Ugir Hossain Sk

Cellular and Pharmacological Selectivity of the Peroxisome Proliferator-Activated Receptor-β/δ Antagonist GSK3787

The availability of high-affinity agonists for peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) has led to significant advances in our understanding of the functional role of PPARβ/δ. In this study, a new PPARβ/δ antagonist, 4-chloro-N-(2-{[5-trifluoromethyl)-2-pyridyl]sulfonyl}ethyl)benzamide (GSK3787), was characterized using in vivo and in vitro models. Orally administered GSK3787 caused antagonism of 4-[2-(3-fluoro-4-trifluoromethyl-phenyl)-4-methyl-thiazol-5-ylmethylsulfanyl]-2-methyl-phenoxy}-acetic acid (GW0742)-induced up-regulation of Angptl4 and Adrp mRNA expression in wild-type mouse colon but not in Pparβ/δ-null mouse colon. Chromatin immunoprecipitation (ChIP) analysis indicates that this correlated with reduced promoter occupancy of PPARβ/δ on the Angptl4 and Adrp genes. Reporter assays demonstrated antagonism of PPARβ/δ activity and weak antagonism and agonism of PPARγ activity but no effect on PPARα activity. Time-resolved fluorescence resonance energy transfer assays confirmed the ability of GSK3787 to modulate the association of both PPARβ/δ and PPARγ coregulator peptides in response to ligand activation, consistent with reporter assays. In vivo and in vitro analysis indicates that the efficacy of GSK3787 to modulate PPARγ activity is markedly lower than the efficacy of GSK3787 to act as a PPARβ/δ antagonist. GSK3787 antagonized GW0742-induced expression of Angptl4 in mouse fibroblasts, mouse keratinocytes, and human cancer cell lines. Cell proliferation was unchanged in response to either GW0742 or GSK3787 in human cancer cell lines. Results from these studies demonstrate that GSK3787 can antagonize PPARβ/δ in vivo, thus providing a new strategy to delineate the functional role of a receptor with great potential as a therapeutic target for the treatment and prevention of disease.

Modulation of Cyclophosphamide-Induced Cellular Toxicity by Diphenylmethyl Selenocyanate In Vivo, an Enzymatic Study

AIM: Cyclophosphamide (CP) is one of the most widely used alkylating antineoplastic agents that damage normal cells while killing cancerous cells in vivo. The use of CP in treating cancer patients is limited due to its severe toxicities induced mainly by oxidative stress. Diphenylmethyl selenocyanate is a synthetic organoselenium compound shown to act as a potent antioxidant in chemically induced murine toxicity and carcinogenesis models in vivo. In the present study, this compound has been evaluated for its protective potential against CP-induced toxicity in Swiss albino mice. METHODS: CP was administered intraperitoneally (50 mg/kg) and diphenylmethyl selenocyanate was given orally (3 mg/kg) in a pretreatment and concomitant treatment schedule, and the effects were assessed by estimating lipid peroxidation level, phase II detoxifying enzyme system, blood hemoglobin level, serum transaminase activity, and nitrite content. RESULTS: Diphenylmethyl selenocyanate significantly (P＜0.05) increased glutathione-S-transferase, glutathione peroxidase, and catalase levels whereas decreased the lipid peroxidation levels in both liver and lung tissues of the animals given CP. Superoxide dismutase was increased significantly in liver (P＜0.05) but not in the lung. The selenium compound also significantly (P＜0.05) increased the blood hemoglobin level whereas decreased the transaminase activity in serum and the nitrite content in peritoneal macrophages. CONCLUSION: Our result suggests that diphenylmethyl selenocyanate has the potential to prevent CP-induced cellular toxicity.

Naphthalimide based novel organoselenocyanates: finding less toxic forms of selenium that would retain protective efficacy.

A series of naphthalimide based organoselenocyanates were synthesized and screened for their toxicity as well as their ability to modulate several detoxifying/antioxidative enzyme levels at a primary screening dose of 3 mg/kg b.w. in normal Swiss albino mice for 30 days. Compound 4d showed highest activity in elevating the detoxifying/antioxidant enzymes levels.

Development of novel naphthalimide derivatives and their evaluation as potential melanoma therapeutics

Synthesis and anti-melanoma activity of various naphthalimide analogs, rationally modified by introducing isothiocyanate (ITC) and thiourea (TU) functionalities, found in well-known anti-cancer agents, is described. The structure-activity relationship comparison of the novel agents in inhibiting cancer cell growth was evaluated in various melanoma cell lines. Both ITC and TU analogs effectively inhibited cell viability and induced apoptosis in various human melanoma cells. Nitro substitution and increase in alkyl chain length, in general, enhanced the apoptotic activity of ITC derivatives. All the new compounds were well tolerated when injected intraperitoneal (i.p.) in mice at effective doses at which both the ITC and TU derivatives inhibited melanoma tumor growth in mice following i.p. xenograft. The nitro substituted naphthalimide-ITC derivative 3d was found to be the most effective in inducing apoptosis, and in inhibiting melanoma cell and tumor growth.

Synthesis and bioactivity of sphingosine kinase inhibitors and their novel aspirinyl conjugated analogs

Sphingosine kinase (SphK) is a lipid kinase with oncogenic activity, and SphK inhibitors (SKIs) are known for their anti-cancer activity. Here, we report highly efficient syntheses of SKIs and their aspirinyl (Asp) analogs. Both SKIs and their Asp analogs were highly cytotoxic towards multiple human cancer cell lines; in several cases the Asp analogs were up to three times more effective. Furthermore, they were equally potent inhibitors of SphK. The pharmacokinetic study indicated that SKI-I-Asp cleaved efficiently to form SKI-I and the half-life of SKI-I was increased from approximately 7 h in SKI-I to approximately 10 h in SKI-I-Asp injected mice, thereby prolonging its effect. In summary, the Asp-conjugated SKIs seem to be promising prodrugs of SKIs where delivery in vivo remains a problem.

Amelioration of cisplatin-induced nephrotoxicity in mice by oral administration of diphenylmethyl selenocyanate

Abstract Cisplatin is one of the most potent and active cytotoxic drug in the treatment of cancer. However, side-effects in normal tissues and organs, notably nephrotoxicity in the kidneys, limit the promising efficacy of cisplatin. The present study was designed to ascertain the possible in vivo protective potential of a synthetic organoselenium compound diphenylmethyl selenocyanate (3 mg/kg.b.w.) against the nephrotoxic damage induced by cisplatin (5 mg/kg.b.w. for 5 days) in Swiss albino mice. Treatment with diphenylmethyl selenocyanate markedly reduced cisplatin-induced lipid peroxidation, serum creatinine and blood urea nitrogen levels. Renal antioxidant defense systems, such as glutathione-S-transferase, glutathione peroxidase, superoxide dismutase, catalase, activities and reduced glutathione level, depleted by cisplatin therapy, were restored to normal by the selenium compound. The selenium compound also reduced renal tubular epithelial cell damage, nitric oxide levels and expression of COX-2, and iNOS in kidneys injured by cisplatin. These results demonstrate the protective effect of diphenylmethyl selenocyanate against cisplatin-induced nephrotoxicity in mice.

Synthesis of isosteric selenium analog of the PPARβ/δ agonist GW501516 and comparison of biological activity

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors and members of the nuclear hormone receptor superfamily. Herein, we describe an efficient synthesis of a novel isosteric selenium analog of the highly specific PPARbeta/delta ligand 2-methyl-4-((4-methyl-2-(4-trifluoromethylphenyl)-1,3-thiazol-5-yl)-methylsulfanyl)phenoxy-acetic acid (GW501516; 1). The study examined the efficiency of the novel selenium analog 2-methyl-4-((4-methyl-2-(4-trifluoromethylphenyl)-1,3-selenazol-5-yl)-methylsulfanyl)phenoxy-acetic acid (2) to activate PPARbeta/delta and the effect of ligand activation of PPARbeta/delta on cell proliferation and target gene expression in human HaCaT keratinocytes. The results showed that similar to GW501516, the Se-analog 2 increased expression of the known PPARbeta/delta target gene angiopoietin-like protein 4 (ANGPTL4); the compound 2 was comparable in efficacy as compared to GW501516. Consistent with a large body of evidence, the Se-analog inhibited cell proliferation in HaCaT keratinocytes similar to that observed with GW501516. In summary, the novel Se-analog 2 has been developed as a potent PPARbeta/delta ligand that may possess additional anti-cancer properties of selenium.

Design and synthesis of coumarin-based organoselenium as a new hit for myeloprotection and synergistic therapeutic efficacy in adjuvant therapy

A newly designed organoselenium compound, methyl substituted umbelliferone selenocyanate (MUS), was synthesized as a primary hit against the myelotoxic activity of carboplatin. MUS was administered at 6 mg/kg b.wt, p.o. in concomitant and pretreatment schedules with carboplatin (12 mg/kg b.wt, i.p. for 10 days) in female Swiss albino mouse. MUS treatment reduced (P < 0.001) the percentage of chromosomal aberrations, micronuclei formation, DNA damage and apoptosis in murine bone marrow cells and also enhanced (P < 0.001) the bone marrow cell proliferation of the carboplatin-treated mice. These activities cumulatively restored the viable bone marrow cell count towards normalcy. Myeloprotection by MUS was achieved, in part, due to a significant reduction in the ROS/RNS formation and restoration of glutathione redox pool. Additionally, MUS synergistically enhanced the cytotoxicity of carboplatin against two human cancer cell lines (MCF-7 and Colo-205). Furthermore, MUS can effectively potentiate the antitumour activity of carboplatin against two murine cancers (Dalton’s Lymphoma and Sarcoma-180) in vivo. These preclinical findings clearly indicate that MUS can improve the therapeutic index of carboplatin and ensures more effective therapeutic strategy against cancer for clinical development.

Abstract 1759: Novel naphthalimide analogs elicit their anti-tumor activity through a dual inhibition of topoisomerase II and Akt signaling pathway

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Naphthalimide derivatives such as mitonafide, a topoisomerase II inhibitor, have shown antitumor activity both in preclinical studies and in phase I and phase II clinical trials. However, it suffers from severe toxicity issues, especially central nervous system (CNS) toxicity. We have recently reported promising naphthalimide analogs designed by incorporating functional groups such as a isothiocyanate (ITC) group which is present in chemicals found in cruciferous vegetables, in place of N, N-dimethyl group in mitonafide. Some of these agents effectively induced apoptosis in melanoma cells and inhibited cell proliferation, and inhibited melanoma tumor growth by ∼60% without any apparent systemic toxicity issues. The ITC group was incorporated in the structure in an attempt to lower the toxicity associated with mitonafide and also because this functionality has been shown to block PI3K/Akt pathway which is activated in majority of cancer types; Akt3 is activated in 70% of sporadic melanomas. The idea was to generate a compound capable of inhibiting not only topoisomerase II similar to mitonafide but also PI3K kinase. In order to evaluate the underlying mechanism of action, we first evaluated topoisomerase-II inhibitory activity of agents relative to mitonafide to check if the new agents retained this property. Relaxation activity of DNA topoisomerase II was determined by measuring the conversion of supercoiled pBR322 plasmid DNA to its relaxed form. Our results indicate that the ITC analogs, NNITC-2 and NITC-6, exhibited significant inhibition for DNA-topoisomerase II similar to mitonafide. Interestingly, corresponding isoselenocyanate (ISC) analog NISC-6 was less effective compared to NITC-6 as a topoisomerase II inhibitor; NNISC-2 on the other hand was equally good as NNITC-2. Furthermore, to test our hypothesis that the addition of ITC group into the structure would make it capable of inhibiting PI3K/Akt pathway, we examined the ability of naphthalimide derivatives to inhibit the total cellular content of human Akt pathway proteins, by Western blot analysis using polyclonal antibodies, in human melanoma cancer cells (UACC903). UACC903 cells treated with ITC derivatives at 0.5, 1 and 5 μM, showed a rapid decrease in Akt pathway proteins level in a dose-dependent manner. In comparison, ISC analogs were less effective. Detailed results of these investigations will be presented. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1759. doi:1538-7445.AM2012-1759

Abstract 5430: Incorporation of selenium into temozolomide elicits superior antitumor activity that involves new mechanisms of action

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Temozolomide (TMZ), an alkylating agent that exerts antitumor activity by interfering with DNA replication, is used commonly in the treatment of two deadly diseases, malignant glioma and melanoma. Although TMZ can improve overall survival of patients with cancer, the therapeutic outcomes remain unsatisfactory: nearly 3/4 of the TMZ-treated glioma patients still die within 2 years. Thus, development of better anticancer drugs remains to be an urgent task. Selenium (Se) compounds, which possess apoptosis-inducing and anti-proliferating effects, have been studied extensively as chemopreventive and chemotherapeutic agents. On the basis of previous structure-activity studies, here we tested the hypothesis that incorporating Se into TMZ would improve the anticancer properties of this drug. A TMZ-Se analog was synthesized by rationally introducing an N-ethylselenocyanate extension to the amide functionality in TMZ structure, which is known not to be involved in its mode of action. In MTT and clonogenic assays, TMZ-Se showed a superior cytotoxicity to TMZ in human glioma cell lines T98G, LN229, U251, and U87MG, and in melanoma cell lines, UACC903 and 1205Lu. TMZ-Se was also effective against a TMZ-resistant glioma cell line, CCF-STTG. In a mouse melanoma xenograft model, TMZ-Se demonstrated a more potent tumor-inhibiting activity than TMZ. To explore the mechanism underlying the superior antitumor activity of TMZ-Se, we compared the effects of TMZ and TMZ-Se on apoptosis and autophagy, two cellular processes that determine death or survival of a cell. Apoptosis was significantly increased in tumor cells treated with TMZ-Se in comparison to those treated with TMZ, as determined by flow cytometric analysis of Annexin V staining, and Western bolts of cleaved caspase-9, caspase-3 and PARP. As compared with TMZ, TMZ-Se also triggered greater autophagic response, as examined by Western blots of the autophagy markers, LC3-II and p62. Interestingly, although qRT-PCR demonstrated that mRNA level of the key autophagy gene, Beclin 1, was increased following TMZ-Se treatment, Beclin 1 protein was down-regulated in the cells subjected to same treatment. TMZ-Se-triggered autophagy appeared to be cytoprotective, as suppressing autophagy by the inhibitors, 3-MA or chloroquine, further enhanced the efficacy of TMZ-Se. Also, TMZ-Se showed stronger down-regulating effect on the amount of O6-methylguanine-DNA methyltransferase, an enzyme that plays a crucial role in protecting cells from cytotoxicity of alkylating agents. These results indicate that incorporation of Se into TMZ can render greater potency to this chemotherapeutic drug via eliciting new mechanisms of action. Thus, incorporating selenium into certain conventional anticancer drugs to improve their effectiveness might represent a new strategy of drug development that warrants further investigation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5430. doi:10.1158/1538-7445.AM2011-5430