Enhancing transporter activity in heterologous expression systems with <scp>SAHA</scp> : a 2500‐times more potent and odorless alternative to butyrate

The functional characterization of plasma membrane transport proteins often relies on their heterologous expression in cultured cells. However, some transporters exhibit low activity, hindering meaningful functional assays. Heterologous expression is usually based on strong viral promoters which in living cells are prone to promoter silencing, a major problem. Here, we investigated the efficacy of low-cost histone deacetylase (HDAC) inhibitors in enhancing transporter activity, comparing the established sodium butyrate (the sodium salt of butyric acid) with valproate/valproic acid (VPA) and suberoylanilide hydroxamic acid (SAHA, also known as vorinostat). Using 293 cells stably transfected with pEBTet plasmids containing the CMV promotor to express the transporters SLC16A9, SLC22A15, and OATP1A2, we measured substrate efflux or uptake via LC-MS/MS following overnight preincubation with the HDAC inhibitors. All three compounds markedly stimulated transporter activity. VPA was less effective than butyrate but still surpassed control conditions. SAHA was cytotoxic at 6 μm, but at 2 μm, the enhancement was consistently comparable to 5 mm butyrate. Additionally, SAHA was more cost-effective and devoid of the repulsive odor characteristic of butyrate. Our findings advocate for replacing butyrate with SAHA to enhance heterologously expressed transporter activity. This offers a more efficient and user-friendly alternative for functional assays.