title: Identification of LDH-A as a therapeutic target for cancer cell killing via (i) p53/NAD(H)-dependent and (ii) p53-independent pathways.
abstract: Most cancer cells use aerobic glycolysis to fuel their growth. The enzyme lactate dehydrogenase-A (LDH-A) is key to cancer's glycolytic phenotype, catalysing the regeneration of nicotinamide adenine dinucleotide (NAD(+)) from reduced nicotinamide adenine dinucleotide (NADH) necessary to sustain glycolysis. As such, LDH-A is a promising target for anticancer therapy. Here we ask if the tumour suppressor p53, a major regulator of cellular metabolism, influences the response of cancer cells to LDH-A suppression. LDH-A knockdown by RNA interference (RNAi) induced cancer cell death in p53 wild-type, mutant and p53-null human cancer cell lines, indicating that endogenous LDH-A promotes cancer cell survival irrespective of cancer cell p53 status. Unexpectedly, however, we uncovered a novel role for p53 in the regulation of cancer cell NAD(+) and its reduced form NADH. Thus, LDH-A silencing by RNAi, or its inhibition using a small-molecule inhibitor, resulted in a p53-dependent increase in the cancer cell ratio of NADH:NAD(+). This effect was specific for p53(+/+) cancer cells and correlated with (i) reduced activity of NAD(+)-dependent deacetylase sirtuin 1 (SIRT1) and (ii) an increase in acetylated p53, a known target of SIRT1 deacetylation activity. In addition, activation of the redox-sensitive anticancer drug EO9 was enhanced selectively in p53(+/+) cancer cells, attributable to increased activity of NAD(P)H-dependent oxidoreductase NQO1 (NAD(P)H quinone oxidoreductase 1). Suppressing LDH-A increased EO9-induced DNA damage in p53(+/+) cancer cells, but importantly had no additive effect in non-cancer cells. Our results identify a unique strategy by which the NADH/NAD(+) cellular redox status can be modulated in a cancer-specific, p53-dependent manner and we show that this can impact upon the activity of important NAD(H)-dependent enzymes. To summarise, this work indicates two distinct mechanisms by which suppressing LDH-A could potentially be used to kill cancer cells selectively, (i) through induction of apoptosis, irrespective of cancer cell p53 status and (ii) as a part of a combinatorial approach with redox-sensitive anticancer drugs via a novel p53/NAD(H)-dependent mechanism.
authors.affiliation: 1] Department of Biology, University of York, York, UK [2] Institute of Cancer Therapeutics, University of Bradford, Bradford, UK.
authors.forename: S J
authors.initials: SJ
authors.lastname: Allison
authors.affiliation: Department of Biology, University of York, York, UK.
authors.forename: J R P
authors.initials: JR
authors.lastname: Knight
authors.affiliation: Dipartimento di Farmacia, Università di Pisa, Pisa, Italy.
authors.forename: C
authors.initials: C
authors.lastname: Granchi
authors.affiliation: Dipartimento di Farmacia, Università di Pisa, Pisa, Italy.
authors.forename: R
authors.initials: R
authors.lastname: Rani
authors.affiliation: Dipartimento di Farmacia, Università di Pisa, Pisa, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Minutolo
authors.affiliation: Department of Biology, University of York, York, UK.
authors.forename: J
authors.initials: J
authors.lastname: Milner
authors.affiliation: Institute of Cancer Therapeutics, University of Bradford, Bradford, UK.
authors.forename: R M
authors.initials: RM
authors.lastname: Phillips