In vitro cytotoxicity, gene expression, bioinformatics, biochemical analysis, and in silico analysis of synthesized carbonitrile derivatives

In this work, an efficient single-step green synthesis protocol of 2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitriles was applied with the purpose of their biological activity studies. It was found that the cytotoxic activities of 2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitriles in the SHSY-5Y cancer cell line were most active after 72 h of incubation. It was determined that 1,6-diamino-4a-methyl-3-oxo-2,8-diphenyl-2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitrile showed higher anticancer activity on SHSY-5Y cancer cells compared to other molecules. The expression levels of Myc-N, Casp2, Tp53, RAD51, BRCA2, MDM2, BAX and NF-kappa B1 genes were analyzed by RT-PCR method by applying 2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitriles to the SHSY-5Y neuroblastoma cancer cell line. It was observed that there were differences in expression levels between study groups and all differences were found to be statistically significant (p < 0.000). Bioinformatics analysis was performed using the STRING v 11 Protein interaction tool. When the molecules in the SHSY-5Y cell line were compared to the control group, an 80% increase in GSH levels was observed in 1,6-diamino-4a-methyl-3-oxo-2,8-diphenyl-2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitrile and 1,6-diamino-8-(4-chlorophenyl)-4a-methyl-3-oxo-2-phenyl-2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitrile groups. SOD enzyme activity increased significantly in the former group compared to the control and other groups. LDH activity was detected at a higher rate in this cell line compared to the control group. Calculations were made for 2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitriles using the Gaussian software package on B3LYP, HF, and M06-2 x levels with the 6-31++g(d,p) basis sets. The activities of the 2,3,4,4a-tetrahydroisoquinoline-5,7-dicarbonitriles found in the study against SH-SY5Y protein (2F37, 3PBL and 5WIV), comparison has been made. MM/GBSA methods are calculated binding free energy for all molecule and 3PBL protein. Afterwards, ADME/T analysis was performed to examine the effects of the molecules on human metabolism.