Analogues of the Potent Nonpolyglutamatable Antifolate N^α-(4-Amino-4-deoxypteroyl)-N^δ-hemiphthaloyl-l-ornithine (PT523) with Modifications in the Side Chain, p-Aminobenzoyl Moiety, or 9,10-Bridge:  Synthesis and in Vitro Antitumor Activity

Abstract

Seven N(alpha)-(4-amino-4-deoxypteroyl)-N(delta)-hemiphthaloyl-L-o rnithine (2, PT523) analogues were synthesized by modifications of the literature synthesis of the corresponding AMT (1) analogues and were tested as inhibitors of tumor cell growth. In growth assays against cultured CCRF-CEM human leukemic cells exposed to drug for 72 h, the IC(50) values of analogues in which N(10) was replaced by CH(2) and CHMe were found to be 0.55 +/- 0.07 and 0.63 +/- 0.08 nM, and thus these analogues are more potent than 1 (IC(50) = 4.4 +/- 1.0 nM) or 2 (IC(50) = 1.5 +/- 0.39 nM). The 10-ethyl-10-deaza analogue of 2 (IC(50) = 1.2 +/- 0.25 nM) was not statistically different from 2 but was more potent than edatrexate, the 10-ethyl-10-deaza analogue of 1, which had an IC(50) of 3.3 +/- 0.36 nM. In contrast, the analogue of 2 with both an ethyl and a CO(2)Me group at the 10-position had an IC(50) of 54 +/- 4.9 nM, showing this modification to be unfavorable. The 4-amino-1-naphthoic acid analogue of 2 had an IC(50) of 1.2 +/- 0.22 nM, indicating that replacement of the p-aminobenzoic acid (pABA) moiety does not diminish cytotoxicity. The analogues in which the (CH(2))(3) side chain was replaced by slightly longer CH(2)SCH(2) and (CH(2))(2)SCH(2) groups gave IC(50) values of 4.4 +/- 1.1 and 5.0 +/- 0.56 nM and thus were somewhat less potent than the parent molecule. However the analogues in which the aromatic COOH group was at the meta and para positions of the phthaloyl ring had IC(50) values of 7.5 +/- 0.47 and 55 +/- 0.07 nM, confirming the low potency we had previously observed with these compounds against other cell lines. Overall, the results in this study support the conclusion that, while the position of the phthaloyl COOH group and the length of the amino acid side chain in 2 are important determinants of cytotoxic potency, changes in the pABA region and 9, 10-bridge are well-tolerated and can even increase potency.

Synthesis and in Vitro Antifolate Activity of Rotationally Restricted Aminopterin and Methotrexate Analogues

“…15 As shown in Table 1, the IC 50 values of 9 and 10 against CCRF-CEM cells during 72 h of exposure were 5.1 ( 0.25 and 140 ( 5.0 nM, as compared with previously determined values of 4.4 ( 0.10 and 14 ( 2.6 nM for AMT and MTX, respectively. It thus appeared that the increase in RFC binding of 9 relative to AMT was able to approximately offset its decrease in DHFR binding, whereas in the case of 10 versus MTX the modest gain in RFC binding was not enough to offset a much larger decrease in DHFR binding.…”

DNA and RNA Synthesis: Antifolates

“…200 Synthesis of a series of analogues of PT 523 with modifications in the side chain, p-aminobenzoyl moiety, or C9-C10 bridge has been reported. 198,201 The growth inhibition values of the selected compounds 40a-40e are shown in the Table 17. The shift of the terminal ortho-carboxyl group to the meta or para position has a detrimental effect on the activity.…”

4.05 Addition of HX Reagents to Alkenes, Alkynes, and Allenes without Transition Metal

“…The resulting solution of 1-chloro-4-methoxy-2-butene was kept at 0-5°C until it could be used in the next step. 37 • OCH 3…”