Synthesis and Evaluation of Co-drugs Derived from Methotrexate and Ibuprofen for the Treatment of Psoriasis.

摘要

The pathophysiology of psoriasis, an autoimmune, inflammatory disease is attributed to abnormal hyperproliferation of keratinocytes and increased levels of certain cytokines which lead to inflammation. To target each of these conditions, a novel class of co-drugs derived from methotrexate (MTX) and ibuprofen (IBU) were synthesized and their inhibition of dihydrofolate reductase (DHFR), an enzyme associated with purine synthesis and cell proliferation was evaluated. The expectation is that these co-drugs hydrolyze at the site of action (skin) and target the two main factors that contribute to the pathophysiology of psoriasis. Co-drugs alpha-MTX-IBU (6a) and gamma-MTX-IBU (6b) were successfully synthesized in moderate yields (20--35%). Spectral analysis (IR, MS and NMR) was used to confirm the formation of the products after reactions to form intermediates and the final co-drugs. The inhibitory activity of these co-drugs, the parent compounds (MTX, IBU) and the combination (1:1 MTX +IBU) was evaluated using a DHFR assay. MTX, MTX in combination with IBU and the co-drugs showed inhibitory activity, while IBU did not inhibit DHFR nor did it interfere with the ability of MTX to inhibit DHFR. At concentrations of 5, 6, 10 and 20nM, combinations of MTX with IBU showed statistically higher observed inhibition than predicted by an additive effect suggesting the combination of MTX and IBU at these concentrations work synergistically. The co-drugs showed some inhibitory activity, although it was significantly lower than that observed for MTX or MTX with IBU. This co-drug activity suggests that either partial hydrolysis occurred to release MTX or the intact co-drug weakly inhibits DHFR. A high pressure liquid chromatography (HPLC) method was also developed to monitor the stability and hydrolysis of the co-drugs for future studies. The retention time for MTX was 1.22 minutes and the two co-drugs 6a and 6b eluted at 9.50 and 9.53 minutes, respectively. These retention times provided a good separation between MTX and the co-drugs and can be used for future studies to monitor for hydrolysis.