka, T.: Peripheral neuropathy related to misonidazole. Incidence and pathology. Br. J. Cancer 37: Suppl. III, 271-275, 1978.