Souvick Roy

Prolonged inflammatory microenvironment is crucial for pro-neoplastic growth and genome instability: a detailed review.

IntroductionChronic inflammation can affect the normal cell homeostasis and metabolism by rendering the cells susceptible to genomic instability that may lead to uncontrolled cellular growth and proliferation ensuing tumorigenesis. The causal agents for inflammation may be pathogenic infections like microbial agents ranging from viruses to bacteria. These infections lead to DNA damage or disruption of normal cell metabolism and alter the genome integrity.FindingsIn this review, we have highlighted the role of recurrent infections in tumor microenvironment can lead to recruitment of pro-inflammatory cells, cytokines and growth factors to the site of inflammation. This makes the environment rich in cytokines, chemokines, DNA-damaging agents (ROS, RNS) and growth factors which activate DNA damage response pathway and help in sustained proliferation of the tumor cells. In any inflammatory response, the production of cytokines and related signaling molecules is self-regulating and limiting. But in case of neoplastic risk, deregulation of these factors may lead to abnormalities and related pathogenesis.ConclusionThe scope of the present review is to explore the probable mechanistic link and factors responsible for chronic inflammation. The relation between chronic inflammation and DNA damage response was further elucidated to understand the mechanism by which it makes the cells susceptible to carcinogenesis.

Systematic analysis of integrated gene functional network of four chronic stress-related lifestyle disorders

Background: Stress is a term used to define factors involved in changes in the physiological balances resulting in disease conditions. Chronic exposure to stress conditions in modern lifestyles has resulted in a group of disorders called lifestyle disorders. Genetic background and environmental factors are interrelated to lifestyle in determining the health status of individuals. Hence, identification of disease-associated genes is the primary step toward explanations of pathogenesis of these diseases. In functional genomics, large-scale molecular and physiological data are used for the identification of causative genes associated with a disease. Aim: The objective of our study was to find a common set of genes involved in chronic stress-related lifestyle diseases such as cardiovascular diseases (CVDs), type 2 diabetes (T2D), hypertension (HTN), and obesity. Materials and Methods: In our study, we have performed a systematic analysis of the functional gene network of four chronic stress-related lifestyle diseases by retrieving genes from published databases. We have tried to systematically construct a functional protein-protein interaction (PPI) network. The goals of establishing this network were the functional enrichment study of interacting partners as well as functional disease ontology annotation (FunDO) of the enriched genes. Results: This study enabled the identification of key genes involved in these stress-related lifestyle diseases by prioritizing candidate genes based on their degree of involvement. In this systematic analysis, we have found key genes for these diseases based on their involvement and association at the gene network level and PPI. Conclusion: We have deciphered a group of genes that in combination play a crucial role and may impact the function of the whole genome in the four lifestyle disorders mentioned.

Role of β-catenin in cisplatin resistance, relapse and prognosis of head and neck squamous cell carcinoma

BackgroundHead and neck squamous cell carcinoma (HNSCC) is one of the most common types of cancer in India with high incidence and rapid recurrence rates. Here, we aimed to investigate the role of β-catenin, a developmental pathway gene, in HNSCC therapy resistance, DNA damage response, recurrence and prognosis.MethodsIn total 80 HNSCC samples were included. Western blot, immunohistochemistry and qRT-PCR analyses were performed to assess β-catenin expression in the cut margin and tumor areas of each sample. Kaplan-Meier analyses were performed to correlate β-catenin expression with the survival and prognosis of HNSCC patients. In addition, chemo-resistance, DNA damage response and DNA repair capacities were evaluated in HNSCC-derived cell lines through LiCl-mediated up-regulation and siRNA-mediated silencing of β-catenin expression.ResultsWe observed β-catenin up-regulation in cut margin areas of recurrent patients compared to their corresponding tumor regions, which subsequently could be associated with poor prognosis. In addition, we found that LiCl-mediated up-regulation of β-catenin in HNSCC-derived cells led to cisplatin resistance, evasion of apoptosis, enhanced DNA repair and enhanced migration. The effects of β-catenin silencing correlated with its putative role in chemo-resistance and DNA damage response.ConclusionFrom our results we conclude that β-catenin may contribute to HNSCC therapy resistance and disease relapse. As such, β-catenin may be explored as a therapeutic target along with conventional therapeutics.

p38 MAPK pathway and its interaction with TRF2 in cisplatin induced chemotherapeutic response in head and neck cancer

TRF2 is a telomere binding protein, a component of the shelterin complex that plays a major role in maintaining the integrity of the genome. TRF2 is over-expressed in a number of human cancers including Head and Neck cancer and might play a key role in tumor initiation and development. p38 MAPK signaling pathway is strongly activated in response to various environmental and cellular stresses and thus overexpressed in most of the Head and Neck cancer cases. In this study, we investigated potential interactions of TRF2 with p38 in HNSCC cells and patient samples. Using in silico experiments, we identified interface polar residue Asp-354 of p38 and Arg-492, Arg-496 of TRF2 as protein–protein interaction hotspots. In addition to these interactions, Arg-49 residue of p38 was also found to interact with Glu-456 of TRF2. A detailed understanding of how phosphorylated and unphosphorylated state of p38 protein can influence the stability, specificity and to some extent a conformational change of p38-TRF2 binding is presented. Silencing of TRF2 significantly decreased the phosphorylation of p38 in HNSCC cells which was confirmed by western blot, immunofluorescence and co-immunoprecipitation and alternatively inhibiting p38 using p38 inhibitor (SB 203580) decreased the expression of TRF2 in HNSCC cells. Furthermore, we checked the effect of TRF2 silencing and p38 inhibition in cisplatin induced chemosensitivity of SCC-131 cells. TRF2 silencing and p38 inhibition chemosensitize HNSCC cells to cisplatin. Thus, targeting TRF2 in combinatorial therapeutics can be a treatment modality for Head and Neck cancer which involves inhibition of p38 MAPK pathway.

Role of p38 MAPK in disease relapse and therapeutic resistance by maintenance of cancer stem cells in head and neck squamous cell carcinoma

AIM This study aimed to investigate the role of p38 MAPK in maintenance of cancer stem cell (CSC) phenotype, therapy resistance, and DNA damage repair and response in head and neck squamous cell carcinoma (HNSCC). METHODS In this study, 104 HNSCC patients were included. Western blot, immunohistochemistry, and qPCR analysis were performed to investigate the expression level of p-p38 and CSC markers in cut margin and tumor area of HNSCC patients. The expression level of p-p38 and CSC markers was also evaluated in HNSCC cells with or without p38 inhibitor. Chemoresistance, wound healing capacity, and multicellular tumor spheroids (MCTS) formation capacity were evaluated in HNSCC-derived cell lines with or without p38 inhibitor. In addition, DNA damage response and repair capacities were also evaluated in HNSCC cells after p38 inhibition using alkaline comet assay and γ-H2 AX immunostaining. RESULT We observed that recurrence could be associated with upregulated status of p-p38 and p38α gene in cut margin area of HNSCC patients as compared to tumor region. p38-inhibited cells showed significantly reduced expression of CSC markers, chemosensitivity toward cisplatin, reduced migration potential, and sphere-forming ability along with increased apoptotic population after treatment with increasing concentration of cisplatin. p38-inhibited cells also exhibited significantly increased comet olive tail moment and accumulation of γ-H2 AX, demonstrating increased DNA damage. CONCLUSION This study demonstrated that p38 MAPK activation may play a role in therapeutic resistance and disease relapse in HNSCC by maintenance of CSCs phenotype.

Role of CDKN2A/p16 expression in the prognostication of oral squamous cell carcinoma

OBJECTIVE CDKN2A/p16 is a known tumor suppressor gene with a homologous deletion in Oral Squamous cell carcinoma. CDKN2A/p16 is found to be inactivated in a broad spectrum of solid tumors and in more than 80% of OSCC. Molecular alteration of CDKN2A/p16 in progression of OSCC can pose an important tool for the prognosis of squamous cell carcinoma. MATERIAL AND METHOD Systematic network analysis was carried out to obtain involvement of CDKN2A/p16 in oral cancer by polysearch and FunDO. In the present study we have screened 104 OSCC patients from eastern region of India for CDKN2A/p16 expression in recurrent and non-recurrent OSCC. The observation was validated by Comparative Genomic Hybridisation and Next generation sequencing in recurrent cases. RESULT Systematic analysis revealed direct involvement of CDKN2A/p16 in oral cancer. There was a consistent downregulated expression of CDKN2A/p16 in the recurrent cases. The gene expression study confirmed a >5-fold downregulation of CDKN2A/p16 in recurrent tumors as compared to non-recurrent ones. Array CGH analysis revealed a copy number deletion in the recurrent case. Furthermore, next generation sequencing validated deletion of CDKN2A/p16 and reported it asa common variant with a nonsense mutation having stop /loss of function of the gene in recurrent cases. Recurrent cases with deleted CDKN2A/p16 expression had poor prognosis and low survival rate. CONCLUSION CDKN2A/p16 frequently alters in oral cancer progression with a deletion/loss of function in the recurrent cases displaying its role in aiding several molecular events for the malignant transformations occurring throughout disease progression.

Role of Telomeric TRF2 in Orosphere Formation and CSC Phenotype Maintenance Through Efficient DNA Repair Pathway and its Correlation with Recurrence in OSCC

The major problem to effective treatment of oral cancer is the presence of therapy resistance. Presence of cancer stem cell in the bulk of tumor have been implicated in therapeutic resistance. In this study, we report a non-telomeric role of TRF2 in formation of oral cancer spheroids and CSC phenotype maintenance via an efficient DNA damage repair mechanism in the presence of chemotherapeutic insult. We report reduced sphere formation efficiency and reduced spheroid size in TRF2 silenced oral cancer cell lines. TRF2 silenced orospheres further reported reduced proliferative capacity as compared to non-silenced orospheres. Furthermore, TRF2 silencing hampered the migratory potential of oral cancer cell line and also reduced the expression of several CSC markers like CD44, Oct4, Sox2, KLF4 and c-Myc along with β-catenin and hTERT molecules both in Cal27 cell line and generated orospheres. TRF2 silencing impaired efficient DNA damage repair capacity of non-orospheric and orospheric cells and repressed ERCC1 expression levels when treated with Cisplatin. TRF2 overexpression was also observed to correlate with poor overall survival and disease relapse of OSCC patients. In silico studies further identified several amino acid residues that show high binding affinity and strong protein-protein interactions among TRF2 and CSC marker KLF4. Hence, our report confirms a non-telomeric role of TRF2 in spheroid generation, maintenance of CSC phenotype and efficient DNA damage repair capacity contributing to chemotherapy resistance in oral cancer cell line. We further iterate the use of TRF2 as a prognostic marker in OSCC for faster detection and improved survival.

The role of p38 MAPK pathway in p53 compromised state and telomere mediated DNA damage response

There is an intricate balance of DNA damage response and repair which determines the homeostasis of human genome function. p53 protein is widely known for its role in cell cycle regulation and tumor suppressor activity. In case of several cancers where function of p53 gene gets compromised either by mutation or partial inactivation, the role of p53 in response to DNA damage needs to be supplemented by another molecule or pathway. Due to sedentary lifestyle and exposure to genotoxic agents, genome is predisposed to chronic stress, which ultimately leads to unrepaired or background DNA damage. p38 MAPK signaling pathway is strongly activated in response to various environmental and cellular stresses. DNA damage response and the repair options have crucial links with chromosomal integrity. Telomere that regulates integrity of genome is protected by a six member shielding unit called shelterin complex which communicates with other pathways for functionality of telomeres. There are evidences that p38 gets activated through ATM in response to DNA damage. Dysfunctional telomere leads to activation of ATM which subsequently activates p38 suggesting a crosstalk between p38, ATM and shelterin complex. This review focuses on activation of p38 in response to genotoxic stress induced DNA damage in p53 mutated or compromised state and its possible cross talk with telomere shelterin proteins. Thus p38 may act as an important target to treat various diseases and in majority of cancers in p53 mutated state.