Enhancement of human prolactin synthesis by sodium butyrate addition to serum-free CHO cell culture.

Sodium butyrate (NaBu) has been used as a productivity enhancer for the synthesis of recombinant proteins in Chinese hamster ovary (CHO) cells. Thus, the influence of NaBu on the production of recombinant human prolactin (hPRL) from CHO cells was investigated for the first time. CHO cell cultures were submitted to a treatment with different concentrations of NaBu (0.25 to 4 mM). Quantitative and qualitative analyses by reverse-phase high-performance liquid chromatography (RP-HPLC) and Western blot or SDS-PAGE, carried out directly on CHO-conditioned medium, showed that the highest hPRL expression was obtained with 1 mM NaBu. In vitro biological assays based on noble rat lymphoma (Nb2) and mouse pro-B lymphoma (Ba/F3-LLP) cells were carried out on purified hPRL. Its bioactivity in the presence of NaBu was not apparently different from that of the First International Reference Reagent of recombinant hPRL (WHO 97/714). Our results show that NaBu increased the synthesis of recombinant hPRL in CHO cells, apparently without compromising either its structure or function.

Terpenes arrest parasite development and inhibit biosynthesis of isoprenoids in Plasmodium falciparum.

Development of new drugs is one of the strategies for malaria control. The biosynthesis of several isoprenoids in Plasmodium falciparum was recently described. Interestingly, some intermediates and final products biosynthesized by this pathway in mammals differ from those biosynthesized in P. falciparum. These facts prompted us to evaluate various terpenes, molecules with a similar chemical structure to the intermediates of the isoprenoids pathway, as potential antimalarial drugs. Different terpenes and S-farnesylthiosalicylic acid were tested on cultures of the intraerythrocytic stages of P. falciparum, and the 50% inhibitory concentrations for each one were found: farnesol, 64 microM; nerolidol, 760 nM; limonene, 1.22 mM; linalool, 0.28 mM; and S-farnesylthiosalicylic acid, 14 microM. All the terpenes tested inhibited dolichol biosynthesis in the trophozoite and schizont stages when [1-(n)-(3)H]farnesyl pyrophosphate triammonium salt ([(3)H]FPP) was used as precursor. Farnesol, nerolidol, and linalool showed stronger inhibitory activity on the biosynthesis of the isoprenic side chain of the benzoquinone ring of ubiquinones in the schizont stage. Treatment of schizont stages with S-farnesylthiosalicylic acid led to a decrease in intensity of the band corresponding a p21(ras) protein. The inhibitory effect of terpenes and S-farnesylthiosalicylic acid on the biosynthesis of both dolichol and the isoprenic side chain of ubiquinones and the isoprenylation of proteins in the intraerythrocytic stages of P. falciparum appears to be specific, because overall protein biosynthesis was not affected. Combinations of some terpenes or S-farnesylthiosalicylic acid tested in this work with other antimalarial drugs, like fosmidomycin, could be a new strategy for the treatment of malaria.