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Preclinical studies suggest that amifostine could protect against toxicities induced by taxoids. We conducted a clinical and pharmacokinetic pilot study to assess the feasibility and toxicity of the docetaxel plus amifostine combination and the absence of influence of amifostine on docetaxel pharmacokinetics. We included 18 previously treated women with metastatic breast cancer (median age, 58.5 years; range, 34-74) in this single-center study. They were to receive a first course of docetaxel at 100 mg/m(2) i.v. as a 1-h infusion, then subsequent courses of docetaxel at 100 mg/m(2) preceded by amifostine at 910 mg/m(2) given as a 15-min i.v. infusion. Treatment was given every 3 weeks until disease progression or occurrence of unacceptable toxicity. We focused on neutrophils nadir (NN), time to nadir occurrence, and time to recovery of a neutrophil count >2 x 10(9)/liter, means of which were compared between cycle 1 and each of the other cycles by paired Student's t test. The total number of administered cycles was 84 (median number/patient, 4; range, 1-8). One patient received only the first course of docetaxel and was therefore not evaluable for amifostine effect. Neutropenia grade >2 occurred in 16 patients and was febrile in only 1. Other hematological and nonhematological toxicities were mild and manageable. Amifostine had no obvious influence on docetaxel-induced myelotoxicity as expressed by NN, time to nadir occurrence, and time to recovery. The mean NN (1 x 10(6)/liter) was 345 (18 patients) for cycle 1, then 318 (17 patients), 330 (13 patients), 340 (11 patients), 470 (8 patients), 200 (8 patients), 680 (5 patients), and 545 (4 patients) for cycles 2, 3, 4, 5, 6, 7, and 8, respectively (P >0.2). Individual pharmacokinetic (PK) parameters of docetaxel estimated in 11 patients undergoing blood sampling showed no influence of amifostine on the PK profile of docetaxel. Mean clearances (liter/h/m(2)) and peak concentration (C(peak); ng/ml) were 29.9 at cycle 1 versus 32.8 at cycle 2 (not significant) and 2849.9 at cycle 1 versus 2542.5 at cycle 2 (not significant), respectively. The population analysis including those 11 patients and 49 additional patients receiving docetaxel in other studies confirmed those findings. This study does not support evidence for amifostine cytoprotection against docetaxel-induced myelosuppression and shows that there is no major influence of amifostine on docetaxel PK parameters. For the other toxicities, which are usually cumulative, the present study has design limitations and further comparative trials are warranted.

Encouraged by preclinical synergism between docetaxel and 5-fluorouracil (5FU), we conducted a Phase I study of docetaxel in combination with continuous i.v. infusion of 5FU in patients with advanced solid tumors to determine the maximum tolerated dose, the recommended dose for Phase II studies, and the safety and pharmacokinetic profiles of this combination. Forty-two patients with advanced solid tumors, most of whom had been previously treated, received docetaxel on day 1 as a 1-h i.v. infusion, immediately followed by a 5-day continuous i.v. infusion of 5FU, every 3 weeks without hematopoietic growth factor support. All patients were premedicated with methylprednisolone. Dose levels of docetaxel/SFU studied were (daily dose, in mg/m2) 60/300, 75/300, 75/500, 75/750, 85/750, 85/1000, and 75/1000. Forty-one patients were assessable for toxicity. The maximum tolerated dose determined during the first cycle was 1000 mg/m2/day for 5 days of 5FU with either 75 or 85 mg/m2 docetaxel. Dose-limiting toxicities at these dose levels were reversible secretory diarrhea (4 of 12 evaluable patients), stomatitis (2 patients), and febrile neutropenia (2 patients). Overall, grade 3/4 neutropenia and febrile neutropenia were seen in 63.4% and 9.8% of the patients, respectively. Four patients experienced grade 3/4 infection, which led to toxic death in one of them. There were five early deaths: (a) one was clearly treatment related; (b) two others were possibly treatment related or remotely treatment related; and (c) two deaths were not related to the study drugs. Partial responses were documented in 5 of 39 evaluable patients. Pharmacokinetic results of both drugs were consistent with those from single-agent studies. The recommended dose of this combination, which showed acceptable toxicity and antitumoral activity at various dose levels, is 85 mg/m2 docetaxel given as a 1-h i.v. infusion on day 1 immediately followed by a 5-day continuous i.v. infusion of 5FU (750 mg/m2/day). This study has been extended by adding cisplatin on day 1 of the combination of docetaxel and 5FU.

The binding of estradiol in extracts of human benign prostatic hypertrophy (BPH) tissue was studied using the agar gel electrophoretic method. Two distinct binding peaks were observed having the same high specificity for natural estrogens and very poor affinity for the other classes of steroids as well as for diethylstilbestrol (DES). Therefore, it is highly probable that these peaks are two different forms of the low affinity binding protein present in human prostate. Cytosols were shown to contain smaller amounts of the binding proteins than the tissular extracts. This difference probably resulted from protein degradation occurring during the time required to prepare the cytosol. The concentrations of the extract and of the radiolabeled ligand, the composition of the buffer, the addition of sodium molybdate, sodium tartrate or proteolytic inhibitors, were evaluated in order to delineate optimal binding conditions. The possible function of these low-affinity high-capacity binding proteins is discussed.

Binding of the natural estrogens, estradiol and estriol, was investigated, in 34 samples of human benign prostatic hypertrophy (BPH) tissue, using Scatchard analysis and agar gel electrophoresis. Saturation binding analysis using a wide range of concentrations of both ligands resulted in curvilinear Scatchard plots. This confirmed the presence of two binding forms for estradiol: a true estrogen receptor, and a protein with lower affinity and higher capacity. Both binding species were also demonstrated and quantified with estriol. The electrophoretic process, after incubation at low and high ligand concentrations also resulted in separation, for both estrogens, of two binding peaks. They are probably two distinct forms of the low affinity, high capacity binding measured by Scatchard. The procedure used in our laboratory was not able to provide accurate determination of the concentrations of these binding forms. Possible modifications to alleviate these drawbacks are discussed.

A pilot study with amphotericin B incorporated in sonicated liposomes (ampholiposomes) made of egg phosphatidylcholine, cholesterol and stearylamine in a molar ratio 4:3:1 was performed in cancer patients with fungal infections. Fifteen patients received a total of 117 intravenous infusions of ampholiposomes. The total dose of amphotericin B administered per patient ranged from 20 to 1004 mg (mean 472 mg). The number of infusions per patient varied from 1 to 20 (mean 8) and the duration of treatment from 1 to 29 days (mean 10 days). Infusion of doses up to 1.8 mg/kg was well tolerated. None of the common side-effects of Fungizone, the colloidal suspension of amphotericin B, occurred; it was noteworthy that patients had no renal function impairment. Serum amphotericin B concentrations given as ampholiposomes were much higher than those obtained with Fungizone. With a daily treatment schedule, peak and trough serum amphotericin B concentrations, as measured by HPLC, were 10 to 20 micrograms/ml and 5 to 10 micrograms/ml respectively; while they did not exceed 2 micrograms/ml and 1 microgram/ml with Fungizone. Amphotericin B given as ampholiposomes had a prolonged serum beta half-life (25.3 +/- 16.0 h). Higher serum antifungal activity was observed with ampholiposomes as compared to Fungizone. We concluded that ampholiposomes have a better therapeutic index than Fungizone.

Five patients with advanced multiresistant neoplasms were infused with NSC-251635 (a water-insoluble antimitotic agent) entrapped in egg yolk lecithin, cholesterol, and stearylamine liposomes. The maximum volume injected was 400 ml containing 8 g of lipids, ie, a dose of 135 mg/kg of body weight. Overall tolerance of the treatment was excellent. This study indicates that liposomes may be used safely as carriers for the iv administration of water-insoluble drugs to humans.

We have searched for tissue-specific binding of 5alpha-androstan-17beta-ol-3-one (5alpha-dihydrotestosterone; 5alpha-DHT) in cytosols prepared from 25 surgically obtained benign prostatic hypertrophy (BPH) samples and in 3 tissue specimens containing prostate cancer cells. The distinction between steroid-receptor complexes and ligand binding to serum sex hormone-binding globulin (SHBG) was facilitated by combination experiments involving both sucrose gradient ultracentrifugation and agar gel electrophoresis. Gradient analysis of a cytosol prepared from a cervical lymph node (CLN) containing metastatic prostate tissue, revealed both 8-S and 4-S forms of high affinity (charcoal stable) 5alpha-[3H]DHT binding. When electrophoresis was performed on gradient fractions from these zones, anodally migrating steroid-receptor complexes were found only in the 8-S peak, the 4-S region containing radioligand bound to cathodally directed SHBG. In similar experiments with two BPH samples heavily invaded with prostate cancer cells only a single 4-S peak of radioligand binding was detected. Its multicomponent nature was uncovered electrophoretically when, in addition to SHBG, saturable, androgen binding molecules appeared anodally. Their incomplete resolution from SHBG on a gradient might have prevented their identification had this been the only method used. In contrast to the cancer-containing tissues, no saturable 5alpha-[3H]-DHT binding, other than that to SHBG, was detected in any of the BPH samples analysed. It is considered that, of the methods currently available, agar gel electrophoresis may be particularly useful for further investigations into the possible multicomponent nature of androgen binding of tissue origin in the human prostate.

Nocodazole, a water insoluble antimitotic drug active on L1210 leukemia was incorporated into liposomes, to investigate whether this procedure could increase cellular uptake. The effects of micronized nocodazole and liposome-entrapped nocodazole were compared on human marrow cells in vitro using a myeloid progenitor cell assay (CFU-C). The human CFU-C were sensitive to the micronized drug in a dose-related fashion. However, contrasting with a previous report on L 1210 leukemia, entrapping of nocodazole into liposomes did not increase its activity of CFU-C of either normal or leukemic subjects.

The use of sonicated phospholipid vesicles (liposomes) as carriers of 2-[3'-(methoxycarbonylamino)-phenyl]-3-phenyl-6-methoxycarbonylamino-4-(3H)- quinazolone (NSC-251635), a water-insoluble antimitotic compound, was investigated in mice. NSC-251635 was incorporated in egg yolk lecithin, cholesterol, and stearylamine (4:3:1) liposomes. In vitro, NSC-251635 in suspension or entrapped in liposomes was not toxic for L1210 cells. In vivo, after ip or iv injections to CDF1 mice bearing intraperitoneal or intravenous L1210 leukemia, NSC-251635 was active only when it was incorporated in the liposomes and not when it was given as a suspension in Klucel or in saline. The NSC-251635 liposome preparation induced significantly prolonged survival of the treated animals.

The use of sonicated phospholipid vesicles (liposomes) as carriers of methyl [5-(2-thienylcarbonyl)-1H-benzimidazol-2-yl] carbamate (Nocodazole), a water insoluble antimitotic compound active on mouse L1210 leukemia was investigated. Nocodazole was incorporated in dipalmitoyl-phosphatidylcholine: cholesterol: stearylamine (4:3:1) liposomes that were stable at room temperature for at least 48 hr. No drug leakage nor lipid exchange occurred after a 4 hr incubation at 37 degrees C with RPMI 1640 medium supplemented with 10% fetal calf serum. L1210 cells preincubated (2 x 10(6) cells/ml) at 37 degrees C for 3 hr with various concentrations of micronized Nocodazole or liposome-entrapped Nocodazole were injected i.p. into normal CDF1 mice (10(5) cells/mouse). Longest mean survival times and long-time survivors were observed in the group inoculated with L1210 cells preincubated with liposomes containing Nocodazole. CDF1 mice bearing i.p. or i.v. L1210 leukemia were treated i.p. on days 1, 5 and 9 with micronized or liposome-entrapped Nocodazole. Administration of this latter preparation induced a 50% increase in animal life span at the dosage (25 mg/kg/day) half the one required with the free compound (50 mg/kg/day). The present data indicate that enclosing Nocodazole, a water insoluble antimitotic compound, in liposomes results in an enhanced therapeutic activity against L1210 murine leukemia.