Design, synthesis and biological evaluation of novel 2,3-dihydroquinazolin- 4(1H)-one derivatives as potential fXa inhibitors

Highlights

• Three series of 2,3-dihydroquinazolin-4(1H)-one derivatives were prepared.

• Most of the compounds had significant fXa inhibitory activity.

• 8e exhibited the strongest potency against fXa and good selectivity versus thrombin.

• 8e also displayed great in vitro and in vivo antithrombotic activity.

• 8e had a similar safety profile on bleeding risk as that of betrixaban.

Abstract

Coagulation factor Xa (fXa) is a particularly attractive target for the development of effective and safe anticoagulants. In this study, novel 2,3-dihydroquinazolin-4(1H)-one derivatives were designed as potential fXa inhibitors based on anthranilamide structure which has been reported in our previous research. The experimental data showed that most of the designed compounds exhibited significant in vitro potency against fXa. Among them, compound 8e displayed the strongest potency against fXa with the IC₅₀ value of 21 nM and highly selectivity versus thrombin (IC₅₀ = 67 μM) and excellent in vitro antithrombotic activity with its 2 × PT value of 1.2 μM and 2 × aPTT value of 0.6 μM. In addition, 8e also displayed excellent in vivo antithrombotic activity in the rat arteriovenous shunt (AV-SHUNT) model. The bleeding risk evaluation showed that 8e had a similar safety profile as that of betrixaban. All results demonstrated that compound 8e could be considered as a potential fXa inhibitor for the prevention and treatment of thromboembolic diseases.

Introduction

Thrombotic disorders, including ischemic stroke, unstable angina, pulmonary embolism (PE), deep venous thrombosis (DVT) and myocardial infarction, constitute a major cause of morbidity and mortality worldwide [1], [2]. Although several anticoagulants such as fondaparinux, warfarin, low molecular weight heparins (LMWHs) and unfractionated heparin are effective in the prevention and treatment of these thrombotic diseases, they also display several undesirable effects [3], [4], [5]. In the past decades, numerous research efforts have been made to identify novel anticoagulants with improved efficacy and safety. After extensive investigation, thrombin (fIIa) and fXa have become the promising targets for the development of potent, selective, and orally bioavailable anticoagulants [6], [7], [8].

Activated serine protease fXa, which is situated at the common pathway of coagulation cascade, catalyzes the conversion of prothrombin to thrombin and plays a central role in various thromboembolic disorders [9], [10]. Compared with thrombin inhibitors, selective fXa inhibitors have better specific antithrombotic effects and lower risk of bleeding due to their ability to reduce the further generation of thrombin without affecting the existing level of thrombin [11]. Currently, there are three oral, direct and selective fXa inhibitors approved by the Food and Drug Administration (FDA): rivaroxaban [12], [13], apixaban [14], [15] and edoxaban [16] (Fig. 1). These novel fXa inhibitors exhibit advantages compared with traditional anticoagulants. However, they still have several disadvantages with the most notable being their narrow indications for use and the lack of an effective antidote for when significant bleeding occurs [17], [18], [19], [20], [21]. The development of an antidote for use with fXa inhibitors is being investigated and would address some of the concerns with the fXa inhibitors, but it would not address the safety and efficacy of the inhibitors themselves. [22], [23], [24], Therefore, the further development of novel and safer fXa inhibitors is desirable to increase use of fXa inhibitors in the clinic.

Most fXa inhibitors bind to the active site in a characteristic L-shaped conformation [8]. They have a three-component system which includes a central scaffold and two hydrophobic fragments (P1 and P4) that provide a non-linear geometry considered to play a crucial role in fXa recognition [25]. The core linkers studied to date can be classified into either highly flexible class or fairly rigid class. In our previous study, we have identified a series of anthranilamide derivatives as fXa inhibitors by a fragment-based strategy integrated with structure-based key pharmacophore and structural consensus docking [26]. Based on the anthranilamide scaffold and structure–activity analysis, novel 2,3-dihydroquinazolin-4(1H)- one as a fairly rigid linker was designed and its derivatives were synthesized and evaluated for in vitro and in vivo antithrombotic potency. Additionally, the compounds reported in this study have a similar structure to anti-tumor molecules designed by our group (unpublished results). Cancer is known to induce a hypercoagulable state, which is a major risk factor for VTE [27], [28], [29], [30]. Thus, compounds possessing both anticoagulant and antitumor activities are desirable. Although this report focused on assessing the anticoagulant properties of these molecules, their anti-tumor activity is also being evaluated by our lab and will be reported in due course.