Metformin Increases Mitochondrial Energy Formation in L6 Muscle Cell Cultures.
Metformin Increases Mitochondrial Energy Formation in L6 Muscle Cell Cultures.
J Biol Chem. 2013 May 29;
Vytla VS, Ochs RS
A popular hypothesis for the action of metformin, the widely used anti-diabetes drug, is the inhibition of mitochondrial respiration, specifically at complex I. This is consistent with metformin stimulation of glucose uptake by muscle and inhibition of gluconeogenesis by liver. Yet, mitochondrial inhibition is inconsistent with metformin stimulation of fatty acid oxidation in both tissues. In the present study we measured mitochondrial energy production in intact cells adapting an in vivo technique of phosphocreatine (PCr) formation following energy interruption (PCr recovery) to cell cultures. Metformin increased PCr recovery from either dinitrophenol (DNP) or azide in L6 cells. We found that metformin alone had no effect on cell viability as measured by total ATP concentration, trypan blue exclusion, or MTT reduction. However, treatments with low concentrations of DNP or azide reversibly decreased ATP concentration. Metformin increased MTT reduction during recovery from either agent. Viability measured by trypan blue exclusion indicated that cells were intact under these conditions. We also found that metformin increased free AMP, and to a smaller extent, free ADP concentrations in cells, an action that was duplicated by a structurally unrelated AMP deaminase inhibitor. We conclude that, in intact cells, metformin can lead to a stimulation of energy formation, rather than an inhibition. HubMed – drug
TLR4 is a novel determinant of the response to paclitaxel in breast cancer.
Mol Cancer Ther. 2013 May 29;
Rajput S, Volk-Draper LD, Ran S
Overexpression of Toll-like Receptor-4 (TLR4) in human tumors often correlates with chemoresistance and metastasis. We found that TLR4 is overexpressed in the majority of clinical breast cancer (BC) samples and in 68% of the examined BC lines. TLR4 is activated by lipopolysaccharide (LPS) and other ligands including the widely-used drug paclitaxel (PXL). LPS is frequently used to show a tumor-promoting role of TLR4 although this bacterial component is unlikely to be found in BC environment. We reasoned that PXL-dependent activation of TLR4 is more relevant to BC chemoresistance that could be mediated by activation of the NF-?B pathway leading to upregulation of pro-survival genes. To test this hypothesis, we correlated TLR4 expression with resistance to PXL in two modified BC lines with either depleted or overexpressed TLR4 protein. Depletion of TLR4 in naturally overexpressing MDA-MB-231 cells downregulated pro-survival genes concomitant with 2-3 fold reduced IC50 to PXL in vitro and a 6-fold decrease in recurrence rate in vivo. Conversely, TLR4 overexpression in a negative cell line HCC1806 significantly increased expression of inflammatory and pro-survival genes along with a 3-fold increase of IC50 to PXL in vitro and enhanced tumor resistance to PXL therapy in vivo. Importantly, both tumor models showed that many PXL-upregulated inflammatory cytokines were co-induced with their receptors suggesting that this therapy induces autocrine tumor-promoting loops. Collectively, these results demonstrate that paclitaxel not only kills tumor cells but also enhances their survival by activating TLR4 pathway. These findings suggest that blocking TLR4 could significantly improve response to chemotherapy. HubMed – drug
An epigenetic antimalarial resistance mechanism involving parasite genes linked to nutrient uptake.
J Biol Chem. 2013 May 28;
Sharma P, Wollenberg K, Sellers M, Zainabadi K, Galinsky K, Moss E, Nguitragool W, Neafsey D, Desai SA
Acquired antimalarial drug resistance produces treatment failures and has led to periods of global disease resurgence. In P. falciparum, resistance is known to arise through genome-level changes such as mutations and gene duplications. We now report an epigenetic resistance mechanism involving genes responsible for the plasmodial surface anion channel, a nutrient channel that also transports ions and antimalarial compounds at the host erythrocyte membrane. Two blasticidin S-resistant lines exhibited markedly reduced expression of clag genes linked to channel activity, but had no genome-level changes. Silencing aborted production of the channel protein and was directly responsible for reduced uptake. Silencing affected clag paralogs on two chromosomes and was mediated by specific histone modifications, allowing a rapidly reversible drug resistance phenotype advantageous to the parasite. These findings implicate a novel epigenetic resistance mechanism that involves reduced host cell uptake and is a worrisome liability for water-soluble antimalarial drugs. HubMed – drug
Selection of Drug-resistant Feline Immunodeficiency Virus (FIV) Encoding FIV/HIV Chimeric Protease in The Presence of HIV-specific Protease Inhibitors.
J Virol. 2013 May 29;
Lin YC, Happer M, Elder JH
An infectious chimeric FIV/HIV encoding six HIV-like mutations in PR (I37V/N55M/V59I/I98S/Q99V/P100N) was subjected to selection in culture against PR inhibitors lopinavir (LPV), darunavir (DRV), or TL-3. LPV selection resulted in the sequential emergence of mutations V99A (S-1X), I59V (S-2X), I108V (S-3X), followed by V37I (S-4X). Mutant PRs were analyzed in vitro and isogenic viruses encoding each mutant PR were analyzed in culture for LPV sensitivity, yielding results consistent with the original selection. IC50 values for the S-1X, S-2X, S-3X and S-4X were 95 nM, 643 nM, 627 nM, and 1543 nM, respectively. The primary resistance mutations, V99(82)A, I59(50)V, and V37(32)I, are consistent with resistance pattern developed by HIV-1 under similar selection conditions. While resistance to LPV emerged readily, similar resistance mutations in PR to either DRV or TL-3 failed to emerge after passage for more than a year. However, a G37D mutation in nucleocapsid was observed in both selections and an isogenic G37D mutant replicated in the presence of 100 nM DRV or TL-3, whereas parental chimeric FIV could not. An additional mutation, L92V, near the PR active site in the folded structure, recently emerged during TL-3 selection. The L92V PR exhibited an IC50 value of 50 nM compared to 35 nM for 6s-98S PR and processed NC-p2 junction more efficiently, consistent with increased viral fitness. The findings emphasize the role of mutations outside the active site of PR in increasing viral fitness against active site inhibitors and suggest additional targets for inhibitor development. HubMed – drug