In vitro synergistic anti-prion effect of cholesterol ester modulators in combination with chlorpromazine and quinacrine

Abstract

In vitro synergistic anti-prion effect of cholesterol ester modulators in combination with chlorpromazine and quinacrine * E-mail: pania@unica.it°These authors contributed equally to this work. Received 06 October 2009; Accepted 16 December 2009Abstract: Our studies on the role of cholesterol in prion infection/replication showed that brains and peripheral cells of sheep susceptible-to or suffering-from Scrapie were characterized by an altered cholesterol homeostasis, and that drugs affecting cholesterol ester pool were endowed with selective anti-prion activity in N2a cell lines infected with the 22L and RML prion strains. In these prion-infected N2a cell lines, we now report increased anti-prion activity of dual-drug combinations consisting of cholesterol ester modulators associated with prion inhibitors. Synergism was obtained with the cholesterol ester modulators everolimus, pioglitazone, progesterone, and verapamil associated with the anti-prion chlorpromazine, and with everolimus and pioglitazone associated with the anti-prion quinacrine. In addition, comparative lipid analyses in prion-infected vs. uninfected N2a cells, demonstrated a derangement of type and distribution of cholesterol ester, free cholesterol, and triglyceride pools in the infected cells. Single-drug treatments differently affected synthesis of the various lipid forms, whereas combined drug treatments appeared to restore a lipid profile similar to that of the untreated-uninfected cells. We conclude that the anti-prion synergistic effects of cholesterol ester modulators associated with the cholesterol-interfering anti-prion drugs chlorpromazine and quinacrine may arise from the ability of combined drugs to re-establish lipid homeostasis in the prion-infected cells. Overall, these data suggest that inhibition of prion replication can be readily potentiated by combinatorial drug treatments and that steps of cholesterol/cholesterol ester metabolism may represent suitable targets.© Versita Sp. z o.o.

Accumulation and aberrant composition of cholesteryl esters in Scrapie-infected N2a cells and C57BL/6 mouse brains

“…These results are in agreement with our previous findings [17,18], as well as with those of a recent systems biology study showing that the SOAT1 gene, encoding for the enzyme (i.e. ACAT1) responsible for cholesterol esterification, is the first of all cholesterol-related genes to be up-regulated in the brain of scrapie-infected mice [25].…”

“…In the current study, qualitative and quantitative lipid analyses by HPLC-MS revealed multiple anomalies associated with prion infection both in ScN2a cell cultures and scrapie-affected C57Bl/6 mouse brains, mainly consisting in abnormal accumulation of CE. These results are in agreement with our previous findings [ 17 , 18 ], as well as with those of a recent systems biology study showing that the SOAT1 gene, encoding for the enzyme (i.e. ACAT1) responsible for cholesterol esterification, is the first of all cholesterol-related genes to be up-regulated in the brain of scrapie-infected mice [ 25 ].…”

“…In prion-infected ScN2a cells, a number of drugs that indirectly targeted cholesterol esterification by affecting steps of cholesterol metabolism/trafficking other than biosynthesis, were associated with a reduction of the CE pool and selective anti-prion activity [ 17 ]. Furthermore, selected combinations of these cholesterol-modulating drugs produced strong synergistic anti-prion effects, apparently by restoring cholesterol homeostasis [ 18 ]. Somewhat contradictory to our findings, however, other research groups reported that the increased content of FC in prion-infected neuronal cell lines was associated with a reduced content of CE [ 19 , 20 ].…”

Cholesterol homeostasis: a key to prevent or slow down neurodegeneration

“…A significant amount of genetic, biochemical, and pharmacological data highlighted that modifications in cholesterol esterification and trafficking are associated with Aβ and PrPsc biogenesis (Puglielli et al, 2001; Hutter-Paier et al, 2004; Bhattacharyya and Kovacs, 2010; Huttunen et al, 2010; Pani et al, 2007a,b,c, 2011; Orrù et al, 2010a,b,c). Moreover, several studies in various cell and animal models of AD evidenced that, genetic or pharmacological inhibition of ACAT activity markedly suppresses Aβ generation.…”

Pharmacologic Cholesterol Homeostasis Affects Prion Generation in a Synergistic Manner

“…In this context, our recent findings in sheep scrapie indicated that modifications of cholesterol metabolism, rather than being limited to the brain, appear to be a systemic trait since also ex vivo dermal fibroblasts and blood lymphocytes of sheep suffering-from or susceptible-to scrapie display similar alterations [12][13][14]. In addition, in prion/cell systems we found that persistent infection is associated to metabolic changes involving, besides cholesterol, all major classes of intracellular lipids, and that treatments with drug combinations triggering lipid homeostasis determine a synergic inhibition of prion generation [13,15,16]. In brains and skin fibroblasts from scrapie susceptible/infected sheep, the accumulation of cholesterol esters is accompanied by altered expression levels of enzymatic activities involved in cholesterol esterification (ACAT-1; up-regulated) and cholesterol trafficking (Cav-1; down-regulated), and by that of PrP C itself (up-regulated), Fig.…”