Inactivators of Ornithine Aminotransferase for the Treatment of Hepatocellular Carcinoma

Abstract

Hepatocellular carcinoma (HCC) is the second or third leading cause of cancer mortality worldwide (depending on which statistics are used), yet there is no effective treatment. Currently, there are nine FDA-approved drugs for HCC, five monoclonal antibodies and four tyrosine kinase inhibitors. Ornithine aminotransferase (OAT) has been validated as a target in preclinical studies, which demonstrates that it is a potential target to treat HCC. Currently, there are no OAT inactivators in clinical trials for HCC. This Innovation describes evidence to support inhibition of OAT as a novel approach for HCC tumor growth inhibition. After the mechanism of OAT is discussed, the origins of our involvement in OAT inactivation, based on our previous work on mechanism-based inactivation of GABA-AT, are described. Once it was demonstrated that OAT inactivation does lead to HCC tumor growth inhibition, new selective OAT inactivators were designed and their inactivation mechanisms were elucidated. A summary of these mechanistic studies is presented. Inactivators of OAT provide the potential for treatment of HCC, targeting the Wnt/β-catenin pathway.

Figure 1

Binding of Wnt to the Frizzled-LRP5/6 co-receptor allows β-catenin to be translocated into the nucleus, where it binds to TCF/LEF transcription factors, leading to transcription of Wnt target genes.

Scheme 1. Conversion of l-Ornithine to l-Glutamate Catalyzed by Ornithine Aminotransferase (OAT)

Figure 2

GABA analogues screened for OAT activity. Figure modified from ref (35). Copyright 2015 American Chemical Society.

Figure 3

(A) Administration of 5 inhibits serum AFP secretion in vivo. Mice were treated for 27 days, 3 times a week, starting 3 weeks following HCC transplantation with two doses of 1 (0.1 mg/kg [2 μg], green bars or 1 mg/kg [20 μg], red bars), compared with untreated controls (blue bars). Levels are normalized to the starting day of therapy. (B) Compound 5 diminishes tumor volume in both treated groups (0.1 mg/kg [2 μg], green bars and 1.0 mg/kg [20 μg], red bars) compared to untreated controls (blue bars). Figure reproduced from ref (35). Copyright 2015 American Chemical Society.

Scheme 2. Inactivation Mechanism for 5 with Human Ornithine Aminotransferase

Scheme reproduced from ref (45). Copyright 2019 American Chemical Society.

Figure 4

X-ray crystal structure of OAT following inactivation by 5 (PDB code: 6OIA). Figure reproduced from ref (45). Copyright 2019 American Chemical Society.

Scheme 3. Mechanism of Inactivation of OAT by 5-Fluoromethylornithine (18)

Scheme 4. Mechanism Proposed for the Inactivation of OAT by 20

Scheme modified from ref (61). Copyright 2021 American Chemical Society.

Figure 5

(A) Crystal structure of OAT resulting from soaking of 20 over 1 h, shown in two alternate conformations (beige): one in which the carboxylate group interacts with Tyr55 (conformation A) and the other in which the carboxylate forms a salt bridge with Arg413 (conformation B); PDB code: 7LK1. (B) Co-crystal structure of OAT with 20; PDB code: 7LK0. Carbon atoms in the residues are colored gray, nitrogen in dark blue, and oxygen in red; the water molecule is shown as a red sphere. Hydrogen bonding distances between atoms are in Ångstroms (Å) and are shown as black dashed lines. Figure reproduced ref (61). Copyright 2021 American Chemical Society.

Figure 6

Crystal structures of 21 (PDB code: 6 V8D) and 22 (PDB code: 6 V8C) cocrystallized with OAT. Amino acid residues are in silver; compounds bound to PLP are in gold; oxygen is red, and nitrogen is blue. Figure reproduced from ref (62). Copyright 2020 American Chemical Society.

Figure 7

(A) X-ray crystal structure of first intermediate (50) from soaking 23 into OAT; PDB code: 7LOM. (B) X-ray cocrystal structure of double covalently bonded 23 (52) with Thr322 and Lys292 in OAT; PDB code: 7LON. Figure reproduced from ref (63). Copyright 2021 American Chemical Society.

Scheme 5. Possible Inactivation Mechanisms by 21 and 22

Scheme reproduced from ref (62). Copyright 2020 American Chemical Society.

Scheme 6. Plausible turnover and Inactivation Mechanisms for the Inactivation of OAT by 23

Scheme modified from ref (63). Copyright 2021 American Chemical Society.