Patrick J. Creaven

Matrix metalloproteinase inhibitors

Matrix metalloproteinases (MMPs) are a family of zincdependent proteinases that are associated with the tumorigenic process. MMPs degrade the extracellular matrix, promoting tumor invasion and metastasis. They also regulate host defense mechanisms and normal cell function; blocking all MMPs may not lead to a positive therapeutic outcome. Most clinical trials of MMP inhibitors (MMPIs) have yielded disappointing results, perhaps due to inappropriate study design or tumor staging, or to lack of selectivity. Positive results have been seen in gastric cancer with marimastat and in Kaposi’s sarcoma with metastat. This review summarizes the current status of MMPIs.

Cytotoxicity, cellular accumulation and DNA binding of oxaliplatin isomers

Oxaliplatin (trans-l-1,2-diaminocyclohexane oxalato Pt(II); 1R,2R-dach, l-OHP), its trans-d isomer (1S,2S-dach) and cis-dach (1R,2S-dach) isomers were compared in in vitro testing against human ovarian carcinoma cell lines A2780, A2780/CP (cisplatin resistant), A2780/l-OHP (oxaliplatin resistant), colon carcinoma cell line HT-29, and murine leukemia cell lines L1210, L1210/CP (cisplatin resistant), and L1210/dach (tetraplatin resistant). The relative molar potency of the three complexes in all the cell lines except A2780/l-OHP and L1210/dach are trans-l > trans-d > cis-dach; in A2780/l-OHP they are trans-l = trans-d > cis-dach; in L1210/dach trans-l > trans-d = cis-dach. The A2780/l-OHP selected for trans-l resistance is 3.6-fold resistant to oxaliplatin, showed no resistance to trans-d isomer and is 6-fold resistant to cis-dach. However, L1210/dach which is selected for carboxyphthalato 1,2-dach (trans-dl) platinum(II) is 140-fold resistant to oxaliplatin, 73-fold resistant to trans-d, and 41-fold resistant to cis-OHP. The accumulation and DNA binding of platinum following a 2-h treatment of A2780 cells with each of the isomers (60 microM) is in the order of trans-l > cis-dach > trans-d which corresponded to the cytotoxicity of trans-l, but not the others. The data suggest that other processes, such as differential formation of specific adducts and/or repair may be involved. Of the three isomers l-OHP is the superior and its accumulation and DNA binding are consistent with its cytotoxicity.

A phase I and pharmacokinetic study of oral uracil, ftorafur, and leucovorin in patients with advanced cancer

Abstract A phase I and pharmacokinetic study of oral uracil, ftorafur, and leucovorin was performed in patients with advanced cancer. Uracil plus ftorafur (UFT) was given in a 4:1 molar ratio in three divided doses for 28 consecutive days. Patient cohorts were treated at 200, 250, 300, and 350 mg/m2 of UFT daily. For all patients, 150 mg of leucovorin was given daily in three oral doses. A 1-week rest period followed each 28-day treatment course. Gastrointestinal toxicity, characterized by diarrhea, nausea, and vomiting, was dose-limiting at 350 mg/m2 UFT in patients who had received prior chemotherapy. Mild fatigue and transient hyperbilirubinemia were also common. In previously untreated patients, UFT at 350 mg/m2 was well-tolerated, suggesting this as an acceptable phase II dose in this schedule with leucovorin. Two of eight previously untreated patients with advanced colorectal cancer had partial responses with UFT (350 mg/m2) plus leucovorin. Pharmacokinetic parameters [ftorafur, uracil, 5-fluorouracil (5-FU), 5-methyltetrahydrofolate] showed wide interpatient variations. Plasma levels of 5-FU (Cmax 1.4±1.9 μM) were comparable to those achieved with protracted venous infusions, and folate levels (Cmax 6.1±3.6 μM) were sufficient for biochemical modulation. Ongoing study will determine if this convenient oral regimen will compare favorably in terms of efficacy, toxicity, and cost with intravenous fluoropyrimidine programs.

Altered glutathione metabolism in oxaliplatin resistant ovarian carcinoma cells

Elevation of glutathione (GSH) is commonly observed in cellular resistance to a number of anticancer agents. Most frequently reported change in GSH metabolism that is associated with the elevated GSH levels is increased mRNA expression and activity of gamma-glutamyl cysteine synthetase (gamma GCS), the first enzyme of the GSH biosynthetic pathway. We have isolated sublines of the A2780 ovarian carcinoma cell line (C10 and C25) that are 8- and 12-fold resistant to oxaliplatin by repeatedly exposing the cells to increasing concentrations of the platinum agent. The GSH levels in C10 and C25 cell sublines are 3.1- and 3.8-fold higher than the parent A2780 cell line. The mRNA levels and activities for gamma GCS and that for gamma-glutamyl transpeptidase (gamma GT), the GSH salvage pathway enzyme, were measured in these cells. The mRNA for gamma GT and gamma GCS were measured by RT-PCR, with quantitation of the PCR product by HPLC; mRNA levels are expressed as ratios to beta-actin mRNA, used as an endogenous standard. GSH and gamma GCS activity were measured by HPLC assays and gamma GT activity by a colorimetric assay. The increase in GSH in C10 and C25 was associated with an elevation in gamma GT mRNA (2.5- and 8-fold) and gamma GT activity (2.7- and 2.8-fold). No changes were observed in gamma GCS mRNA levels or activity. The data indicate that alterations in GSH metabolism leading to elevations in cellular GSH in A2780 ovarian carcinoma cells selected for low levels of resistance to oxaliplatin are mediated by gamma GT, the salvage" pathway, rather than an increase in GSH biosynthesis.

Plasma and tumor tissue pharmacology of high-dose intravenous leucovorin calcium in combination with fluorouracil in patients with advanced colorectal carcinoma

Plasma pharmacokinetics of high-dose (500 mg/m2) leucovorin calcium (dl-5-formyltetrahydrofolic acid [dl-CF]) and fluorouracil (FUra) have been evaluated in patients with advanced colorectal cancer treated with the combination of FUra and dl-CF by two different intravenous (IV) schedules: (A) In patients with no prior chemotherapy, dl-CF was administered by a two-hour IV infusion and FUra by rapid IV injection one hour after the start of the dl-CF infusion and (B) in previously treated patients, dl-CF and FUra were administered by five-day continuous IV infusion (CI). Following the two-hour infusion of dl-CF, mean peak plasma concentration and elimination half-life of I-5-formyltetrahydrofolic acid (I-CF) were 24 +/- 6 mumol/L and 0.8 +/- 0.1 hour, respectively. CI of dl-CF over five days yielded a mean steady-state plasma level of I-CF of only 1.2 +/- 0.5 mumol/L. Peak and steady-state plasma concentrations of the metabolite 5-methyl tetrahydrofolic acid were comparable in the two schedules (17 +/- 8 mumol/L for the two-hour infusion and 12 +/- 5 mumol/L for the CI). Areas under the concentration v time curve (AUC) of total reduced folates were significantly greater under conditions of CI: 89.0 v 16.7 mmol/L/min for the two-hour infusion. In tumor tissue, 5,10-methylenetetrahydrofolate increased eight-fold two to four hours following the two-hour infusion and two-fold during the CI of dl-CF and FUra. Inhibition of thymidylate synthase (dTMP-S) by the two-hour and CI infusion schedules were 66% v 39%, respectively. The observed differences in the intracellular dTMP-S folate cofactor pools and the degree of inhibition of dTMP-S achieved in patients treated by two different schedules may be due to differences in the biochemical properties and/or to differences in the modulation of FUra metabolism by folate of tumor tissues obtained from newly diagnosed and previously treated patients.

Pharmacokinetics of methotrexate following intravenous and intraventricular administration in acute lymphocytic leukemia and non-Hodgkin's lymphoma.

A pharmacokinetic study of methotrexate (MTX) administered by the intravenous (IV) and intraventricular (via an Ommaya reservoir) route was performed in 16 children, 13 with acute lymphocytic leukemia (ALL) and three with non‐Hodgkins lymphoma. Five children with ALL were treated “prophylactically” for presumed subclinical central nervous system (CNS) leukemia. The remaining 11 patients were treated for overt meningeal involvement. MTX was administered intravenously at a dose of either 500 mg/m2 or 1500 mg/m2 with one third by rapid intravenous infusion and two thirds intravenously over 24 hours with leucovorin rescue. Intraventricular MTX was given in some treatment courses at a dose of 12 mg/m2. At either 500 mg/m2 or 1500 mg/m2, when given only IV, MTX results in a 100 fold higher concentration in plasma compared to cerebrospinal fluid (CSF). The plasma levels are three times higher with the 1500 mg/m2 dose compared to the 500 mg/m2 dose. After intraventricular administration of MTX, patients with overt CNS leukemia retained MTX in the CSF significantly longer than patients treated prophylactically. This may be due to abnormal transport of MTX out of the CSF in patients with meningeal disease. Cancer 50:1676‐1682, 1982.

Unusual central nervous system toxicity in a phase I study of N1N11diethylnorspermine in patients with advanced malignancy

The objectives of this study were to determine the dose limiting toxicity (DLT) and other major toxicities, the maximum tolerated dose (MTD) and the human pharmacokinetics of N1N11diethylnorspermine (DENSPM), a new polyamine analog which in experimental systems inhibits the biosynthesis of intracellular polyamines and promotes their degradation by inducing the enzyme spermine/spermidine N-acetyl transferase.These objectives were incompletely achieved because of the occurrence of an unusual syndrome of acute central nervous system toxicity which forms the basis of the present report. Fifteen patients with advanced solid tumors were entered into a phase I study of DENSPM given by a 1h i.v. infusion every 12h for 5 days (10 doses). The starting dose was 25 mg/m2/day (12.5 mg/m2/dose) with escalation by a modified Fibonacci search.Doses of 25 and 50 mg/m2/day were tolerated with only minor side effects of facial flushing, nausea, headache and dizziness (all grade I). At doses of 83 and 125 mg/m2/day, a symptom complex of headache, nausea and vomiting, unilateral weakness, dysphagia, dysarthria, numbness, paresthesias, and ataxia, was seen in 3 patients, one after 2 courses of 83 and 2 after 1 course of 125 mg/m2/day. This syndrome occurred after drug administration was complete and the patients had returned home. Lesser CNS toxicity was seen in 2 other patients at lower daily doses. Preliminary pharmacokinetics of DESPM measured in plasma by HPLC in 8 patients showed linearity with dose and a rapid plasma decay with a t2 of 0.12h.We conclude that great caution is warranted in administering DENSPM on this schedule at doses of ≥ 83 mg/m2/day.

Bowel rest, intravenous hydration, and continuous high‐dose infusion of octreotide acetate for the treatment of chemotherapy‐induced diarrhea in patients with colorectal carcinoma

Background. A prospective trial was conducted involving 16 patients with colorectal adenocarcinoma using a regimen of continuous‐infusion octreotide acetate (Sandostatin [octreotide acetate], Sandoz, East Hanover, NJ) for the treatment of severe diarrhea induced by the weekly schedule of 5‐fluorouracil (5‐FU) in combination with leucovorin who were refractory to opiate therapy.

Intracellular glutathione and cytotoxicity of platinum complexes

Although there have been a number of reports correlating cellular GSH levels with cytotoxicity of platinum agents, none has examined the relationship between GSH concentrations and cytotoxicity. In this study, using a highly specific HPLC method for measuring GSH and expressing GSH as concentration and also per cell number, we evaluated the correlation between GSH levels and the cytotoxicity to five agents in ten human tumor cell lines. The five platinum agents included the platinum(II) complexes cisplatin, carboplatin and oxaliplatin and platinum(IV) complexes iproplatin and tetraplatin. The correlation between intracellular GSH concentration and cytotoxicity was highly significant only for iproplatin (P=0.002) followed by tetraplatin, which demonstrated a trend toward statistical significance (P=0.06). Cytotoxicity of the other platinum complexes showed no relation to GSH concentration, cisplatin itself showing aP-value of 0.09. In contrast, the GSH levels normalized to cell number showed a statistically significant correlation with the cytotoxicity of four of the five platinum agents, the exception being carboplatin; the strongest correlation observed was that for iproplatin and tetraplatin. Glutathione-S-transferase (GST) activity in these cell lines showed no correlation with cytotoxicity of any of the platinum complexes. Our results, from the analyses of both GSH concentration as well as GSH per cell number, suggest a significantly higher interaction between GSH and iproplatin compared with the other platinum agents. Moreover, our data suggest that relationships between cytotoxicity and GSH levels on a per-cell basis may not persist when differences in cell volume are taken into account.

Intrathecal methotrexate overdose without neurotoxicity. Case report and literature review

A 24‐month‐old female developed a central nervous system relapse, while in bone marrow remission, eight months after the diagnosis of acute lymphocytic leukemi was made. Inadvertently, she received 14 times the standard dose of intrathecal methotrexate (170 mg/m2 vs. 12 mg/m2). Treatment with intravenous leucovorin and oral dexamethasone was given. Her only symptomatology was mild headaches. No neurological abnormalities developed. Her cerebrospinal fluid methotrexate levels (5.2 × 10−6 at 23 hours and 5.9 × 10−7M at 47 hours) and half‐life (t 1/2 = 8 hours) were within the range previously reported in patients following standard doses of intrathecal methotrexate who did not develop neurotoxicity. The various manifestations of neurotoxicity associated with the use of intrathecal methotrexate as well as those factors considered to play a part in the development of neurotoxicity are reviewed.