Edward F. Elslager

Biochemical pharmacology of the lipophilic antifolate, trimetrexate

Trimetrexate is a novel lipophilic folate antagonist that causes growth inhibition, inhibition of nucleic acid biosynthesis, and cytotoxicity at nanomolar concentrations in tissue cultures. The potency of trimetrexate cytotoxicity against most cell lines is greater than that of methotrexate. Trimetrexate has antitumor activity in vivo in several murine leukemia and solid tumor systems, including tumors in which methotrexate is inactive. Antitumor activity was seen following oral, intravenous, or intraperitoneal administration. Trimetrexate causes a pronounced and early depression in incorporation of deoxyuridine into DNA. In tumor cell lines resistant to methotrexate because of a drug transport defect, trimetrexate retains activity. In many such cases the methotrexate-resistant tumors show collateral sensitivity to trimetrexate. In methotrexate-resistant cells with impaired drug transport, trimetrexate sensitivity was even more pronounced when cells were grown in folate-free medium supplemented with physiological levels of tetrahydrofolate cofactor. In the human tumor stem cell colony assay, trimetrexate, at concentrations achievable in vivo, gave activity against many human tumors, including samples that were unresponsive to methotrexate. Trimetrexate crosses the blood-brain barrier, and at very high doses may cause neurotoxicity. At conventional doses the primary toxic effects in mice are gastrointestinal. This toxicity is reversible at therapeutic doses. Unlike earlier lipophilic antifolates, trimetrexate has rapid plasma clearance (t1/2 in mice of 45 minutes). Trimetrexate is a tight-binding competitive inhibitor of dihydrofolate reductase. The Ki,slope for inhibition of the human enzyme was 4 X 10(-11) M. A dose-dependent decrease in cellular purine ribonucleotide pools is given by trimetrexate. Pyrimidine ribonucleotide pools tend to increase in treated cells. Trimetrexate caused a marked depression of cellular pools of dTTP and dGTP, and a lesser depression in dATP. Cytotoxicity of trimetrexate in vitro was prevented by leucovorin. Leucovorin also protected mice from trimetrexate toxicity. Thymidine protected cells from lethal effects of low concentrations of trimetrexate, but not from high concentrations. The combination of thymidine and hypoxanthine completely protected cells from low and high concentrations of trimetrexate. A new, stable and highly water-soluble formulation of trimetrexate has been developed. Because of the interesting biochemical and pharmacological properties of trimetrexate, and its experimental antitumor activity, clinical trials are planned.

In Vitro activity of the novel antitumor antibiotic fostriecin (CI-920) in a human tumor cloning assay☆

A human tumor cloning assay was utilized to evaluate the antineoplastic activity of the novel antitumor antibiotic fostriecin (CI-920). Initial screening with 10.0 mcg/ml continuous exposure against a variety of histologic tumor types resulted in 14/51 (27%) in vitro responses (defined as greater than 50% decrease in TCFUs). Further investigation of the compound was performed in 1-hr preincubation experiments. The in vitro response rate at a concentration of 1.0 mcg/ml (which was considered to correspond to a clinically achievable concentration) was 15/43 (35%). Response rates for specific tumor types included: 5/15 in ovarian cancer, 5/12 in breast, and 4/11 in human lung cancer. The impression of significant antitumor activity of the compound at this dose was further substantiated by comparing its in vitro activity with a variety of simultaneously tested standard anticancer agents. In addition, these data indicated the possibility of non-cross resistance of CI-920 to several established cytostatics. CI-920 is a compound with good in vitro activity which should be further developed for clinical trials.

Folate antagonists. 12. Antimalarial and antibacterial effects of 2,4-diamino-6-[(aralkyl and alicyclid)thio-, sulfinyl-, and sulfonyl]quinazolines.

A series of 2,4-diamino-6-[(aralkyl and alicyclic)thio-, sulfinyl-, and sulfonyl]quinazolines was prepared via condensation of 5-chloro-2-nitrobenzonitrile or 5,6-dichloro-2-nitrobenzonitrile with the appropriate aralkyl or alicyclic thiopseudourea, reduction of the resulting 2-nitro-5-[(aralkyl or alicyclic)thio]benzonitrile with stannous chloride to the amine, and cyclization with chloroformamidine hydrochloride. Oxidation was effected with hydrogen peroxide or the bromine complex of 1,4-diazabicyclo[2.2.2]octane. These analogues when examined for suppressive activity against drug-sensitive lines of Plasmodium berghei in mice were not as active as 2,4-diamino-6-[3,4-dichlorobenzyl)amino]quinazoline (Ia).

New perspectives on the chemotherapy of malaria, filariasis, and leprosy.

The great advances made in chemotherapy during this century have enabled some measure of control of the major killing diseases of the tropics. Notwithstanding past progress, man today is confronted by a catalog of challenging problems, and the incentive to develop novel and improved antiinfective agents has not diminished. Adequate drugs are still lacking for the treatment of many chronic and debilitating diseases including leishmaniasis, Chagas’ disease, filariasis, schistosomiasis, clonorchiasis, trichuriasis, strongyloidiasis, and leprosy. Moreover, reports of drug resistance have often followed advances in chemotherapy like a ‘faithful shadow’, and this shadow will no doubt lengthen in the future. The philosophy of global eradication of communicable diseases [1] and the evolution of techniques for its achievement have added a new dimension to research in parasite chemotherapy, namely the urgent need for drugs with protracted action. While the clinician interested in treating an individual often has a variety of good drugs at his disposal and may feel little need for new agents, the public health worker, whose aim is the ultimate eradication of a disease from an entire community, faces problems of a magnitude and type not encountered by the clinician and is frequently unable to achieve his objective of total coverage with currently available drugs. Future progress in parasite chemotherapy will depend mainly on an increased awareness of such critical problems and needs and on appropriate recognition of biochemical processes within both parasite and host. Intensive efforts to develop useful agents for the treatment of parasitic diseases of livestock, poultry, and other domestic animals have also yielded a variety of promising new substances, and the benefits derived from interplay between human and veterinary research are already apparent.

SYNTHESIS AND ANTIMALARIAL EFFECTS OF 2‐(3,4‐DICHLOROANILINO)‐7‐(((DIALKYLAMINO)ALKYL)AMINO)‐5‐METHYL‐S‐TRIAZOLO(1,5‐A)PYRIMIDINES(1,2)

3,4-Dichlorophenylisothioeyanate (10) was allowed to react with 2-methy1-2-thiopseudourea to give methyl 4-(3,4-dichlorophenyl)(dithioaltophanimidate (11) (41%), which upon treatment with hydrazine afforded 3-amino-5-(3,4-dichloroanilino)-s-triazole (12) (54-91%). Ring-closure with ethyl acetoacetale in acetic acid afforded 2-(3,4-dichloroanilino)-5-methyl-s-triazolo[ 1,5-α ]-pyrimidin-7-ol (13) (81%). Chlorination with phosphorus oxychloride gave 7-chloro-2-(3,4-dichloroanilino)-5-methyl-s-triazolo[1,5-α ]pyrimidine (14) (98%), which was condensed with various amines to yield the desired 2-(3,4-diehloroanilino)-7-¶[(dialkylamino)alkyl]arnino¶-5-methyl-s-triazolo[ 1,5-α]pyrimidines (6a-d). The structures of the s-triazolo[ 1,5-α ]pyrimidines were based on nmr spectroscopy and ring stability considerations. Several of the amino-s-triazolo[ 1,5-α ]pyrimidines possessed antimalarial activity against P. berghei in mice.

Progress in Malaria Chemotherapy

Among the ills that have plagued mankind, few have exacted a higher toll than malaria. The advent of insecticides simplified the logistics of malaria control, and following World War II many nations took up this approach in earnest. The goal of global eradication was formally adopted at the eighth World Health Assembly in 1955.