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RPR 130401, a Nonpeptidomimetic Farnesyltransferase Inhibitor with in Vivo Activity

P. VRIGNAUD,a M.C. BISSERY, P. MAILLIET, AND F. LAVELLE

Rhône-Poulenc Rorer S.A., Centre de recherche de Vitry-Alfortville, 13 quai Jules Guesde, B.P. 14, 94403 Vitry sur Seine, France

A nonpeptidomimetic farnesyltransferase inhibitor (FTI) was identified throughan enzymatic Ras screening, leading to the synthesis of benzo[f]perhydroisoindolederivatives, a new class of competitive inhibitors with the farnesyl pyrophosphatesubstrate.1 One compound of this family, exhibiting enzymatic and cellular activitiesat a micromolar level, was evaluated in vivo, but was found inactive.2 Thereafter,new chemical modifications were performed. RPR 130401 was selected for its cyto-static in vitro activity in a colony formation assay against human Ki-Ras carcinomacells, inhibiting concentration 50% (IC50) ranging from 0.045 to 0.48 µM. The goalof the work presented herein was to evaluate its in vivo activity.

According to the chemotherapy methods previously described,3 the in vivo anti-tumor efficacy of RPR 130401 was evaluated against human Ki-Ras carcinomas ob-tained from the American Type Culture Collection and xenografted in Swiss nudemice: two colons (HCT 116 and SW620), one pancreas (MIA PaCa-2), and one lung(H460). Swiss nude mice were grafted sc with tumor implants on day 0. Tumorswere measured with a caliper 2–3 times a week, and tumor weight was evaluated us-ing the following formula: tumor weight (mg) = length (mm) × width2 (mm2)/2. RPR130401 was suspended in water containing 0.5–1% polysorbate 80 and 0.5% methylcellulose. It was administered orally twice a day for 19 consecutive days to micebearing palpable sc tumors at dosages of 248, 400, and 645 mg/kg/administration. Adosage producing 20% body weight loss or drug-related death was declared toxic.The end points were tumor growth delay (T-C ��difference in median times requiredfor treatment and control groups to reach a predetermined size) and log10 cell kill net(lck net =�tumor growth delay − treatment duration/3.32 × tumor doubling time). Cy-tostatic activity was obtained for a tumor growth delay corresponding at least to thetreatment duration. A negative lck net indicated that tumor grew under treatment.

Pharmacokinetics of RPR 130401 was determined following a single administra-tion of 250 mg/kg in nontumor-bearing B6D2F1 mice (FIG. 1). The area under thecurve (AUC0–24h) was 820.3 h.µg/ml, with a 2.8-hour half-life of elimination. Peakplasma concentration was 142.1 µg/ml 2 hours postdose, a concentration 100 timesgreater than the in vitro IC50 (∼0.091 µg/ml in HCT 116 colony formation assay),and 19.9 µg/ml 8 hours postdose. Therefore, for efficacy studies, this compound wasadministered orally twice a day, 7 hours apart, for 3 consecutive weeks.

aPhone, 33 1 55 71 36 29; fax, 33 1 55 71 34 71.e-mail, Patricia.VRIGNAUD@RP-RORER.FR

250 ANNALS NEW YORK ACADEMY OF SCIENCES

Against the most in vitro sensitive tumor, colon HCT 116, tumor size at the startof therapy ranged between 80 and 180 mg in the various treatment and controlgroups. The highest dosage administered was well tolerated with no bodyweight loss and no adverse effects. Cytostatic activity was achieved with a tumorgrowth delay of 22.8 days and an 0.3 lck net (FIG. 2). There was a clear dose responseeffect in the tumor growth delays at the two dosages below 400 and 248 mg/kg/ad-ministration with a transient cytostatic effect. Tumor growth delays were 17.3 and11.7 days, respectively, with a negative lck net.

In mice bearing colon carcinoma SW620 with a 0–179 mg tumor weight at thestart of therapy, the highest dosage tested of 645 mg/kg/administration was well tol-erated and produced a tumor growth delay of 15.7 days for a treatment duration of19 days and a negative lck net (−0.3). This indicates that no cytostatic activity wasobtained. The two lowest doses of 400 and 248 mg/kg/administration were alsoinactive.

At the highest dose tested, 645 mg/kg/dam, RPR 130401 was toxic in lung H460-bearing mice, with two of eight drug-related deaths associated with excessive bodyweight loss. Tumor size at the start of therapy ranged between 100 and 200 mg. Theoptimal dosage of 400 mg/kg/administration produced no cytostatic activity with atumor growth delay of 15.2 days for a treatment duration of 19 days and a negativelck net (−0.4). The lowest dosage was also inactive.

In the same way, three of eight drug-related deaths occurred at the highest dosagetested of 645 mg/kg/administration in mice bearing pancreas MIA PaCa-2, with tu-mor size at the start of therapy ranging from 80–170 mg. Although we observed aslow down in tumor growth in mice treated at the optimal dosage of 400 mg/kg/ad-

FIGURE 1. Pharmacokinetics of oral RPR 130401. RPR 130401 was suspended inwater containing 0.5% polysorbate 80 and 0.5% methyl cellulose and administered toB6D2F1 female mice at 250 mg/kg. Plasma was collected at 5, 15, and 30 minutes and 1, 2,3, 4, 6, 8, 12, and 24 hours postdose. Plasma levels were determined by HPLC with UV de-tection (LC-MS/MS, limit of quantification was 0.05 µg/ml).

251VRIGNAUD et al.: RPR 130401

ministration, RPR 130401 did not produce any cytostatic activity (T − C = 10.6 days;−0.7 lck net).

In conclusion, RPR 130401 was bioavailable by the oral route and produced cy-tostatic activity against one Ki-Ras activated carcinoma, colon HCT 116, xenograft-ed in nude mice. Further in vivo evaluation of RPR 130401 will be performed inhuman and murine tumors exhibiting no Ki-Ras mutations, the main point being toevaluate this type of compound in combination with chemotherapy.

REFERENCES

1. MAILLIET, P. et al. 1997. Proc. Am. Assoc. Cancer Res. 39: 2347.2. VRIGNAUD, P. et al. 1997. Proc. Am. Assoc. Cancer Res. 39: 2348.3. SCHABEL, F.M., JR. et al. 1977. Quantitative evaluation of anticancer agents in experi-

mental animals. Pharmacol. Ther. Part A 1: 411–435.

FIGURE 2. RPR 130401 in vivo efficacy against human colon carcinoma HCT116.RPR 130401 was administered orally twice daily for 19 consecutive days at dosages of 248(�), 400 (�), and 645 mg/kg/administration (�) to Swiss nu/nu mice bearing measurablehuman colon HCT116 (80–180 mg). Each curve represents the median tumor growth of eachgroup. Cytostatic activity = (tumor growth delay ≥ treatment duration), corresponding to apositive log10 cell kill net.