The posttranslational addition of palmitate to cysteines occurs ubiquitously in eukaryotic cells, where it functions in anchoring target proteins to membranes and in vesicular trafficking. Here we show that the Saccharomyces cerevisiae palmitoyltransferase Pfa4 enhanced heterochromatin formation at the cryptic mating-type loci HMR and HML via Rif1, a telomere regulatory protein. Acylated Rif1 was detected in extracts from wild-type but not pfa4Δ mutant cells. In a pfa4Δ mutant, Rif1-GFP dispersed away from foci positioned at the nuclear periphery into the nucleoplasm. Sir3-GFP distribution was also perturbed, indicating a change in the nuclear dynamics of heterochromatin proteins. Genetic analyses indicated that PFA4 functioned upstream of RIF1. Surprisingly, the pfa4Δ mutation had only mild effects on telomeric regulation, suggesting Rif1's roles at HM loci and telomeres were more complexly related than previously thought. These data supported a model in which Pfa4-dependent palmitoylation of Rif1 anchored it to the inner nuclear membrane, influencing its role in heterochromatin dynamics.transcriptional silencing | chromosome architecture

Background: NKP-2235 is a first-in-class, orally bioavailable gallium based anti-cancer compound. The intracellular targets for NKP-2235 include the endoplasmic reticulum (ER). NKP-2235 treatment of tumor cells causes ER stress, intracellular Ca2+ release and induction of pro-apoptotic factors. In single agent in vitro cytotoxicity assays, NKP-2235 has shown activity against tumor lines resistant to many standard anti-cancer agents. The aims of this study were to evaluate the anti-tumor effects of NKP-2235 in combination with standard cytotoxics or targeted agents and assess its single agent cytotoxicity pattern in the NCI 60 tumor cell line anti-cancer drug screen and COMPARE analysis. Methods: NKP-2235 testing in the NCI 60 tumor cell line anticancer drug screen was performed by the NCI. COMPARE analysis was conducted using cytotoxicity data from 170 compounds in the NCI compound library. In vitro combination studies were performed in breast, colon, glioma, lung and prostate tumor cell lines. NKP-2235 was tested in combination with 5-FU, temozolomide, gemcitabine, paclitaxel, erlotinib, temsirolimus, or docetaxel either simultaneously or sequentially. For simultaneous combinations, cells were exposed to NKP-2235 and a combination agent, at their respective ED50, for 72hrs. For sequential NKP-2235 combinations cells were exposed to different dosing sequences, all agents at their respective ED50, for a total of 96 hrs. Cytotoxicity was determined using the MTT assay and combination index (CI) values were calculated using the Chou-Talalay method. Results: Screening against the NCI 60 tumor cell line panel and COMPARE analysis show that single agent NKP-2235 has a markedly different pattern of cytotoxicity from the 170 anti-cancer agents previously tested by the NCI. In addition, the screening confirmed that the NKP-2235 cytotoxicity is distinct from that of gallium salts such as gallium nitrate. The combination studies demonstrated that NKP-2235 has synergistic activity when combined simultaneously with: 5-FU (CI 0.6), temozolomide (CI 0.8), gemcitabine (CI 0.7), temsirolimus (CI 0.7), and docetaxel (CI 0.7) in the cell lines tested. Interestingly, while simultaneous combination is synergistic with docetaxel in prostate cancer cells, the NKP-2235-paclitaxel combinations required sequential dosing for synergistic activity (CI 0.09) in lung cancer cells. Similarly, the NKP-2235-temsirolimus combination is synergistic in simultaneous setting, while NKP-2235-erlotinib combination required sequential dosing for synergistic activity (CI 0.3) in lung cancer cells. Conclusions: This study demonstrates that NKP-2235 has a novel pattern of cytotoxicity and marked synergism in combination with a broad range of anti-tumor agents in many tumor types. These data support the study of this novel agent in single agent and combination clinical trials. 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 3838. doi:1538-7445.AM2012-3838

Background: NKP-1339 is a novel transferrin targeted ruthenium based compound that has shown promising single agent anti-cancer activity in preclinical models and in man. The compound has a novel mechanism of action, with GRP78 down-regulation included in its target pathways. In an ongoing Phase I study, the side effects are all mild and do not include those associated with standard cytotoxics or targeted agents, suggesting that NKP-1339 could be safely combined with standard anticancer therapies. The aim of this study was to evaluate the anti-tumor effects of NKP-1339 combination with these agents. Methods: In vitro combination studies were performed in breast, colon, lung, gastric, prostate, pancreatic and liver tumor cell lines with cisplatin, oxaliplatin, 5-FU, docetaxel, doxorubicin, gemcitabine, erlotinib or sorafenib as appropriate for that tumor type. Cells were exposed to NKP-1339 and a combination agent, both agents at their respective ED50, for 72hrs. Cytotoxicity was determined using the MTT assay and combination index (CI) values were calculated using the Chou-Talalay method. The effect of the NKP-1339 and cisplatin combination was further studied in vivo in a gastric carcinoma xenograft model. Results: The in vitro combination studies demonstrated NKP-1339 synergism with other anti-cancer agents in varied tumor types. The CI values ranged from 0.8 (synergism) to 0.1 (strong synergism): cisplatin (CI 0.1), doxorubicin (CI 0.2), 5-FU (CI 0.3), oxaliplatin (CI 0.4), sorafenib (CI 0.5), docetaxel (CI 0.5), erlotinib (CI 0.6), gemcitabine (0.8). In vivo studies on the combination of NKP-1339 and cisplatin in gastric carcinoma xenografts showed significant (P<0.05) tumor growth delay and extended survival when compared to the single agent activity for either compound. Conclusions: NKP-1339's novel mechanism of action and its favorable safety profile in humans suggest that it could be safely combined with standard anti-cancer therapies. The marked synergistic effects of NKP-1339 in combination with a broad range of anti-tumor agents in vitro and in vivo supports the study of this novel agent with standard anticancer therapies in Phase I combination clinical trials. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B223.

Background Invasive mucormycosis (IM) is associated with high mortality and morbidity. MAT2203 is an encochleated oral formulation of amphotericin B which has been shown to be safe and effective against murine aspergillosis and murine cryptococcal meningoencephalitis. We sought to compare the efficacy of MAT2203 to liposomal amphotericin B (LAMB) treatment in a neutropenic mouse model of IM. Methods ICR mice were immunosuppressed with cyclophosphamide and cortisone acetate on days -2, -3 and +8, relative to infection with intratracheally instilled Rhizopus delemar 99-880 or M. circinelloides f. jenssenii DI15-131. Treatment with placebo (diluent control), oral MAT2203 (5 to 45 mg/kg, given qd or bid for 7 days) or LAMB (10 mg/kg, iv, qd for 4 days), began 16 h post infection. Survival (n=10-20/group from 1/2 experiments) through Day +21 and tissue fungal burden of lungs or brain (n=10/group) euthanized on Day +4 post infection (conidial equivalents using qPCR) served as a primary and secondary endpoint, respectively. Results For Rhizopus delemar infection, doses of MAT2203 5,15 mg/kg qd or 7.5 mg/kg bid, significantly prolonged median survival time (MST) and enhanced overall survival vs. placebo-treated mice (MST of 9 Days and 0% survival for placebo-treated mice vs. 13 Days, and 20-40% for MAT2203 doses, P< 0.05 by Log-Rank). Importantly, MAT2203 treatments were as effective as LAMB (MST of 16 days and overall survival of 45%). For mice infected with M. circinelloides, MAT2203 at 15 mg/kg, qd significantly prolonged MST and enhanced overall survival vs. placebo-treated mice (MST of 5 Days and 0% survival, for placebo-treated mice vs. 13.5 Days and 50% survival for MAT2203, P< 0.05). In both infection models MAT2203 treatment of 15 mg/kg or LAMB resulted in significant ∼1.0-1.5 log reduction and ∼2.0-2.2 log reduction in lung and brain fungal burden vs. placebo, respectively (Wilcoxon Rank Sum). Conclusion MAT2203 demonstrated in vivo efficacy in treating R. delemar or M. circinelloides pulmonary infection in immunosuppressed mice, which was equivalent to the LAMB current standard of care. Continued investigation and development of MAT2203 as a novel, and oral formulation of amphotericin antifungal agent against mucormycosis is warranted. Disclosures Theresa Matkovits, PhD, Matinas BioPharma: Employee Jenel Cobb, PhD, Matinas BioPharma: Employee Raphael J. Mannino, PhD, Matinas BioPharma: Employee Ashraf S. Ibrahim, PhD, Matinas BioPharma: Advisor/Consultant|Matinas BioPharma: Grant/Research Support|SFunga: Grant/Research Support.