SIDE-CHAIN MODIFIED 5-DEAZAFOLATE AND 5-DEAZATETRAHYDROFOLATE ANALOGSAS MAMMALIAN FOLYPOLYGLUTAMATE SYNTHETASE AND GYLCINAMIDE RIBONUCLEOTIDE FORMYLTRANSFERASE INHIBITORS - SYNTHESIS AND IN-VITRO BIOLOGICAL EVALUATION .48. SYNTHESIS AND BIOLOGICAL-ACTIVITY OF N-OMEGA-HEMIPHTHALOYL-ALPHA,OMEGA-DIAMINOALKANOIC ACID ANALOGS OF AMINOPTERIN AND 3',5-DICHLOROAMINOPTERIN

Analogues of -deoxypteroyl)-N-delta-(hemiphthaloyl)-L-ornithine (PT523 ) with 3',5'-dichloro substitution in the p-aminobenzoyl moiety or wit h one less or one more CH2 group in the amino acid moiety were synthes ized and tested as inhibitors of dihydrofolate reductase (DHFR) activi ty and cell growth. Replacement of L-ornithine in PT523 by L-2,4-diami nobutanoic acid or L-lysine did not decrease binding to human recombin ant DHFR but resulted in some loss of activity against SCC25 human and SCC VII murine squamous cell carcinoma and against MCF-7 human breast carcinoma in culture. PT523 was several times more potent than methot rexate (MTX), aminopterin (AMT), or trimetrexate (TMQ). 3',5'-Dichloro substitution did not decrease either DHFR binding or cytotoxicity. A new synthetic route to PT523 from 2,4-diamino-6-(hydroxymethyl)pteridi ne and methyl a-(4-aminobenzoyl)-N-delta-phthaloyl-L-ornithinate was i nvestigated but was not found superior to previously described methods . In comparative experiments on the ability of PT523 and MTX to compet itively inhibit the influx of (GR)-5,10-dideazatetrahydrofolate (DDATH F, lometrexol), used here as a surrogate for MTX and reduced folates, the K-i of PT523 was lower than that of MTX in both wild-type CCRF-CEM human leukemic lymphoblasts and the transport- and polyglutamylation- defective subline CEM/MTX. The CCRF-CEM cells were 10-fold more sensit ive to PT523 than to MTX, whereas the CEM/MTX cells were 240-fold more sensitive. However, in contrast to other MTX-resistant cells where co llateral sensitivity to PT523 has been seen. CEM/MTX cells still showe d substantial cross resistance to PT523 which may reflect an unusual h eightened ability to utilize exogenous folic acid. The good correlatio n observed with both cell lines between the cytotoxicity of PT523 and MTX and the ability to inhibit DDATHF influx supported the view that P T523 and MTX share, at least in part, a common protein carrier for mem brane transport.