Sujit Suklabaidya

TLR4 activation by lipopolysaccharide confers survival advantage to growth factor deprived prostate cancer cells

Prostate cancer (PCa) cells express Toll‐like receptor‐4 (TLR4), a known pro‐tumorigenic molecule for different cancer cells. The cancer cells residing in the avascular region of the tumor confront various metabolic stresses and continuously adapt mechanisms to overcome them. We hypothesized that TLR4 activation might provide direct survival advantage to metabolically stressed PCa cells.

Chitohexaose protects against acetaminophen-induced hepatotoxicity in mice

Acetaminophen (N-acetyl-para-aminophenol (APAP)) toxicity causes acute liver failure by inducing centrilobular hepatic damage as a consequence of mitochondrial oxidative stress. Sterile inflammation, triggered by hepatic damage, facilitates gut bacterial translocation leading to systemic inflammation; TLR4-mediated activation by LPS has been shown to have a critical role in APAP-mediated hepatotoxicity. In this study, we demonstrate significant protection mediated by chitohexaose (Chtx) in mice challenged with a lethal dose of APAP (400 mg/kg b.w.). Decreased mortality by Chtx was associated with reduced hepatic damage, increased peritoneal migration of neutrophils, decreased mRNA expression of IL-1β as well as inhibition of inflammasome activation in liver. Further, an alternate mouse model of co-administration of a sublethal doses of APAP (200 mg/kg b.w.) and LPS (5 mg/kg b.w.) operating synergistically and mediating complete mortality was developed. Overwhelming inflammation, characterized by increased inflammatory cytokines (TNF-α, IL-1β and so on) in liver as well as in circulation and mortality was demonstrable in this model. Also, Chtx administration mediated significant reversal of mortality in APAP+LPS co-administered mice, which was associated with reduced IL-1β in liver and plasma cytokines in this model. In conclusion, Chtx being a small molecular weight linear carbohydrate offers promise for clinical management of liver failure associated with APAP overdose.

Characterization and use of HapT1-derived homologous tumors as a preclinical model to evaluate therapeutic efficacy of drugs against pancreatic tumor desmoplasia

Desmoplasia in human pancreatic cancer (PC) promotes cancer progression and hinders effective drug delivery. The objectives of this study were to characterize a homologous orthotopic model of PC in Syrian golden hamster and investigate the effect of anti-fibrotic (pirfenidone), antioxidant (N-acetyl cysteine, NAC) and anti-addiction (disulfiram, DSF) drugs on desmoplasia and tumor growth in this model. The HapT1 PC cells when implanted orthotopically into hamsters formed tumors with morphological, cellular and molecular similarities to human PC. Protein profiling of activated hamster pancreatic stellate cells (ha-PSCs) revealed expression of proteins involved in fibrosis, cancer cells growth and metastasis. Pirfenidone, suppressed growth of HapT1 cells and the desmoplastic response in vivo; these effects were enhanced by co-administration of NAC. Disulfiram alone or in combination with copper (Cu) was toxic to HapT1 cells and PSCs in vitro; but co-administration of DSF and Cu accelerated growth of HapT1 cells in vivo. Moreover, DSF had no effect on tumor-associated desmoplasia. Overall, this study identifies HapT1-derived orthotopic tumors as a useful model to study desmoplasia and tumor-directed therapeutics in PC. Pirfenidone in combination with NAC could be a novel combination therapy for PC and warrants investigation in human subjects.

Experimental models of pancreatic cancer desmoplasia

Desmoplasia is a fibro-inflammatory process and a well-established feature of pancreatic cancer. A key contributor to pancreatic cancer desmoplasia is the pancreatic stellate cell. Various in vitro and in vivo methods have emerged for the isolation, characterization, and use of pancreatic stellate cells in models of cancer-associated fibrosis. In addition to cell culture models, genetically engineered animal models have been established that spontaneously develop pancreatic cancer with desmoplasia. These animal models are currently being used for the study of pancreatic cancer pathogenesis and for evaluating therapeutics against pancreatic cancer. Here, we review various in vitro and in vivo models that are being used or have the potential to be used to study desmoplasia in pancreatic cancer.

Plant-derived SAC domain of PAR-4 (Prostate Apoptosis Response 4) exhibits growth inhibitory effects in prostate cancer cells

The gene Par-4 (Prostate Apoptosis Response 4) was originally identified in prostate cancer cells undergoing apoptosis and its product Par-4 showed cancer specific pro-apoptotic activity. Particularly, the SAC domain of Par-4 (SAC-Par-4) selectively kills cancer cells leaving normal cells unaffected. The therapeutic significance of bioactive SAC-Par-4 is enormous in cancer biology; however, its large scale production is still a matter of concern. Here we report the production of SAC-Par-4-GFP fusion protein coupled to translational enhancer sequence (5′ AMV) and apoplast signal peptide (aTP) in transgenic Nicotiana tabacum cv. Samsun NN plants under the control of a unique recombinant promoter M24. Transgene integration was confirmed by genomic DNA PCR, Southern and Northern blotting, Real-time PCR, and Nuclear run-on assays. Results of Western blot analysis and ELISA confirmed expression of recombinant SAC-Par-4-GFP protein and it was as high as 0.15% of total soluble protein. In addition, we found that targeting of plant recombinant SAC-Par-4-GFP to the apoplast and endoplasmic reticulum (ER) was essential for the stability of plant recombinant protein in comparison to the bacterial derived SAC-Par-4. Deglycosylation analysis demonstrated that ER-targeted SAC-Par-4-GFP-SEKDEL undergoes O-linked glycosylation unlike apoplast-targeted SAC-Par-4-GFP. Furthermore, various in vitro studies like mammalian cells proliferation assay (MTT), apoptosis induction assays, and NF-κB suppression suggested the cytotoxic and apoptotic properties of plant-derived SAC-Par-4-GFP against multiple prostate cancer cell lines. Additionally, pre-treatment of MAT-LyLu prostate cancer cells with purified SAC-Par-4-GFP significantly delayed the onset of tumor in a syngeneic rat prostate cancer model. Taken altogether, we proclaim that plant made SAC-Par-4 may become a useful alternate therapy for effectively alleviating cancer in the new era.

Pancreatic fibroblast exosomes regulate survival of cancer cells

With high enthusiasm, we enjoyed reading the recent article published by Richards et al. where they have shown that exosomes secreted by gemcitabine (GEM)-treated cancer-associated fibroblasts (CAFs) promote proliferation and drug resistance of pancreatic cancer cells (PCCs).1 The exosomes released from GEM-treated CAFs when added to recipient PCCs, it upregulated Snail expression, which is known to confer resistance to GEM.2 At the same time, GEM is already known to upregulate Snail expression in PCCs.2 Here an important point that deserves clarification is ‘exosomes isolated from GEM-treated CAFs might carry some amount of GEM and/or its active metabolites and transfer it to PCCs’ (Figure 1). In recent past, it has been shown that conditioned medium (CM) of pancreas-derived mesenchymal stromal cells primed with GEM-induced cytotoxicity in CFPAC-1 PCCs.3 Moreover, exosomal release of another chemotherapeutic drug, paclitaxel from treated mesenchymal cells has already been reported.4 Hence, it is highly possible that after GEM treatment to CAFs, some amount of GEM and/or its active metabolites will be retained inside CAFs and later on gets released to the CM (Figure 1). As a consequence, GEM and/or its metabolites present in CAFs’ exosomes will be directly released into PCCs and induce Snail expression in PCCs.2 Though the mechanism(s) behind GEM-mediated Snail expression in PCCs is not clear, but GEM-induced Hif1α upregulation in PCCs5 followed by Hif1α-mediated Snail overexpression6 might be a potential mechanism (Figure 1). In our ongoing study, we have observed that CM (GEM treatment for 24 h followed by conditioning for 24 h) collected from pancreatic stellate cells (GEM-PSCs) confers less survival advantage to GEM-treated PCCs compared with 24 h CM collected from GEM-PSCs with a second round of media replacement after 8 h of first removal. Together, it indicates that CM from GEM-treated CAFs might contain some level of drug that either sensitizes or alters PCCs property toward additional GEM treatment. To induce cytotoxicity, the GEM concentration present in the exosomes should be at the optimal level; however, GEM concentration that is required for HIF1α and/or Snail upregulation might be below the cytotoxic dose.5 Together, it is potentially possible that in the current study the exosomes obtained from 5 × 105 numbers of CAFs treated with 1 μM of GEM might have GEM concentration that is not cytotoxic but good enough to induce Snail expression in PCCs. Therefore, transfer of GEM through CAF-derived exosomes into PCCs and its role in induction of chemoresistance warrants further investigation.

MAGEA3 (melanoma antigen family A, 3)

In the year 1991 Van der Bruggen P et al. cloned and named MAGE-1 gene that encodes MZ2E antigen, which is expressed in melanoma tissues and cell lines (van der Bruggen et al., 1991).Since then based on sequence similarity MAGE family has expanded to more than 60 genes (Chomez et al., 2001).According to their chromosomal location and tissue-specific expression pattern, all the members of this family are categorized into two groups; type I (cancer and testis specific) and type II (ubiquitous) MAGE.MAGEA sub-family has 12 members starting from MAGEA1 to MAGEA12, among them MAGEA7 is a pseudo-gene (Doyle et al., 2010).The current review summarizes the information specifically on MAGEA3s DNA/RNA, protein structure, function and where the gene is implicated.

Dual role of Par-4 in abrogation of EMT and switching on Mesenchymal to Epithelial Transition (MET) in metastatic pancreatic cancer cells

Epithelial‐mesenchymal transition (EMT) is a critical event that occurs during the invasion and metastatic spread of cancer cells. Here, we conceive a dual mechanism of Par‐4‐mediated inhibition of EMT and induction of MET in metastatic pancreatic cancer cells. First, we demonstrate that 1,1′‐β‐D‐glucopyranosyl‐3,3′‐bis(5‐bromoindolyl)‐octyl methane (NGD16), an N‐glycosylated derivative of medicinally important phytochemical 3,3′‐diindolylmethane (DIM) abrogates EMT by inducing pro‐apoptotic protein Par‐4. Induction of Par‐4 (by NGD16 or ectopic overexpression) strongly impedes invasion with inhibition of major mesenchymal markers viz. Vimentin and Twist‐1 epithelial marker‐ E‐cadherin. Further, NGD16 triggers MET phenotypes in pancreatic cancer cells by augmenting ALK2/Smad4 signaling in a Par‐4‐dependent manner. Conversely, siRNA‐mediated silencing of endogenous Par‐4 unveil reversal of MET with diminished E‐cadherin expression and invasive phenotypes. Additionally, we demonstrate that intact Smad4 is essential for Par‐4‐mediated maintenance of E‐cadherin level in MET induced cells. Notably, we imply that Par‐4 induction regulates E‐cadherin levels in the pancreatic cancer cells via modulating Twist‐1 promoter activity. Finally, in vivo studies with syngenic mouse metastatic pancreatic cancer model reveal that NGD16 strongly suppresses metastatic burden, ascites formation, and prolongs the overall survival of animals effectively.

Enhanced anti-metastatic and anti-tumorigenic efficacy of Berbamine loaded lipid nanoparticles in vivo

Research on metastasis is gaining momentum for effective cancer management. Berbamine (BBM) has the potency to act as a therapeutic in multiple cancers and cancer metastasis. However, the major limitation of the compound includes poor bioavailability at the tumor site due to short plasma half-life. Here, our major objective involved development of lipid based nanoparticles (NPs) loaded with BBM with an aim to circumvent the above problem. Moreover its, therapeutic potentiality was evaluated through various in vitro cellular studies and in vivo melanoma primary and experimental lung metastatic tumor model in C57BL/6 mice. Results of different cellular experiments demonstrated enhanced therapeutic efficacy of BBM-NPs in inhibiting metastasis, cell proliferation and growth as compared to native BBM in highly metastatic cancer cell lines. Further, in vivo results demonstrated suppression of primary B16F10 melanoma tumor growth in C57BL/6 mice model treated with BBM-NPs than that of native BBM. Importantly, a moderately cytotoxic dose of BBM-NPs was able to significantly suppress the incidence of B16F10 cells lung metastasis in vivo. Results indicated development of an effective approach for aggressive metastatic cancer.

Urinary bladder stone associated with seminal vesicle and prostate infection in a Copenhagen rat

We report a very rare case of urinary bladder stone in a laboratory rat, which was associated with severe prostatitis and seminal vesiculitis. Importantly, the histopathological analysis revealed the rare variety of keratinizing desquamative squamous metaplasia of bladder, prostate, and seminal vesicle epithelium. Immunohistochemistry for alpha smooth muscle actin protein and aniline blue staining for collagen clearly showed interstitial prostate fibrosis. The detail information about these findings and subsequent discussion are provided here.