SarCNU, a new chloroethylnitrosourea with a methylglycinamide (sarcosinamide) carrier has enhanced in vitro cytotoxicity as compared to BCNU, against human glioma specimens and human glioma cell lines, when utilized at clinically achievable plasma concentrations. The increased in vivo antitumor activity of SarCNU in athymic mice bearing the U-251 glioma cell line intracranially confirmed the in vitro results. SarCNU is transported into SK-MG-1 glioma cells by a carrier mediated mechanism in contrast to BCNU which enters via passive diffusion. The Km of sarcosinamide uptake by this carrier is 284muM. Dixon plot analysis of SarCNU inhibition of [3H]sarcosinamide uptake reveals a KI of 3.26 muM. This carrier has a greater affinity for sarcosinamide than SarCNU. The carrier does not appear to accomodate any natural substrate tested to date except leucine at high concentrations. Amino acids at physiological concentrations do not alter the cytotoxicity of SarCNU. This carrier mediated uptake of SarCNU may explain the 2-fold greater intracellular accumulation of SarCNU as compared to BCNU. In order to examine the increased antitumor activity of SarCNU and define parameters associated with nitrosourea antitumor activity in patients with malignant glioma, we will determine: (1) the activity of 06-alkyl-quanine- DNA-alkyl-transferase (GATase) activity in 20 primary glioma specimens and correlate this with tumor response in these patients to BNCU; (2) the activity of Gatase in 3 glioma cell lines of varying sensitivities to SarCNU; (3) if SarCNU inactivates Gatase more effectively than BCNU; 94) if the kinetic parameters of [3H]sarcosinamide transport, KI of SarCNU and intracellular accumulation of SarCNU correlate with the decreased sensitivities to SarCNU of SK-MG-4 and SKI-1 glioma cell lines; (5) if fibroblasts differ from glioma cell lines in the above kinetic parameters of sarcosinamide transport; (6) [3H]SarCNU transport in glioma cell lines and fibroblasts; (7) the affinity of other amino acid amides and their corresponding nitrosoureas for the sarcosinamide carrier; (8) the DNA crosslink kinetics in the 3 glioma cell lines in order to correlate these results with cytotoxicity. Investigations of the carries mediated amino acid amide transport systems in glioma cells may lead to the development of a new class of anticancer agents with more selective antitumor activity. GRANTRO1NS22404 Nerve injury, and subsequent repair, is a major problem confronting modern medicine. It has been estimated that more than 200,000 nerve repair procedures are performed yearly in the United states alone. The extent of this medical problem is exacerbated by the generally poor results of current peripheral nerve repair procedures. Our proposed studies will test the ability of collagen-based nerve guide tubes to promote nerve regeneration in both the peripheral and central nervous system. In non- human primates we show that such prostheses perform as well as, and in instances better than, a standard nerve autograft to repair transected median nerve. It is thus becoming increasingly evident that such studies have direct relevance to human peripheral nerve repair.
The specific aim of this proposal can be divided into three different categories: 1) The rat experiments (Specific Aims I-V) address a long- standing question of the ability of mammalian peripheral nerves to selectively reinnervate appropriate target muscle, 2) the primate experiments (Specific Aim VI) will test and compare the long-term efficacy of peripheral nerve repair with new biodegradable nerve guide tube vs a standard nerve autograft, and 3) the rabbit experiments (Specific Aim VII) will begin to identify individual non-neuronal cells which may be responsible for promoting regeneration within the central nervous system. The rodent and primate peripheral nerve experiments represent a logical progression of steps toward the goal of improving the clinical management of human nerve injuries. The rat studies will uncover manipulations that effect the number of individual motor and sensory neurons which regenerate to an appropriate muscle, thus increasing the possibility of functional recovery. The monkey studies will take the possibility of using currently available collagen based nerve guide tubes closer to the realm of human peripheral nerve repair by definitely establishing the long-term usefulness and safety of such prostheses for promoting the nerve regeneration in primates. Finally, the experiments with the rabbit optic nerve indicate the ability of nerve guide tubes to test, one by one, the individual cellular elements which may be responsible for promoting central nervous system regeneration.
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