Bcl-2 inhibitors: emerging drugs in cancer therapy

Dose-limiting toxicity to healthy tissues is among the major hurdles in anticancer treatment along with intrinsic or acquired multi-drug resistance. Development of small molecule inhibitors (SMI) specific for antiapoptotic Bcl-2 proteins is a novel approach in a way that these antagonists are aimed to interfere with specific protein-protein interactions unlike conventional chemo-/radiotherapies. SMIs of antiapoptotic Bcl-2 proteins are assumed to compete with proapoptotic Bcl-2s to occupy BH3 docking grooves on the surfaces of antiapoptotic family members. Instead of directly initiating cell death, these inhibitors are intended to decrease apoptotic threshold in tumor cells that were already primed to death. In this regard, antiapoptotic Bcl-2 protein SMIs have the advantage of lower normal tissue toxicity relative to conventional anticancer therapies that interfere with general mechanisms including DNA synthesis, mitosis and tyrosine kinase activity. Besides, Bcl-2 antagonists were shown to potentiate efficacies of established drugs in several hematological malignancies and solid tumors which render them promising candidates for combination anticancer therapy. Utilizing these SMIs in such a way may prove to decrease the patient drug load by diminishing the required chemo-/radiotherapy dose. This review summarizes and compares BH3 mimetics on the basis of specificity, mode of action and efficacy, as well as providing remarks on their therapeutical potential and routes of development in near future

Pramanicin analog induces apoptosis in human colon cancer cells: critical roles for Bcl-2, Bim, and p38 MAPK signaling

Pramanicin (PMC) is an antifungal agent that was previously demonstrated to exhibit antiangiogenic and anticancer properties in a few in vitro studies. We initially screened a number of PMC analogs for their cytotoxic effects on HCT116 human colon cancer cells. PMC-A, the analog with the most potent antiproliferative effect was chosen to further interrogate the underlying mechanism of action. PMC-A led to apoptosis through activation of caspase-9 and -3. The apoptotic nature of cell death was confirmed by abrogation of cell death with pretreatment with specific caspase inhibitors. Stress-related MAPKs JNK and p38 were both activated concomittantly with the intrinsic apoptotic pathway. Moreover, pharmacological inhibition of p38 proved to attenuate the cell death induction while pretreatment with JNK inhibitor did not exhibit a protective effect. Resistance of Bax -/- cells and the protective nature of caspase-9 inhibition indicate that mitochondria play a central role in PMC-A induced apoptosis. Early post-exposure elevation of cellular Bim and Bax was followed by a marginal Bcl-2 depletion and Bid cleavage. Further analysis revealed that Bcl-2 downregulation occurs at the mRNA level and is critical to mediate PMC-A induced apoptosis, as ectopic Bcl-2 expression substantially spared the cells from death. Conversely, forced expression of Bim proved to significantly increase cell death. In addition, analyses of p53-/- cells demonstrated that Bcl-2/Bim/Bax modulation and MAPK activations take place independently of p53 expression. Taken together, p53-independent transcriptional Bcl-2 downregulation and p38 signaling appear to be the key modulatory events in PMC-A induced apoptosis

Effects of Tooth Brushing and MouthWashing on Leaching Bisphenol A Levels From an Orthodontic Adhesive: An In Vitro Study

Objective: To assess the levels of bisphenol A (BPA) released from an orthodontic adhesive with respect to the effects of tooth brushing and mouth washing.Methods: Three groups, each containing fifteen adhesive samples were prepared. In Group 1, samples were polymerized according to manufacturer instructions. In Group 2, after the same polymerization protocol, each sample was brushed with a fluoride-containing toothpaste. For Group 3, samples were immersed in a mouth washing solution after polymerization. Later, all samples were placed into glass tubes containing 5 mL distilled water. High-performance liquid chromatography (HPLC) measurements were performed to assess the leaching amount of BPA. Intergroup comparison was performed by one way ANOVA test.Results: Mean amounts of BPA were found to be 0.2674 μg/L, 0.2692 μg/L, and 0.2705 μg/L, respectively. Only a significant difference was found between Group 1 and 3 (P < .01), revealing higher BPA levels with the mouth washing solution.Conclusion: Measurable amounts of BPA release were observed in all groups of orthodontic adhesive samples, but the detected amounts were below the toxic levels. From a clinical point of view, alcohol-containing mouth washing solutions might increase the amount of leaching monomer, since alcohol is solvent of BPA

Balarısı toplumlarının genetik yapılarının mikrosatelitler kullanılarak incelenmesi

We analyzed the genetic structures of 11 honeybee (Apis mellifera) populations from Türkiye and one population from Cyprus using 9 microsatellite loci. Average gene diversity levels were found to change between 0,542 and 0,681. Heterozygosity levels, mean number of alleles per population, presence of diagnostic alleles and pairwise FST values confirmed the mitochondrial DNA finding that Anatolian honeybees belong to north Mediterranean (C) lineage. We detected a very high level of genetic divergence among populations of Türkiye and Cyprus based on pairwise FST levels (between 0,0 and 0,2). Out of 66 population pairs 52 were found to be genetically different significantly. This level of significant differentiation has not been reported yet in any other study conducted on European and African honeybee populations. High allelic ranges, and high divergence indicate that Anatolia is a genetic centre for C lineage honeybees. We suggest that certain precautions should be taken to limit or forbid introduction and trade of Italian and Carniolan honeybees to Türkiye and Cyprus in order to preserve genetic resources formed in these territories in thousands of years. Effectivity at previously isolated regions in Artvin, Ardahan and Kırklareli was confirmed by the high genetic differentiation in honeybees of these regions. Genetically differentiated Karaburun and Cyprus honeybees v and geographical positions of the regions make these zones first candidates as new isolation areas.Ph.D. - Doctoral Progra

Inactivation of Bcl-2 through I kappa B kinase (IKK)-dependent phosphorylation mediates apoptosis upon exposure to 4-hydroxynonenal (HNE)

Apoptosis of macrophage foam cells loaded with modified/oxidized lipids is implicated in destabilization of advanced atherosclerotic plaques in humans. Concentration of HNE, main aldehydic product of plasma LDL peroxidation, elevates in atherosclerotic lesions as well as in cultured cells under oxidative stress. Although this reactive aldehyde has been shown to promote apoptosis with the involvement of p38 MAPK and JNK in various mammalian cell lines, roles of B-cell lymphoma 2 (Bcl-2) family proteins remain to be deciphered. We demonstrated that HNE-induced apoptosis was accompanied by concurrent downregulations of antiapoptotic Bcl-xL and Mcl-1 as well as upregulation of proapoptotic Bak. Furthermore, phoshorylation of Bcl-2 at Thr56, Ser70, and probably more phosphorylation sites located on N-terminal loop domain associated with HNE-induced apoptosis in both U937 and HeLa cells while ectopic expression of a phospho-defective Bcl-2 mutant significantly attenuated apoptosis. In parallel to this, HNE treatment caused release of proapoptotic Bax from Bcl-2. Pharmacological inhbition of IKK inhibited HNE-induced Bcl-2 phosphorylation. Similarly, silencing IKKa and -beta both ended up with abrogation of Bcl-2 phosphorylation along with attenuation of apoptosis. Moreover, both IKKa and -beta coimmunoprecipitated with Bcl-2 and in vitro kinase assay proved the ability of IKK to phosphorylate Bcl-2. In view of these findings and considering HNE inhibits DNA-binding activity of nuclear factor-?B (NF-?B) through prevention of I?B phosphorylation/ubiquitination/proteolysis, IKK appears to directly interfere with Bcl-2 activity through phosphorylation in HNE-mediated apoptosis independent of NF-?B signaling

SIRT1 contributes to aldose reductase expression through modulating NFAT5 under osmotic stress: in vitro and in silico insights

So far, a myriad of molecules were characterized to modulate NFAT5 and its downstream targets. Among these NFAT5 modifiers, SIRT1 was proposed to have a promising role in NFAT5 dependent events, yet the exact underlying mechanism still remains obscure. Hence, the link between SIRT1 and NFAT5-aldose reductase (AR) axis under osmotic stress, was aimed to be delineated in this study. A unique osmotic stress model was generated and its mechanistic components were deciphered in U937 monocytes. In this model, AR expression and nuclear NFAT5 stabilization were revealed to be positively regulated by SIRT1 through utilization of pharmacological modulators. Overexpression and co-transfection studies of NFAT5 and SIRT1 further validated the contribution of SIRT1 to AR and NFAT5. The involvement of SIRT1 activity in these events was mediated via modification of DNA binding of NFAT5 to AR ORE region. Besides, NFAT5 and SIRT1 were also shown to co-immunoprecipitate under isosmotic conditions and this interaction was disrupted by osmotic stress. Further in silico experiments were conducted to investigate if SIRT1 directly targets NFAT5. In this regard, certain lysine residues of NFAT5, when kept deacetylated, were found to contribute to its DNA binding and SIRT1 was shown to directly bind K282 of NFAT5. Based on these in vitro and in silico findings, SIRT1 was identified, for the first time, as a novel positive regulator of NFAT5 dependent AR expression under osmotic stress in U937 monocytes

Role of Bcl-2 family members in 4-hydroxynonenal (HNE) mediated apoptosis

4-Hydroxynonenal (HNE), a very reactive lipid peroxidation product, was shown to induce oxidative stress and apoptosis in a dose-dependent manner in a variety of cells Our recent data showed that FINE triggered caspase-9 and 3 activation and PARP cleavage in monocytic cell line U937. Following 10 mu M HNE treatment, levels of phosphorylated JNK, ERK1/2 and p38 MAP kinases increased significantly in 2 and 4 h Our results indicate a time-dependent decrease in Bcl-2 expression in response to HNE treatment. We observed a slight decrease in Bcl-X-L expression, while Mcl-1 amount remained unchanged Pro-apoptotic Bax and Bak expressions did not change; but truncated form of Bax disapperared Just after HNE treatment Our preliminary studies conducted by specific inhibitors of MAP kinase proteins indicate that phosphorylation of p38, JNK or ERK 1/2 proteins regulate the activation of Bcl-2 proteins, effecting the cellular outcome such as apoptosi