Exotica in rotating compact stars

The determination of mass and radius of a single neutron star EXO 0748-676 has been reported recently. Also, the estimate of radius from the measurement of moment of inertia of pulsar A in double pulsar system PSR J0737-3039 would be possible in near future. Here we construct models of static and uniformly rotating neutron stars involving exotic matter and compare our theoretical calculations with the recent findings from observations to probe dense matter in neutron stars.Comment: 8 pages, 3 figures Plenary talk at the 5th International Conference on Physics and Astrophysics of Quark Gluon Plasma (ICPAQGP), Kolkata, India, 8-12 February, 200

Enhancing the effect of immunotherapy by inhibiting tumor promoting effect of HDAC6

Histone deacetylases (HDAC) perform diverse functions beyond remodeling of chromatin landscape. It ranges from regulating the cellular-health to immune-diseases like cancer, positioning the HDAC inhibitors (HDACi) at a crucial junction of immunotherapy. The toxicity among pan-HDACi has led to the development of selective inhibitors, which helped to understand the roles of specific HDACs in immune responses. For example, HDAC6 promotes the pro-tumorigenic STAT3 pathway. By using specific HDAC6i, the downstream immune-modulatory pathways of STAT3, such as PDL1, could be targeted. HDAC6 has been associated with numerous structural functions, including cellular motility, shape, and intracellular transport through the acetylation of tubulin and cortactin. This function suggests that HDAC6 could also be a regulator of metastasis. We found that the selective HDAC6i NextA was able to reduce the primary tumor growth and the appearance of spontaneously metastatic nodules in the murine Triple Negative Breast Cancer (TNBC) tumor model 4T1. Additionally, the in vitro use of NextA in multiple murine and human breast cancer cell lines was found to reduce invasion and modulate multiple EMT-specific genes without exhibiting excessive cytotoxicity. PDL1 expression was also reduced, as described previously in melanoma models. In vivo, NextA reduced both the primary and secondary tumor progression. Given that 4T1 is a TIL-infiltrated tumor, we tested the efficacy of ÔÅ°PD-1 immune checkpoint inhibitors (ICI) and found a lower dose of ÔÅ°PD-1 to be more effective than the higher doses to reduce primary and secondary tumor growth. However, the expression of IFNÔÅß & PD-L1 were enhanced with the monotherapy. In an in vitro setting, we were able to nullify the upregulation of PD-L1 by ÔÅ°PD-1 with either NextA or IFNÔÅß neutralization. To apply this insight in vivo, we tested the combination of NextA and ÔÅ°PD-1 to find a significant reduction in tumor growth, both in primary & secondary nodules. Analysis of the effector molecules revealed a reduction in intra-tumoral PD-L1 and IDO1 as well as a reduction in several key EMT signature genes, including cMYC, MMP9, vimentin, and twist. We identified E-cadherin, a negative regulator metastasis, upregulated in vivo with NextA. This was corroborated by in vitro observations in several murine and human breast cancer cell lines, irrespective of hormonal receptor status. In conclusion, we propose that combining HDAC6i along with ICI such as ÔÅ°PD-1 may offer a novel avenue to enhance the efficacy of immunotherapy, and alter tumor-intricate physiology, without incurring unnecessary toxicity

Immunoepigenetics Combination Therapies: An Overview of the Role of HDACs in Cancer Immunotherapy

Long-standing efforts to identify the multifaceted roles of histone deacetylase inhibitors (HDACis) have positioned these agents as promising drug candidates in combatting cancer, autoimmune, neurodegenerative, and infectious diseases. The same has also encouraged the evaluation of multiple HDACi candidates in preclinical studies in cancer and other diseases as well as the FDA-approval towards clinical use for specific agents. In this review, we have discussed how the efficacy of immunotherapy can be leveraged by combining it with HDACis. We have also included a brief overview of the classification of HDACis as well as their various roles in physiological and pathophysiological scenarios to target key cellular processes promoting the initiation, establishment, and progression of cancer. Given the critical role of the tumor microenvironment (TME) towards the outcome of anticancer therapies, we have also discussed the effect of HDACis on different components of the TME. We then have gradually progressed into examples of specific pan-HDACis, class I HDACi, and selective HDACis that either have been incorporated into clinical trials or show promising preclinical effects for future consideration. Finally, we have included examples of ongoing trials for each of the above categories of HDACis as standalone agents or in combination with immunotherapeutic approaches

Systematic Screening of Histone Deacetylase Inhibitors: Enhancing Immunological Function Against Cancer.

Cancer is the result of a compromised immune system lacking the ability to recognize and eliminate transformed cells. These transformed cells induce an upregulation of negative regulatory tumor cells, which suppress T-cell activation and proliferation while inducing tumor tolerance. Immunotherapy is a novel therapeutic modality that strengthens the immune system against cancer cells, which in turn avoid immunological tolerance of cancer cells and the impedance of transformed cells survival. Histone deacetylase inhibitors (HDACinh) are specific drug compounds that are largely used as anti-cancer drugs. HDACinh have demonstrated control of apoptosis, cell survival, along with immune functionalities. HDAC inhibition also modulates the expression of tumor associated antigens (TAA) and immunosuppressive proteins. Our group has reported that the genetic and pharmacological abrogation of HDACs results in both, a decline in proliferation of tumor cells and important changes in immune regulatory pathways. For this reason, our group is currently focusing on the standardization of methodologies to identify changes in immunological markers without affecting other cellular mechanisms involved in cell survival: such as cell viability, necrosis, and apoptosis, which could interfere with the evaluation of the immune function. In vitro cultured murine SM1 cells were subjected to eight concentrations from clinical grade compounds – Tubastatin A, Nexturastat A, MS275 and LBH589 – to be analyzed within HDAC-Glo and ApoTox-Glo assays. The HDAC-Glo assay provided relative HDAC activity data, while the ApoTox-Glo assay specified the trends of Digitonin (viability control), Ionomycin (cytotoxicity control) and Mitomycin (apoptosis control). Upon inspection, the ApoTox-Glo recommended manufacturer controls compared to a pan-HDAC inhibitor LBH589 were deemed unreliable; LBH589 proved to be a universal control for the quantitative multiplexing protocol outlined. These results were then validated by Western blot protein analysis. Traditional approaches involve the time consuming and costly execution of individual assays. However, multiplex assays allow for quantifiable evaluation of anti-tumor HDACinh affects within cellular pathways by minimizing execution time and maximizing comprehensive statistical analysis of signal outputs from a spectrophotometer. Furthermore, an assay in tandem with our systematic high-throughput platform can accurately evaluate therapeutic doses and innovate treatments for melanoma clinical trials, along with other varieties of cancerous cell lines. The established reproducible and reliable protocol developed from this study will be important for identifying the dose range necessary to achieve immunological effects with minimal toxicity, thus improving the quality of life for treated subjects

Interferon regulatory factor-8 is important for histone deacetylase inhibitor-mediated antitumor activity.

The notion that epigenetic alterations in neoplasia are reversible has provided the rationale to identify epigenetic modifiers for their ability to induce or enhance tumor cell death. Histone deacetylase inhibitors (HDACi) represent one such class of anti-neoplastic agents. Despite great interest for clinical use, little is known regarding the molecular targets important for response to HDACi-based cancer therapy. We had previously shown that interferon regulatory factor (IRF)-8, originally discovered as a leukemia suppressor gene by regulating apoptosis, also regulates Fas-mediated killing in non-hematologic tumor models. Furthermore, we and others have shown that epigenetic mechanisms are involved in repression of IRF-8 in tumors. Therefore, in our preclinical tumor model, we tested the hypothesis that IRF-8 expression is important for response to HDACi-based antitumor activity. In the majority of experiments, we selected the pan-HDACi, Trichostatin A (TSA), because it was previously shown to restore Fas sensitivity to tumor cells. Overall, we found that: 1) TSA alone and more so in combination with IFN-γ enhanced both IRF-8 expression and Fas-mediated death of tumor cells in vitro; 2) TSA treatment enhanced IRF-8 promoter activity via a STAT1-dependent pathway; and 3) IRF-8 was required for this death response, as tumor cells rendered IRF-8 incompetent were significantly less susceptible to Fas-mediated killing in vitro and to HDACi-mediated antitumor activity in vivo. Thus, IRF-8 status may underlie a novel molecular basis for response to HDACi-based antitumor treatment

TSA-mediated IRF-8 transcription is STAT1-dependent.

<p>(<i>A</i>) STAT1 mRNA levels in CMS4 or CMS4.met.sel cells after the indicated treatments, as in Fig. 1. <i>(A)</i> Top, real-time PCR. Data presented as fold-change, as in Fig. 1. (<i>A</i>) Bottom, RT-PCR. *<i>P</i><0.05, based on comparing the single agent treatment to the vehicle-treated control. **<i>P</i><0.05, based on comparing the combination regimen to the single treatment counterparts. (<i>B</i>) CMS4 or CMS4.met.sel cells were transfected with an IRF-8 promoter reporter construct, followed by treatment with the indicated agents for 6 hr. Results are reported as the mean ± SEM of the fold-change relative to the vehicle-treated cells from three separate experiments. *<i>P</i><0.05, based on comparing treatment to matched vehicle control. No activity was observed using the pGL3 vector lacking a promoter. (<i>C</i>) Similar to <i>B</i>, except that CMS4 cells were silenced for STAT1 expression. *<i>P</i><0.05, based on comparing the indicated treatment group to the matched vehicle-treated control. **<i>P</i><0.05, based on comparing the STAT1-deficient groups to their matched STAT1-expressing vector controls. (<i>D</i>) Phosphorylated STAT1 (pSTAT1) and total STAT1 protein levels in CMS4.met.sel cells after treatment with the indicated treatments (TSA, 500 nM; IFN-γ, 200 U/ml) for 15 min, as measured by Western blot. This experiment is representative of one of three with similar results. (<i>E</i>) Similar to <i>D</i>, except that acetylated STAT1 and total STAT1 levels were measured by IP-Western blot (i.e., IP with anti-STAT1 antibody, followed by Western blot with anti-acetyl-lysine antibody) after treatment with or without TSA (500 nM for 6 hr). Band intensities were quantified, and the data presented as fold-change of TSA-treated vs. untreated samples (mean ± SEM of triplicate experiments). *<i>P</i><0.05, based on the TSA-treated group relative to the matched vehicle-treated control.</p

HDACi enhances IRF-8 expression in tumor cells.

<p>(<i>A</i>) CMS4 cells were treated with TSA, IFN-γ (100 U/ml) or a combination of both at the indicated concentrations and then analyzed by real-time PCR (top). Representative RT-PCR is shown in the bottom panel, which shares the same treatment labels. Data in top panel are presented as fold-change (shown above each bar) of the treated samples relative to the vehicle-treated controls. (<i>B</i>) Similar to <i>A</i>, except that CMS4 cells were treated with DP (25 ng/ml) instead of TSA. (<i>C</i>) CMS4-met.sel cells were treated with TSA (500 nM), IFN-γ or a combination of both and then analyzed real-time PCR (top) or RT-PCR (bottom), as in <i>A</i>. (<i>D</i>) Similar to <i>C</i>, except that CMS4 cells were treated with DP instead of TSA. All data are expressed as the mean ± SEM of triplicate determinations (shown above each bar). *<i>P</i><0.05, based on comparing the single agent treatment to the vehicle-treated control. **<i>P</i><0.05, based on comparing the combination regimen to the single treatment counterparts.</p