L-Carnitine Is an Endogenous HDAC Inhibitor Selectively Inhibiting Cancer Cell Growth In Vivo and In Vitro

Abstract

L-carnitine (LC) is generally believed to transport long-chain acyl groups from fatty acids into the mitochondrial matrix for ATP generation via the citric acid cycle. Based on Warburg's theory that most cancer cells mainly depend on glycolysis for ATP generation, we hypothesize that, LC treatment would lead to disturbance of cellular metabolism and cytotoxicity in cancer cells. In this study, Human hepatoma HepG2, SMMC-7721 cell lines, primary cultured thymocytes and mice bearing HepG2 tumor were used. ATP content was detected by HPLC assay. Cell cycle, cell death and cell viability were assayed by flow cytometry and MTS respectively. Gene, mRNA expression and protein level were detected by gene microarray, Real-time PCR and Western blot respectively. HDAC activities and histone acetylation were detected both in test tube and in cultured cells. A molecular docking study was carried out with CDOCKER protocol of Discovery Studio 2.0 to predict the molecular interaction between L-carnitine and HDAC. Here we found that (1) LC treatment selectively inhibited cancer cell growth in vivo and in vitro; (2) LC treatment selectively induces the expression of p21cip1 gene, mRNA and protein in cancer cells but not p27kip1; (4) LC increases histone acetylation and induces accumulation of acetylated histones both in normal thymocytes and cancer cells; (5) LC directly inhibits HDAC I/II activities via binding to the active sites of HDAC and induces histone acetylation and lysine-acetylation accumulation in vitro; (6) LC treatment induces accumulation of acetylated histones in chromatin associated with the p21cip1 gene but not p27kip1 detected by ChIP assay. These data support that LC, besides transporting acyl group, works as an endogenous HDAC inhibitor in the cell, which would be of physiological and pathological importance.

Methamphetamine promotes α-tubulin deacetylation in endothelial cells: The protective role of acetyl-l-carnitine

“…As a result, in the present work, we show that 1 mM ALC was able to prevent METH-induced deacetylation of a-tubulin. A previous report postulated that ALC had the potential to directly inhibit HDACs activity in several cell types possibly by chelating and removing Zn2+ from the active site of the HDAC enzyme (Huang et al, 2012). However, here, we could not observe a significant effect of ALC on HDACs activity.…”

“…Under TSA treatment, chromatin relaxation is known to increase the expression of CDKN1A (cyclin-dependent kinases inhibitor, or p21) gene (Blagosklonny et al, 2002;Hrabeta et al, 2014;Ocker and Schneider-Stock, 2007;Xu et al, 2007). As such, CDKN1A (p21) expression is frequently used as a marker of HDAC inhibition (Harrison and Dexter, 2013;Huang et al, 2012;Xu et al, 2007). As expected our results show that TSA promotes the overexpression of CDKN1A.…”

“…However, here, we could not observe a significant effect of ALC on HDACs activity. Of note, the doses used by Huang et al (2012) were 5 and 10 fold higher than the 1 mM dose shown here to efficiently prevent tubulin deacetylation. In addition, neither METH nor ALC led to altered levels of HDAC6 gene expression, which seems to indicate that both ALC-and METH-modulation of HDAC6 activity may occur in a post-translation way, as described for TSA (Xu et al, 2007).…”

“…TSA was used in a 100 nM concentration. The selected TSA and ALC doses were previously shown to be safe for the cells and are below the range of doses previously used in similar works (Huang et al, 2012;Hubbert et al, 2002;Tu et al, 2014).…”

“…Therefore, we hypothesized that METH-induced regulation of HDACs activity, and in particular of HDAC6, may also mediate the structural loss observed in METH-exposed endothelial cells. Moreover, since we recently showed that a pretreatment with acetyl-L-carnitine (ALC) was able to prevent METH-induced activation of MMP-9, preserving the actin structural arrangement in the endothelial cells (Fernandes et al, 2014), and ALC was shown to have the potential to interact with HDAC activity (Huang et al, 2012), we reasoned that ALC could also preserve the acetylation of microtubules under METH action. ALC is a natural occurring compound that was seen to be protective by different mechanisms in several neurological conditions, including BBB dysfunction (Alves et al, 2009;Haorah et al, 2011;Muneer et al, 2011;Pettegrew et al, 2000).…”