1,2,3,4,6-Penta-O-galloylglucose within Galla Chinensis Inhibits Human LDH-A and Attenuates Cell Proliferation in MDA-MB-231 Breast Cancer Cells

Abstract

A characteristic feature of aggressive malignancy is the overexpression of lactic acid dehydrogenase- (LDH-) A, concomitant to pericellular accumulation of lactate. In a recent high-throughput screening, we identified Rhus chinensis (Mill.) gallnut (RCG) (also known as Galla Chinensis) extract as a potent (IC50 < 1 µg/mL) inhibitor of human LDH-A (hLDH-A). In this study, through bioactivity guided fractionation of the crude extract, the data demonstrate that penta-1,2,3,4,6-O-galloyl-β-D-glucose (PGG) was a primary constituent responsible for hLDH-A inhibition, present at ~9.95 ± 0.34% dry weight. Theoretical molecular docking studies of hLDH-A indicate that PGG acts through competitive binding at the NADH cofactor site, effects confirmed by functional enzyme studies where the IC50 = 27.32 nM was reversed with increasing concentration of NADH. Moreover, we confirm protein expression of hLDH-A in MDA-231 human breast carcinoma cells and show that PGG was toxic (LC50 = 94.18 µM), parallel to attenuated lactic acid production (IC50 = 97.81 µM). In a 72-hour cell proliferation assay, PGG was found to be a potent cytostatic agent with ability to halt cell division (IC50 = 1.2 µM) relative to paclitaxel (IC50 < 100 nM). In summary, these findings demonstrate that PGG is a potent hLDH-A inhibitor with significant capacity to halt proliferation of human breast cancer cells.

Stable shRNA Silencing of Lactate Dehydrogenase A (LDHA) in Human MDA-MB-231 Breast Cancer Cells Fails to Alter Lactic Acid Production, Glycolytic Activity, ATP or Survival

“…From the existing data, it was not predicted that we should see absolutely no change in any metabolic parameter tested by a near complete silencing of LDHA, however this could account for similar observation in our previous work in which we identified PGG as a potent LDHA inhibitor, having no influence on the production of lactic acid itself (18). Future studies must be carried out to determine the effects of a double knockdown of LDHA/B to find out specifically how this protein (all isoforms) controls pivotal events in metabolism and production of acid.…”

“…Further analysis showed that 1,2,3,4,6-Penta-O-galloylglucose was the compound in Chinese gallnut with capacity to inhibit LDHA in the nM range (18). The latter study confirmed that LDHA inhibitors seem to slow the growth or cell division of TNBC cells, but we did not find concrete evidence to show that production of lactate was hampered, nor did we find evidence of toxicity.…”

Gallnuts: A Potential Treasure in Anticancer Drug Discovery

“…According to the literature reported ( Figure 3 ), twelve chemical components have been identified from gallnuts to the date, including gallic acid (1), syringic acid (2), ellagic acid (3), methyl gallate (4), β -sitosterol (5), 1,2,3,4,6-penta- O -galloyl- β -D-glucose (6), 1,2,3,4,6-tetra- O -galloyl- β -D-glucose (7), amentoflavone (8), purpurogallin (9), hexamethyl ether (10), isocryptomerin (11), and methyl betulate (12) [ 22 , 32 , 39 – 42 ]. Among them, gallic acid (1), ellagic acid (3), methyl gallate (4), β -sitosterol (5), 1,2,3,4,6-penta- O -galloyl- β -D-glucose (6), amentoflavone (8), and purpurogallin (9) have been reported to present anticancer activities in either in vitro or in vivo studies [ 22 , 23 , 34 , 43 – 48 ]. However, the current researches of identification and isolation of gallnuts compounds were very few and mainly focused on the gallotannin only.…”

The attenuating effects of 1,2,3,4,6 penta-O-galloyl-β- d -glucose on inflammatory cytokines release from activated BV-2 microglial cells

“…Studies have shown that PGG exhibits an anti-inflammatory effect through the inhibition of the expression of many cytokines, such as TNF-α, IL-6, and IL-8 in different cell models (Oh et al, 2004; Feldman et al, 2011;Wu and Gu, 2009). PGG presented an anti-cancer potential by inhibiting the enzyme hLDH-A (Deiab et al, 2015), and inhibiting epidermal growth factor (EGF)-induced cell invasiveness and MMP-9 expression in a dose- and time-dependent manner by reducing the MMP-9 transcriptional activity, suggesting that PGG may be a therapeutic drug candidate for the treatment of advanced metastatic prostate cancer (Zhang et al, 2009). …”

“…PGG has been shown to exhibit anti-cancer effects, anti-angiogenesis and anti-diabetic activities (Yu et al, 2011; Bruno et al, 2013; Hu et al, 2008; Zhang et al, 2009). Also, in our laboratory we were able to establish that PGG is a potent inhibitor for hLDH-A, which is an anti-cancer therapeutic target with an established role in tumor initiation, progression, and metastasis (Deiab et al, 2015). …”