Triiodothyronine-induced thyrotoxicosis increases mononuclear leukocyte beta-adrenergic receptor density in man.

beta-Adrenergic receptors are increased in some tissues of experimentally thyrotoxic animals but are reported to be unchanged in mononuclear leukocytes of spontaneously thyrotoxic humans. We examined the effects of triiodothyronine (100 mug/d for 7 d) and placebo on high-affinity mononuclear leukocyte beta-adrenergic receptors in 24 normal human subjects, using a double-blind design. beta-Adrenergic receptors were assessed by specific binding of the antagonist (-)[(3)H]dihydroalprenolol. Triiodothyronine administration resulted in objective evidence of moderate thyrotoxicosis and an increase in mean (-)[(3)H]dihydroalprenolol binding from 25+/-3 to 57+/-9 fmol/mg protein (P < 0.001). The latter was attributable, by Scatchard analysis, to an increase in beta-adrenergic receptor density (967 +/- 134 to 2250 +/- 387 sites per cell, P < 0.01); apparent dissociation constants did not change. Placebo administration had no effects. Marked inter- and intraindividual variation in mononuclear leukocyte beta-adrenergic receptor density was also noted. Because this was approximately threefold greater than analytical variation, it is largely attributable to biologic variation. Thus, we conclude: (a) The finding of a triiodothyronine-induced increase in mononuclear leukocyte beta-adrenergic receptor density in human mononuclear leukocytes, coupled with similar findings in tissues of experimentally thyrotoxic animals, provides support for the use of mononuclear leukocytes to assess receptor status in man. (b) There is considerable biologic variation in beta-adrenergic receptor density in man. (c) The findings of thyroid hormone-induced increments in beta-adrenergic receptor density provide a plausible mechanism for the putative enhanced responsiveness to endogenous catecholamines of patients with thyrotoxicosis.

Lack of ranitidine effects on cyclophosphamide bone marrow toxicity or metabolism: a placebo-controlled clinical trial.

We previously reported that cimetidine but not ranitidine significantly enhances cyclophosphamide-induced bone marrow toxic effects and the appearance of cyclophosphamide alkylating species in a murine leukemia mouse model, and we advised caution in the use of cimetidine with microsomally metabolized anticancer drugs. Both drugs have been used for the treatment of gastric complications of chemotherapy. Using a randomized, double-blind, crossover study design, we have now evaluated the potential interaction of ranitidine with cyclophosphamide in seven cancer patients, who received two courses of cyclophosphamide, one with ranitidine and one with placebo. Four patients received ranitidine in the first course, and three received placebo. Ranitidine or placebo was started 3 days before a single dose of cyclophosphamide and given for 17 consecutive days. Ranitidine or placebo was given orally (300 mg/d), and cyclophosphamide (600 mg/m2) was given intravenously with [3H]cyclophosphamide (1000 muCi). Cyclophosphamide treatment was repeated at 4 weeks plus or minus 4 days. Blood samples were collected at intervals from 5 minutes to 24 hours after cyclophosphamide treatment and analyzed by thin-layer chromatography and radioassay for the drug and its metabolites. On days 0, 7, 14, and 21 after cyclophosphamide administration, complete blood cell counts, white blood cell differential counts, platelet counts, and SMA-17 were determined. The differences in mean nadir white blood cell counts, granulocyte counts, hemoglobin levels, and hematocrit values during ranitidine versus placebo treatment were not statistically significant. In a statistical but not a clinical sense, mean nadir platelet counts were significantly lower with ranitidine. There was a statistically significant increase in area under the curve for drug concentration in plasma x time (AUC) with ranitidine as well as a statistically significant decrease in the total-body clearance rate of the cyclophosphamide molecule. However, the effect on AUC for the major oncolytic metabolites 4-hydroxycyclophosphamide and phosphoramide mustard was not statistically significant. The lack of toxicologic or metabolic interaction between ranitidine and cyclophosphamide suggests that ranitidine can be used safely with cyclophosphamide.

Phase I trial of granulocyte-macrophage colony-stimulating factor plus high-dose cyclophosphamide given every 2 weeks: a Cancer and Leukemia Group B study.

BACKGROUND: Chemotherapy-induced myelosuppression often limits escalation of cancer chemotherapy doses. Cyclophosphamide, an alkylating agent, is an ideal candidate for dose escalation: A log-linear relationship between cell kill and dose has been demonstrated, and the drug spares hematopoietic stem cells. In addition, studies suggest that granulocyte-macrophage colony-stimulating factor (GM-CSF) can enhance the ability to achieve optimal dose intensity as well as ameliorating chemotherapy-induced myelosuppression. PURPOSE: The purpose of this study was to determine the maximum tolerated dose and the toxic effects of cyclophosphamide administered every 2 weeks with GM-CSF support. METHODS: For this trial by the Cancer and Leukemia Group B (CALGB), cohorts of patients were treated with cyclophosphamide as a 1-hour intravenous infusion every 14 days; GM-CSF was given subcutaneously on days 3-10. Four dose levels of cyclophosphamide (1.5, 3.0, 4.5, and 6.0 g/m2) and three dose levels of GM-CSF (2.5, 5.0, and 10.0 micrograms/kg per day) were evaluated. There was no dose escalation in individual patients. Fifty-one patients with solid tumors who had CALGB performance status 0 or 1 and minimal prior radiotherapy were eligible for analysis. Drug clearance and area under the curve for plasma drug concentration x time (AUC) were estimated at completion of the infusion and at 4 and 24 hours after the start of the infusion. RESULTS: Ninety-five courses of therapy were analyzed. Treatment with cyclophosphamide at 3.0 g/m2 or more resulted in neutropenia (absolute neutrophil counts < 100/microL) in all cycles of therapy. At those doses, blood cell count recovery adequate for re-treatment occurred in 67%-85% of cycles (median, 16 days). Doses of 6.0 g/m2 were associated with the greatest degree of myelosuppression and frequent hospitalization (88% of cycles); requirements for blood transfusion prohibited further dose escalation. Nonhematologic toxic effects were tolerable, with two episodes of reversible cardiotoxicity and four episodes of hemorrhagic cystitis that precluded further therapy. Degree of myelosuppression was not correlated with cyclophosphamide AUC or clearance. CONCLUSIONS: The recommended phase II dose of cyclophosphamide is 4.5 g/m2 administered every 2 weeks with GM-CSF given at 5.0 micrograms/kg per day of GM-CSF. Our results suggest that, with GM-CSF support, high cumulative doses of cyclophosphamide can be given to achieve optimal dose intensity, with reproducible blood cell count recovery and without the need for autologous bone marrow transplantation. IMPLICATIONS: Phase II studies of this intensive regimen in malignant diseases sensitive to alkylating agents are currently being done in CALGB.

Effects of recombinant human granulocyte-macrophage colony-stimulating factor on myelosuppression induced by multiple cycles of high-dose chemotherapy in patients with advanced breast cancer.

In this study, 18 patients with advanced breast cancer were treated with multiple cycles of doxorubicin (75 or 90 mg/m2) plus cyclophosphamide (750 or 1000 mg/m2) every 21 days. Granulocyte-macrophage colony-stimulating factor (GM-CSF) (250 micrograms/m2 per day) was administered by continuous infusion during 10 days (days 2-12), starting in the first or second cycle of chemotherapy. Sixteen (89%) of 18 patients (95% confidence interval, 65%-99%) achieved an objective remission, five (28%) of which were complete. The median duration of response was 7 months. When GM-CSF was used for the first time, it had an effect on the kinetics of all blood cells, including neutrophils, lymphocytes, thrombocytes, and reticulocytes. However, in subsequent cycles of chemotherapy, the stimulatory effect of GM-CSF on hematopoiesis was substantially diminished. World Health Organization grade 3 and 4 neutropenia and thrombocytopenia necessitated dose reductions of doxorubicin and cyclophosphamide from cycle 2 onward in all patients treated with the highest dose. Side effects of GM-CSF included fever, general weakness, and hypotension. These toxic effects mimicked sepsis, and hospital admission for treatment with intravenous antibiotics was required for 73 days in 61 cycles of chemotherapy that included GM-CSF. Dose-intensive chemotherapy produced a high response rate in patients with advanced breast cancer. However, GM-CSF administered from day 2 to day 12 at a dose of 250 micrograms/m2.

Biologic activity of tamoxifen at low doses in healthy women.

BACKGROUND: Results of a clinical trial recently completed in the United States indicate that administration of tamoxifen (20 mg/day) to women at risk can reduce breast cancer incidence by approximately 50% but is associated with an increased risk of developing endometrial cancer and venous thromboembolic events. Since these adverse effects may be dose related, we investigated the effect of tamoxifen on several biomarkers when the drug was given at doses lower than those currently in use. METHODS: In two sequential experiments, 127 healthy hysterectomized women aged 35-70 years were randomly assigned to one of the following four treatment arms: placebo (n = 31) or tamoxifen at 20 mg/day (n = 30) (first experiment); or tamoxifen at 10 mg/day (n = 34) or tamoxifen at 10 mg/ alternate days (n = 32) (second experiment). Baseline and 2-month measurements of the following parameters were compared: 1) total cholesterol (primary end point) and other surrogate markers of cardiovascular disease, e.g., low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, triglycerides, and lipoprotein(a); 2) blood cell count; 3) fibrinogen; 4) antithrombin III; 5) osteocalcin; and, 6) in a subgroup of 103 women, insulin-like growth factor-I (IGF-I), a possible surrogate marker for breast cancer. RESULTS: After adjustment for the baseline values, there were reductions in circulating levels of total cholesterol and IGF-I of the same magnitude in all three tamoxifen treatment arms. A similar pattern was observed for most of the other parameters. In the placebo arm, fibrinogen level, which showed a decrease, was the only parameter exhibiting change. CONCLUSIONS: Up to a 75% reduction in the conventional dose of tamoxifen (i.e., 20 mg/day) does not affect the activity of the drug on a large number of biomarkers, most of which are surrogate markers of cardiovascular disease. This study was hypothesis generating, and larger studies are warranted to assess the efficacy of tamoxifen at low doses.

Phase I/pharmacokinetic study of topotecan by 24-hour continuous infusion weekly.

Topotecan (SK&F 104864, hycamptamine, NSC 609699) is believed to exert its cytotoxic effects through inhibition of topoisomerase I, the activity of which recovers rapidly on removal of the drug in vitro. In vivo studies show that the activity of topotecan is schedule dependent, favoring repeated doses. Early human studies showed that topotecan (the active lactone) had a short half-life in plasma. To prolong drug exposure, we administered topotecan as a 24-h i.v. infusion and repeated it weekly. We treated 32 patients with doses of 1.0-2.0 mg/m2. Median performance status was 1, and all but four patients had received prior chemotherapy. Dose-limiting neutropenia occurred at doses > or = 1.75 mg/m2; nadirs were observed after 1-3 doses. The recommended phase II dose is 1.5 mg/m2/week. One patient with metastatic colon cancer had a partial response. Both plasma topotecan (lactone) and total topotecan (measured by converting the hydroxyacid form to the lactone by acidification of the sample) were measured by high-performance liquid chromatography in 21 patients. During infusion, mean topotecan plasma steady-state concentrations ranged from 4.7-11.4 nM. Plasma elimination was best fit to a one-compartment model with a mean t1/2 of 3.5 h. The mean total body clearance was 388 ml/min/m2. Concentrations of the inactive form approximated those of the lactone throughout. No evidence for dose-dependent pharmacokinetics was observed in this dose range. Pharmacodynamic analysis, using the sigmoid Emax model, revealed that the pharmacokinetic parameters of both lactone and total drug were positively correlated with bone marrow toxicity. Total drug steady-state plasma concentration provided a good estimate of neutropenia, suggesting a simple, easily monitored, pharmacokinetic parameter for adaptive dosing using this schedule. Phase II evaluation of this weekly schedule is indicated in solid tumors.

Pharmacokinetics of recombinant human interleukin 3 administered subcutaneously and by continuous intravenous infusion in patients after chemotherapy for ovarian cancer.

Twenty chemotherapy-naive patients with ovarian carcinoma received 1, 5, 10, or 15 micrograms/kg/day (five patients per dose step) of recombinant human interleukin 3 (rhIL-3) over 7 days after carboplatin/cyclophosphamide in Cycles 1 and 3. Patients received rhIL-3 by continuous i.v. infusion or once daily s.c. injection in Cycle 1 and the alternate route in Cycle 3. Plasma rhIL-3 samples were obtained once daily on Days 1 to 6 and serially over a 24-h period on Day 7 for pharmacokinetic assessment of s.c. and i.v. administered rhIL-3 in 16 and 17 patients, respectively. Concentrations were assayed by a time-resolved fluorescence sandwich immunoassay. Pharmacokinetic parameters were derived by noncompartmental methods. Mean steady-state concentrations during continuous i.v. infusion ranged from 117 pg/ml (1 microgram/kg/day) to 2217 pg/ml (15 micrograms/kg/day) and were linearly related to dose (r = 0.87, P < 0.001). When dose normalized, the mean steady-state concentrations were comparable at all doses. The total-body clearance was approximately 4 to 5 ml/min/kg. Elimination half-life (t1/2 i.v.) could be assessed for the 5- to 15-micrograms/kg/day dose levels and was 53, 41, and 26 min for the 5-, 10-, and 15-micrograms dose levels, respectively (not significant between dose levels). Following s.c. injection, the maximum rhIL-3 plasma concentration ranged from 206 pg/ml (1 microgram/kg/day) to 6930 pg/ml (15 micrograms/kg/day). Both the maximum measured plasma concentration (r = 0.89, P < 0.0001) and the area under the plasma concentration/time curve (r = 0.93, P < 0.0001) were related to dose. Dose-normalized values were comparable over the entire dose range. Elimination t1/2s.c. was 4.8 h at the 1-microgram dose level and roughly half this time for the 5- to 15-micrograms/kg/day dose levels. The systemic clearance of approximately 5 to 6 ml/min/kg was comparable at all dose levels. Based on trough levels of the 7-day s.c. course, no rhIL-3 accumulation occurred. Bioavailability of s.c. administered rhIL-3 was nearly 100%. No correlation between creatinine clearance and pharmacokinetic parameters of rhIL-3 could be demonstrated. Since there was also no difference in hematological efficacy between the two routes of rhIL-3 administration, we conclude that the s.c. route of administration appears to have no disadvantages over the i.v. route and may facilitate its clinical application.

Phase I and immunomodulatory study of a muramyl peptide, muramyl tripeptide phosphatidylethanolamine.

Muramyl tripeptide phosphatidylethanolamine (MTP-PE; CGP 19835A from Ciba Geigy) is a synthetic muramyl tripeptide structurally related to bacterial cell wall constituents. MTP-PE activates monocytes in vitro to a tumoricidal state and has in vivo antitumor effects in animal models. We studied the toxicity and immunomodulatory effects of once weekly i.v. administration of liposomal-encapsulated MTP-PE for 8 weeks in 27 patients with advanced malignancies. Doses ranged from 0.1 to 2.7 mg/m2. No major tumor responses were seen; 11 patients had stable disease after 8 weeks of therapy and 3 continued on maintenance therapy because of minor tumor regressions and/or clinical improvement. MTP-PE at these doses was well tolerated. Shaking chills and fevers were the most common toxicities and occurred at all dose levels. There was no treatment-induced loss of performance status. Immunomodulatory studies revealed evidence of a biological effect on monocytes. C-reactive protein levels rose in the majority of patients with end-of-treatment values 2 to 10 times higher than baseline. Serum neopterin levels were consistently increased 24 h after MTP-PE administration and significant decreases in expression of two different types of Fc receptors on peripheral blood monocytes were noted 6 h after treatment. Although no major tumor responses were seen in this group of patients with advanced malignancies, MTP-PE was well tolerated and exerted biological effects on monocytes. Serum neopterin levels may be a useful marker for the biological effects of MTP-PE.

Antitumor activity and toxicity in animals of BMY-25282, a new mitomycin derivative.

BMY-25282, 7-N-(dimethylamino methylene)mitomycin C, is one of a novel series of amidino mitomycin derivatives. Some of these were discovered as intermediates in a synthetic program being conducted to find improved procedures for modifying the structure of mitomycin C (MMC). Markedly superior in vivo antitumor effects have been observed with BMY-25282 compared to MMC in initial tests against i.p.-implanted P388 leukemia and B16 melanoma. When administered i.v. to mice bearing s.c. B16 melanoma, BMY-25282 was also superior to MMC. The derivative was fully active against a line of L1210 leukemia which was partially resistant to MMC treatment but had little or no activity against a line of L1210 fully resistant to MMC. It is also 2 to 4 times more potent than MMC based on a comparison of doses required to achieve optimum antitumor effects. The superior antitumor efficacy of BMY-25282 over MMC against both i.p. and s.c. B16 melanoma was maintained when the drug was given in pluronic acid formulation. Against P-388 leukemia, however, the efficacy of the drugs was equivalent when BMY-25282 was administered in the pluronic vehicle. In an in vitro clonogenic assay involving freshly explanted human tumors, BMY-25282 was consistently more potent in cytotoxic effects than MMC. With human colorectal carcinoma samples, BMY-25282 was 13.8 times more potent than MMC. The i.v. 50% lethal dose values of BMY-25282 and MMC in C57BL/6 X DBA/2 F1 mice were 2.1 mg/kg and 8.6 mg/kg, respectively. Leukopenic effects of the drugs in mice were comparable at doses up to their respective 50% lethal dose values. Hematology studies in ferrets revealed a similar pattern of depression and recovery of lymphocytes, neutrophils, and platelets for BMY-25282 and MMC; however, with BMY-25282 there was earlier recovery of platelet counts. BMY-25282 is being further developed toward possible clinical trial.

Protective effect of ICRF-187 against normal tissue injury induced by adriamycin in combination with whole body hyperthermia.

The use of [(+)-1,2-bis(3,5-dioxopiperazinyl-1-yl)]propane (ICRF-187) as a protective agent against normal tissue toxicity caused by combined Adriamycin (ADR) and whole body hyperthermia (WBH; 2 h at 41.5 degrees C) was assessed in a rat model. The effect of ICRF-187 on the antitumor response induced by the combination of ADR and WBH was also investigated in order to assess alterations in the therapeutic index of this combined therapeutic modality treatment. ICRF-187 significantly reduced ADR-mediated body weight loss, renal toxicity, and cardiomyopathy under both normothermic and hyperthermic conditions as shown by morphological and functional assays. ADR-induced neuropathy (seen only in normothermic rats) was also ameliorated by ICRF-187. Although this study did not show a pronounced effect of ICRF-187 on ADR-induced acute myelosuppression, ADR-mediated chronic anemia, leukocytosis, and thrombocytosis were reduced by ICRF-187 in both normothermic and WBH-treated rats. The effect of ICRF-187 on antitumor response was evaluated with a tumor growth delay assay using an in vivo transplantable fibrosarcoma. ICRF-187 caused no significant change in tumor growth delay induced by either ADR alone or ADR combined with WBH. Indeed, the only complete tumor regression following treatment resulted from the combination of ICRF-187 plus ADR plus WBH. Thus, ICRF-187 significantly increases the therapeutic index of the combined modality treatment of ADR and WBH by selectively reducing normal tissue toxicity without interfering with antitumor efficacy.

Combined interleukin 1 beta/interleukin 2 treatment in mice: synergistic myelostimulatory activity and protection against cyclophosphamide-induced myelosuppression.

We have studied the effects of single and combined treatment with interleukin 1 beta (IL-1 beta) and interleukin 2 (IL-2) on spleen and bone marrow hematopoiesis in normal mice. Injection of IL-1 beta alone was followed by a significant increase in the number of granulocytes in spleen and progenitors (burst-forming units-erythroid and colony-forming units-granulomonocytic) in both spleen and bone marrow, s compared to control mice. In contrast, IL-2 alone induced only a slight increase in the number of marrow colony-forming units-granulomonocytic and had no significant effect on spleen progenitors. Repeated injections of both IL-1 beta and IL-2 resulted in a synergistic increase in spleen weight and splenocyte number, as compared to mice treated with the single cytokine regimen; in particular, the combined treatment induced a marked rise in the number of neutrophilic granulocytes and erythroblasts, whereas splenic lymphocytes were not affected. This regimen also caused a synergistic increase in the number of spleen and marrow progenitor cells: a time-course analysis showed an elevation in numbers of both burst-forming units-erythroid and colony-forming units-granulomonocytic, first in marrow (day 10) and subsequently in spleen (day 18). Combined IL-1 beta/IL-2 treatment dampened the decrease and accelerated the recovery of myeloid cells after cyclophosphamide injection, whereas the single cytokine regimen was not effective. Similarly, the rebound of WBC (especially neutrophilic granulocytes) after cyclophosphamide treatment was markedly enhanced by the combined treatment, whereas the single cytokine regimen was ineffective. These results, indicating a myelostimulatory effect by the combined cytokine regimen, together with our previous observations showing a synergistic antitumor activity by IL-1/IL-2 treatment in experimental mouse tumors (V. Ciolli et al., J. Exp. Med., 173: 313-322, 1991), may provide the basis for the development of new combination therapies with cytokines and antiblastic agents in the treatment of cancer patients.

Biochemically directed therapy of leukemia with tiazofurin, a selective blocker of inosine 5'-phosphate dehydrogenase activity.

Tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide, NSC 286193), a selective inhibitor of the activity of IMP dehydrogenase (EC 1.1.1.205), the rate-limiting enzyme of de novo GTP biosynthesis, provided in end stage leukemic patients a rapid decrease of IMP dehydrogenase activity and GTP concentration in the blast cells and a subsequent decline in blast cell count. Sixteen consecutive patients with end stage acute nonlymphocytic leukemia or myeloid blast crisis of chronic granulocytic leukemia were treated with tiazofurin. Allopurinol was also given to inhibit xanthine oxidase activity to decrease uric acid excretion and to elevate the serum concentration of hypoxanthine, which should competitively inhibit the activity of hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8), the salvage enzyme of guanylate synthesis. Assays of IMP dehydrogenase activity and GTP concentration in leukemic cells provided a method to monitor the impact of tiazofurin and allopurinol and to adjust the drug doses. In this group of patients with poor prognosis, five attained a complete hematological remission and one showed a hematological improvement. A marked antileukemic effect was seen in two other patients. All five evaluable patients with myeloid blast crisis of chronic granulocytic leukemia reentered the chronic phase of their disease. Five patients with acute nonlymphocytic leukemia were refractory to tiazofurin and three were unevaluable for hematological effect because of early severe complications. Responses with intermittent 5- to 15-day courses of tiazofurin lasted 3-10 months. Tiazofurin had a clear antiproliferative effect, but the pattern of hematological response indicated that it appeared to induce differentiation of leukemic cells. In spite of toxicity with severe or life-threatening complications in 11 of 16 patients, tiazofurin was better tolerated in most patients than other antileukemic treatment modalities and provided a rational, biochemically targeted, and biochemically monitored chemotherapy which should be of interest in the treatment of leukemias and as a paradigm in enzyme pattern-targeted chemotherapy.

Effects of high-dose methotrexate and leucovorin on murine hemopoietic stem cells.

This study compares the effects of a high dose of methotrexate (HDMTX) to that of a high dose of methotrexate plus leucovorin protection on the hemopoietic stem cells in a murine model. C57BL X C3H F1 mice were treated with a single large bolus (500 mg/kg body weight) of methotrexate or with the same dose of the drug plus leucovorin administered in fractionated doses during the following 24 hr. At 1 to 2 days after the administration of HDMTX, there was a large bone marrow and spleen depopulation of pluripotent stem cells and of committed and recognizable progenitors. At 2 to 3 days, a severe fall of white blood cells and reticulocytes ensued. The recovery process of hemopoietic precursors followed alternate phases of overshooting and secondary falls. Leucovorin administration appeared to protect all stages of hemopoiesis and prevented the severe drops of bone marrow cellularity and stem cell content which followed the HDMTX bolus. However, the effect of leucovorin on peripheral blood cell reduction was less significant. After treatment with HDMTX, the recovery of bone marrow cells and the burst of splenic hemopoietic activity followed a pattern similar in both leucovorin-protected and unprotected animals, but in the former, the increase in stem cells and hemopoietic progenitors appeared to reach higher values and to last longer. In particular, the overshooting of colony-forming units, culture and erythroid, reached a higher peak in leucovorin-treated mice and was more prolonged. Our results indicate that, in HDMTX-treated mice, leucovorin protection involves the earliest stages of hemopoiesis, assuring the maintenance of a satisfactory endowment of stem and progenitor cells.

Interleukin 6 perfusion stimulates reconstitution of the immune and hematopoietic systems after 5-fluorouracil treatment.

Effects of interleukin 6 (IL-6) on the functional capacity of the immune and hematopoietic systems in 5-fluorouracil (5-FU)-treated mice were determined. IL-6 (5 x 10(4) units/mouse/day) was administered s.c. for 7 days by implantation of an osmotic pump, since it was demonstrated that a much higher increase in the primary response to sheep RBC was observed by administration of slowly released rather than daily s.c. injection of IL-6. IL-6 perfusion significantly augmented anti-sheep RBC antibody responses depressed by 5-FU (150 mg/kg) treatment. IL-6 also was shown to stimulate hematological recovery in mice treated with 5-FU. Namely, IL-6 perfusion accelerated the recovery of the number of hematopoietic stem cells, granulocyte-macrophage progenitors, and mature neutrophils in the spleen, although IL-6 did not stimulate the recovery of the neutrophil count in blood. Recovery of the platelet count in blood was stimulated by IL-6. Furthermore, it was found that the endogenous IL-6 level in serum increased after 5-FU treatment, which suggests that IL-6 may play some role in the recovery of the immune and hematopoietic systems. Finally, we examined the effect of IL-6 on the survival of mice treated with a higher dosage of 5-FU (300 mg/kg). IL-6 perfusion produced a distinct increase in survival rate at Day 30 (74% versus 28%). It is of note that the number of bacteria (identified as Escherichia coli) cultured from the spleen and the liver decreased in IL-6-perfused mice. This IL-6-induced effect was accompanied by enhancement of an oxidative burst response. Moreover, the anti-E. coli antibody titer in serum was higher in IL-6-perfused mice than in control mice. These results suggest the possible use of IL-6 for stimulating the reconstitution of the immune and hematopoietic systems after chemotherapy treatment.

Role of thromboxane in interleukin 2-induced lung injury in sheep.

Interleukin (IL)-2 administration leads to respiratory dysfunction due to increased vascular permeability. This study examines the role of thromboxane (Tx)A2 in IL-2 induced lung injury in sheep with chronic lung lymph fistulae. This preparation enables evaluation of permeability prior to the development of gross edema. IL-2, 10(5) units/kg (n = 6), or its excipient control (n = 5) was given as an i.v. bolus over 2 min. After 2 h of IL-2 administration, plasma TxB2 increased from 168 to 388 pg/ml (P less than 0.05) and lung lymph TxB2 from 235 to 694 pg/ml (P less than 0.05). Mean pulmonary artery pressure (MPAP) rose from 13 to 29 mm of Hg (P less than 0.05) at 30 min and remained elevated for 4 h while the pulmonary artery wedge pressure was unchanged at 4 mm of Hg. Arterial oxygen tension (PaO2) fell from 88 to 77 mm of Hg (P less than 0.05). Lung lymph flow (QL) rose from 2.2 to 3.8 ml/30 min (P less than 0.05) at 1 h and to 6.4 ml/30 min at 3 h. This rise coincided with an increase in the lymph/plasma (L/P) protein ratio from 0.67 to 0.77 (P less than 0.05). In contrast, the non-IL-2-infused sheep (n = 3) recruitment of the lung vasculature by left atrial balloon inflation led to a rise in QL from 2.4 to 8.2 ml/30 min, whereas the L/P ratio declined from 0.62 to 0.25, suggesting that the protein-rich lymph flow after IL-2 administration reflected increased microvascular permeability. In further proof of an increase in permeability, IL-2 administration into sheep (n = 2) with an inflated left atrial balloon led, after a pressure-independent L/P protein ratio had been achieved, to an increase in L/P protein ratio and decrease in protein reflection coefficient. At 2 h after IL-2, the blood leukocyte count fell from 8156 to 4375/mm3 (P less than 0.05) primarily due to a 73% drop in lymphocytes. The platelet count declined from 292 to 184 x 10(3)/mm3 (P less than 0.05). Body temperature rose from 38.9-40.3 degrees C (P less than 0.05), and shaking chills were common. Pretreatment with the Tx synthetase inhibitor OKY 046 (n = 7) lowered baseline plasma and lymph TxB2 levels to 22 and 52 pg/ml (P less than 0.05) and prevented the IL-2-induced increase in plasma and lung lymph TxB2 (P less than 0.05).(ABSTRACT TRUNCATED AT 400 WORDS)

Chronic daily administration of oral etoposide--a phase I trial.

In this phase I study, we administered etoposide (VP-16) orally for 21 consecutive days to patients with advanced refractory cancers. All patients had received previous chemotherapy, and 50% of patients had received more than one combination regimen. When given for 21 consecutive days, the maximum-tolerated dose of oral VP-16 was 50 mg/m2/d. Myelosuppression was the dose-limiting toxicity, and occurred between days 21 and 28. In most patients, blood counts had recovered sufficiently by day 35 to begin another 3-week course. WBC count nadirs of less than 1,000/microL occurred in four of 20 courses at this dose, and three patients required hospitalization for treatment of neutropenia and fever. Alopecia occurred in most patients; gastrointestinal (GI) and other toxicities were uncommon. Five of 16 patients with measurable tumor had partial responses of 3 to 4 months duration. Four of these five patients had malignancies that are usually unresponsive to VP-16 when administered by previously investigated schedules. This method of VP-16 administration is well tolerated, convenient, and may optimize antitumor efficacy by exploiting the schedule dependency of this drug. Phase II studies are necessary to define the level of activity of this schedule of VP-16.

Plasma pharmacokinetics and pharmacodynamics of a new prodrug N-l-leucyldoxorubicin and its metabolites in a phase I clinical trial.

PURPOSE: N-l-leucyldoxorubicin (Leu-Dox) was developed as a prodrug of doxorubicin (Dox) to circumvent the cardiotoxicity associated with repeated administration of Dox. Our purpose was to assess the pharmacokinetics of Leu-Dox, Dox, doxorubicinol (Dol) and four other metabolites for pharmacokinetically guided dose-escalation and to verify the prodrug character of Leu-Dox. PATIENTS AND METHODS: Blood and urine of 14 patients were sampled during the phase I clinical trial and analyzed by high-performance liquid chromatography. Dose levels of Leu-Dox ranged from 18 mg/m2 to 225 mg/m2, the maximum-tolerated dose (MTD). Hematologic parameters were monitored regularly in each patient. RESULTS: Leu-Dox was rapidly distributed (half-life at alpha phase [t1/2 alpha] = 2.5 +/- 0.6 minutes) followed by a biphasic elimination (half-life at beta phase [t1/2 beta] = 17.4 +/- 7.3 minutes; half-life at gamma phase [t1/2 gamma] = 1.5 +/- 0.5 hours), as measured over the first 12 hours after administration. In three patients, in whom Leu-Dox was found in the plasma for up to 48 hours after injection, a final elimination half-life (t1/2,elim) of 16 hours was observed. The t1/2,elim of Leu-Dox was short (0.6 to 16.5 hours) compared with the t1/2,elim of Dox (38 +/- 11 hours). The mean residence time and apparent volume of distribution were 23 +/- 5 minutes and 19 +/- 6 L/m2, respectively. Only 1.5% to 5% of the dose was excreted in the urine over 48 hours, with Dox as major constituent. Dox was rapidly formed, reaching its maximum concentration within 10 minutes after the end of Leu-Dox infusion. Areas under the plasma concentration versus time curve (AUC infinity, mean +/- SD, n = 16) of Leu-Dox, Dox, and Dol were 115 +/- 27 mumol.min/L, 41 +/- 12 mumol.min/L, and 33 +/- 14 mumol.min/L after a dose of 60 mg/m2 Leu-Dox (= 86 mumol/m2). After the same molar dose of Dox (50 mg/m2 = 86 mumol/m2), the AUC infinity of Dox was 179 mumol.min/L, indicating that Leu-Dox was converted into Dox for 23% in the plasma compartment. The AUCs infinity of Leu-Dox, Dox, and Dol increased linearly with the dose. Negligible AUCs were observed for the other four metabolites. The AUCs infinity of Leu-Dox and Dox at the MTD (517 and 145 mumol.min/L, respectively) were lower than those in mice at the LD10 (1,930 and 798 mumol.min/L, respectively), which means that the MTD could not be predicted from the preclinical pharmacokinetics in mice. Hematologic toxicity, especially the WBC count, appeared to correlate much better with the AUC of Dox (r = .91) than with the AUC of Leu-Dox (r = .74), thus confirming the prodrug character of Leu-Dox. CONCLUSIONS: Dox is rapidly formed from Leu-Dox, and seems causative in the observed myelotoxicity. The MTD could not be predicted from the AUC at the LD10 in mice.

Phase I study of paclitaxel in combination with cyclophosphamide and granulocyte colony-stimulating factor in metastatic breast cancer patients.

PURPOSE: In vitro data suggest that prolonged exposure to paclitaxel enhances breast cancer cytotoxicity. Our objective in this phase I study was to determine the tolerability of paclitaxel administered by 72-hour continuous intravenous (i.v.) infusion (CIVI) in combination with high-dose cyclophosphamide and granulocyte colony-stimulating factor (G-CSF) in the ambulatory setting to metastatic breast cancer patients. PATIENTS AND METHODS: Paclitaxel was administered over 72 hours by CIVI and cyclophosphamide was given daily by i.v. bolus on days 1, 2, and 3, followed by G-CSF every 21 days. The availability of ambulatory infusion pumps and paclitaxel-compatible tubing permitted outpatient administration. RESULTS: Fifty-five patients with metastatic breast cancer who had been previously treated with a median of two prior chemotherapy regimens were entered onto the study. Dose-limiting toxicity of grade 4 neutropenia for longer than 5 days and grade 4 thrombocytopenia occurred in three of five patients treated with paclitaxel 160 mg/m2 CIVI and cyclophosphamide 3,300 mg/m2 followed by G-CSF. The maximum-tolerated dose (MTD) was paclitaxel 160 mg/m2 CIVI and cyclophosphamide 2,700 mg/m2 in divided doses with G-CSF. Nonhematologic toxicities were moderate and included diarrhea, mucositis, and arthalgias. Although hemorrhagic cystitis developed in six patients, recurrence was prevented with i.v. and oral mesna, which permitted continued outpatient delivery. One hundred seventy-four cycles were safely administered in the ambulatory setting using infusional pumps and tubing. Objective responses occurred in 23 (one complete and 22 partial) of 42 patients with bidimensionally measurable disease (55%; 95% confidence interval, 38% to 70%), with a response rate of 73% (11 of 15) seen at the highest dose levels. CONCLUSION: Paclitaxel by 72-hour CIVI with daily cyclophosphamide followed by G-CSF can be administered safely in the ambulatory setting, has acceptable toxicity, and is an active regimen in the treatment of metastatic breast cancer.

Phase I trial of subcutaneous interleukin-6 in patients with advanced malignancies.

PURPOSE: Based on preclinical evidence in murine models that interleukin-6 (IL-6) mediates regression of metastatic tumors, we performed a phase I study of recombinant human IL-6 in patients with refractory advanced malignancies to determine its pharmacokinetics, toxicities, and possible immunologic and antitumor effects. PATIENTS AND METHODS: Recombinant IL-6 was administered as a single subcutaneous dose daily for 7 days, with 7 days off therapy followed by another 7 days of IL-6. Doses were escalated in cohorts of three patients starting at 3 micrograms/kg/d, provided that toxicity at the preceding dose level was not dose-limiting. Dose-limiting toxicity was defined as grade III or IV major organ toxicity that did not resolve to grade II or less in 24 hours after stopping IL-6, using the National Cancer Institute Common Toxicity Criteria. Patients were treated with 3, 10, and 30 micrograms/kg/d IL-6 subcutaneously. RESULTS: Three patients each were treated at the 3- and 10-micrograms dose levels. Two of five patients treated with 30 micrograms/kg/d IL-6 subcutaneously had grade III major organ toxicity that required IL-6 therapy to be discontinued. All patients experienced fever, chills, and minor fatigue. Significant increases in C-reactive protein (CRP), fibrinogen, platelet counts, and lymphocyte IL-2 receptor levels were seen in patients at the 10- and 30-micrograms/kg dose levels. Decreases in albumin and hemoglobin were observed, particularly at the 30-micrograms/kg dose level. The half-life (T1/2 beta) was 4.2 hours, with a peak IL-6 level at 5 hours. No antitumor responses were seen. CONCLUSION: A safely tolerated dose of daily subcutaneous IL-6 is 10 micrograms/kg, with hepatotoxicity and cardiac arrhythmia being the dose-limiting toxicities at 30 micrograms/kg. Phase II trials of IL-6 administered subcutaneously daily for at least 7 days for two cycles with an intervening week of rest are recommended for phase II trials. However, patients with extensive replacement of liver by tumor and abnormal liver functions should receive IL-6 therapy with caution.

Peripheral-blood stem-cell collections after paclitaxel, cyclophosphamide, and recombinant human granulocyte colony-stimulating factor in patients with breast and ovarian cancer.

PURPOSE: Here we evaluate Taxol (paclitaxel; Bristol-Myers Squibb, Princeton, NJ) and cyclophosphamide (CY) with recombinant human granulocyte colony-stimulating factor (rhG-CSF) for mobilization of peripheral-blood stem cells (PBSCs) for autologous stem-cell transplantation in patients with breast and ovarian cancer. PATIENTS AND METHODS: PBSCs were collected from 17 patients with breast (n = 11), ovarian (n = 5), and gastric (n = 1) cancer after administration of Taxol (170 mg/m2 x 1) and CY (4 g/m2 x 1) followed by rhG-CSF (10 micrograms/kg/d). PBSC collections after Taxol and CY were compared with PBSC collections from nine patients with stage IV breast (n = 8) or stage III ovarian (n = 1) cancer who had received CY (4 g/m2 x 1) followed by rhG-CSF (16 micrograms/kg/d) for mobilization. RESULTS: Mean WBC and platelet counts on the first day of apheresis were 6.3 x 10(9)/L (range, 1.9 to 22.1) and 35 x 10(9)/L (range, 19 to 77), respectively. The median numbers of CD34+ cells, peripheral-blood mononuclear cells (PBMNC), and peripheral-blood total nucleated cells (PBTNC) collected were 13.02 x 10(6)/kg (range, 5.4 to 57.8; mean, 16.02), 6.86 x 10(8)/kg (range, 1.9 to 51.2), and 17.41 x 10(8)/kg (range, 2.4 to 106.6), respectively. In the comparison group, the median yield of CD34+ cells was 6.39 x 10(6)/kg (range, 0.2 to 28; mean, 10.01; P = .01). The mean daily yield of CD34+ cells/kg/collection was 3.5 (range, 0.8 to 28.9) after Taxol and CY, as compared with 1.3 (range, 0.1 to 7.0) for patients who received CY alone (P = .01). All patients who received CY and Taxol reached a target level of 5 x 10(6) CD34+ cells/kg, as compared with five of nine patients (55.5%) who received CY alone (P = .03). CONCLUSION: These data suggest that Taxol and CY followed by rhG-CSF mobilizes PBSCs in patients with advanced breast and ovarian cancer more effectively than this regimen without Taxol.