Department of Chemistry, University of Southern California, Los Angeles, CA 90089-0744, USA.
Eur J Med Chem. 2011 Oct;46(10):4820-6. doi: 10.1016/j.ejmech.2011.04.063. Epub 2011 May 7.
Phosphonocarboxylate (PC) analogues of bisphosphonates are of interest due to their selective inhibition of a key enzyme in the mevalonate pathway, Rab geranylgeranyl transferase (RGGT). The dextrarotatory enantiomer of 2-hydroxy-3-(imidazo[1,2-a]pyridin-3-yl)-2-phosphonopropanoic acid (3-IPEHPC, 1) is the most potent PC-type RGGT inhibitor thus far identified. The absolute configuration of (+)-1 in the active site complex has remained unknown due to difficulties in obtaining RGGT inhibitor complex crystals suitable for X-ray diffraction analysis. However, we have now succeeded in crystallizing (-)-1 and here report its absolute configuration (AC) obtained by X-ray crystallography, thus also defining the AC of (+)-1. An Autodock Vina 1.1 computer modeling study of (+)-1 in the active site of modified RGGT binding GGPP (3DSV) identifies stereochemistry-dependent interactions that could account for the potency of (+)-1 and supports the hypothesis that this type of inhibitor binds at the TAG tunnel, inhibiting the second geranylgeranylation step. We also report a convenient (31)P NMR method to determine enantiomeric excess of 1 and its pyridyl analogue 2, using α- and β-cyclodextrins as chiral solvating agents, and describe the synthesis of a small series of 1 α-X (X = H, F, Cl, Br; 7a-d) analogues to assess the contribution of the α-OH group to activity at enzyme and cellular levels. The IC(50) of 1 was 5-10× lower than 7a-d, and the LED for inhibition of Rab11 prenylation in vitro was 2-8× lower than for 7a-d. However, in a viability reduction assay with J774 cells, 1 and 7b had similar IC(50) values, ~10× lower than those of 7a and 7c-d.
膦羧酸酯（PC）类似物的双膦酸盐由于其选择性抑制甲羟戊酸途径中的关键酶，法尼基焦磷酸合酶（RGGT）。右旋异构体的 2-羟基-3-（咪唑并[1,2-a]吡啶-3-基）-2-膦酸丙酸（3-IPEHPC，1）是迄今为止鉴定的最有效的 PC 型 RGGT 抑制剂。由于难以获得适合 X 射线衍射分析的 RGGT 抑制剂复合物晶体，因此活性部位复合物中（+）-1 的绝对构型（AC）仍然未知。然而，我们现在已经成功地结晶了（-）-1，并在此报告了通过 X 射线晶体学获得的绝对构型（AC），从而也确定了（+）-1 的 AC。计算机建模研究表明，（+）-1 在经修饰的 RGGT 结合 GGPP（3DSV）的活性部位中，通过 Autodock Vina 1.1 进行了研究，确定了立体化学依赖性相互作用，这可以解释（+）-1 的效力，并支持这种类型的抑制剂结合在 TAG 隧道中，抑制第二个 geranylgeranylation 步骤。我们还报告了一种方便的（31）P NMR 方法，用于确定 1 及其吡啶类似物 2 的对映体过量，使用α-和β-环糊精作为手性溶剂，并描述了一小系列 1α-X（X = H，F，Cl，Br; 7a-d）类似物的合成，以评估α-OH 基团在酶和细胞水平上的活性的贡献。1 的 IC（50）比 7a-d 低 5-10 倍，体外抑制 Rab11 prenylation 的 LED 比 7a-d 低 2-8 倍。然而，在 J774 细胞的细胞活力降低测定中，1 和 7b 的 IC（50）值相似，比 7a 和 7c-d 低约 10 倍。
