Synthesis and evaluation of anti-apoptotic activity of L-carnitine cyclic analogues and amino acid derivatives.

Two series of derivatives were synthesised. In one series (R)-4-hydroxy-2-pyrrolidinone was used as a mimic of cyclic L-carnitine analogue and in the second series 3-amino-2-piperidinone was used as a cyclic ornithine analogue. N-Benzyloxycarbonyl derivatives of some amino acids were also prepared. The newly synthesised compounds were tested for their ability to inhibit Fas-activated apoptosis of human Jurkatt T-cell line. The results confirm the previously described anti-apoptotic activity of carnitine and indicate new carnitine and amino acid analogues (1, 3, 6, 7, 20) that inhibit Fas-induced apoptosis.

1,4-benzothiazine analogues and apoptosis: structure-activity relationship.

We have previously shown 1,4-benzothiazine (1,4-B) derivatives induce thymocyte apoptosis in vitro and thymus cell loss in vivo. Apoptosis is mediated through a complex of biochemical events including phosphatidylcholine specific-phospholipase C (PC-PLC) activation, acidic sphingomyelinase (aSMase) activation and ceramide generation, caspase-8 and caspase-3 activation. As preliminary analysis of the structure-activity relationship (SAR) suggested some structural features were responsible for apoptosis, we synthesised several derivatives and tested for apoptosis activity at equimolar concentrations. In particular, we synthesised analogues that differed in the nature of skeleton (1,4-benzothiazine, 1,4-benzoxazine and 1,2,3,4-tetrahydroquinoline) and in the nature of side chain (imidazole, benzimidazole or piperazine as azole substituent; presence, absence or transformation of alcoholic group). Results of apoptosis induction indicate that transforming the 1,4-benzothiazine skeleton into 1,2,3,4-tetrahydroquinoline does not result in significant change. Transformation into 1,4-benzoxazine decreased activity. Replacing imidazole at the side chain with different piperazines also decreased activity while replacing it with benzimidazole does not change apoptotic activity. Finally, removal of the alcoholic group by dehydration to olefin, or by transforming it into ether, increased activity. Moreover, in an attempt to analyse further the SAR characteristics that are responsible for 1,4-B-activated apoptosis we tested the effect on caspase-8,-9 and-3 activation. 1,4-B analogues activate caspases and the structural requirements correlate with those responsible for apoptosis induction.