Adequacies and Inadequacies in Assessing Murine Toxicity Data with Antineoplastic Agents

Home , Clearance (pharmacology), Drug metabolism

[CANCER RESEARCH 39, 2204-2210, June 1979] 0008-5472/79/0039-0000$02.00 Adequacies and Inadequacies in Assessing Murine Toxicity Data with Antineoplastic Agents

Anthony M. Guarino,1Marcel Rozencweig,Ira Kline, John S. Penta, John M. Venditti, Harris H. Lloyd, Donald A. Holzworth, and Franco M. Muggia

Division of Cancer Treatment, National Cancer Institute, NIH, Bethesda, Maryland 20205 [A. M. G., M. R., I. K., J. S. P., J. M. V., F. M. M]; Southern Research Institute, Birmingham, Alabama 35205 (H. H. L.]; Toxicology Program Office, Battelle Columbus Laboratories, Vienna, Virginia 22 180 (0. A. H.]

ABSTRACT regardless of the number of treatments. Finally, implicit and/or explicit assumptions were made regarding the reproducibility Previous retrospective analyses have suggested a very pos of these experimental results. itive correlation in toxic doses of antineoplastic agents between This retrospective study takes advantage of the accumulation mice and humans. Additional toxicological information has now of additional data to further assess problems associated with been accumulated and reveals a noticeable variability in the toxicological results in mice and their potential for use in dose existing data base. Nevertheless, it is likely that mouse toxi prediction in humans. cological studies will become a principal determinant for esti mating initial doses to be used in humans. Recognition of the factors responsible for differences in determinations of toxic MATERIALS AND METHODS dose levels in mice will enhance the proper utilization of this approach. The cytotoxic antitumor agents selected for analysis were confined to those for which clinical toxicological data were available for i.v. administration by at least one of the following INTRODUCTION schedules: high intermittent doses, weekly doses, and daily Antitumon agents are commonly administered in humans at administration for 5 or 7 days. These limitations were necessary or near their MTD.2 When a new agent is first introduced into because other routes of administration, particularly the p.o. clinical trials, its MTD is reached by progressive increments of route, may introduce further bias in correlation studies; the an initial dose that has been derived from toxicological data schedules chosen are those most frequently investigated in obtained in animals (4, 5, 9, 24). Significant interspecies dif humans. ferences in the rate of drug metabolism and excretion make it Toxicological studies in mice were performed under contract difficult to accurately extrapolate to humans the iatrogenic to the Laboratory of Toxicology and Drug Evaluation Branch of effects observed in animals (1, 6, 7). Nevertheless, based on the Division of Cancer Treatment in the National Cancer Insti the clinical experience with 37 drugs, Homan (20) has esti tute, according to previously described methods (13, 25). mated that an initial dose of one-third the MTD (in mg/sq m), Lethality was chosen as a nonspecific but easily measurable as determined in the most sensitive large animal species (bea toxicological end point. The lethal dose categories are LD10, gle dog or rhesus monkey), would be tolerated in humans in LD50,and LDso;in this study, this period ranged from 14 to 60 about 94% of the cases. Whether such a determination con days after the last dose administration, but more than 90% of stitutes the most efficient method for rapidly and safely reach all deaths actually occurred within 14 days. ing the effective dose in humans or whether rodent data may Statistical analyses were performed to assess true differ be used to advantage has been the subject of recent analysis ences between the related parameters of animal strain, injec (14). tion site, and drug-dissolving or suspending vehicles. Probit The value of combined animal toxicological data, including analyses and tests for parallelism of response were performed findings in rodents, for predictive purposes in humans has been wherever possible (3, 10, 11, 16). Slopes of log probit lines repeatedly advocated (12, 14, 26). This approach has been should be parallel if: (a) there is no statistically significant hampered by the availability of only limited rodent data not difference between the data (11); and (b) the cause of death expressly obtained for toxicological purposes. In fact, previous (i.e. , mechanism of toxicity) in the 2 treatment groups is the correlations had to be based on various assumptions. Specifi same, that is, both groups show the same â€˜â€˜specificactionâ€• cally, assumptions were made regarding the route of adminis (8). tration, considering i.p. and i.v. injections equivalent and ne glecting the effect of schedule. The toxicity pen course was RESULTS considered to be a function of the total dose administered, Clinical toxicity data were available for 58 drugs that have I To whom requests for reprints should be addressed, at Building 37, Room been administered i.v. in humans by at least one of the 3 5B22, Laboratory of Toxicology, National Cancer Institute, NIH, Bethesda, Md. 20205. schedules of interest (Table 1). The daily for 5 or 7 days 2 The abbreviations used are: MTD, maximum tolerated dose; LD10, drug dose schedule has been most widely used, for 49 (84%) of the that kills 10% of the non-tumor-bearing animals during the observation period; LD@,dose that kills 50% of the non-tumor-bearing animals during the observation drugs, whereas high-dose intermittent administration has been period; LD@,dose that kills 90% of the non-tumor-bearing animals during the studied for only 41 % of the drugs. observation period; 6-MP, 6-mercaptopurine; ara-C, 1-f@-n-arabinofuranosylcy Of the 3 categories of lethal dose, the mouse LD50,estimated tosine; BCNU, 1,3-bis(2-chioroethyi)-1-nitrosourea; FU, 5-fluorouracil; TIC mus tard, lmidazole-4-carboxamide, 5-(3,3-bis(2-chloroethyl)-1 -triazeno). by the probit model, is statistically more reliable than is the Received November 27, 1978; accepted March 2, 1979. LD10or LDso (11, 15) and was most often available for our

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Downloaded from cancerres.aacrjournals.org on September 28, 2021. © 1979 American Association for Cancer Research. Drug Toxicity in Rodents and Humans Table1 for FU. The slopes of the probit response lines were parallel for Availability of mouse lethality data for 58 drugs used i. v. in humans all of the drugs except daunomycin, where the LD50and LD10 on the most common schedules differed by 52 and 275%, respectively. â€” No. of drugs for which mouse Is The variation in the total equitoxic dose per course could be thecorrespondingthality data are available for schedule in humans studied for 6 drugs, each given on 1 equitoxic dose on Day 1 No. of drugs and 1 equitoxic dose on Days 1 to 5 schedules in a single LD,0Schedule given iv. in hu- LD@+ mouse strain using one vehicle and injection route. The L050or LD@Single mans LD@ and/or LD10is shown for each drug on both schedules in Table 5. The 13(54)Weeklydose 24 20 (83)a differences in total equitoxic dose per course apparently vary (6)Daily 34 11 (32) 2 (33)afor 5 (or 7) days 49 23 (47) 16 greatly according to the drug in question. Thus, the difference dataareNumbers in parentheses, number of drugs for which mouse lethality was modest for BCNU (7%), intermediate for ama-C(23%) and available for the corresponding schedule in humans. isophosphamide (28%), large for cis-diamminedichlono platinum (77%) and 6-MP (150%), and extreme for 2'-deoxy thioguanosine (1440%). study. LD50's were known for 83, 32, and 47% of the drugs Toxicity studies have been repeated, usually at the same tested at the single, weekly, and daily schedules, respectively laboratory, with 16 drugs under the same conditions with (Table 1). Knowledge of other lethal dose levels does provide regard to the animal species and strain, route of administration, additional information of obvious importance in the analysis of vehicle, and schedule. The data for the vast majority of these toxicological data. However, fully usable lethality data (LD50 agents show striking discrepancies (Table 6). The average plus LD10or LOso)for the 3 schedules were only available for difference between each pair of lethality studies at LOsowas 54, 6, and 33% of the drugs. 42%. The LD50was identical for 3 drugs (thiotepa, pseudourea, Variations in the lethal dose levels according to the mouse and BCNU) and differed by 20% or less for azasenine, actino strain relate to data obtained largely in Swiss and C57BL x mycin 0, cyclophosphamide, porfinomycin, and daunomycmn. DBA/2 F1 (hereafter called BD2F1) mice (Table 2). Occasion Differences between 21 and 50% were noted in the LD50of ally, results were available for other strains, i.e. , AKA, BALB/ methotrexate, mitomycin C, ama-C,and TIC mustard. The LD50 c X DBA/2 F1(hereafter called CD2F1), and DBA2. differed by 71% with nitrogen mustard, by 82% with vincnistine, At least one comparison could be made for each of 16 drugs by 85% with cis-diamminedichloroplatinum, and by 245% with that used identical routes, vehicles, and schedules. For a camptothecin. number of these agents, noticeable variations existed in one on It is also noteworthy that there was a lack of parallelism in more of the lethal dose levels (L010, LD50, and LDso). The probit slopes for more than one-half (9 of 16) of the compani average difference between the LD50obtained in a given pair sons. Occasionally, the slopes diverged so dramatically within of mouse strains receiving the same drug was 45% and ranged the LD10to LD@range that reversals of results were observed between 0% (fluorouracil deoxyriboside) and 145% (vincnis according to the lethal dose category considered. For example, tine). For 5 drugs (6-MP, cyclophosphamide, pseudourea, yin the LD50of cyclophosphamide (58 mg/kg) in one study is 1@% cnistine, and camptothecin), the LD50differed by more than smaller than that in the other (67 mg/kg). However, when the 50% between strains. In one case (fluonouracil deoxyniboside), LD10 in the same pain of experiments is compared, the first the LD50's were exactly the same, but the LD10's differed by study shows a 24% higher toxicity (26 mg/kg) than does the 17%. In another case (6-MP), the LD10's were virtually the latter one (21 mg/kg). In the case of pseudoumea,the L050 of same in both strains, but the LD50differed by 52%. both studies was equal, but reversal was evident when the LD10 The last column of Table 2 shows the results of standard and LDso were considered. Similarly, examination of the tests for parallelism of response to the drugs in each pair of lethal doses for ara-C and daunomycin showed reversals de strains tested. Of the 16 drugs in this table, 13 strain pairs had pending upon which lethal dose category was considered. parallel slopes. For the other 3 (6-MP, pseudourea, and ama A summary of the lethality data by factors of variability C), either the 2 strains are not truly comparable from a statis appears in Table 7. Of the 49 separate drug comparisons tical point of view or it may be assumed that there was a strain included in the analyses, 38.8% showed a difference of @s20% difference in the cause of death. in LD50.This 20% variability corresponds to the range tradi Because the primary screening protocols of the National tionally accepted among toxicologists (19). Cancer Institute (13) involve essentially i.p. administration, data Differences exceeding 100% occurred in 7 (14.3%) of 49 comparing the i.p. and i.v. routes were available for only 3 comparisons. The mouse strain used was associated with drugs (Table 3). None of the lethal dose levels of Adniamycin differences of 128% for camptothecin, and 125 and 145% in varied significantly with the route of injection. On the other 2 tests of vincnistine (Table 2). A change in the route of injection hand, both vincristine and daunomycin were severalfold more (Table 3) for vincnistine and daunomycin apparently modified toxic via the i.p. route. The well-known local toxicity (17) of the results by 115 and 300%, respectively, between groups of these agents apparently shifted the dose-response curves in mice receiving the same treatment. Alternate vehicles seemed favor of greater i.p. toxicity, but the slopes of these curves responsible for a 250% difference in the LD50of FU (Table 4). remained parallel. This observation contrasts with the usual Unknown variables led to a 245% difference in a pair of expectation that the route carrying the substance most rapidly experiments where camptothecin toxicity was tested under to the bloodstream is invariably the most toxic (27). apparently similar conditions (Table 6). Finally, the 3 known The average LD50difference was 72% between groups of variables (strain, route, and vehicle) yielded a greater percent animals receiving 10 separate drugs in 2 different drug vehicles age of parallel probit response lines (84 to 100%) than the (Table 4). This difference ranged from 9% for BCNU to 250% 47% that was obtained when none of these factors could be

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Table 2 Variation of toxicity according to mouse strain Foreachdrug,thecomparedstudieswereperformedbythe i.p. routeof administrationinthesamelaboratorybutnotnecessarilyatthesame time. Lethal (mg/kg/dose) of probit re strainVehicleaSchedulebAlkylating@!@:@! LD,0doses LD@ LD@Parallelismsponse linesc agents @,CyciophosphamideSwissAlanine mustardSwiss sulfoxideDaily for 7 days5.2 BD2F,Dimethyl 5.96.6 8.98.4 13pd

VNitrogen NaCI solutionDaily on Days 1 and 850 38 BD2F,0.9% 1471 266>200 479P.

VNitrominSwissmustardSwiss NaCI solutionDay 1 only3.1 BD2F,0.9% 3,53.5 4.84.0 6.5P.

VAntimetabolites for 7 days22 BD2FOtherDaily 1960 55>100 >120P.

Vara-CBD2FCycloleucineSwiss for 7 days74 18 88 BD2F,OtherDaily 881 1401 223P,

NaCI solutionDay 1 only C C5FUSwiss CD2F,0.9% Day 1 only2829 39623779 43365047 4745NP, NP,

VFluorouracil for 7 days39 BD2F,CarboxymethyicelluloseDaily 3442 4446 56P.

C6-MPSwiss for 7 days1 28 78 deoxyribosideSwiss BD2F,OtherDaily 1091 178248 291P,

VAntibiotics for 7 days43 BD2F,OtherDaily 4691 60>146 77NP,

Actinomycin DBD2F, VBD2F, NaCI solutionDaily for 7 daysP, 0.070 0.089 0.114 AKR0.9% 0.0680.4320.099 0.144

VAzaserineSwiss NaCI solutionDaily on Days 1 and 4P, 0.485 0.546 AKR0.9% 0.2827.4 0.563 >0.680

VMitomycin for 7 daysP. 17 38 BD2FOtherDaily 141.6 23 37

VOthers CSwiss NaCI solutionDaily for 7 daysP. 2.1 2.8 BD2F,0.9% 2.021 2.2 2.3

BCNUSwiss VSwiss NaCI solutionDay 1 onlyP. 42 86 BD2F0.9% 3012 56 101

CCamptothecinSwiss NaCI solution for 7 daysP, 17 23 BD2F0.9% + 2% ethanolDaily 1455 18 23

CPseudoureaSwiss NaCi solutionDay 1 onlyP, 64 74 BD2F,0.9% 12714 146 169

VVincristineSwiss for 5 daysNP, 35 86 BD2FCarboxymethylcelluloseDaily 60 68 76

NaCI solutionDay 1 only0.826 V (all 3) BD2F 2.7 4.5 7.6 DBA20.9% 2.22.0 4.95.1 6.5P. aThevehiclewasthesameforeachcomparison.Other,actualvehiclenotrecorded(datafrom1961),butitwasidenticalforbothstrains. b Unless otherwise stated, doses were daily for Indicated number of consecutive days.

@ C See â€˜Materials and Methods.â€• d p probit lines parallel; NP, probit lines not parallel; C, computer-generated result; V, visual observation when data were insufficient for computer fit (1 1).

considered a possible contributor to variable results in toxicity and the route of administration. In addition, the cumulative total studies. dose per equitoxic course also appeared to vary greatly ac cording to schedule. DISCUSSION This latter observation seems to contrast with findings re ported by Griswold eta!. (18). Their study showed no consistent Three parameters were found to modify markedly the toxicity difference in the toxicity of a wide variety of agents in random of anticancer agents in mice, i.e. , the strain, the drug vehicle, bred Swiss mice and inbred BO2F1 mice. Furthermore, when

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Table 3 Variation of toxicity with route of administration Foreachdrug,the comparedstudieswereperformedin the samelaboratorybut notnecessanilyatthe sametime. dose (mg/kg/dose)Parallel ism of probit re sponse llnesbLD,0LD@[email protected] strainVehicle8ScheduleLethal CDaunomycini.p. NaCi solutionDay 1 only0.8 iv.Swiss0.9% 3.52.0 4.35.1 5.2p,C

CAdriamycini.p. NaCI solutionDay 1 only5.4 2 iv.BD2F0.9% 298.0 321 34P.

1 only1 0 8 V iv.BD2FBuffered 0.9% NaCI solutionDay 121 1825 26P, a The vehicle was the same for each comparison. Buffered 0.9% NaCI solution, 6.5 g Na2HPO4-4 g NaH2PO4 in 1 lIter 0.9% NaCI solution at final pH 6.9. b See Table 2.

C p probit lines parallel; C, computer-generated result.

Table 4 Variation of toxicity with drug vehicle For each pair of vehicles,studywas performedby the i.p. route in the samemousestrainat the samelaboratorybut not necessarilyat the same time. dose (mg/kg/dose)Parallelism of probit re sponse llnesbLD,0LD@LD@AlkylatingDrugVehicleaMouse strainScheduleLethaI agents VTICMelphalan (L- 1 onlyI 1 phenylalanine Hydroxypropylcellulose 1920 2936 46pC mustard)Carboxymethylcellulose(klucel)BD2FDay

CAntimetabolltesmustardCarboxymethylcellulose/ 1 only520 ethanol 262871 4571458 796P. CarboxymethylcelluloseBD2F,Day

VMethotrexateWater/iFUCarboxymethylceilulose 1 only62 02 67 OtherSwissDay 2401 3571 560P.

VAntibiotics N NaOH on Days 1 and 872 14 81 NaHCO3BD2FDaily 421 631 112P.

ActinomycinVAdriamycinBuffered DOther for 7 days0.098 18 0.9% NaCI solutionBD2FDaily 0.0700.1 0.0890.142 0.114P,

VDaunomycin0.9% 0.9% NaCI solution 1 only1 0 8 0.9% NaCI solutionBD2FDay 141 2336 37P.

COthers NaCI solution on Days 1 and 43.0 WaterBD2F,Daily 0.83.5 2.34.1 6.5NP,

VDecarbazineCarboxymethylcelluloseBCNU0.9% NaCI solution/ethanol 1 only30 24 0.9% NaCI solutionBD2F,Day 3061 561 101P.

V5-Hydroxypicoilnaide 1 only626 177 Hydroxypropylcellulose 856859 14001 2020P, (kiucel)BD2F,Day

NaCI solution/IN NaOH for 5 days91 C hyde 0.9% NaCI solution/Tween 289283 467884 756P. thiosemicarbazone0.9% 80BD2F1Daily

@ a additional definitions in previous tables. @ b Table 2, Footnote c.

C See Tabe 2, Footnote d.

the LD10in BD2F1mice for 70 anticancer agents was compared the total dose per equitoxic course was independent of the for 2 schedules (daily for 7 and 11 days), there appeared to be schedule. Griswold et a!. (18) also reported that in more than evidence of cumulative drug toxicity. These findings with 2 200 lethal dose determinations (daily for 7 days in Swiss mice; similar subacute dose schedules led to the assumption that all daily for 7 and I 1 days in BD2F1) the median range between animal dose schedules could be converted by considering that the lower and upper confidence limits at the 0.05 probability

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Downloaded from cancerres.aacrjournals.org on September 28, 2021. © 1979 American Association for Cancer Research. A. M. Guarino et a!. Table 5 Variation of total equitoxic doseper course accordingto scheduleDose/course (mgi kg)DrugMouse

1 for (%)6-MP strainRouteVehicleToxic dose levelDayonlyDaily 5 daysVariation NaCI solution ara-C Swiss iv. 0.9% NaCI solution LD@ 2050 1660 23 2'-Deoxythioguanosine BALB/c i.p. Water LD@ 6i 6 40 1440 lsophosphamide Swiss p. 0.9% NaCi solution LD@ 565 725 28 cis-Diamminedichloroplatinum BD2F p. 0.9% NaCI solution LD0 13 23 77 Tween 80 BCNUSwiss BD2Fi.p. i.p.0.9% 0.9 NaCI solutionLD@ LD@200 56500 60150 7

Table 6 Variability of toxicity data in repeated studies For a givendrug, the comparedstudieswereperformed,usuallyin the samelaboratory,usingthe samemousestrain,vehicle,and routeof administration but at time intervals ranging from 1 month to several years apart. Lethal dose (mg/kg/dose) Parallelism of probitre sponse Drug Mouse strain Vehicle8 Schedule LD,0 L0@ LD@ linesb Alkylating agents Cyclophosphamide BD2F 0.9% NaCI solution Daily for 5 days 26 58 131 NP,CV 21 67 >200

Nitrogen mustard Swiss Other Daily 7 days 0.7 1.2 2.2 P, C 0.4 0.7 1.2

Thiotepa BD2F 0.9% NaCI solution Day 1 only 24 27 31 P. C 16 27 46

TIC mustard Swiss Carboxymethylcelluiose/ Daily for 5 days 49 88 158 P. V ethanol 31 63 131

Antimetabolites ara-C BD2F, 0.9% NaCi solution Day 1 only 1.872 4,689 >10,000 NP, V 2,828 3,779 5,049 CD2F 0.9% NaCI solution Day I only 3,961 4,336 4,747 NP, V 1,925 6.275 >6,700

Methotrexate BD2F Water/i N NaOH Daily on Days 1 and 8 72 114 118 P. V 54 86 139

Antibiotics Actinomycin D Swiss Other Daily 7 days 0.046 0.095 0. 195 NP, V 0.053 0.079 0.116

Azaserine BD2F Other Daily 7 days 11 25 55 NP, C 21 29 40

Daunomycin BD2F1 0.9% NaCI solution Day 1 only 3.0 9.2 14 NP, V 5.4 8.0 12

Mitomycin C BD2F 0.9% NaCI solution Day 1 only 9 14 24 P. C 6 11 19

Porfiromycin Swiss Other Day 1 only 26 49 89 P, V 25 50 100

Others BCNU Swiss 0.9% NaC1solution/ Day 1 only 21 42 86 P. V alcohol 23 42 78

Camptothecin BD2F 0.9% NaCi solution Day 1 only 58 73 91 NP, V 141 252 449

cis-diamminodichloro- BD2F 0.9% NaCI solution/ Daily for 5 days 4.6 8.9 16 NP, V platinum Tween 80 4.5 4.8 5.2

Pseudourea Swiss Carboxymethyicellulose Daily for 5 days 14 35 86 NP, V 9 35 135

Vincristine BD2F 0.9% NaCi solution Day 1 only 2.7 4.5 7.6 P, V 4.2 8.2 15.4 a See definitions in previous tables.

b See Table 2, Footnote c.

C See Table 2, Footnote d.

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Table 7

miceNo.Effect of variables 0fl L050 levels in in LD@sNo. of compari sons yielding of drugDifference parallel probit re linesStrain1Variable factoracomparisons<20%21 -50% 51-1 00%>1 00%sponse (84)bRoute310 994 331 6 (100)Vehicle1013 023 519(90)Noneofabove1785 318(47)Totals491912

ii7C%10038.824.5 22.414.3 @ a Tables2to4and6. b Numbers in parentheses, number of comparisons yielding parallel probit response lines.

C Includes 3 of >200%: daunomycin (route), 300%: FU (vehicle), 250%; camptothecin (unknown variable), â€”245%.

level corresponds to about 22% variability. Institute for most drug tests in rodents, has involved an ap Further analysis of our data showed wide variation in lethal proximate 20% chance that part of the drug â€˜â€˜wasnotinjected dose levels even when using the same mouse strain, drug into the peritoneal cavity' â€˜(22). vehicle, route of administration, and schedule, which indicates The findings in the present analysis point out variables to that, in fact, the actual impact ot these parameters is difficult to consider while establishing retrospective correlations between assess accurately in retrospective analyses. animal and human quantitative toxicity. The wide mangeof The variability of the data does not necessarily reflect a lack experimental values available precludes firm conclusions from of reproducibility of such determinations. Some of the discrep retrospective toxicological studies for prospectively choosing ancies in this paper could be accounted for by the heteroge starting doses for clinical trials. neity of study circumstances and sources. It should be empha In the earlier report of Goldsmith et a!. (14), one-third LD10, sized that part of the available toxicity data in nontumor-beaning expressed in mg/sq m, was greater than the human MTD for rodents was extrapolated from experiments treating groups of 2 of 28 drug schedules (bleomycin and camptothecin). This mice in parallel to tumor-bearing mice or in pilot studies to updated survey of rodent toxicology would caution that use of determine the potential therapeutic range for the screening of one-third LD10might, according to the data base used (high or new drugs. Thus, the toxicity results often were not obtained in low LD10),potentially lead to exceeding the human MTD for 6 experiments primarily designed (e.g. , with respect to number additional drugs: dacanbazine, vincristine, methotmexate, thi of animals per test group on selection of dosage levels for otepa, L-phenylalanine mustard, and nitmomin.Six drug sched determining dose-response curves) to define LD10,LD50or LDso ules in our study were also reported by Goldsmith et a!. (14) so per se by the usual mathematical methods, such as that of that the combined studies represent 51 distinct drug schedules Litchfield and Wilcoxon (23). with available LD10's. For at least 43 (84%) of these, one-third Moreover, it is likely that different laboratories and expeni LD10in mice would have been a tolerated dose in humans. This mentalists might have used different standards for animal sup 84% mateof safe estimation for clinical trials is very similar to plies, number of animals involved per experiment, type of diet, that found by using one-third toxic dose low in larger animals degree of fasting, concentration of drug solution, size of needle (14) as a starting dose in Phase 1 trials in humans. Thus, from used for injection, and the like. However, even when such this point of view, neither lange animal or rodent quantitative factors are rigidly controlled, as in collaborative studies, a high toxicometric results present overWhelming differences. degree of variability is not uncommon in the results of toxicity The application of dose levels toxic to animals in determining tests. For example, Swoap (28) has reported that there was a tolerable starting doses for clinical trials in humans should be difference of as much as 71% in the LD50 obtained by 6 evaluated further in the light of the dose escalation scheme different laboratories giving injections of the same drug. Even chosen to reach the human MTD, as well as the number of replicates of one drug in a single laboratory differed by up to such escalations required. Decreasing the fraction of a specific 63%. Similarly, in a study involving 8 laboratories testing 4 toxic dose level obtained in a particular animal species to drugs (2), themewas a range of 12 to 58% difference between determine the initial dose in humans will always improve the LDso'sfor the same drug. tolerability of the so-defined clinical starting dose, but at the Perhaps toxicologists, in their haste to elevate their field to expense of a greaten number of dose escalations necessary to a quantitative science, have created what Ipsen (21) refers to reach a therapeutic dose. @ as an â€˜LD50-fixation.â€˜â€˜Fromthis analysis, we must caution the The potential of mouse data for selecting starting doses in clinician, who is anxiously awaiting completion of animal toxi Phase 1 clinical trials has held considerable appeal even cological studies, not to accept LDso'5 as readily as one ac though the strain was not stipulated (14, 26). The selection of cepts melting points on pure organic compounds. More than a fraction of a mouse toxic dose level must allow a margin of 60% of the drug comparisons (Table 7) varied by >20%, and safety for the range of data which may be observed with a there was >1 00% variability in 14%, including 3 examples number of drugs. However, studies designed primarily for (daunomycin, FU, and camptothecin) where the LD50values in toxicology with the specific purpose of determining LD10's, mice receiving the same drug differed by >200%. LDso'S,and LD@'smay in the future prove to be more accurate. Another source of variation arises when we consider that the This represents an added factor favoring the use of mouse i.p. route of injection, which is used by the National Cancer toxicology data as a principal determinant of the starting dose

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2210 CANCERRESEARCHVOL. 39

Downloaded from cancerres.aacrjournals.org on September 28, 2021. © 1979 American Association for Cancer Research. Adequacies and Inadequacies in Assessing Murine Toxicity Data with Antineoplastic Agents

Anthony M. Guarino, Marcel Rozencweig, Ira Kline, et al.

Cancer Res 1979;39:2204-2210.

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