Murano Kenji, Yamanaka Toshio, Toda Ayako, Ohki Hidenori, Okuda Shinya, Kawabata Kohji, Hatano Kazuo, Takeda Shinobu, Akamatsu Hisashi, Itoh Kenji, Misumi Keiji, Inoue Satoshi, Takagi Tatsuya
Lead Discovery Research Labs, Astellas Pharma Inc., 21 Miyukigaoka, Tsukuba, Ibaraki 305-8585, Japan.
Bioorg Med Chem. 2008 Mar 1;16(5):2261-75. doi: 10.1016/j.bmc.2007.11.074. Epub 2007 Dec 3.
AmpC beta-lactamase is one of the leading causes of Pseudomonas aeruginosa (P. aeruginosa) resistance to cephalosporins. FR259647 is a cephalosporin having a novel pyrazolium substituent at the 3-position and exhibits excellent activity (MIC=1 microg/mL) against the AmpC beta-lactamase overproducing P. aeruginosa FP1380 strain in comparison with the third-generation cephalosporins FK518 [Abstracts of Papers, 30th Interscience Conference on Antimicrobial Agents and Chemotherapy, Atlanta, GA, October 21-24, 1990, Abs. 454; Abstracts of Papers, 30th Interscience Conference on Antimicrobial Agents and Chemotherapy, Atlanta, GA, October 21-24, 1990, Abs. 455; Abstracts of Papers, 30th Interscience Conference on Antimicrobial Agents and Chemotherapy, Atlanta, GA, October 21-24, 1990, Abs. 456; Abstracts of Papers, 30th Interscience Conference on Antimicrobial Agents and Chemotherapy, Atlanta, GA, October 21-24, 1990, Abs. 457] (MIC=16 microg/mL) and ceftazidime (CAZ) (MIC=128 microg/mL). The stability of FR259647 and FK518 to AmpC beta-lactamase was evaluated using MIC assays against both the P. aeruginosa PAO1 strain and a PAO1 mutant strain overproducing AmpC beta-lactamase as a differential assay, which indicates that the main difference derives from their stability to AmpC beta-lactamase. A structural analysis using computer simulations indicated that the difference in stability may be due to steric hindrance of the 3-position substituents causing differential affinity. This steric hindrance may disturb entry of the cephalosporins into the binding pocket. We predicted the possibility of inhibition of entry as a potential means of enhancing stability by conformational analysis. In order to validate this speculation, novel FR259647 derivatives 4-9 were designed, calculated, synthesized, and evaluated. As a result, we demonstrated that their probability of entry correlated with the MIC ratio of the mutant strain to the parent strain and supports the validity of our model.
AmpCβ-内酰胺酶是铜绿假单胞菌对头孢菌素耐药的主要原因之一。FR259647是一种在3位具有新型吡唑鎓取代基的头孢菌素，与第三代头孢菌素FK518[《论文摘要》，第30届抗菌药物和化疗跨学科会议，佐治亚州亚特兰大，1990年10月21日至24日，摘要454；《论文摘要》，第30届抗菌药物和化疗跨学科会议，佐治亚州亚特兰大，1990年10月21日至24日，摘要455；《论文摘要》，第30届抗菌药物和化疗跨学科会议，佐治亚州亚特兰大，1990年10月21日至24日，摘要456；《论文摘要》，第30届抗菌药物和化疗跨学科会议，佐治亚州亚特兰大，1990年10月21日至24日，摘要457]（MIC = 16μg/mL）和头孢他啶（CAZ）（MIC = 128μg/mL）相比，对过量产生AmpCβ-内酰胺酶的铜绿假单胞菌FP1380菌株表现出优异的活性（MIC = 1μg/mL）。使用针对铜绿假单胞菌PAO1菌株和过量产生AmpCβ-内酰胺酶的PAO1突变菌株的MIC测定法作为差异测定法，评估了FR259647和FK518对AmpCβ-内酰胺酶的稳定性，这表明主要差异源于它们对AmpCβ-内酰胺酶的稳定性。使用计算机模拟进行的结构分析表明，稳定性的差异可能是由于3位取代基的空间位阻导致亲和力不同。这种空间位阻可能会干扰头孢菌素进入结合口袋。我们通过构象分析预测了抑制进入作为增强稳定性的潜在手段的可能性。为了验证这一推测，设计、计算、合成并评估了新型FR259647衍生物4 - 9。结果，我们证明了它们进入的可能性与突变菌株与亲本菌株的MIC比值相关，并支持了我们模型的有效性。
