human pharmacokinetics, excretion, and metabolism...

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(CANCER RESEARCH 46, 1513-1520, March 1986] Human Pharmacokinetics, Excretion, and Metabolism of the Anthracycline Antibiotic Menogaril (7-OMEN, NSC 269148) and Their Correlation with Clinical Toxicities1 Merrill J. Egorin,2 David A. Van Echo, Margaret Y. Whitacre, Alan Forrest, Lawrence M. Sigman, Kathrin L. Engisch, and Joseph Aisner Divisions of Developmental Therapeutics [M. J. E., K. L. E., A. F., L. M. S.] and Medical Oncology [D. A. V. E., M. Y. W., J. A.], University of Maryland Cancer Center, Baltimore, Maryland 21207 ABSTRACT In a Phase I study, menogaril (7-OMEN) was administered daily for 5 days/course, every 21-28 days. Dosages of 3.5, 7, 11.5, 17, and 31.5 mg/m2 were infused over 1 h, and dosages of 42,50, and 56 mg/m2 were infused over 2 h. Pharmacokinetics was studied at all dosages. Plasma and urine samples were collected from 24 patients, and bile samples were also collected from 2 patients. 7-OMEN and metabolites were measured by high performance liquid chromatography. 7-OMEN was the major plasma fluorescent species at all times, with only trace amounts of N-demethyl menogaril observed. 7-OMEN disappeared from plasma biexponentially with f./,a 0.19 ±0.04 (mean ±SE) h and tv,ß13.22 ±1.54 h. Plasma pharmacokinetics of 7-OMEN was linear from 3.5-56 mg/m2; area under the curve increased pro portionally with dosage. Total body clearance of 7-OMEN was 28.18 ±3.33 liter/nf/h, Vcwas 224 ±30.8 liter/m2, and Vsswas 370 ±25.7 liter/m2. Plasma pharmacokinetics of 7-OMEN stud ied on multiple days of a given course were similar. Urinary excretion of 7-OMEN and fluorescent metabolites accounted for 5.4 ±0.4% of the daily dose. Parent compound still represented >80% of urinary drug fluorescence after 24 h. W-demethyl men ogaril was the only other fluorescent drug species detected in urine. In two patients with biliary tract drains, biliary excretion of drug fluorescence accounted for 2.2-4.2% of the daily dose. Only 7-OMEN and A/-demethyl menogaril were detected in bile by high performance liquid chromatography and thin layer chro matography. 7-OMEN was the major fluorescent biliary species, but, by 24 h, A/-demethyl menogaril accounted for approximately 40% of biliary drug fluorescence. When considered in light of each patient's observed toxicities, excellent relationships were observed between the plasma area under the curve of 7-OMEN and the percentage of decreases in WBC and absolute neutrophil count. These latter findings should be useful in developing more precise and intelligent dosing schemes for 7-OMEN. INTRODUCTION Anthracycline antibiotics, of which doxorubicin and daunorub- icin (Fig. 1) are the most important representatives, are a major class of antitumor agents. However, these two agents produce Received 6/27/85; revised 11/8/85; accepted 11/11/85. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 'This work was supported in part by Contract NO1CM27541, and Grant IP50CA32107 awarded by the National Cancer Institute, Department of Health and Human Services, Bethesda, MD 20205. 2To whom requests for reprints should be addressed. considerable toxicity and have limited efficacy against many neoplasms. Therefore, the search continues for anthracycline antibiotics with improved therapeutic indices, altered spectra, or activity of both (1). 7-OMEN3 is a semisynthetic derivative of the anthracycline antibiotic nogalamycin (Fig. 1), a drug with significant experimen tal antitumor activity but prohibitive toxicity in animals (2). 7- OMEN has been introduced into Phase I clinical trials based on its broad spectrum of in vivo activity against animal tumors, its demonstrated activity after p.o. as well i.v. administration (2, 3), and its reduced cardiotoxic potential (4). The mechanism of action of 7-OMEN may differ from that of doxorubicin. 7-OMEN is most toxic to cells in d phase, whereas doxorubicin is most toxic to cells in S phase (2). In addition, while doxorubicin interacts strongly with DNA, 7-OMEN interacts only weakly with DNA and is highly cytotoxic at concentrations which do not inhibit nucleic acid synthesis (2, 5). Finally, when various tumor lines are incubated in vitro with 7-OMEN or doxorubicin, 7-OMEN fluorescence is localized mainly in the cytoplasm, whereas doxorubicin fluorescence accumulates pri marily in the nucleus (2, 6). Previous studies have described the plasma pharmacokinetics of 7-OMEN in beagle dogs (7), rabbits (8), and mice (9, 10), while our laboratory has defined the organ distribution of 7-OMEN in mice and rabbits (8, 9), and the urinary and biliary excretion of 7-OMEN by rabbits (8). During the performance of a Phase I study at our center, we had the opportunity to define the human plasma pharmacokinet ics and urinary excretion of 7-OMEN and its metabolites at multiple dosages, and to investigate the biliary excretion of these compounds in two patients with biliary drains. Finally, when we analyzed these plasma pharmacokinetic data and compared them with the toxicity observed in each patient, an excellent relationship was apparent between the plasma pharmacokinetics of 7-OMEN in any patient and the pharmacodynamics resulting from 7-OMEN therapy of that patient. This paper presents these results. MATERIALS AND METHODS Patient Selection. All patients entering this trial had to fullfill the following criteria: histological proof of malignant disease which had failed conventional chemotherapy or for which no conventional therapy existed, recovery from all toxicities from prior treatments and passage of at least 4 weeks from any prior chemotherapy or radiotherapy, a minimal life 3The abbreviations used are: 7-OMEN, menogaril; HPLC, high performance liquidchromatography;TLC, thin layer chromatography;AUC, area under the curve of plasma drug concentration versus time; Vc,volume of the central compartment; V„, steady-state volume of distribution. CANCER RESEARCH VOL. 46 MARCH 1986 1513 on July 15, 2018. © 1986 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from

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(CANCER RESEARCH 46, 1513-1520, March 1986]

Human Pharmacokinetics, Excretion, and Metabolism of the AnthracyclineAntibiotic Menogaril (7-OMEN, NSC 269148) and Their Correlation withClinical Toxicities1

Merrill J. Egorin,2 David A. Van Echo, Margaret Y. Whitacre, Alan Forrest, Lawrence M. Sigman,

Kathrin L. Engisch, and Joseph Aisner

Divisions of Developmental Therapeutics [M. J. E., K. L. E., A. F., L. M. S.] and Medical Oncology [D. A. V. E., M. Y. W., J. A.], University of Maryland Cancer Center,Baltimore, Maryland 21207

ABSTRACT

In a Phase I study, menogaril (7-OMEN) was administereddaily for 5 days/course, every 21-28 days. Dosages of 3.5, 7,11.5, 17, and 31.5 mg/m2 were infused over 1 h, and dosagesof 42,50, and 56 mg/m2 were infused over 2 h. Pharmacokinetics

was studied at all dosages. Plasma and urine samples werecollected from 24 patients, and bile samples were also collectedfrom 2 patients. 7-OMEN and metabolites were measured byhigh performance liquid chromatography. 7-OMEN was the major

plasma fluorescent species at all times, with only trace amountsof N-demethyl menogaril observed. 7-OMEN disappeared from

plasma biexponentially with f./, a 0.19 ±0.04 (mean ±SE) h andtv, ß13.22 ±1.54 h. Plasma pharmacokinetics of 7-OMEN waslinear from 3.5-56 mg/m2; area under the curve increased pro

portionally with dosage. Total body clearance of 7-OMEN was28.18 ±3.33 liter/nf/h, Vcwas 224 ±30.8 liter/m2, and Vsswas370 ±25.7 liter/m2. Plasma pharmacokinetics of 7-OMEN stud

ied on multiple days of a given course were similar. Urinaryexcretion of 7-OMEN and fluorescent metabolites accounted for

5.4 ±0.4% of the daily dose. Parent compound still represented>80% of urinary drug fluorescence after 24 h. W-demethyl men

ogaril was the only other fluorescent drug species detected inurine. In two patients with biliary tract drains, biliary excretion ofdrug fluorescence accounted for 2.2-4.2% of the daily dose.Only 7-OMEN and A/-demethyl menogaril were detected in bile

by high performance liquid chromatography and thin layer chromatography. 7-OMEN was the major fluorescent biliary species,but, by 24 h, A/-demethyl menogaril accounted for approximately

40% of biliary drug fluorescence. When considered in light ofeach patient's observed toxicities, excellent relationships were

observed between the plasma area under the curve of 7-OMEN

and the percentage of decreases in WBC and absolute neutrophilcount. These latter findings should be useful in developing moreprecise and intelligent dosing schemes for 7-OMEN.

INTRODUCTION

Anthracycline antibiotics, of which doxorubicin and daunorub-

icin (Fig. 1) are the most important representatives, are a majorclass of antitumor agents. However, these two agents produce

Received 6/27/85; revised 11/8/85; accepted 11/11/85.The costs of publication of this article were defrayed in part by the payment of

page charges. This article must therefore be hereby marked advertisement inaccordance with 18 U.S.C. Section 1734 solely to indicate this fact.

'This work was supported in part by Contract NO1CM27541, and Grant

IP50CA32107 awarded by the National Cancer Institute, Department of Health andHuman Services, Bethesda, MD 20205.

2To whom requests for reprints should be addressed.

considerable toxicity and have limited efficacy against manyneoplasms. Therefore, the search continues for anthracyclineantibiotics with improved therapeutic indices, altered spectra, oractivity of both (1).

7-OMEN3 is a semisynthetic derivative of the anthracycline

antibiotic nogalamycin (Fig. 1), a drug with significant experimental antitumor activity but prohibitive toxicity in animals (2). 7-

OMEN has been introduced into Phase I clinical trials based onits broad spectrum of in vivo activity against animal tumors, itsdemonstrated activity after p.o. as well i.v. administration (2, 3),and its reduced cardiotoxic potential (4).

The mechanism of action of 7-OMEN may differ from that ofdoxorubicin. 7-OMEN is most toxic to cells in d phase, whereas

doxorubicin is most toxic to cells in S phase (2). In addition,while doxorubicin interacts strongly with DNA, 7-OMEN interactsonly weakly with DNA and is highly cytotoxic at concentrationswhich do not inhibit nucleic acid synthesis (2, 5). Finally, whenvarious tumor lines are incubated in vitro with 7-OMEN ordoxorubicin, 7-OMEN fluorescence is localized mainly in the

cytoplasm, whereas doxorubicin fluorescence accumulates primarily in the nucleus (2, 6). Previous studies have described theplasma pharmacokinetics of 7-OMEN in beagle dogs (7), rabbits

(8), and mice (9, 10), while our laboratory has defined the organdistribution of 7-OMEN in mice and rabbits (8, 9), and the urinaryand biliary excretion of 7-OMEN by rabbits (8).

During the performance of a Phase I study at our center, wehad the opportunity to define the human plasma pharmacokinetics and urinary excretion of 7-OMEN and its metabolites at

multiple dosages, and to investigate the biliary excretion of thesecompounds in two patients with biliary drains. Finally, when weanalyzed these plasma pharmacokinetic data and comparedthem with the toxicity observed in each patient, an excellentrelationship was apparent between the plasma pharmacokineticsof 7-OMEN in any patient and the pharmacodynamics resultingfrom 7-OMEN therapy of that patient. This paper presents these

results.

MATERIALS AND METHODS

Patient Selection. All patients entering this trial had to fullfill thefollowing criteria: histological proof of malignant disease which had failedconventional chemotherapy or for which no conventional therapy existed,recovery from all toxicities from prior treatments and passage of at least4 weeks from any prior chemotherapy or radiotherapy, a minimal life

3The abbreviations used are: 7-OMEN, menogaril; HPLC, high performanceliquidchromatography;TLC, thin layerchromatography;AUC,areaunder the curveof plasmadrug concentration versus time; Vc,volume of the central compartment;V„,steady-state volumeof distribution.

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7-OMEN PHARMACOKINETICS AND TOXICITIES

DounorublcinCH3Doxorubicin

CH2OHNoqolomycin

CH3Menogoril

CH3N-DemethylMenogonlHNogorol0

CH]7-Deoxynogarol

CH3N-Demethyl-7-Deoxynogarol

HNogalose

COOCH3OCHj

HOCH3

HOH

HH

HH

H

Fig. 1. Structures of doxorubicin, daunorubicin, nogalamycin, 7-OMEN, andsome metabolites of 7-OMEN. Nogarene (Nogarol') corresponds to 7,8-9,10-

bisanhydronogarol.

expectancy of 12 weeks, a Karnofsky performance status >60%, adéquate bone marrow function (WBC >3,500 cells//»!),and platelet count>100,000/ííl,adequate liver function (bilirubin <2.0 mg/100 ml), andadequate renal function (creatinine <2.0 mg/100 ml, and a 24-h creatinine

clearance of >50 ml/min). Previous anthracycline administration did not,

by itself, exclude patients from this study; however, all patients musthave had an ejection fraction, as assessed by MUGA scan, >45% at

rest. Objective measurable disease was desirable, but not required.Written informed consent in accordance with Federal and institutionalpolicies was obtained from all patients prior to their entry onto study.

Before entry onto study, each patient had a detailed history taken anda physical examination performed. Tumor measurements were madeand performance status was assessed. Laboratory studies relevant tothis paper included pretreatment and weekly complete and differentialblood counts and platelet counts.

Dosage Preparation and Administration. 7-OMEN (NSC 269148),was supplied in freeze-dried dosage form by the Investigational DrugBranch, National Cancer Institute, Bethesda, MD. 7-OMEN was recon

stituted by adding 10 ml of sterile water for injection (DSP) to each vial.The resulting solution contained 50 mg of 7-OMEN, 100 mg of mannitol,and 16.6 mg of L-lactic acid. This was further diluted with sterile 5%dextrose in water. Dosages of 3.5 to 31.5 mg/m2 were diluted to 150 mland were infused i.v. over 1 h, while dosages of 42, 50, and 56 mg/m2

were diluted to 250 ml and were infused over 2 h. 7-OMEN was admin

istered i.v. on 5 consecutive days with courses repeated after 4 weeks.Sample Acquisition. Heparinized blood samples were obtained before

and at multiple times during and after the 7-OMEN infusion. Specifically,on day 1 of the 5-day course, blood was obtained prior to and at 0.5-hintervals during the 7-OMEN infusion. In addition, samples were obtained

at 5, 10, 20, 30, and 60 min, and at 2, 4, 6, 12, 18, and 24 h aftercessation of the infusion. Blood samples were also obtained before the7-OMEN infusion on days 2,3,4, and 5 of each course. In some patients,

a repeat of the intensive sampling scheme performed on day 1 was alsoperformed on day 4 or 5 of a course to investigate whether the phar-

macokinetics on later days of a course were similar to that on the firstday. Blood samples were immediately centrifuged at 1000 x g for 10min, and the resulting plasma supernatant was immediately removedand frozen at -20°C until analyzed. During the first 24 h after drug

administration, urine was collected as voided and stored as 4-h aliquotsin opaque containers at 4°C. Subsequently, a portion of each 4-hcollection was frozen and stored at -20°C until analyzed.

Two of the patients on this study had indwelling biliary tract drainsand provided the opportunity to collect and analyze the biliary excretionof 7-OMEN and its metabolites. The first of these patients had completelynormal serum liver function tests (bilirubin, 0.8 mg/dl; aspartate amino-

transferase, 35 units/liter; alanine aminotransferase, 7 units/liter; lactate

dehydrogenase, 136 units/liter; alkaline phosphatase, 292 units/liter;total protein, 6.9 g/dl; and albumin, 4.2 g/dl). The second patient hadmoderate hepatic dysfunction as assessed by serum liver function tests(bilirubin, 5.4 mg/dl; aspartate aminotransferase, 56 units/liter; alanineaminotransferase, 38 units/liter; lactate dehydrogenase, 143 units/liter,alkaline phosphatase, 1102 units/liter, protein, 6.2 g/dl, and albumin, 2.9g/dl). Each patient had bile collected on ice and stored frozen as 4-haliquots during the first 24 h after initiation of 7-OMEN therapy. The

second of these patients also had the subsequent 4 days of biliaryexcretion collected and stored frozen as 24-h aliquots.

Extraction and Chromatographie Analysis of 7-OMEN and Metabolites. 7-OMEN and metabolites were analyzed by the method of Mc-Govren ef al. (11). Briefly, 1-ml samples of plasma, urine, or bile weredeproteinized with 0.2 ml of 0.05 M glucuronic acid and 2.4 ml acetoni-trile:methanol, 1:1 by volume, containing 0.25 MM7-con-O-ethyl nogarol

as an internal standard. Internal standard was graciously provided by Dr.J. P. McGovren of the Upjohn Co., Kalamazoo, Ml. Samples weresubsequently centrifuged at 27,000 x g for 15 min at 4°C.The resulting

supernatant fraction was transferred to prechilled, screw-cap vials,shielded from light, and stored at 4°Cuntil injection onto the HPLC.

The HPLC system used in these studies consisted of a Waters M45pump (Waters Associates, Milford, MA) and a C8 radial compressioncartridge (Waters Associates) held in a Z module (Waters Associates).7-OMEN and metabolites were eluted with a mobile phase of 0.2 M

acetic acid, 0.015 M methane sulfonic acid:acetonitrile, 3:2 by volume,pumped at 1.5 to 4 ml/min. Fluorescent materials were detected with anAminco fluoromonitor (SLM Instruments, Urbana, IL), fitted with 470 nmexcitation and 550 nm cut-off emission filters. 7-OMEN concentrationswere determined by comparison of the areas of 7-OMEN peaks with that

of the corresponding internal standard.In addition to HPLC, bile and urine samples were also analyzed by

TLC. TLC analyses utilized 250 firn Silica Gel 60 TLC plates (E. Merck,Darmstadt, West Germany), that were developed for 15 cm, in anascending fashion, in chloroform:methanol:acetic acid:water, 80:20:14:6by volume. Fluorescent spots were identified under 254 nm light (UVS-

54 Mineralight, Ultraviolet Products, San Gabriel, CA). Known standardsof 7-OMEN, W-demethyl menogaril, 7-deoxynogarol, nogarol, and nogar-

ene were also run on each TLC plate.Pharmacokinetic Analysis. Computer modeling of the decline in

plasma concentrations of 7-OMEN was performed with the MLAB pro

gram (12), and pharmacokinetic parameters were calculated from themodeled data. The relationships between changes in WBC and AUCwere modeled with ADAPT (13), another program which uses iterative,weighted, nonlinear, least squares regression.

RESULTS

At all dosages, plasma concentrations of 7-OMEN rose pro

gressively during the drug infusion, after which they decayed inbiexponential fashion with tv, a 0.19 ±0.04 h (mean ±SE) andtv, ß13.22 ±1.54 h (Fig. 2; Table 1). The pharmacokinetics of7-OMEN was linear over the dosage range studied, the AUC

increasing linearly with increased dosage (Fig. 3; Table 1). Ascan be inferred from this, the total body clearance of 7-OMEN,28.18 ±3.33 liter/h/m2, was not altered by increasing the dose

(Table 1). Similarly, neither the volume of the central compartment, 224 ±30.8 liter/m2, nor the steady-state volume of distribution, 370 ± 25.7 liter/m2, were altered by increasing the

amount of drug infused. These plasma pharmacokinetic parameter values did not change with repeated dosing since thosedetermined on day 4 or 5 of a course were unchanged fromthose observed in that same patient on day 1.

At all times, 7-OMEN was the major fluorescent species pres-

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7-OMEN PHARMACOKINETICS AND TOXICITIES

zIO

1.000-

0.7000.500-

0.300-

0.100-

0.070-

0.050-

0.030

0.010-

0.007-

0.005-

0.003-

0.001

oB

!§Ml 5(2)

a 175(1)o 7 (S)

A3.5(3)

/O 2 IO 12 14 16 18 20 22 24TIME (hr)

1.000-

0.700-

0.500-

0.300-

3. 0.1 00-"~Z 0.070-

^ 0.050-

o,1. 0.030-

in3 o.oio°- 0.007-

0.005-

0003-

0.001

o 50(5)A56(4)«42(5)

B

0 2 4 6 8 10 12 14 16 18 20 22 24

TIME (hr)Fig. 2. Concentrations of 7-OMEN in plasma of patients treated with varying

dosages of 7-OMEN. Dosages of 3.5 to 31.5 mg/m2 (A) were infused over 1 h whiledosages of 42 to 56 mg/m2 (B) were infused over 2 h. Concentrations of 7-OMENwere determined by HPLC, as described in "Materials and Methods." Points, mean

values. The number of courses studied at each dosage are indicated in parenthesesnext to the dosage.

ent in plasma. Although small HPLC peaks corresponding to themono- and didemethyl metabolites of menogaril were observed,

the plasma concentrations of these materials were negligible andrepresented <5% of plasma drug fluorescence at any time.

Urinary excretion of 7-OMEN and its metabolites increased

progressively over the 24 h after drug injection, and accountedfor approximately 5% of the administered dose (Fig. 4). Only twofluorescent drug species were observed in urine (Fig. 5). Ofthese, 7-OMEN was easily the major species, still accounting for

greater than 70% of drug fluorescence at 24 h after drug administration. The remainder of drug fluorescence was accounted for

by A/-demethyl menogaril. Although the proportion of total fluo

rescence in urine due to this metabolite increased with time, itnever represented more than 20 to 30% of urinary drug fluorescence (Fig. 5). There were no measurable amounts of didemethylmenogaril or any aglycones in the urine at any time during the24 h period of study.

As the urinary excretion, biliary excretion of 7-OMEN and

metabolites increased progressively in the two patients studied(Fig. 6). In the first patient, the 24-h biliary excretion accountedfor 4.2% of the dose of drug administered on day 1 of the 5-day

course. In the second patient, this value was 1.9%. Not only wasthere reasonable agreement between these two patients, butthere was also internal consistency for the second patient, sincethe percentage of daily dose of 7-OMEN excreted in the bile on

days 2, 3, 4, and 5 was 2.2, 1.1, 2.7, and 1.3%, respectively.Only two fluorescent species were present in the bile (Fig. 7). Inboth patients, 7-OMEN was the major biliary fluorescent drug

species. However, the percentage of drug fluorescence accounted for by A/-demethyl menogaril increased progressively,so that from 16 to 24 h, parent compound and its A/-demethyl

metabolite were present in bile at ratios of approximately 6:4,respectively. In addition to analysis by HPLC all bile sampleswere analyzed by TLC. This was to confirm the presence of thetwo drug forms in the bile and to rule out the presence of verynonpolar drug forms, such as aglycones, which might not elutefrom the HPLC column, or very polar drug forms, such asconjugates, which might elute with the solvent front and therebyescape detection. Only two fluorescent drug spots, corresponding to 7-OMEN and A/-demethyl menogaril, were observed. There

were no other fluorescent metabolites observed on the TLCplate, at the origin, or at the solvent front.

In addition to defining the pharmacokinetic behavior of 7-

OMEN in humans, the availability of toxicity data from our PhaseI study allowed us to attempt relating the pharmacokinetics andpharmacodynamics of this agent. It was apparent that analysesusing peak plasma concentrations were of no use. Similarly,analyses using dosage and change in WBC were suboptimal.The first of these variables neglected interpatient variability withregard to drug disposition, and the latter neglected the fact thatindividual patients were treated with pretreatment WBC of 3,500to 18,500/^1, which by definition put a limit on the maximumchange in WBC that could occur. On the other hand, an excellentrelationship was obtained when area under the curve of 7-OMEN

plasma concentration versus time was related to the percentageof decrease in WBC, i.e.,

Pretreatment WBC - nadir WBC

Pretreatment WBCx 100

(Figs. 8 and 9). This relationship was well described by twodifferent models.

The first of these, taking a form similar to that of a onecompartment model approaching steady state during continuousinfusion, was described by the equation:

% of decrease in WBC = 100 (1 - e-0067AUC)

(Fig. 8). When this relationship was evaluated by comparing the

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7-OMEN PHARMACOKINETICS AND TOXICITIES

Table 1Summaryof menogarilplasma pharmacokineticparameters

Dosage(mg/m2)3.5711.517.531.5425056MeanSDSEDosage(pmd/m2)6.312.9521.27532.37558.27577.791.7103.6No.

ofstudies23312544«(h-)12.087.0611.097.3611.126.323.664.23.694.132.241.971.404.84

±2.527.465.201.06f*

«(h)0.06

±0.220.17+0.080.08±0.020.090.06

±0.010.20±0.090.33±0.190.32

±0.130.190.200.04ß

(h-1)0.24

±0.220.113±0.060.093

+0.0160.1210.075

+0.0040.057±0.0060.057±0.0160.049±0.0120.0710.0440.01f»0(h)13.2

±11.611.4±5.98.0

±1.55.79.3

±0.512.7+1.216.2

±5.217.2±4.313.227.391.54AUC^M-h)0.158

±0.0840.95+0.330.79±0.180.8131.95

±0.263.46±0.676.16+2.074.8

±1.01Total

bodyclearance

(liter/h/m*)57.7

±30.418.0+6.928.4±6.439.830.4

±4.025.5+4.122.4±8.026.3±6.028.1816.303.33Vc

(liter/rtfl135

+50174±24123+1798100

+33212±13362±48355

±12622414830.80V»

(liter/m2)183

±24350±49362+

13373384

±48375±66391±46465±9937012625.70

Fig.3. Relationshipof dosage to area underthe curve of plasma concentration of 7-OMENversus time A, data on individual patients andcourses of therapy. B, means + SD of allcourses studied at each dosage.

I2-

II-

IO-

9-

8

7-

. 6-

5- O

10

9

8

7-

6-

. 5-

4-

3-

2-

IO 20 30 40 50DOSAGE (mg/m2)

60

I

I B

IO

8-1

7-

5-

3-

2-

20 30 40 50 60DOSAGE (mg/m2)

The other relationship which fit our data was the Hill equation(14) which produced the equation:

CUMULATIVE

% of decrease in WBC =(100) (AUC163).

(10.14)163 + (AUC)'

8 16 20 2412TIME (hr)

Fig. 4. Urinary excretion of 7-OMEN and metabolites by patients treated with7-OMEN. Points, means ±SD of 24 courses.

predicted percentage of decrease in WBC with that actuallyobserved (Fig. 9), the data described a line very close to the lineof identity, with an r value of 0.889.

(Fig. 8). In this case, percentage of decrease in WBC equals theeffect measured, 100 represents the maximum effect possible,10.14 represents the AUC that produces 50% of the maximumeffect, and 1.63 is a constant derived from our set of data. Whenthis relationship was evaluated by comparing the predicted percentage of decrease in WBC with that actually observed (Fig. 9),the data were very similar to those produced by the previousmodel, i.e., it was close to the line of identity with an r value of0.893. There was no statistical difference between the twomodels with regard to their suitability in describing the data (15).

When we applied the same two models to the relationship ofarea under the curve and the percentage of decrease in absoluteneutrophil count, we observed results very similar to those seenwith total WBC (Figs. 10 and 11). Once again, each modelproduced an excellent description of the data, with the secondmodel producing a slightly, but not statistically significantly,

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7-OMEN PHARMACOKINETICS AND TOXICITES

20 244 6 12 16TIME (hr)

Fig. 5. Fluorescent species present in urines at 4-h intervals after i.v. administration of 7-OMEN. Points, means ±SD of 24 courses.

5-

4-

ouil-uacoXluUJto

8

3-

2-

I-

8 16 20 2412TIME (hr)

Fig. 6. Biliary excretion of 7-OMEN and metabolites by two patients treatedwith i.v. 7-OMEN. Patient 1 received 7-OMEN at a dosage of 56 mg/m2, and patient2 received 7-OMEN at a dosage of 42 mg/m2. Both patients had urines collectedas 4-h aliquots on day 1 of their 5-day course of therapy. Patient 2 also had 24-h

bile collections on days 2, 3, 4, and 5 of therapy.

better fit. Comparison of the data for total WBC with that forneutrophils revealed that neutrophils were more sensitive to 7-

OMEN, as demonstrated by both a lower AUC producing a 50%decrease in cell count (7.023 versus 10.14) and a steeper slope,as reflected by a larger value for the constant in the exponentialterm of each equation (1.72 versus 1.63).

IOO-

90-

80

UJO 60-

3

o

30-

IO-

MENOGARIL

N-DEMETHYLMENOGARIL

8 12 2416 20

TIME (hr)Fig. 7. Fluorescent species present in bile of a patient treated with i.v. menogaril.

Bile was collected on ice as 4-h aliquots and was stored frozen. 7-OMEN metabolites were separated and quantitated by HPLC as described in "Materials andMethods." The HPLC results were confirmed by TLC as described in "Materialsand Methods."

DISCUSSION

The introduction of 7-OMEN into clinical trials represents an

other attempt to find an anthracycline antitumor antibiotic withtrue advantages over doxorubicin and daunorubicin, the mostcommonly used members of this major class of antineoplasticdrugs. When considered in light of previous animal and clinicalstudies of doxorubicin and daunorubicin, as well as animal studies of 7-OMEN, the data presented in this manuscript may

provide a better appreciation of the effects of species andstructural differences on anthracycline metabolism and disposition. In addition, the clinical pharmacological data and the datacorrelating the pharmacokinetics and pharmacodynamics of 7-

OMEN should provide a foundation for more intelligent use ofthis drug in subsequent Phase II and Phase III clinical trials.

The major metabolic alteration of 7-OMEN by humans involves

demethylation of the tertiary amino sugar attached to the D ringof the anthracycline nucleus. This is in contrast to the biotransformation of doxorubicin and daunorubicin by humans and animals, where cytoplasmic NADPH-requiring aldo-keto reducíase

reduces the carbonyl function attached to the A anthracyclinering to produce the glycosides doxorubicinol and daunorubicinol(16-22). Moreover, with time these reduced glycosidic metabo

lites of doxorubicin and daunorubicin appear in plasma at concentrations that approach or exceed those of the parent compound. In contrast there is a very small percentage of drugpresent in plasma as 7-OMEN metabolites. Our data on the

fluorescent drug species present in plasma of humans treatedwith 7-OMEN are similar to those in our previously publishedstudies of 7-OMEN metabolism and disposition in rabbits (8) and

mice (9). More precisely, we observed no plasma fluorescentdrug species other than 7-OMEN in rabbit plasma (8), but diddetect measurable concentrations of A/-demethyl menogaril in

mouse plasma (9). In humans, although small peaks corresponding to A/-demethylated metabolites of 7-OMEN were observed,

their concentrations were negligible when compared to those of

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7-OMEN PHARMACOKINETICS AND TOXICITES

Fig. 8. Relationship of area under the 7-

OMEN plasma concentration versus time curvefor individual patients to the percentage decrease in WBC observed in that same course.The curves and equations displayed were modeled as described in "Materials and Methods."

I00-|

95-

90-

85-

80-

7570-

65-60-

55-

50-

45-

40-

35-

30-

25-

20-

15-

10-

5

O10 20 25 30 35 40

AUC (yuM-hr )45 50 55 60

100-

90-

80-

70-

60-

jt 50-o 40-S 30-z 20-u, IO-IS o

x0 lOO-i3 90-

DC80-

% 7°-§ 60-

50-40-30-20-

10-

0

•/.AWBC-ioo(i-e-°OS7AUC)

OBSERVED•0.998 EXPECTED-0«r •0.884

20 40 60 80 100

,(10.14) + AUC

OBSERVED-0.945 EXPECTED+ 5 06r-0893

=1—i—i—i—20 40

—i—i600 20 40 60 80 100

PREDICTED CHANGE IN WBC(%)Fig. 9. Relationship of observed percentage decrease of WBC to that predicted

by the two models described in "Results" and displayed in Fig. 8. Points, individual

courses of therapy..

the parent compound, mimicking the situation in mice. The 13.22-h terminal plasma half-life of 7-OMEN in humans is similar to the10.16-h terminal half-life observed in mice (9, 10), but is muchlonger than the 2.66-h terminal half-life observed in rabbits (9).On the other hand, the total body clearance of 28.18 liter/hr/m2

observed in our patients corresponded more closely to that of43.2 liter/h/m2 observed in rabbits (9), than to that of 5.5 liter/hr/m2 observed in mice (9, 10). We recognize that our sampling

schedule on day 1 was limited to 24 h after drug administrationby the treatment administered on day 2. Although a more idealsampling schedule would have included times at three to four

half-lives after drug injection, such a schedule was unworkable

in our patient population with the treatment schedule used. Still,sampling at 24 h after drug injection approximates two half-liveswhich should allow reasonable assessment of the plasma phar-macokinetic parameters presented.

In humans, as in rabbits (8), neither urinary or biliary excretionof 7-OMEN and its fluorescent metabolites represented a major

route of drug clearance. This again is in contrast to the behaviorof doxorubicin and daunorubicin, in which biliary excretion playsa major role in drug elimination (23, 24). The fluorescent drugspecies present in urine and bile of humans treated with 7-OMENwere limited to parent compound and /V-demethyl menogaril. We

observed no didemethylated forms, conjugates, or aglycones ineither urine or bile. This is in contrast to rabbits in which threeand five 7-OMEN metabolites besides A/-demethyl menogaril

were observed in urine and bile, respectively (8). The metabolitesof 7-OMEN observed in rabbit bile included the 7-deoxyagly-cones of both parent compound and /V-demethyl menogaril. Thisimplied that 7-OMEN can serve as a substrate for microsomal,NADPH-dependent cytochrome P-450 reducíase, an enzyme

known to play a major role in the biotransformation of daunorubicin, doxorubicin, and other anthracycline compounds (25, 26).We are unsure why such deoxyaglycones were not observed inany of the multiple bile samples collected from our patients. Thefact that only approximately 10% of an administered dose of 7-

OMEN can be accounted for in bile and urine raises the issue ofwhich processes play major roles in drug clearance. In animalstudies, little of the administered dose of drug could be accounted for as fluorescent drug forms when tissues were extracted and analyzed (8). Metabolism to nonfluorescent metabolites may therefore represent the major route of drug clearance.The existence of this metabolic pathway for other anthracyclineshas already been documented by our laboratory (27). Unfortunately, suitable radiolabeled 7-OMEN, which would greatly facil

itate further investigation of this question, is not available atpresent.

We believe that the data presented provide more than additional information about anthracycline pharmacology. The ability

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7-OMEN PHARMACOKINETICS AND TOXICITES

Fig. 10. Relationship of area under the 7-OMEN plasma conœntration versus time curvefor individual patients to the percentage decrease in absolute neutrophil (PMN) count observed in that same course. The curves andequations displayed were modeled as described in 'Materials and Methods."

100-

95

9085

80

75Z 70

E 65

Z 60ü 55z 50< 45ü 40S* 35

30292019109

20 25 30

AUC

35

hr)

40 49 SO 55 60

'/.¿pMN=ioo(i-e-°095AUC)

OBSERVED-0.984 EXPECTED + 0.47r. 0.915

20 40 60 80 100PREDICTED CHANGE IN PMN CM

Fig. 11. Relationship of observed percentage decrease in absolute neutrophil(PMN) count to that predicted by the two models described in "Results" and

displayed in Fig. 10. Pomfs, individual courses of therapy.

to correlate pharmacokinetics and pharmacodynamics, while anestablished concept for many other types of drugs, is still arelatively uncommon occurrence for antineoplastic agents. Tosome extent this reflects the relatively long temporal lag betweenestablishment of a pharmacokinetic "milieu" by drug administra

tion and the pharmacodynamic manifestations resulting from thatadministration. Our approach to the modeling of our data wasempiric and used models known to produce curves the shape ofwhich seemed appropriate for our data. As such, our choices ofmodels do not necessarily reflect concepts of the mechanisticrelationship of 7-OMEN pharmacokinetics to pharmacodynamics. Rather, they provide a mathematical description which might

prove useful in dosing of patients. At present, our mathematicalmodels do not fulfill this promise since a critical piece is stillmissing, i.e., the ability to predict the pharmacokinetic behaviorof 7-OMEN in an individual patient. Our pharmacokinetic data

lead us to believe that assessment of renal function will not beas important as assessment of hepatic drug-metabolizing capa

bility. However, we are currently attempting to correlate bothrenal function, as assessed by serum creatinine and creatinineclearance, and hepatic function, as assessed by liver functiontests and indocyanine green and antipyrine clearances, with theliver pharmacokinetics and pharmacodynamics of 7-OMEN.

ACKNOWLEDGMENTS

We thank H. Chlewicki and B. Knickman for excellent secretarial assistance.

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1986;46:1513-1520. Cancer Res   Merrill J. Egorin, David A. Van Echo, Margaret Y. Whitacre, et al.   Their Correlation with Clinical ToxicitiesAnthracycline Antibiotic Menogaril (7-OMEN, NSC 269148) and Human Pharmacokinetics, Excretion, and Metabolism of the

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