Structure simplification of GNE-140 and discovery of a pyridone based analogue as a potential lead for hLDHA inhibitors

doi:10.21203/rs.3.rs-3807509/v1

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Structure simplification of GNE-140 and discovery of a pyridone based analogue as a potential lead for hLDHA inhibitors

https://doi.org/10.21203/rs.3.rs-3807509/v1

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In this paper, taking GNE-140, a structurally complicated hLDHA inhibitor, as a lead compound, 19 simple analogues with a pyrimidone, pyranone or pyridone core were designed and conveniently synthesized. Subsequent biological evaluation indicated that although the pyrimidone core had a weak contribution to hLDHA inhibition, compounds with pyranone or pyridone core exhibited better anti-hLDHA activity. Notably, pyridone-bearing compound 1c showed significant selective inhibition of hLDHA (IC₅₀ = 19.5 ± 2.9 µM) over hLDHB (IC₅₀ = 117.6 ± 13.2 µM). Meanwhile, 1c showed inhibition on the lactate production of lung cancer cell A549. Thus, the result of this study indicated 1c is worthy of further study owing to its convenient synthesis and strong selective hLDHA inhibition.

hLDHA inhibitor

GNE-140

Pyrimidone

Pyranone

Pyridone

Human lactate dehydrogenase A (hLDHA), the critical enzyme in the last step of glycolysis catalyzing the conversion of pyruvate to lactate, is overexpressed in a variety of human cancer tissues and promotes acidification of extracellular matrix (ECM), tumor invasion and metastasis^1–4. Thus, hLDHA is considered as a promising target for the development of antitumor drugs^5–8. At present, many hLDHA inhibitors have been discovered^9–14. However, most of them suffer from either weak anti-tumor proliferative activities or poor pharmacokinetic properties or no selective inhibition of hLDHA over hLDHB^15–16. GNE-140 (Fig. 1, top) is a strong hLDHA inhibitor (IC₅₀ = 3 nM) discovered by Purkey in 2016 and entered clinical trial Phase I/II¹⁴. GNE-140 can inhibit the proliferation and lactate production of MiaPaCa-2 cells at low concentration, and inhibit tumor growth in MiaPaCa-2 mice xenotransplantation model.¹⁷ However, GNE-140 with a complicated chemical structure and large molecular weight (M.W.: 499.04) may lead to its unsatisfactory ADME and pharmacokinetic properties¹⁸. In addition, due to the complex chiral dihydropyridone based structure, synthesis of GNE-140 is relatively difficult, which hinders its further structure activity relationship exploration.

The cocrystal structure of GNE-140 and hLDHA (PDB: 4ZVV) revealed the dihydropyridone core of GNE-140 formed critical multi-hydrogen-bonds with amino acids Arg168, His192 and Thr247 of hLDHA (Fig. 1a). By molecular modeling, we found that much simpler pyrimidone, pyranone or pyridone based analogues would maintain these main interactions and most likely to possess significant hLDHA inhibiting ability (Fig. 1b-d). Thus, herein 19 compounds were designed, conveniently synthesized, and evaluated for its hLDHA inhibiting activity.

Chemistry

As shown in Scheme 1, compounds 1a, 2a, 8a and 9a with pyrimidinone core were easily obtained by a two-step manipulation, with readily available materials.

But compounds 3a and 4a failed to give decent yields with the same synthetic strategy. Alternatively, 3a-7a, and 10a were synthesized through coupling of diphenyldisulfane with simple pyrimidone intermediates 1, 12 (Scheme 2).

Compounds with pyranone core (1b-3b) and pyridone core (1c-2c) were also smoothly synthesized using similar coupling reactions of diphenyldisulfane 4, 5 with pyranone intermediate 13, 14 and pyridone intermediate 15, 16, respectively (scheme 3).

hLDHA and hLDHB Inhibiting activity

hLDHA inhibiting activity of these analogues were evaluated according to the method by Cao et al¹⁹ and the results are listed in Table 1. Firstly, pyrimidinone-bearing compounds 1, 12 and 1a-10a (entries 1–12) showed weak hLDHA inhibiting activity (less than 50% inhibition at 100µM). Secondly, pyranone-bearing compounds 1b-3b showed better hLDHA inhibiting activity (entries 13–15). However, these compounds only showed slightly selective inhibition of hLDHA over hLDHB. Finally, to our delight, although the pyridone-bearing compound 2c (entry 17) with a chlorine substitution at the C3-position showed low hLDHA inhibiting activity, 1c (entry 16) showed both strong inhibition on hLDHA (IC₅₀ = 19.5 ± 2.7 µM) and selectively over hLDHB (IC₅₀ = 117.6 ± 13.2 µM).

Table 1

anti-hLDHA and anti-hLDHB activities of synthesized compounds.
Entry	Compds	hLDH inhibiting rate (%) at 100 µM		IC₅₀ (µM)
Entry	Compds	hLDHA	hLDHB	hLDHA	hLDHB
1	1	17.6	18.9	> 100	> 100
2	12	14.6	29.9	> 100	> 100
3	1a	40.3	39.8	> 100	> 100
4	2a	31.0	37.2	> 100	> 100
5	3a	34.6	25.1	> 100	> 100
6	4a	27.6	35.1	> 100	> 100
7	5a	32.4	45.7	> 100	> 100
8	6a	32.6	44.1	> 100	> 100
9	7a	31.8	46.8	> 100	> 100
10	8a	43.1	36.6	> 100	> 100
11	9a	37.3	39.3	> 100	> 100
12	10a	22.5	22.6	> 100	> 100
13	1b	87.7	74.9	13.6 ± 1.4	27.0 ± 6.5
14	2b	45.8	36.6	> 100	> 100
15	3b	100	77.2	12.3 ± 1.8	21.9 ± 4.1
16	1c	100	45.5	19.5 ± 2.7	117.6 ± 13.2
17	2c	43.7	31.3	> 100	> 100
18	GNE-140	88.5 ^a	57.8 ^a	0.0022	0.0121

^a hLDHA and hLDHB inhibiting rates (%) at 10 nM.

However, although compound 1c showed potent hLDHA inhibitory activity, there is still a big gap to that of GNE-140. Considering of the easy preparation and derivatization, pridone-core based analogues still has a large room for structural optimization, which deserves further study.

Lactate production inhibiting activity

hLDHA is the key enzyme in glycolysis, catalyzing the conversion of pyruvate to lactate. Thus, compounds 1b, 3b and 1c with good hLDHA inhibiting activity were subjected to lactate production inhibiting test using human lung cancer cell line A549. As shown in Fig. 2, compound 1b, 3b and especially 1c significantly inhibited lactate formation of human lung cancer A549 cells at concentrations of 50 and 25 µM.

In this paper, the potent but structurally complicated hLDHA inhibitor GNE-140 was used as a lead compound, and a total of 19 compounds with simple pyrimidone, pyranone or pyridone core were designed, synthesized and evaluated for hLDHA/hLDHB inhibiting activity. It’s worth noting that the pyridone-bearing compound 1c showed selective inhibition of hLDHA over hLDHB and good inhibition on lactate production in lung cancer A549 cells. Considering of its simple structure and decent hLDHA inhibiting activity, 1c is worthy of further exploration as a potential lead compound.

Acknowledgements

This work was supported by the grants from the Natural Science Foundation of Hunan Province (No. 2021JJ30903), the National Natural Science Foundation of China (No. 22177137) and Central South University Research Program of Advanced Interdisciplinary Studies (Grant No. 2023QYJC032).

Data availability

The data that support the findings of this study are available in the supplementary material of this article.

Conflict of interest

The authors declare no competing interests.

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Scheme 1 to 3 are available in the Supplementary Files section.

GA.png
S1.png
Scheme 1. Syntheses of compounds 1a, 2a, 8a and 9a
S2.png
Scheme 2. Syntheses of compounds 3a-7a and 10a
S3.png
Scheme 3. Syntheses of compounds 1b-3b and1c-2c
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Structure simplification of GNE-140 and discovery of a pyridone based analogue as a potential lead for hLDHA inhibitors

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Abstract

Figures

Introduction

Results and discussion

Chemistry

hLDHA and hLDHB Inhibiting activity

Lactate production inhibiting activity

Conclusion

Declarations

References

Scheme

Supplementary Files

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