**4.1. Direct synthetic FXa inhibitors**

Indirect FXa inhibitor development led to the advance of direct oral FXa inhibitors, such as rivaroxaban [100] (**Figure 7c**) and apixaban [98] (**Figure 7d**). Interestingly, the "Xa" suffix comes from FXa and "ban" indicating inhibition [101].

These latest FXa direct oral inhibitors comprised a group of small molecules. Taking into account the chronological order, these new direct oral anticoagulants (DOACs) are razaxaban, rivaroxaban, apixaban, darexaban, edoxaban, and betrixaban; however, some of them did not obtain the approval during clinical trials and they are not commercialized [48].

## *4.1.1. Razaxaban*

This novel FXa synthetic and orally active compound was developed by Bristol Myers Squibb in 2004. The production of razaxaban was carried out through a seven synthetic pathway (**Figure 9**) [56]. It acts as a selective and reversible direct FXa inhibitor which was the first synthetic direct FXa inhibitor developed. Moreover, razaxaban has demonstrated FXa selectivity in venous thrombosis in human beings and arterial thrombosis prevention in animal models [94].

novel synthetic route was design by using only a seven-step procedure diminishing the environmental impact and increasing the reaction yields (**Figure 11**) [103]. This novel DOAC is a selective and reversible FXa inhibitor which shows 100-fold greater selectivity for FXa over any other serine protease. Rivaroxaban inhibits the complex between FXa and prothrombinase with an IC50 2.1 nM; moreover, it shows nanomolar inhibitory constant [Ki = 0.4 nM] [33, 93, 104]. As it is shown in **Figure 12**, the two-ringed moiety, including the morpholinone and bencenic moieties, produced S4 hydrophobic interactions with Phe174 and Tyr99 residues. Moreover, the oxazolidone ring interacts with Gly219 through hydrogen bonds and the chlorothiophene moiety produces necessary interactions with Asp189, Ala190 and Tyr228 in the profound S1

**Figure 10.** Razaxaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are

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This first commercially available DOAC doesn't show food interactions and it is prescribed as one dose-per-day drug after heart attack or stroke [105, 106]. Furthermore, rivaroxaban was

**Figure 11.** Chemical structures: Commercial starting material and rivaroxaban. The moiety that interacts with S1 is

site (**Figure 12**) [33, 68, 82].

labeled.

approved for prophylaxis after knee or hip surgery [96].

shown in blue and the portion involved in the S4 interaction is shown in red [594].

The razaxaban structural L shape allowed it to fit within the FXa S1 pocket where the nitrogen atom of the benzoisoxazole moiety interacts with Ala190 and Asp189 (**Figure 10**) [48, 100]. Moreover, the development was discontinued in Phase II at the end of 2004.

#### *4.1.2. Rivaroxaban*

Rivaroxaban (Xarelto®), was the first direct oral FXa inhibitor developed by Bayer Schering Pharma AG and it obtained the clinical approved in 2008 [102]. At the beginning, the production of rivaroxaban was carried out through a nine synthetic pathway [68]. In recent years, a

**Figure 9.** Chemical structures: Commercial starting material and Razaxaban. The moiety that interacts with S1, is shown in blue, and the portion involved in the S4 interaction is shown in red [50].

**Figure 10.** Razaxaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are labeled.

novel synthetic route was design by using only a seven-step procedure diminishing the environmental impact and increasing the reaction yields (**Figure 11**) [103]. This novel DOAC is a selective and reversible FXa inhibitor which shows 100-fold greater selectivity for FXa over any other serine protease. Rivaroxaban inhibits the complex between FXa and prothrombinase with an IC50 2.1 nM; moreover, it shows nanomolar inhibitory constant [Ki = 0.4 nM] [33, 93, 104].

As it is shown in **Figure 12**, the two-ringed moiety, including the morpholinone and bencenic moieties, produced S4 hydrophobic interactions with Phe174 and Tyr99 residues. Moreover, the oxazolidone ring interacts with Gly219 through hydrogen bonds and the chlorothiophene moiety produces necessary interactions with Asp189, Ala190 and Tyr228 in the profound S1 site (**Figure 12**) [33, 68, 82].

This first commercially available DOAC doesn't show food interactions and it is prescribed as one dose-per-day drug after heart attack or stroke [105, 106]. Furthermore, rivaroxaban was approved for prophylaxis after knee or hip surgery [96].

**Figure 11.** Chemical structures: Commercial starting material and rivaroxaban. The moiety that interacts with S1 is shown in blue and the portion involved in the S4 interaction is shown in red [594].

**Figure 9.** Chemical structures: Commercial starting material and Razaxaban. The moiety that interacts with S1, is shown

approved by the Food and Drug Administration (FDA) in December 2012 with an indication of reducing the risk of stroke and dangerous blood clots (systemic embolism) in patients with

Based on these discoveries, in the mid-1990s, it was assumed that small-molecule, direct factor Xa inhibitors could most likely become a better option than the antithrombotic therapies

Indirect FXa inhibitor development led to the advance of direct oral FXa inhibitors, such as rivaroxaban [100] (**Figure 7c**) and apixaban [98] (**Figure 7d**). Interestingly, the "Xa" suffix

These latest FXa direct oral inhibitors comprised a group of small molecules. Taking into account the chronological order, these new direct oral anticoagulants (DOACs) are razaxaban, rivaroxaban, apixaban, darexaban, edoxaban, and betrixaban; however, some of them did not obtain the approval during clinical trials and they are not commercialized [48].

This novel FXa synthetic and orally active compound was developed by Bristol Myers Squibb in 2004. The production of razaxaban was carried out through a seven synthetic pathway (**Figure 9**) [56]. It acts as a selective and reversible direct FXa inhibitor which was the first synthetic direct FXa inhibitor developed. Moreover, razaxaban has demonstrated FXa selectivity in venous thrombosis in human beings and arterial thrombosis prevention in animal models [94]. The razaxaban structural L shape allowed it to fit within the FXa S1 pocket where the nitrogen atom of the benzoisoxazole moiety interacts with Ala190 and Asp189 (**Figure 10**) [48, 100].

Rivaroxaban (Xarelto®), was the first direct oral FXa inhibitor developed by Bayer Schering Pharma AG and it obtained the clinical approved in 2008 [102]. At the beginning, the production of rivaroxaban was carried out through a nine synthetic pathway [68]. In recent years, a

Moreover, the development was discontinued in Phase II at the end of 2004.

atrial fibrillation (AF) [95–98].

**4.1. Direct synthetic FXa inhibitors**

comes from FXa and "ban" indicating inhibition [101].

used in those days [99].

22 Anticoagulant Drugs

*4.1.1. Razaxaban*

*4.1.2. Rivaroxaban*

in blue, and the portion involved in the S4 interaction is shown in red [50].

**Figure 12.** Rivaroxaban bound to FXa (PDB ID 2w26) [68]. The binding site is shown in surface mode. Specificity sites and important residues are labeled.

#### *4.1.3. Apixaban*

Apixaban (Eliquis®) was the second FXa oral inhibitor approved by the European Medicines Agency (EMA) in 2011 and by the Food and Drug Administration (FDA) in 2012 [98]. This novel molecule is a design evolution of razaxaban, and it was developed by Bristol Myers Squibb [53]. Moreover, apixaban is a reversible and selective FXa inhibitor which is prescribed in thromboembolic prophylaxis events such as preventing thrombus production and strokes in persons with atrial fibrillation. Furthermore, apixaban is prescribed to prevent blood clots in deep vein thrombosis (DVT) and pulmonary embolus formation according to the United States regulations [52, 107, 108].

Currently, apixaban has FXa inhibitory activity showing a Ki = 0.08 nM with 50% oral bioavailability [104, 107]. In addition, its action showed selectivity for clot-bound (IC50 1.3 nM) vs. free FXa (IC50 7.6 nM) [107]. Besides, this DOAC induces hepatotoxicity as its adverse effect (**Figure 13**) [109].

It showed an inhibition constant Ki 0.031 μM and IC50 40 nM for free FXa and IC50 80 nM for blood clots [111]. However, darexaban development was discontinued in September 2011, after a phase II in Australia, Canada, and the European Union (EU) because the clinical trial showed that the combination of darexaban with an antiplatelet agent such as acetyl salicylic acid (ASA) caused a fourfold increase in bleeding rates and had no effect on acute coronary

**Figure 14.** Apixaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are

**Figure 13.** Chemical structures: Commercial starting material and apixaban. The moiety that interacts with S1 is shown

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in blue, and the portion involved in the S4 interaction is shown in red [53].

Darexaban establishes the same interactions with Asp189, Ala190, and Tyr228 in the S1 pocket

Edoxaban (Savaysa® in USA and Lixiana ® in Canada and outside the USA) was developed by Daiichi Sankyo and it was approved in Japan (2011) and by the FDA (2015) [71, 113]. This

syndrome (ACS) (**Figure 15**) [112].

as other DOACs (**Figure 16**) [100].

*4.1.5. Edoxaban*

labeled.

The characteristic FXa inhibitors L shape is produced by the peptide bond present between the two ring pyrazole linked to a phenyl piperidinone (**Figure 14**). Apixaban shows the same interactions than Rivaroxaban in the S1 pocket by using the methoxyphenyl portion at the bottom of the S1 pocket [100].

#### *4.1.4. Darexaban*

Darexaban was designed by Astellas Pharma in 2007 for venous and arterial thromboembolic disease prophylaxis such as venous thrombosis, myocardial infarction, and ischemic stroke [110].

**Figure 13.** Chemical structures: Commercial starting material and apixaban. The moiety that interacts with S1 is shown in blue, and the portion involved in the S4 interaction is shown in red [53].

**Figure 14.** Apixaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are labeled.

It showed an inhibition constant Ki 0.031 μM and IC50 40 nM for free FXa and IC50 80 nM for blood clots [111]. However, darexaban development was discontinued in September 2011, after a phase II in Australia, Canada, and the European Union (EU) because the clinical trial showed that the combination of darexaban with an antiplatelet agent such as acetyl salicylic acid (ASA) caused a fourfold increase in bleeding rates and had no effect on acute coronary syndrome (ACS) (**Figure 15**) [112].

Darexaban establishes the same interactions with Asp189, Ala190, and Tyr228 in the S1 pocket as other DOACs (**Figure 16**) [100].

#### *4.1.5. Edoxaban*

*4.1.3. Apixaban*

24 Anticoagulant Drugs

and important residues are labeled.

regulations [52, 107, 108].

bottom of the S1 pocket [100].

(**Figure 13**) [109].

*4.1.4. Darexaban*

Apixaban (Eliquis®) was the second FXa oral inhibitor approved by the European Medicines Agency (EMA) in 2011 and by the Food and Drug Administration (FDA) in 2012 [98]. This novel molecule is a design evolution of razaxaban, and it was developed by Bristol Myers Squibb [53]. Moreover, apixaban is a reversible and selective FXa inhibitor which is prescribed in thromboembolic prophylaxis events such as preventing thrombus production and strokes in persons with atrial fibrillation. Furthermore, apixaban is prescribed to prevent blood clots in deep vein thrombosis (DVT) and pulmonary embolus formation according to the United States

**Figure 12.** Rivaroxaban bound to FXa (PDB ID 2w26) [68]. The binding site is shown in surface mode. Specificity sites

Currently, apixaban has FXa inhibitory activity showing a Ki = 0.08 nM with 50% oral bioavailability [104, 107]. In addition, its action showed selectivity for clot-bound (IC50 1.3 nM) vs. free FXa (IC50 7.6 nM) [107]. Besides, this DOAC induces hepatotoxicity as its adverse effect

The characteristic FXa inhibitors L shape is produced by the peptide bond present between the two ring pyrazole linked to a phenyl piperidinone (**Figure 14**). Apixaban shows the same interactions than Rivaroxaban in the S1 pocket by using the methoxyphenyl portion at the

Darexaban was designed by Astellas Pharma in 2007 for venous and arterial thromboembolic disease prophylaxis such as venous thrombosis, myocardial infarction, and ischemic stroke [110].

Edoxaban (Savaysa® in USA and Lixiana ® in Canada and outside the USA) was developed by Daiichi Sankyo and it was approved in Japan (2011) and by the FDA (2015) [71, 113]. This

**Figure 15.** Darexaban chemical structure. The moiety that interacts with S1 is shown in blue, and the portion involved in the S4 interaction is shown in red.

novel DOAC is used for stroke and VTE prophylaxis in patients with atrial fibrillation [114]. Edoxaban was synthesized through a twelve-step procedure (**Figure 17**) [113].

Edoxaban bioavailability is 62%, and it is prescribed at 15–150 mg daily. It has been shown a nanomolar value for its Ki (0.56 nM) and IC50 (3 nM) [115–117]. Currently, Daiichi Sankyo is developing a phase III trial for cardiovascular disorders during February 2018.

This DOAC interacts with the same amino acidic residues in the S1 serine enzyme pocket as the other FXa direct inhibitors (**Figure 18**). The chloropyridine moiety is responsible for the S1 pocket interaction meanwhile the tetrahydrothiazolo-pyridine moiety interacts with the S4 pocket [100].

### *4.1.6. Betrixaban*

Betrixaban (Bevyxxa®) is the newest DOAC developed by Portola Pharmaceuticals and it was designed through structure activity relationship (SAR) studies [80, 118]. It was approved by the FDA in June 2017 for prevention of venous thromboembolism in acute hospitalized medical adult patients by using an initial single dose of 80 mg (**Figure 19**) [119].

**Figure 16.** Darexaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are labeled.

This new DOAC is a competitive and reversible FXa inhibitor and it has a Ki 0.117 pM and IC50 1.5 nM [119]. Betrixaban may therefore have several potential advantages over the other

**Figure 19.** Chemical structures: commercial starting material and Betrixaban. The moiety that interacts with S1 is shown

**Figure 18.** Edoxaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are

**Figure 17.** Chemical structures: commercial starting material and Edoxaban. The moiety that interacts with S1 is shown

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in blue, and the portion involved in the S4 interaction is shown in red [113].

FXa inhibitors (**Figure 20**).

in blue, and the portion involved in the S4 interaction is shown in red.

labeled.

**Figure 17.** Chemical structures: commercial starting material and Edoxaban. The moiety that interacts with S1 is shown in blue, and the portion involved in the S4 interaction is shown in red [113].

**Figure 15.** Darexaban chemical structure. The moiety that interacts with S1 is shown in blue, and the portion involved

novel DOAC is used for stroke and VTE prophylaxis in patients with atrial fibrillation [114].

Edoxaban bioavailability is 62%, and it is prescribed at 15–150 mg daily. It has been shown a nanomolar value for its Ki (0.56 nM) and IC50 (3 nM) [115–117]. Currently, Daiichi Sankyo is

This DOAC interacts with the same amino acidic residues in the S1 serine enzyme pocket as the other FXa direct inhibitors (**Figure 18**). The chloropyridine moiety is responsible for the S1 pocket interaction meanwhile the tetrahydrothiazolo-pyridine moiety interacts with the S4 pocket [100].

Betrixaban (Bevyxxa®) is the newest DOAC developed by Portola Pharmaceuticals and it was designed through structure activity relationship (SAR) studies [80, 118]. It was approved by the FDA in June 2017 for prevention of venous thromboembolism in acute hospitalized medi-

**Figure 16.** Darexaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are

Edoxaban was synthesized through a twelve-step procedure (**Figure 17**) [113].

developing a phase III trial for cardiovascular disorders during February 2018.

cal adult patients by using an initial single dose of 80 mg (**Figure 19**) [119].

in the S4 interaction is shown in red.

26 Anticoagulant Drugs

*4.1.6. Betrixaban*

labeled.

**Figure 18.** Edoxaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are labeled.

**Figure 19.** Chemical structures: commercial starting material and Betrixaban. The moiety that interacts with S1 is shown in blue, and the portion involved in the S4 interaction is shown in red.

This new DOAC is a competitive and reversible FXa inhibitor and it has a Ki 0.117 pM and IC50 1.5 nM [119]. Betrixaban may therefore have several potential advantages over the other FXa inhibitors (**Figure 20**).

and Medical Engineering. This work was financially supported to F.C.Z. by the CONICYT/ FONDECYT Fondecyt Iniciación N° 11130595 and Fondecyt Regular N° 1181408 project.

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The author would like to express a sincere gratitude to Nicolás E. Núñez-Navarro for his valuable assistance with the bibliographical search and to Fabián M. Santana for the images

[2] Pérez-Gómez F, Bover R. La nueva cascada de la coagulación y su posible influencia en el difícil equilibrio entre trombosis y hemorragia. Revista Española de Cardiología

[3] Riddel J, Aouizerat B, Miaskowski C, Lillicrap D. Theories of blood coagulation. Journal

[4] Smith S. The cell-based model of coagulation. Journal of Veterinary Emergency and

[5] Hoffman M, Monroe D III. A cell-based model of hemostasis. Thrombosis and

[6] Hoffman M, Monroe D. Rethinking the coagulation cascade. Current Hematology

[7] Palta S, Saroa R. Overview of the coagulation system. Indian Journal of Anaesthesia.

[8] Vojacek J. Should we replace the terms intrinsic and extrinsic coagulation pathways with tissue factor pathway? Clinical and Applied Thrombosis/Hemostasis. 2016;**1**:1-6

**Conflict of interest**

**Thanks**

improvement.

**Author details**

Flavia C. Zacconi

**References**

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The author declares no conflict of interest.

Address all correspondence to: fzacconi@uc.cl

Pontificia Universidad Católica de Chile, Santiago de Chile, Chile

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**Figure 20.** Betrixaban bound to FXa (PDB ID 2w26). The binding site is shown in surface mode. Important residues are labeled.
