Pharmacokinetic Approach to Rational Therapeutic Doses for Human Tumor-bearing Nude Mice

To improve clinical predictability from therapeutic results of various antitnmor agents in human tumor/nude mouse models it seems to be important to use a dose pharmacokinetically equivalent to the clinical dose. Thus, we attempted to find the dose of a given drug that can reproduce in the nude mouse a plasma level similar to that seen in human patients treated with an effective dose of the drug based on comparative pharmacokinetic studies between man and nude mouse. As a result, those of 3 alkylating agents, mitomycin C, 3-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-l-(2-chloroethyl)-l-nitrosourea (ACNU) and cyclophosphamide, and those of 2 antimitotic agents, vincristine and vinblastine, were estimated to be one-fourth or one-fifth of their maximum tolerated doses (MTD's). On the other hand, in the case of adriamycin, its MTD was approxmately equivalent to its clinical dose pharmacokinetically. In contrast, clinically equivalent doses of 2 antimetabolites tested, 5-fluorouracil and methotrexate, were significantly greater than their MTD's; i.e., their plasma levels did not reach the effective clinical ones even when their MTD's were administered to the nude mice. These results suggest that the antitumor effects of mostv antitumor agents are over- or underestimated in this model when MTD's are used as a therapeutic dose, and indicate that the use of clinically equivalent doses determined pharmacokinetically isdesirable.     

Pharmacokinetic approach to the improvement of clinical predictability in the preclinical test for antitumor agents

Clinical predictability of preclinical test for antitumor agents has not been significantly improved even after the use of a human tumor/nude mouse model. Such different antitumor activities between preclinical and clinical tests probably due to the fact that therapeutic used in both tests usually each maximum tolerated dose (MTD), are pharmacokinetically not equivalent. Therefore, we introduced a new concept of "clinically equivalent dose (CED)", which can reproduce in the nude mouse the blood level of a given drug observed with human patients received its therapeutic dose. Treatment of human tumors implanted in the nude mice with CEDs of several drugs exhibited much better correlation with their clinical efficacies than those with MTDs. The feasibility of use of CED predicted by animal scale-up procedure as a therapeutic dose in the preclinical test was discussed.     

Responsiveness of human gastric tumors implanted in nude mice to clinically equivalent doses of various antitumor agents

To reproduce clinical effects of various antitumor agents in the human tumor/nude mouse model, we investigated the responsiveness of 11 lines of human gastric tumor xenografts to doses of the agents pharmacokinetically equivalent to the respective clinical doses, which we designated the "rational dose" (RD). We found that the response rates to mitomycin C, 3-[(4-amino-2-methyl-5-pyrimidinyl]methyl-1-[2-chloroethyl]-1- nitrosourea (ACNU), adriamycin, 5-fluorouracil were 18%, and that to vinblastine was 30%; on the other hand, those to vincristine, methotrexate, and cyclophosphamide were poor. In contrast, in our previous study using the maximum tolerated doses, response rates to mitomycin C, ACNU, and vinblastine were as high as 64-82%, and those to adriamycin and 5-fluorouracil were 18%. When these results were compared with the clinical response rates of gastric tumors, as a whole, the results with RD's exhibited much better coincidence with the clinical data in terms of relative therapeutic potency, indicating the validity of the use of clinically equivalent doses instead of maximum tolerated doses in the human tumor model.     

Responsiveness of Human Gastric Tumors Implanted in Nude Mice to Clinically Equivalent Doses of Various Antitumor Agents

To reproduce clinical effects of various antitumor agents in the human tumor/nude mouse model, we investigated the responsiveness of 11 lines of human gastric tumor xenografts to doses of the agents pharmacokinetically equivalent to the respective clinical doses, which we designated the "rational dose" (RD). We found that the response rates to mitomycin C, 3-[(4-amino-2-methyl-5-pyrimidinyl]methyl-1-[2-chloroethyl]-1-nitrosourea (ACNU), adriamycin, 5-fluorouracil were 18%, and that to vinblastine was 30%; on the other hand, those to vincristine, methotrexate, and cyclophosphamide were poor. In contrast, in our previous study using the maximum tolerated doses, response rates to mitomycin C, ACNU, and vinblastine were as high as 64–82%, and those to adriamycin and 5-fluorouracil were 18%. When these results were compared with the clinical response rates of gastric tumors, as a whole, the results with RD's exhibited much better coincidence with the clinical data in terms of relative therapeutic potency, indicating the validity of the use of clinically equivalent doses instead of maximum tolerated doses in the human tumor model.     

Evaluation of antitumor activity in a human breast tumor/nude mouse model with a special emphasis on treatment dose

Eight lines of human breast tumors implanted in nude mice were treated with various antitumor agents at two different doses, maximum tolerated doses (MTD) and rational doses (RD) that were pharmacokinetically equivalent to the clinical doses; the response rates to both doses were compared. With MTD, the response rates to mitomycin C and vinblastine were 100%, and those to other agents including cyclophosphamide, nimustine (a water-soluble nitrosourea), vincristine, Adriamycin (doxorubicin; Adria Laboratories, Columbus, OH), 5-fluorouracil (5-FU), and methotrexate were 30%-50%, indicating high responsiveness to the former two agents. In contrast, when the RD were used, the response rates to the majority of these agents were 25%-40%, and those to vincristine and nimustine were 13% and 0%, respectively. These results agree with the reported clinical results compared with those with MTD, suggesting the importance of the use of clinically equivalent doses in the evaluation of antitumor efficacy in a human tumor/nude mouse system.     

Evaluation of response rates to various antitumor agents of human gastric tumors implanted in nude mouse

To ascertain the clinical predictability and, in the long run, the usefulness of the human tumor/nude mouse model as a screening tool for antitumor agents, it seems particularly important to use as many tumor lines as possible and to evaluate the therapeutic effectiveness of antitumor agents in terms of the overall response rate of a range of tumors. In this study, using 11 strains of established human gastric tumor xenografts with various histological characteristics and proliferation behavior, the experimental response rates to 8 typical antitumor drugs (mitomycin C, cyclophosphamide, ACNU, cisplatin, adriamycin, vincristine, vinblastine and 5-fluorouracil) were determined and compared with the clinical values. The experimental response rates to adriamycin, vincristine and 5-fluorouracil were in good accordance with the clinical results. However, with the other drugs, significantly higher response rates were observed with the nude mouse model as compared to clinical therapy, indicating that this model tends to overestimate the responsiveness of human tumor to a number of antitumor agents. These results strongly suggest the importance of using appropriate dose levels in the nude mouse to reproduce clinically equivalent effects in this model.     

Responsiveness of human lung cancer/nude mouse to antitumor agents in a model using clinically equivalent doses

Summary The responses of 14 lines of human lung cancer xenografts in BALB/c-nu/nu mice to eight known antitumor agents were investigated. These xenografts consisted of four small-cell carcinomas (SCLC) and ten non-small-cell carcinomas (four large cell, three squamous cell, and three adenocarcinomas; NSCLC). The doses used in the experiments were the maximum tolerated dose (MTD) in nude mice and the rational dose (RD), the latter considered to be pharmacokinetically equivalent to the clinical dose. When given at MTDs, all drugs except 5-fluorouracil (5-FU) and methotrexate (MTX) were extremely effective against NSCLC as well as SCLC. The response rates of drug-sensitive SCLC to mitomycin C (MMC), ACNU, and vinblastine (VLB) were 100%, and those to Adriamycin (ADR) and vincristine (VCR) were 75%. In addition, the response rates of even drug-resistant NSCLC to MMC and VLB were 70% and 90%, respectively. In contrast, the response rates of NSCLC to RDs of the drugs were reduced to <40% and corresponded well to the respective clinical rates. In SCLC, a good correlation of experimental and clinical response rates was observed with four drugs [cyclophosphamide (CPM), ACNU, VLB, and 5-FU]. As a result, we emphasize that a more reasonable prediction of the clinical effectiveness of antitumor agents can be made by a protocol using clinically equivalent doses.Abbreviations SCLC small-cell lung cancer - NSCLC non-small-cell lung cancer - MTD maximum tolerated dose - RD rational dose - MMC mitomycin C - CPM cyclophosphamide - ACNU 1-(4-amino-2-methylpyrimidin-5-yl)-methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride - ADR Adriamycin - VCR vincristine - VLB vinblastine - 5-FU 5-fluorouracil - MTX methotrexate This study was supported in part by Grants-in-Aid for New Drug Development Research from the Ministry of Health and Welfare, Japan     

Responsiveness of subcutaneous human glioma xenografts to various antitumor agents

Responsiveness of seven human glioma xenografts to seven antitumor agents was investigated by an sc-iv system and the efficacies of these agents against human glioma were evaluated in terms of response rate. When their maximum tolerated doses were used, experimental response rates of nimustine (ACNU), vincristine (VCR), adriamycin (ADR) and vinblastine (VLB) were as high as 86-100%, while that of mitomycin (MMC) was 57%, and those of 5-fluorouracil (5-FU) and methotrexate (MTX) were 0%. On the other hand, when the doses pharmacokinetically equivalent to their clinical doses were employed, the response rate of ADR was the highest, followed by VCR and ACNU in this order. These results suggest that gliomas are significantly responsive to various antitumor agents in this sc-iv system.     

Preclinical and phase I studies with rhizoxin to apply a pharmacokinetically guided dose-escalation scheme.

BACKGROUND: Rhizoxin is a new macrocyclic lactone isolated from the fungus Rhizopus chinensis which displays broad-spectrum antitumor activity against murine and human tumor xenografts and has activity against a number of vincristine-resistant tumors in vitro and in vivo. PURPOSE: This study describes the preclinical and clinical pharmacology of rhizoxin to apply a pharmacokinetically guided dose-escalation (PGDE) strategy during the phase I trial. METHODS: Rhizoxin was administered by a single intravenous bolus injection to female BALB/c mice over the dose range 7.5-18 mg/m2 from which we derived the dose that was lethal to 10% and 50% of the mice (i.e., LD10 and LD50, respectively). The LD10 was 11.7 +/- 0.7 mg/m2 (mean +/- SD), and the LD50 was 14.7 +/- 0.6 mg/m2. Pharmacokinetic studies were integrated with the toxicity study in female BALB/c mice at one-tenth the LD10, one-half the LD10, and the LD10 (i.e., 1.2, 6, and 12 mg/m2, respectively). From these data, a target area under the plasma drug concentration versus time curve (AUC) (i.e., 40% of the LD10 AUC) was calculated for clinical studies. Phase I studies were initiated at 0.8 mg/m2 (one-tenth the equivalent LD10 in male CD1 mice), with the intent of escalating the dose by an extended factor-of-two method until the target AUC and/or maximum tolerated dose (MTD) was reached. RESULTS: The major drug toxic effects in mice were body weight loss, sluggishness, ataxia, transient changes in hematological parameters, and hematuria. Diarrhea was universal at doses greater than 9 mg/m2, and hind limb paralysis was observed in one of 10 mice, but only at supralethal doses (18 mg/m2). Rhizoxin pharmacokinetics were best described by a two-compartment open model (half-life [t 1/2] alpha = 4.4 minutes +/- 0.9 minute [mean +/- SD], and t 1/2 beta = 84 minutes +/- 20 minutes at 12 mg/m2) and found to be nonlinear with respect to dose. At doses of 1.2, 6, and 12 mg/m2, the respective AUC values were 1.3, 22.4, and 70.6 microM x minute. From these data, a target AUC value of 28 microM x minute (40% of the LD10 AUC) was derived. Rhizoxin was not detectable in patient plasma (less than 5 ng/mL at 0.8 and 1.6 mg/m2), and doses had to be escalated by conventional methods. Myelosuppression was dose limiting in patients: Seven of eight treated at 2.6 mg/m2 experienced World Health Organization grade 3-4 neutropenia, and five of eight developed mucositis. The AUC values at the human MTD (2.6 mg/m2) were in the range of 0.41-1.01 microM x minute, considerably lower than the target AUC of 28 microM x minute. CONCLUSION AND IMPLICATIONS: Although PGDE schemes have been successfully employed for other antitumor agents, this methodology could not be applied during the phase I trial of rhizoxin. PGDE studies in the future may incorporate comparative murine versus human metabolism studies in vitro with phenotyped liver microsomes. It may also be useful to assess the comparative myelotoxicity of a new drug by performing in vitro cytotoxicity studies on mouse and human bone marrow stem cells.     

Improvement of the antitumor activity of intraperitoneally and orally administered 5,6-dimethylxanthenone-4-acetic acid by optimal scheduling.

PURPOSE: 5,6-Dimethylxanthenone-4-acetic acid (DMXAA), a new anticancer drug that has recently completed Phase I clinical trial, is effective against transplantable murine tumors with established vasculature. We wished to determine the relationship between administration schedule and antitumor activity. EXPERIMENTAL DESIGN: C57Bl/6 mice with s.c. implanted Colon 38 tumors were used for determination of maximal tolerated doses and tumor growth delay. Plasma and tissue DMXAA concentrations were measured by high-performance liquid chromatography. RESULTS: Continuous infusion (30 mg/kg/day for 3 days) and daily i.p. administration schedules (7.5 mg/kg) were ineffective. A pharmacokinetically guided schedule was developed to increase tumor tissue drug concentrations without increasing the maximal plasma concentration. A schedule comprising a loading dose (25 mg/kg, i.p.) followed by supplementary doses (5 mg/kg after 4 and 8 h) provided a 1.6-fold increase in tumor tissue area under the concentration-time curve, no increased toxicity, and superior antitumor activity (100% cure rate, as compared with 55% for a single i.p. dose of 25 mg/kg). A similar strategy was developed for oral administration with a loading dose (30 mg/kg) and supplementary doses (15 mg/kg after 4 and 8 h). It provided a 90% cure rate, in contrast to a single oral dose (0% cure rate). CONCLUSIONS: The antitumor action of DMXAA is schedule dependent, and the achievement of an adequate tumor tissue DMXAA concentration above a threshold value appears to be critical for activity. The use of a pharmacokinetically guided schedule provides excellent oral activity against Colon 38 tumors and provides a basis for developing more effective administration schedules in clinical trials.     

Pharmacology and clinical toxicity of 4'-iodo-4'-deoxydoxorubicin: an example of successful application of pharmacokinetics to dose escalation in phase I trials

In a prospective phase I trial involving 35 patients with metastatic carcinoma, we tested a pharmacokinetic strategy for guiding dose escalation of the anthracycline 4'-iodo-4'-deoxydoxorubicin (I-DOX), a new analogue reported to be more potent and less toxic than doxorubicin. This strategy is potentially a safe and more rapid way of determining the maximum tolerated dose (MTD) of anticancer agents. Retrospective studies have shown that the total plasma drug exposure after a dose lethal to 10% of mice (LD10) is approximately equivalent to the total exposure produced in humans by the MTD. Thus, we intended to aim dose escalation in humans to achieve the area under the curve for I-DOX plasma concentration x time (AUC) equivalent to that produced in mice by an LD10. However, differences in I-DOX pharmacokinetics and metabolism in BDF1 mice and humans at the initial dose prevented immediate application of this strategy. Therefore, we escalated the dose by the modified Fibonacci scheme while investigating the pharmacology of I-DOX and its major plasma metabolite 4'-iodo-4'-deoxy-13-dihydrodoxorubicin (I-DOXOL). Plasma pharmacokinetics was characterized by rapid elimination and extensive metabolism of I-DOX to I-DOXOL. The ratio of I-DOXOL to I-DOX plasma AUC was 12.8 +/- 7.3 SD. The plasma pharmacokinetics of I-DOX and I-DOXOL were linear in the range of tested doses (2-90 mg/m2). The LD10 in mice was 6.8 mg/kg for I-DOXOL and 6 mg/kg for I-DOX, and the concentration of drug that inhibited by 50% (IC50) the growth of human granulocyte-macrophage colony-forming units (CFU-GM) was 80 nM for I-DOXOL and 50 nM for I-DOX. From these findings, we concluded that the toxic effects of I-DOX and I-DOXOL are equivalent and reset the pharmacokinetic target of escalation to the sum of I-DOX and I-DOXOL AUCs at I-DOX LD10. Then we safely applied pharmacokinetically guided escalation to determine the MTD (80 mg/m2). The plasma AUC of I-DOX and I-DOXOL at the human MTD is 71% of the AUC at mouse LD10. The only dose-limiting toxic effect was severe granulocytopenia.     

Development of the novel biologically targeted anticancer agent gefitinib: determining the optimum dose for clinical efficacy.

The emergence of novel, biologically targeted anticancer agents such as gefitinib ('Iressa', ZD1839) has raised the question of how the dose for later-stage clinical development and clinical use is best determined. For cytotoxic drugs, because toxic effects and antitumor activity often fall within the same dose range and are dose dependent, the clinically used dose will depend on the therapeutic window. Therefore, the maximum tolerated dose identified in Phase I trials is typically used to determine the dose level for Phase II and III trials. However, because biologically targeted agents are expected to provide clinical benefits that are not predicted by surrogate end points of toxicity to normal replicating tissue, new Phase I trials have been designed to determine the optimum biological dose for use in further studies. A large, multifaceted Phase I program was designed to evaluate the pharmacokinetics, safety, efficacy, and targeted biological activity of a once-daily oral dose of gefitinib. The maximum tolerated dose was >or=700 mg/day, although doses as low as 150 mg/day provided (a). plasma concentrations sufficient for pharmacological activity, (b). evidence of targeted biological effect, and (c). antitumor activity. From these observations, two large Phase II trials ('Iressa' Dose Evaluation in Advanced Lung Cancer 1 and 2) evaluated 250- and 500-mg/day doses of gefitinib in patients with advanced non-small cell lung cancer (NSCLC). As predicted from the Phase I trials, doses >250 mg/day provided no additional efficacy benefit, whereas adverse effects increased in a dose-dependent manner. Consequently, the recommended dose of gefitinib in NSCLC is 250 mg/day. The early clinical trial development of gefitinib provides a model for the development of novel, noncytotoxic anticancer agents.     

Clinically relevant suramin dosing regimen in mice with no effects against PC-3 prostate xenografts

The goal of these studies was to develop a suramin dosing schedule that would maintain suramin plasma concentrations in mice in the 150-250 mu g/ml range. A high pressure liquid chromatography method was used to determine suramin plasma concentrations in mice. For pharmacokinetic studies CD2F(1) mice were treated intraperitoneally with 140 mg/kg of suramin. These pharmacokinetic data were used to develop a clinically relevant dosing regimen. To test the efficacy of this dosing regimen, athymic nude mice were implanted orthotopically with PC-3 prostate carcinoma cells, randomized, and treated intraperitoneally. The pharmacokinetically derived dosing regimen resulted in no antitumor effect against PC-3 prostate tumors. Suramin plasma concentrations ranged from 155 to 258 mu g/ml over the 14-day therapy period with tumor concentrations in the 53-241 mu g/g wet weight range.     

Synergistic antitumor activity of irinotecan in combination with 5-fluorouracil in rats bearing advanced colorectal cancer: role of drug sequence and dose

The basis for current clinical trials in the treatment of colorectal cancer with the combination of irinotecan (CPT-11) and 5-fluorouracil (FUra) with or without leucovorin (LV) is their proven activity as single agents, their different mechanisms of action, and lack of CPT-11 cross-resistance to previous FUra/LV treatment. The role of drug dose and administration sequence in this combination was studied in vivo using a rat colon tumor model (Ward colon carcinoma); we administered CPT-11 and FUra by i.v. push once a week for four consecutive weeks (weekly x 4), a clinically relevant schedule. The maximum tolerated doses (MTDs) of CPT-11 and FUra administered as single agents were 100 mg/kg/week for both agents. Three different combination administration sequences were evaluated: (a) CPT-11 administered simultaneously with FUra (sequence I); (b) FUra administered 24 h before CPT-11 (sequence II); and (c) CPT-11 administered 24 h before FUra (sequence III). When combining the two drugs at 50% of their respective MTD, the antitumor efficacy was sequence dependent with 62, 38, and 95% complete tumor regression rate for sequences I, II, and III, respectively. For sequences I and II, dose escalation to 75% of the MTD for each drug was paralleled by reversible host toxicity with no significant increase in the antitumor activity of the combination. With sequence III, however, the combination was lethal in 100% of treated animals when the doses of both drugs were at 75% of the MTD or higher. With the sequential combination of CPT-11 followed 24 h later by FUra (sequence III), the high complete tumor regression rate (cure) could be maintained, even when the dose of CPT-11 was reduced to 12.5% of the MTD as long as the doses of FUra was kept at 50 -75 % of the MTD. The data demonstrate that the antitumor activity and toxicity of combining CPT-11 with FUra is highly sequence dependent and that a sequence of CPT-11 preceding FUra is superior with a significant increase in the therapeutic index over the other sequences tested. In addition, the data also demonstrate that toxicity associated with high dose of CPT-11 can be eliminated without loss of the antitumor efficacy by reducing the dose of CPT-11 to at least 50% of its MTD, whereas the dose of FUra is kept at 50-75 % of its MTD.     

Phase I and pharmacokinetic study of a stable, polyethylene-glycolated liposomal doxorubicin in patients with solid tumors: the relation between pharmacokinetic property and toxicity

BACKGROUND: Compared with free drug, sterically stabilized liposomal drug has prolonged circulation time and, thereby, higher tumor selectivity and antitumor activity. The stability in plasma is an important consideration in the formulation of clinically useful liposomal drug. A Phase I study of a stable liposomal doxorubicin with polyethylene glycol (PEG) coating and phospholipid component of distearoyl phosphatidylcholine (DSPC) was performed to characterize its pharmacokinetic properties, toxicity profile, and maximal tolerated dose. METHODS: The starting dose was 30 mg/m(2) every 3 weeks with an increment of 10 mg/m(2) for each level. A cohort of at least three patients was entered for each level. Dose escalation stopped when more than one-third of patients had dose limiting toxicity (DLT), which was equal to or more than Grade 3 nonhematologic toxicity. Blood was sampled immediately before and at 5 minutes, 2 hours, 4 hours, 10 hours, 24 hours, 48 hours, 72 hours, and 168 hours after the completion of PEGylated liposomal doxorubicin (PLD) infusion. Plasma level of doxorubicin was determined with fluorometry, and the pharmacokinetic properties were analyzed. RESULTS: Twenty-six patients were entered, and 101 courses were studied. This DSPC PLD had a steady-state distribution volume (Vss) of 2.4 +/- 0.9 liters (mean +/- standard deviation), a clearance of 0.027 +/- 0.010 liters per hour, and a beta half-life of 65.0 +/- 17.8 per hour. These characteristics were dose independent, and the Vss and clearance were smaller than those of a well characterized PLD comprised of hydrogenated soybean phosphatidylcholine (HSPC). At the dose level of 50 mg/m(2), its plasma area under the concentration time curve was approximately twice that of HSPC PLD. Attenuation of acute toxicity, such as nausea, emesis, and alopecia, was noted in all dose levels. However, stomatitis was common from the dose level of 30 mg/m(2), and its incidence and severity increased with dosage and became dose limiting at 50 mg/m(2). A dose of 45 mg/m(2) every 3 weeks was then given in eight patients, and the side effects were acceptable. This dose was recommended for Phase II clinical trials. Fourteen of 17 patients with a dose level > or = 40 mg/m(2) were evaluable for response, but none achieved partial remission. CONCLUSIONS: This DSPC PLD had the characteristics of second-generation liposomal drug pharmacokinetically and toxicologically. The incidence of severe stomatitis was higher than that of HSPC PLD, corresponding to the difference in pharmacokinetics. Only limited antitumor activity was observed, although defining its therapeutic application will need further Phase II studies. Further prolongation of plasma stability of PLD may not be clinically beneficial considering the increased stomatitis and the reduced achievable dose intensity.     

Preclinical screening of chemotherapeutic and endocrine agents using human tumor xenografts--nude mouse system

Although newly developed antitumor agents had been screened using cultured human cell lines and rodent tumor systems, it was apparent that the positive antitumor activity of the agents on these experimental tumors is not a sufficient but a necessary condition to elucidate the antitumor activity on clinical human cancers. We have clarified that the chemosensitivity of human gastric, breast colon carcinoma xenografts could reproduce the essentially identical chemosensitivity of clinical carcinomas when the maximum tolerable doses of the conventionally available agents were administered to tumor bearing nude mice, and human breast carcinoma xenografts had a similar estrogen dependency and endocrine sensitivity to clinical human breast cancer. These findings indicated that the human tumor xenografts--nude mouse is an appropriate model to predict the clinical antitumor activity of newly developed antitumor agents.     

Clinical pharmacokinetics of the anthrapyrazole CI-941: factors compromising the implementation of a pharmacokinetically guided dose escalation scheme.

The pharmacokinetics of the anthrapyrazole CI-941 has been investigated in conjunction with the Phase I evaluation of the drug with the intent of applying a pharmacokinetically guided dose escalation strategy. A starting dose of 5 mg/m2 was chosen, based on one-tenth the 10% lethal dose in mice. Due to the steep dose lethality relationship and nonlinear pharmacokinetics in mice, a target area under the CI-941 plasma concentration x time curve (AUC) of 110 microM x min (i.e., 40% of the mouse 10% lethal dose AUC) was chosen. This AUC was achieved in mice at 40 mg/m2. A total of 37 patients received 74 courses of CI-941 (5 to 55 mg/m2), with 26 patients consenting to pharmacokinetic monitoring. CI-941 was rapidly cleared from plasma, and a triexponential open model could be fitted to the data (t1/2 alpha = 7.6 +/- 2 min, t1/2 beta = 65 +/- 27 min, t1/2 zeta = 21 +/- 9 h). CI-941 was subjected to only limited urinary elimination, accounting for 5.2 +/- 2.8% of the administered dose. Wide interpatient variability in plasma CI-941 clearance at the starting dose and subsequent doses precluded the implementation of a pharmacokinetically guided dose escalation scheme, and the dose was escalated in 5-mg/m2 increments until the maximally tolerated dose was achieved. A number of investigations were performed to study potential reasons for variability in CI-941 clearance. However, CI-941 plasma protein binding (95 +/- 1%) and measures of pretreatment renal (51Cr-EDTA clearance), hepatic (plasma alanine transaminase and alkaline phosphatase levels), or cardiac function (left ventricular ejection fractions) did not relate strongly to CI-941 clearance. In patients treated at 40 mg/m2, the AUC values (156 to 415 microM x min) approximated or exceeded the target AUC. Fifty mg/m2 was the Phase II recommended dose. Further prospective studies are warranted to assess the utility of pharmacokinetically guided dose escalation strategies and to determine whether or not the variability encountered in clearance is unique to CI-941.     

Preclinical evaluation of the anticancer activity and toxicity of 9-nitro-20(S)-camptothecin (Rubitecan).

The purpose of this study is to establish the maximum tolerated dose of rubitecan in mice, dogs and men and to establish the anticancer activity of such dose against human tumors xenografted in nude mice. Nude mice received increasing doses of Rubitecan by intrastomach injection until the maximum tolerated dose (MTD) had been established for both the single dose and the multiple doses at the schedule of 5 days on, 2 days off. Extrapolating from the mouse data, MTD was determined for oral administration in dogs and man. Levels of the drug in plasma were determined by high pressure liquid chromatography (HPLC). Using maximum tolerated multiple doses, the sensitivity of human cancer xenografts in nude mice to Rubitecan was determined. MTD of Rubitecan in mice for multiple doses intrastomach at the schedule of 5+,2- was 1 mg/kg/day. MTD in dogs was also 1 mg/kg/day, administered orally but at the schedule of 4+,3-. In man, it was 1 mg/m2/day at the schedule of 5+,2-. Treatment of human cancer xenografts in nude mice with MTD of Rubitecan resulted in 100% growth inhibition of 30/30 tumors tested and in 24/30 in their total disappearance. These 30 tumors comprised all the most common human cancers: lung, colorectal, breast, pancreatic, ovarian, prostate, stomach, melanoma and a leukemia. From the data collected, it appears that rubitecan is a very promising anticancer drug with high potency against a wide spectrum of human cancers. These cancers growing as xenografts in nude mice are always growth inhibited (30/30) and frequently (24/30) totally destroyed by the administration of non-toxic doses of Rubitecan.     

Increased plasma vascular endothelial growth factor (VEGF) as a surrogate marker for optimal therapeutic dosing of VEGF receptor-2 monoclonal antibodies

A major obstacle compromising the successful application of many of the new targeted anticancer drugs, including angiogenesis inhibitors, is the empiricism associated with determining an effective biological/therapeutic dose because many of these drugs express optimum therapeutic activity below the maximum tolerated dose, if such a dose can be defined. Hence, surrogate markers are needed to help determine optimal dosing. Here we describe such a molecular marker, increased plasma levels of vascular endothelial growth factor (VEGF), in normal or tumor-bearing mice that received injections of an anti-VEGF receptor (VEGFR)-2 monoclonal antibody, such as DC101. Rapid increases of mouse VEGF (e.g., within 24 hours) up to 1 order of magnitude were observed after single injections of DC101 in non-tumor-bearing severe combined immunodeficient or nude mice; similar increases in human plasma VEGF were detected in human tumor-bearing mice. RAFL-1, another anti-VEGFR-2 antibody, also caused a significant increase in plasma VEGF. In contrast, increases in mouse VEGF levels were not seen when small molecule VEGFR-2 inhibitors were tested in normal mice. Most importantly, the increases in plasma VEGF were induced in a dose-dependent manner, with the maximum values peaking when doses previously determined to be optimally therapeutic were used. Plasma VEGF should be considered as a possible surrogate pharmacodynamic marker for determining the optimal biological dose of antibody drugs that block VEGFR-2 (KDR) activity in a clinical setting.     

Antitumor activity of poly(L-glutamic acid)-paclitaxel on syngeneic and xenografted tumors.

Poly(L-glutamic acid)-paclitaxel (PG-TXL) is a new water-soluble paclitaxel derivative that has shown remarkable antitumor activity against both ovarian and breast tumors. The purpose of this study was to test whether the antitumor efficacy of PG-TXL depends on tumor type, as is the case for paclitaxel, and to test whether paclitaxel-resistant tumors could be responsive to PG-TXL. We evaluated the therapeutic activity of PG-TXL against four syngeneic murine tumors (MCa-4, MCa-35, HCa-1, and FSa-II) inoculated i.m. into C3Hf/Kam mice, a human SKOV3ip1 ovarian tumor injected i.p. into nude mice, and a human MDA-MB-435Lung2 breast tumor grown in the mammary fat pad of nude mice. Two paclitaxel-responsive murine tumors, MCa-4 and MCa-35, showed significant growth delay with PG-TXL given as a single i.v. injection at its maximum tolerated dose of 160 mg of equivalent paclitaxel/kg or even at a lower dose of 120 mg of equivalent paclitaxel/kg. The other two murine tumors, HCa-1 and FSa-II, did not respond particularly well to either of the two agents, although significant growth delay was observed for both tumors with PG-TXL. In mice with SKOV3ip1 tumors, the median survival times for mice treated with PG alone and PG-TXL at doses of 60 or 120 mg of equivalent paclitaxel/kg were 43, 61, and 75 days, respectively; no survival difference was found between paclitaxel-treated and Cremophor vehicle-treated mice. In mice with MDA-MB-435Lung2 tumor, PG-TXL at a dose of 120 mg of equivalent paclitaxel/kg produced regression of the tumor in 50% of the animals, and in the remaining mice, micrometastases in the lung were found only in 25% of the animals. In comparison, treatment with paclitaxel at 60 mg/kg did not result in tumor regression, and the rate of lung metastases was 42%. These results clearly demonstrate that PG-TXL has significant therapeutic activity against breast and ovarian tumors tested in this study. Future studies to elucidate the mechanism of action of PG-TXL and to assess its clinical applications are warranted.