LDHA/lactate dehydrogenase A (Ki =8 μM)

Acetone-CoA pyruvylase, which is the substrate of FX-11 (9 μM) [2], is phosphorylated to demonstrate the activation of AMP. In P493 cells, FX-11 suppresses glycolysis and modifies cellular energy supplementing. In BxPc-3 and MIA PaCa-2 cells, FX-11 (0-100 μM, 72 h) limits cell growth [3].

FX-11 (42 μg/mouse; IP, once daily for 10–14 days) suppresses the formation of P493 tumors [2]. FX-11 (0–2 mg/kg, IP, once daily for three weeks) Standing

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In this study, researchers investigated whether the PKM2 activator, TEPP-46, and the LDHA inhibitor, FX-11, can be combined to inhibit in vitro and in vivo tumor growth in preclinical models of pancreatic cancer. They assessed PKM2 and LDHA expression, enzyme activity, and cell proliferation rate after treatment with TEPP-46, FX-11, or a combination of both. Efficacy was validated in vivo by evaluating tumor growth, PK and LDHA activity in plasma and tumors, and PKM2, LDHA, and Ki-67 expression in tumor tissues following treatment. Dual therapy synergistically inhibited pancreatic cancer cell proliferation and significantly delayed tumor growth in vivo without apparent toxicity. Treatment with TEPP-46 and FX-11 resulted in increased PK and reduced LDHA enzyme activity in plasma and tumor tissues and decreased PKM2 and LDHA expression in tumors, which was reflected by a decrease in tumor volume and proliferation. The targeting of glycolytic enzymes such as PKM2 and LDHA represents a promising therapeutic approach for the treatment of pancreatic cancer.[2]

Western Blot Analysis [2]
Cell Types: P493 Cell
Tested Concentrations: 9 μM
Incubation Duration: 24 hrs (hours), 48 hrs (hours)
Experimental Results: ATP levels diminished, accompanied by activation of AMP kinase and phosphorylation of its substrate acetyl-CoA carboxylase.

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Cell proliferation assay [3]
Cell Types: BxPc-3 and MIA PaCa-2 Cell
Tested Concentrations: 0-100 µM
Incubation Duration: 72 hrs (hours)
Experimental Results: diminished cell metabolic activity in a concentration-dependent manner, showing significant reduction in cell proliferation, BxPc- The IC50 values for 3 and MIA PaCa-2 cells were 49.27 µM and 60.54 µM, respectively.

Animal/Disease Models: Male SCID (severe combined immunodeficient) mouse and RH-Foxn1nu (nude) mice (human P493 B cell xenografts) [2]
Doses: 42 μg /mouse (2.1 mg/kg)
Route of Administration: IP; delays tumor growth [3]. one time/day for 10-14 days.
Experimental Results: Significant inhibition of tumor growth and inhibition of tumor xenograft progression.

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Animal/Disease Models: Immunocompromised CD-1 mice (6-8 weeks; 20-25 g, n=5 per group) [3]
Doses: 2 mg/kg, 1 mg/kg+15 mg/kg TEPP- 46. 2 mg/kg+30 mg/kg TEPP-46
Route of Administration: intraperitoneal (ip) injection (100 µL), daily, for 3 weeks
Experimental Results: LDHA activity in plasma and tumor lysates was Dramatically diminished; proliferation markers were Dramatically diminished The expression of Ki-67; a significant decrease in proliferation index was observed in tumor sections; and a significant delay in tumor growth.

[1]. EC Calvaresi. Small molecule inhibitors of lactate dehydrogenase a as an anticancer strategy. 2014.

[2]. Inhibition of lactate dehydrogenase A induces oxidative stress and inhibits tumor progression. Proc Natl Acad Sci U S A. 2010 Feb 2;107(5):2037-42.

[3]. Targeting Pyruvate Kinase M2 and Lactate Dehydrogenase A Is an Effective Combination Strategy for the Treatment of Pancreatic Cancer. Cancers (Basel). 2019 Sep 16;11(9):1372.

As the result of genetic alterations and tumor hypoxia, many cancer cells avidly take up glucose and generate lactate through lactate dehydrogenase A (LDHA), which is encoded by a target gene of c-Myc and hypoxia-inducible factor (HIF-1). Previous studies with reduction of LDHA expression indicate that LDHA is involved in tumor initiation, but its role in tumor maintenance and progression has not been established. Furthermore, how reduction of LDHA expression by interference or antisense RNA inhibits tumorigenesis is not well understood. Here, we report that reduction of LDHA by siRNA or its inhibition by a small-molecule inhibitor (FX11 [3-dihydroxy-6-methyl-7-(phenylmethyl)-4-propylnaphthalene-1-carboxylic acid]) reduced ATP levels and induced significant oxidative stress and cell death that could be partially reversed by the antioxidant N-acetylcysteine. Furthermore, we document that FX11 inhibited the progression of sizable human lymphoma and pancreatic cancer xenografts. When used in combination with the NAD(+) synthesis inhibitor FK866, FX11 induced lymphoma regression. Hence, inhibition of LDHA with FX11 is an achievable and tolerable treatment for LDHA-dependent tumors. Our studies document a therapeutical approach to the Warburg effect and demonstrate that oxidative stress and metabolic phenotyping of cancers are critical aspects of cancer biology to consider for the therapeutical targeting of cancer energy metabolism.[2]

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Exploiting cancer cell metabolism as an anticancer therapeutic strategy has garnered much attention in recent years. As early as the 1920s, German scientist Otto Warburg observed cancer tissues’ avid glucose consumption and high rates of aerobic glycolysis, a phenomenon now known as the Warburg effect. Today, we understand the Warburg effect is mediated by a number of complex factors, including overexpression of the insulin-independent glucose transporter GLUT-1 and overexpression of various glycolytic enzymes, including lactate dehydrogenase A (LDH-A). As the terminal enzyme of glycolysis, LDH-A catalyzes the reversible conversion of pyruvate to lactate, and in doing so, oxidizes NADH to NAD+ . The lactate produced by this reaction is largely excreted into the tumor microenvironment, where it acidifies surrounding tissues and helps the tumor evade destruction by immune cells. The oxidation of NADH to NAD+ allows for continued ATP production through glycolysis by replenishing NAD+ in the absence, or reduced function, of oxidative metabolism. Cell culture and in vivo studies of LDH-A knockdown (using RNA interference) have been shown to lead to substantial decreases in cell and tumor proliferation, thus providing evidence that LDH-A would be a viable anticancer target. While various in vitro LDH-A inhibitors exist, there is a need for a potent and selective small molecule inhibitor that functions both in cells and in vivo. Here, the development and biological assessment of the N-hydroxyindole class of LDH-A inhibitors, including a series of novel dual-Warburg targeting glucose-conjugated LDH-A inhibitors, developed through a collaboration between the Hergenrother and Minutolo laboratories, is reported. The development of novel assays to assess the relative cell uptake, cell lactate production, and competition with 13C glucose for cellular entry, of NHI series compounds are also discussed. Head-to-head cellular assessments of the most promising NHI series compounds alongside literature-reported in vitro inhibitors of LDH-A are reported. Finally, efforts to directly probe the interactions of compounds with LDH-A in cell lysate and whole cells using CETSA and DARTS techniques are discussed[1]

C22H22O4MO

350.4077

350.152

C, 75.41; H, 6.33; O, 18.26

213971-34-7

10498042

White to off-white solid powder

4.8

3

4

5

26

473

0

LVPYVYFMCKYFCZ-UHFFFAOYSA-N

InChI=1S/C22H22O4/c1-3-7-16-17-10-13(2)15(11-14-8-5-4-6-9-14)12-18(17)19(22(25)26)21(24)20(16)23/h4-6,8-10,12,23-24H,3,7,11H2,1-2H3,(H,25,26)

7-Benzyl-2,3-dihydroxy-6-methyl-4-propylnaphthalene-1-carboxylic acid

FX 11; FX11; LDHA Inhibitor FX11; FX11; 7-benzyl-2,3-dihydroxy-6-methyl-4-propyl-naphthalene-1-carboxylic Acid; 2,3-Dihydroxy-6-methyl-7-(phenylmethyl)-4-propyl-1-naphthalenecarboxylic Acid; CHEMBL126519; 7-benzyl-2,3-dihydroxy-6-methyl-4-propylnaphthalene-1-carboxylic acid; FX-11

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~250 mg/mL (~713.45 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (5.94 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (5.94 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)