Targeting the S2 Subsite Enables the Structure-Based Discovery of Novel Highly Selective Factor XIa Inhibitors

Abstract

FXIa inhibition has been a promising strategy for treating thrombotic diseases. Up to date, many small-molecule FXIa inhibitors have been identified; however, most of them exhibit undesirable selectivity over the homologous plasma kallikrein (PKal). By employing structure-based drug design strategies, we identified many novel selective FXIa inhibitors that have extra interactions with the S2 subsite of FXIa. Among them, compound 35 displayed good inhibitory activity against FXIa and high selectivity over PKal and even several other serine proteases. Additionally, 35 showed significant anticoagulant activity toward the intrinsic pathway without affecting the extrinsic pathway. In vivo, 35 exhibited significant antithrombotic activity without increasing the bleeding risk and obvious toxicity in mice, demonstrating that it could be a promising candidate for further research. This study first demonstrates the importance of the S2 subsite of FXIa, paving the way to design highly selective FXIa inhibitors for clinical uses.

Factor XIa Inhibitors in Anticoagulation Therapy: Recent Advances and Perspectives

“…However, because of their highest homology, it is challenging to enhance the selectivity of SMFIs over PKal, and it should be noted that the requirement for high selectivity over PKal remains controversial. For example, we found that dual inhibition against FXIa and PKal slightly increased the bleeding risk in mice, 65 whereas Xu et al proposed that the dual inhibition may contribute to a synergistic anticoagulant effect. 119 Therefore, the precise benefits and drawbacks of dual FXIa/PKal inhibition must be determined in the future.…”

“…Because of the importance of the S2 subsite in increasing potency and selectivity, a series of aspartate derivatives modified from 8 were identified as selective SMFIs by our group. 65 Among them, compound 14 with an N-phenylpiperizine moiety demonstrated good FXIa potency (IC 50 = 14 nM) and more than 1000-fold selectivity over other relevant serine proteases, including PKal (IC 50 > 14 000 nM). The docking model of 14 with FXIa revealed that the phenyl ring in the N-phenylpiperazine fills the narrow pocket of the S2 subsite and establishs a hydrophobic interaction with Tyr58b, possibly explaining the excellent selectivity for PKal.…”

“…Mild bleeding caused by FXI deficiency during trauma or surgery is not correlated with FXI level but may be linked to a reduction in PKal, which is involved in the bypass pathway of PKal-FIX activation . In a tail bleeding assay, synergistic inhibition of FXIa and PKal at relatively high doses led to a somewhat longer bleeding time than selective inhibition of FXIa . Moreover, in addition to the coagulation pathway, PKal is involved in the kallikrein-kinin system, the complement pathway of host defense, and the renin-angiotensin system in humans, all of which may contribute to the potential side effects of dual FXIa/PKal inhibitors.…”

“…Because of the importance of the S2 subsite in increasing potency and selectivity, a series of aspartate derivatives modified from 8 were identified as selective SMFIs by our group . Among them, compound 14 with an N -phenylpiperizine moiety demonstrated good FXIa potency (IC 50 = 14 nM) and more than 1000-fold selectivity over other relevant serine proteases, including PKal (IC 50 > 14 000 nM).…”

“…64 In a tail bleeding assay, synergistic inhibition of FXIa and PKal at relatively high doses led to a somewhat longer bleeding time than selective inhibition of FXIa. 65 Moreover, in addition to the coagulation pathway, PKal is involved in the kallikrein-kinin system, the complement pathway of host defense, and the reninangiotensin system in humans, 31 all of which may contribute to the potential side effects of dual FXIa/PKal inhibitors. Unfortunately, due to the high homology between FXIa and PKal, it is very challenging to achieve a high selectivity for FXIa over PKal (selectivity index (SI) > 500).…”