18 research outputs found
Evaluation Of Toxicity, Pharmacokinetic And Clastogenicity Profile Of Ethyl 2-[4[(Piperidin-1-yl) Phenyl]-1h-benzimidazole-5-carboxylate (Bzd9l1), Novel Sirtuin Inhibitor
The mammalian sirtuins (SIRTs) have been linked to various diseases including cancer, diabetes, neurodegenerative and cardiovascular diseases. Thus, SIRTs are attractive targets for the development of pharmaceuticals. The lack of potential SIRT inhibitors in cancer clinical trials highlights the need to develop a potent SIRT modulator as an anti-cancer therapeutic. Studies in the lab have highlighted the capability of a lead sirtuin inhibitor (BZD9L1) to reduce colorectal tumour growth in vitro and in vivo by modulating different cancer pathways in colorectal cancer with different mutation profiles. Also, synergistic effects of BZD9L1 in in vitro assays and tumour xenograft study when used in adjunct with 5- Fluorouracil (5-FU) further support the promising therapeutic effects of BZD9L1 as anticancer regime. Nevertheless, the toxicology profile of this compound remains unknown. To our knowledge, no prior work has reported the regulation of SIRTs in the liver and kidney by a small molecule inhibitor in a drug toxicity study. Therefore, this project aims to determine the toxicology, molecular regulation of toxicity, pharmacokinetics and clastogenicity profiles of BZD9L1 in in vitro or in vivo model
BZD9L1 Differentially Regulates Sirtuins in Liver-Derived Cells by Inducing Reactive Oxygen Species
Growing evidence has highlighted that mitochondrial dysfunction contributes to drug-induced toxicities and leads to drug attrition and post-market withdrawals. The acetylation or deacetylation of mitochondrial proteins can affect mitochondrial functions as the cells adapt to various cellular stresses and other metabolic challenges. SIRTs act as critical deacetylases in modulating mitochondrial function in response to drug toxicity, oxidative stress, reactive oxygen species (ROS), and energy metabolism. We previously showed that a recently characterised SIRT inhibitor (BZD9L1) is non-toxic in rodents in a short-term toxicity evaluation. However, the impact of BZD9L1 on mitochondrial function is unknown. This work aims to determine the effects of BZD9L1 on mitochondrial function in human normal liver and kidney-derived cell lines using the Agilent Seahorse Cell Mito Stress Test to complement our short-term toxicity evaluations in vivo. The Mito Stress assay revealed that BZD9L1 could potentially trigger oxidative stress by inducing ROS, which promotes proton leak and reduces coupling efficiency in liver-derived THLE cells. However, the same was not observed in human kidney-derived HEK293 cells. Interestingly, BZD9L1 had no impact on SIRT3 protein expression in both cell lines but affected SOD2 and its acetylated form at 72 h in THLE cells, indicating that BZD9L1 exerted its effect through SIRT3 activity rather than protein expression. In contrast, BZD9L1 reduced SIRT1 protein expression and impacted the p53 protein differently in both cell lines. Although BZD9L1 did not affect the spare respiratory capacity in vitro, these findings call for further validation of mitochondrial function through assessment of other mitochondrial parameters to evaluate the safety of BZD9L1
Cancer-Associated Fibroblasts: Epigenetic Regulation and Therapeutic Intervention in Breast Cancer
Breast cancer is the leading cause of cancer-related mortality in women worldwide. Cancer-associated fibroblasts (CAFs) are a heterogeneous population of cells in the solid tumour microenvironment. These cells are positively linked to breast cancer progression. Breast CAFs can be categorised into distinct subtypes according to their roles in breast carcinogenesis. Epigenetic modifications change gene expression patterns as a consequence of altered chromatin configuration and DNA accessibility to transcriptional machinery, without affecting the primary structure of DNA. Epigenetic dysregulation in breast CAFs may enhance breast cancer cell survival and ultimately lead to therapeutic resistance. A growing body of evidence has described epigenetic modulators that target histones, DNA, and miRNA as a promising approach to treat cancer. This review aims to summarise the current findings on the mechanisms involved in the epigenetic regulation in breast CAFs and discusses the potential therapeutic strategies via targeting these factors
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BZD9L1 benzimidazole analogue hampers colorectal tumor progression by impeding angiogenesis.
BackgroundThe development of new vasculatures (angiogenesis) is indispensable in supplying oxygen and nutrients to fuel tumor growth. Epigenetic dysregulation in the tumor vasculature is critical to colorectal cancer (CRC) progression. Sirtuin (SIRT) enzymes are highly expressed in blood vessels. BZD9L1 benzimidazole analogue is a SIRT 1 and 2 inhibitor with reported anticancer activities in CRC. However, its role has yet to be explored in CRC tumor angiogenesis.AimTo investigate the anti-angiogenic potential of BZD9L1 on endothelial cells (EC) in vitro, ex vivo and in HCT116 CRC xenograft in vivo models.MethodsEA.hy926 EC were treated with half inhibitory concentration (IC50) (2.5 μM), IC50 (5.0 μM), and double IC50 (10.0 μM) of BZD9L1 and assessed for cell proliferation, adhesion and SIRT 1 and 2 protein expression. Next, 2.5 μM and 5.0 μM of BZD9L1 were employed in downstream in vitro assays, including cell cycle, cell death and sprouting in EC. The effect of BZD9L1 on cell adhesion molecules and SIRT 1 and 2 were assessed via real-time quantitative polymerase chain reaction (qPCR). The growth factors secreted by EC post-treatment were evaluated using the Quantibody Human Angiogenesis Array. Indirect co-culture with HCT116 CRC cells was performed to investigate the impact of growth factors modulated by BZD9L1-treated EC on CRC. The effect of BZD9L1 on sprouting impediment and vessel regression was determined using mouse choroids. HCT116 cells were also injected subcutaneously into nude mice and analyzed for the outcome of BZD9L1 on tumor necrosis, Ki67 protein expression indicative of proliferation, cluster of differentiation 31 (CD31) and CD34 EC markers, and SIRT 1 and 2 genes via hematoxylin and eosin, immunohistochemistry and qPCR, respectively.ResultsBZD9L1 impeded EC proliferation, adhesion, and spheroid sprouting through the downregulation of intercellular adhesion molecule 1, vascular endothelial cadherin, integrin-alpha V, SIRT1 and SIRT2 genes. The compound also arrested the cells at G1 phase and induced apoptosis in the EC. In mouse choroids, BZD9L1 inhibited sprouting and regressed sprouting vessels compared to the negative control. Compared to the negative control, the compound also reduced the protein levels of angiogenin, basic fibroblast growth factor, platelet-derived growth factor and placental growth factor, which then inhibited HCT116 CRC spheroid invasion in co-culture. In addition, a significant reduction in CRC tumor growth was noted alongside the downregulation of human SIRT1 (hSIRT1), hSIRT2, CD31, and CD34 EC markers and murine SIRT2 gene, while the murine SIRT1 gene remained unaffected, compared to vehicle control. Histology analyses revealed that BZD9L1 at low (50 mg/kg) and high (250 mg/kg) doses reduced Ki-67 protein expression, while BZD9L1 at the high dose diminished tumor necrosis compared to vehicle control.ConclusionThese results highlighted the anti-angiogenic potential of BZD9L1 to reduce CRC tumor progression. Furthermore, together with previous anticancer findings, this study provides valuable insights into the potential of BZD9L1 to co-target CRC tumor vasculatures and cancer cells via SIRT1 and/or SIRT2 down-regulation to improve the therapeutic outcome
Autophagy pathway is required for IL-6 induced neuroendocrine differentiation and chemoresistance of prostate cancer LNCaP cells.
Prostate cancer (PCa) cells undergoing neuroendocrine differentiation (NED) are clinically relevant to the development of relapsed castration-resistant PCa. Increasing evidences show that autophagy involves in the development of neuroendocrine (NE) tumors, including PCa. To clarify the effect of autophagy on NED, androgen-sensitive PCa LNCaP cells were examined. Treatment of LNCaP cells with IL-6 resulted in an induction of autophagy. In the absence of androgen, IL-6 caused an even stronger activation of autophagy. Similar result was identified in NED induction. Inhibition of autophagy with chloroquine (CQ) markedly decreased NED. This observation was confirmed by beclin1 and Atg5 silencing experiments. Further supporting the role of autophagy in NED, we found that LC3 was up-regulated in PCa tissue that had relapsed after androgen-deprivation therapy when compared with their primary tumor counterpart. LC3 staining in relapsed PCa tissue showed punctate pattern similar to the staining of chromogranin A (CgA), a marker for NED cells. Moreover, autophagy inhibition induced the apoptosis of IL-6 induced NE differentiated PCa cells. Consistently, inhibition of autophagy by knockdown of beclin1 or Atg5 sensitized NE differentiated LNCaP cells to etoposide, a chemotherapy drug. To identify the mechanisms, phosphorylation of IL-6 downstream targets was analyzed. An increase in phospho-AMPK and a decrease in phospho-mTOR were found, which implies that IL-6 regulates autophagy through the AMPK/mTOR pathway. Most important to this study is the discovery of REST, a neuronal gene-specific transcriptional repressor that is involved in autophagy activation. REST was down-regulated in IL-6 treatment. Knockdown experiments suggest that REST is critical to NED and autophagy activation by IL-6. Together, our studies imply that autophagy is involved in PCa progression and plays a cytoprotective role when NED is induced in PCa cells by IL-6 treatment. These results reveal the potential of targeting autophagy as part of a combined therapeutic regime for NE tumors
Anticancer activities of a benzimidazole compound through sirtuin inhibition in colorectal cancer
Opening the doors of memory: Is declarative memory a natural kind?
Klein's target article argues that autonoetic consciousness is a necessary condition
for memory; this unusually narrow view of the scope of memory implies that only
episodic memory is, strictly speaking, memory. The narrow view is opposed to the
standard broad view, on which causal connection with past experience is sufficient
for memory; on the broad view, both declarative (i.e., episodic and semantic) and
procedural memory count as genuine forms of memory. Klein mounts a convincing
attack on the broad view, arguing that it opens the "doors of memory" too far, but this
commentary contends that the narrow view does not open them far enough. It may
be preferable to adopt an intermediate view of the scope of memory, on which
causal connection is sufficient for memory only when it involves encoding, storage,
and retrieval of content. More demanding than the simple causal condition but less
demanding than the autonoesis condition, the encoding-storage-retrieval condition
implies that both episodic and semantic memory count as genuine forms of memory
but that procedural memory does not
A comprehensive RT-qPCR analysis of microarray data of IL-6 treated LNCaP cells.
<p>A comprehensive RT-qPCR analysis of microarray data of IL-6 treated LNCaP cells.</p
Up-regulation of the autophagy related genes LC3, the NE marker Chromogranin A (CgA), and the down-modulation of REST in relapsed PCa tissue compared to their primary tumor counterpart.
<p>Representative IHC staining images of two pairs of tissue samples, namely primary and relapsed PCa specimens from the same individual, were stained using anti-LC3, anti-CgA, and anti-REST antibodies.</p
Inhibition of autophagy by CQ results in the cell death of IL-6-induced NE differentiated LNCaP cells.
<p>(A) LNCaP cells were cultured in 2.5% CDT supplemented RPMI 1640 and then treated with 100 ng/ml IL-6 or vehicle control in the presence or absence of 50 µM CQ. Cell numbers were analyzed using a MTT cell viability assay at the indicated time points. <i>Points</i>, mean; <i>bars</i>, SD. (B) LNCaP cells were treated as described in (A) and then assayed for caspase-3/7 activity at the indicated time points. Each data point shown is the mean of three independent experiments; <i>bars</i>, SD.</p