Chemical synthesis, NMR analysis and evaluation on a cancer xenograft model (HL-60) of the aminosteroid derivative RM-133

NMR-Assisted Structure Elucidation of an Anticancer Steroid-β-Enaminone Derivative

“…In fact, the only disparities are related to the presence of a CH 3 instead of the steroid nucleus on a piperazine nitrogen, which affects the chemical shifts of the two CH 2 -1 . Analysis of the 13 C NMR data also makes it possible to completely confirm the 5α-androstan-3β,17β-diol backbone, as well as the presence of an ethynyl group (73.9 and 87.6 ppm) at C-17α (79.8 ppm) [7]. Finally, mass analysis ([M+H] + = 679.5 m/z) is in agreement with the proposed structure for compound 1.…”

“…In NMR spectra of both compounds, there is no evidence of peak splitting for the N-CH 3 of piperazine, the CH of proline and some quinoline signals. This duplication is typical of the presence of the two rotamers observed for a side chain like 6 [7], especially obvious for the CH-2 ( Figure 3A). New signals (a and b) integrating for 2H were also detected in the aromatic or vinylic region (6.80 and 7.91 ppm for 7 and 6.45/6.70 and 7.90/8.08 ppm for 8; correlations in correlation spectroscopy (COSY) spectra).…”

“…The major difficulty for the characterization of unknown compound 1 was the complex nature of the starting aminosteroid derivative RM-128 [8], which is well-known to exist as a mixture of two conformers in different proportions according to the solvent used for NMR analysis [7]. In fact, the presence of two amide bonds linking a piperazine, a proline, and a quinoline explain these two conformers, but the combinaison of this side chain and a steroid backbone with 19 carbons greatly complexifies the identification of such steroid derivatives by NMR.…”

“…Steroid derivatives having a (1-quinolin-2-ylcarbonyl)-L-proline-piperazine side chain at position C2 of estra-1,3,5(10)-trien-3,17β-diol [5] or 5α-androstane-3α,17β-diol [6-8] represent a new family of anticancer agents. They were found to reduce the proliferation of various human cancer cell lines [8][9][10], as well as to block tumor growth when tested in mouse tumor (xenografts) models of human cancers (pancreas, ovary, breast, and leukemia) [7,11]. During our work to increase their metabolic stability, especially for oral administration, one of our strategies was to introduce an ethynyl group at position C17 of the steroid nucleus to generate a tertiary alcohol instead of a secondary alcohol [12].…”

“…This more metabolic stable compound was alternatively obtained by another strategy that introduced the ethynyl group earlier in the reaction sequence, thus providing RM-133 in good yield and purity [7], but the negative results of the first chemical strategy provided an unexpected finding. In fact, the addition of lithium (trimethylsilyl)acetylide to the C17-carbonyl of RM-128 generated a new compound, which after purification was found to be very active as an antiproliferative agent on a variety of human cancer cell lines (OVCAR-3, PANC-1, MCF-7, T-47D), being about two times more potent than RM-133 in a comparative assay (unpublished results).…”