RESUMO
BACKGROUND: This study aims to evaluate the role of cancer stem cell marker, CD44, and its ligand HA as potential molecular biomarker for early detection of HNSCC. METHODS AND RESULTS: The expression profile (mRNA/Protein) of CD44 variants were analysed in primary HNSCC lesions and plasma of the patients. Then, prevalence of HA variants was analysed in plasma of the patients. The mRNA expression of CD44 variants, CD44S and CD44v3, were significantly high in both early (stage I/II) and late (stage III/IV) invasive lesions, with predominant expression of CD44v3 in the late-stage lesions. In plasma of HNSCC patients, increased levels of SolCD44, CD44-ICD and unique 62 KD CD44 variants with respect to standard CD44S were seen, in comparison to their prevalence in plasma of normal individuals. The abundance of CD44-ICD and 62 KD variants were significantly high in plasma of late stage HNSCC patients. Interestingly, significantly high level of low molecular weight HA(LMW HA) with respect to high molecular weight HA(HMW HA) was seen in plasma of HNSCC patients irrespective of clinical stages. On the contrary, high HMW HA level in plasma of normal individuals was seen. The high level of LMW HA in plasma of HNSCC patients might be due to combinatorial effect of increased mRNA expression of HA synthesizing enzyme HAS1/2/3 and HA degrading enzyme HYAL1/2, as seen in the primary HNSCC samples. CONCLUSION: Thus, our data revealed the importance of specific CD44 and HA variants in plasma of HNSCC patients during its development as potential non-invasive molecular biomarker of the disease.
Assuntos
Neoplasias de Cabeça e Pescoço , Ácido Hialurônico , Humanos , Relevância Clínica , Prevalência , Ligantes , Peso Molecular , Carcinoma de Células Escamosas de Cabeça e Pescoço , RNA Mensageiro , Neoplasias de Cabeça e Pescoço/genética , Biomarcadores , Receptores de Hialuronatos/genéticaRESUMO
Cervical cancer is one of the leading causes of cancer death among females, worldwide. The contributory role of different cellular pathways in the process of carcinogenesis is still poorly understood. Our study was focused here to understand the functional evaluation of key regulatory genes of FA-BRCA pathway in the development of CACX and their role in chemo-tolerance of the disease by analyzing the molecular profile of the genes both in normal and tumour tissue of our sample pool, also validated in in silico datasets. Later on, prognostic importance of the genes was further evaluated in plasma DNA and cisplatin-treated in vitro system. We found that expression profile of FA-BRCA pathway genes was gradually reduced from undifferentiated basal-parabasal layers of normal tissue towards the progression of the disease. Further analysis revealed that frequent promoter methylation [32-55%] and deletion [34-52%] events were the plausible reasons for their reduced expression in CACX. Noticeably, invasion of promoter methylation of the genes [11-17%] in plasma CTCs of CACX patients was positively correlated [p < 0.001] with poor prognosis among patients. On the other hand, functional upregulation of these genes at higher concentrations [IC50-70] of cisplatin was a predictor for the development of drug tolerance, as evaluated in our in vitro study. This finding was supported further by low prevalence of γ-H2X foci formation and reduced expression of DNMT1 at higher concentrations of cisplatin. In totality, we discovered that the FA-BRCA pathway must be inactivated for cancer formation. In contrast, elevated gene expression played a substantial role in building of chemo-tolerance and might be associated with developing increased risk of disease recurrence among patients.
Assuntos
Neoplasias do Colo do Útero , Feminino , Humanos , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismo , Cisplatino/farmacologia , Cisplatino/uso terapêutico , Regulação para Baixo , Recidiva Local de Neoplasia , PrognósticoRESUMO
Cervical cancer (CACX) is one of the top causes of cancer death in women globally. The involvement of several cellular pathways in carcinogenesis is still poorly understood. Here, we focused to evaluate the contributory role of Mismatch Repair (MMR) pathway genes-MLH1 and MSH2 in CACX and their association with chemo-tolerance of the disease. For this purpose, molecular profiles (expression/promoter methylation/deletion) of the genes were analysed in both normal cervical epithelium and tumour tissue, also validated in in-silico dataset as well. Later on, prognostic importance of the genes was identified through analysis of their methylation/expression status in plasma DNA of circulating tumour cells (CTCs) and cisplatin-tolerant CACX cell lines respectively. It was found that the expression profile of MLH1 and MSH2 genes was considerably reduced from undifferentiated basal-parabasal layers of normal cervical epithelium towards progression of the disease. Further analysis showed that frequent deletion [34-48%] and promoter methylation events [28-46%] of the genes were the plausible reasons for their reduced expression during tumorigenesis. Incidentally, the prevalence of MLH1 [32%] and MSH2 [27%] promoter methylation found in CTCs of plasma of the clinically advanced CACX patients implicated their prognostic importance of the disease. In addition, the patients having high alterations of those genes resulted in poor patient outcomes even after the therapy. In in-depth analysis of this result in cisplatin-tolerant CACX cell lines, we discovered that increased promoter methylation frequency of those genes at higher concentrations of cisplatin and gradual accumulation of the cells in the G2/M phase of the cell cycle were the rational causes for their reduced expression and MMR deficiency in the system. Hence, it is possible to conclude that the gradual down-regulation of MLH1 and MSH2 proteins may be a key event for MMR pathway inactivation in CACX. This might also be associated with chemo-tolerance and overall poor survival among the patients.
Assuntos
Neoplasias do Colo do Útero , Humanos , Feminino , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismo , Proteína 2 Homóloga a MutS/genética , Proteína 2 Homóloga a MutS/metabolismo , Cisplatino/farmacologia , Cisplatino/uso terapêutico , Colo do Útero/patologia , Carcinogênese/genética , Carcinogênese/metabolismo , Transformação Celular Neoplásica/metabolismo , Proteína 1 Homóloga a MutL/genética , Proteína 1 Homóloga a MutL/metabolismoRESUMO
Gamma-papillomaviruses, though traditionally classified as cutaneotropic, actual tissue tropism is largely unexplored. This study aimed to evaluate the tissue-specific prevalence of two novel-HPV 223 and 225 in samples of oral mucosa and keratinized epithelium of varied skin parts from 226 female and male subjects, with or without neoplastic/dysplastic lesions in oral cavity or cervix. The gamma-human papillomavirus (gamma-HPV) 223 and 225 DNA presences were determined by polymerase chain reaction (PCR) ursing the HPV type-specific primers and confirmed by Sanger sequencing. Viral load in the HPV 223 and HPV 225 positive samples were determined by absolute real-time quantification method. Alpha-HPV DNA prevalence was also checked in oral mucosa to ascertain coinfection status. Novel HPV 223 was present in 4.4% (10/226) oral mucosal samples of the study population; interestingly all were females with no prevalence in their corresponding skin swab samples. Whereas, the prevalence of HPV 225 was found both in the skin and oral mucosa of 28.2% (N = 37/131) female and 17.9% (N = 17/95) male participants. Alongside, HPV 223 viral load was found to be significantly higher (p = 0.02 < 0.05) in the oral mucosa of diseased participants, whereas, HPV 225 viral load was higher in the oral mucosa of normal participants. Our results suggest that gamma-HPV 223 has its prevalence only in the oral mucosal epithelium, whereas, HPV 225 has its prevalence on both mucosal and keratinized skin epithelium, indicating its dual tropism nature.
Assuntos
Infecções por Papillomavirus , Humanos , Masculino , Feminino , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/epidemiologia , Boca , Mucosa Bucal , Papillomaviridae/genética , Pele , Papillomavirus Humano , DNA Viral/genética , DNA Viral/análiseRESUMO
Cervical carcinoma (CACX) is still a dreadful threat to women in developing countries. Available conventional chemo-radiation therapies are not sufficient to restrict the disease recurrence. To unravel the mechanism of the disease recurrence, alteration of hedgehog self-renewal pathway was evaluated during development of CACX and in chemo-tolerance of the tumor. We have analyzed the alterations (expression/methylation/deletion) of some key regulatory genes (HHIP/SUFU/SHH/ SMO/GLI1) of hedgehog self-renewal pathway in cervical lesions at different clinical stages and compared with different datasets, followed by their clinico-pathological correlations. The changes in expression/methylation of the genes were then evaluated in two CACX cell lines (SiHa/HeLa) after treatment with chemotherapeutic drug cisplatin at different concentrations. Down regulation (mRNA/protein) of the antagonists HHIP and SUFU due to promoter methylation and/or deletion along with upregulation (protein) of agonists SHH, SMO and GLI1 was seen in early invasive lesions and subsequent clinical stages. Reduced protein expression of HHIP and SUFU showed significant association with high/intermediate expression of agonists SHH, SMO, GLI1 in the tumors and also poor prognosis of the patients. It was evident that cisplatin could restrict the growth of HeLa and SiHa cells through significant upregulation of antagonists HHIP and SUFU due to their promoter hypomethylation and down regulation of SHH in a concentration dependent manner without any significant changes in expression of SMO and GLI1, leading to the tumor cells in a dormant state. Thus, interplay of the agonists and antagonists has important role in activation of hedgehog pathway during development of CACX, whereas inactivation of the pathway due to upregulation of the antagonists is an important phenomenon in chemo-tolerance of the tumor. This suggests importance of epigenetic modification in chemo-resistance of CACX.
Assuntos
Carcinoma , Neoplasias do Colo do Útero , Humanos , Feminino , Proteínas Hedgehog/genética , Transdução de Sinais/genética , Proteína GLI1 em Dedos de Zinco/genética , Proteína GLI1 em Dedos de Zinco/metabolismo , Cisplatino/farmacologia , Recidiva Local de Neoplasia , Neoplasias do Colo do Útero/genéticaRESUMO
PURPOSE: Cervical Squamous Cell Carcinoma (CSCC) is one of the significant causes of cancer deaths among women. Distinct genetic and epigenetic-altered loci, including chromosomal 11p15.5-15.4, have been identified. CDKN1C (Cyclin-Dependent Kinase Inhibitor 1C, p57KIP2), a member of the CIP/KIP family of cyclin-dependent kinase inhibitors (CDKIs), located at 11p15.4, is a putative tumor suppressor. Apart from transcriptional control, S-Phase Kinase Associated Protein 2 (SKP2), an oncogenic E3 ubiquitin ligase, regulates the protein turnover of CDKN1C. But the molecular status of CDKN1C in CSCC and the underlying mechanistic underpinnings have yet to be explored. METHODS: TCGA and other publicly available datasets were analyzed to evaluate the expression of CDKN1C and SKP2. The expression (transcript/protein) was validated in independent CSCC tumors (n = 155). Copy number alteration and promoter methylation were correlated with the expression. Finally, in vitro functional validation was performed. RESULTS: CDKN1C was down-regulated, and SKP2 was up-regulated at the transcript and protein levels in CSCC tumors and the SiHa cell line. Notably, promoter methylation (50%) was associated with the downregulation of the CDKN1C transcript. However, high expression of SKP2 was found to be associated with the decreased expression of CDKN1C protein. Independent treatments with 5-aza-dC, MG132, and SKP2i (SKPin C1) in SiHa cells led to an enhanced expression of CDKN1C protein, validating the mechanism of down-regulation in CSCC. CONCLUSION: Collectively, CDKN1C was down-regulated due to the synergistic effect of promoter hyper-methylation and SKP2 over-expression in CSCC tumors, paving the way for further studies of its role in the pathogenesis of the disease.
Assuntos
Carcinoma de Células Escamosas , Neoplasias do Colo do Útero , Feminino , Humanos , Carcinoma de Células Escamosas/genética , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Inibidor de Quinase Dependente de Ciclina p57/metabolismo , Regulação para Baixo/genética , Metilação , Proteínas Quinases Associadas a Fase S/genética , Proteínas Quinases Associadas a Fase S/metabolismo , Neoplasias do Colo do Útero/genéticaRESUMO
In this study, we aimed to understand the interplay of the epigenetic modifier genes DNMT1 and TET1 along with HPV infection in the cervical epithelium and how it changes during tumorigenesis. For this purpose, initially the bioinformatical analysis (methylation and expression profile) of DNMT1 and TET1 was analyzed in the TCGA dataset. Next genetic (deletion) and epigenetic profiling (promoter methylation) of DNMT1 and TET1 were done in our sample pool and also validated in CACX cell lines as well. The results were further correlated with different clinicopathological parameters. Our data revealed that HPV infection in basal/parabasal layers of cervical epithelium actually disrupts the epigenetic homeostasis of DNMT1 and TET1 proteins which ultimately leads to the high expression of DNMT1 along with further reduction in TET1 protein during the development of carcinoma. Further, in-depth look into the results revealed that comparatively low methylation frequency of DNMT1 coupled with high promoter methylation and deletion frequency [22-46%] of TET1 were the plausible reasons of their antagonistic expression profile during the progression of the disease. Interestingly, the prevalence of DNMT1 [9.1%] and TET1 promoter methylation [22.7%] found in both the plasma DNA of the respective CACX patients implicated its diagnostic importance in this study. Lastly, molecular alteration of TET1 alone or in combination with DNMT1 showed the worst overall survival among the patients. Hence, it may be concluded that an inverse molecular profile of DNMT1 and TET1 genes seen in the proliferative basal-parabasal layers of the cervical epithelium was aggravated during the development of CACX along with genetic and epigenetic changes due to HPV infection.
Assuntos
Carcinoma , Infecções por Papillomavirus , Neoplasias do Colo do Útero , Feminino , Humanos , Carcinoma/genética , Metilação de DNA , Regulação Neoplásica da Expressão Gênica , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Infecções por Papillomavirus/genética , Prognóstico , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , DNA (Citosina-5-)-Metiltransferase 1/genética , DNA (Citosina-5-)-Metiltransferase 1/metabolismoRESUMO
We aimed to understand the crosstalk between mismatch repair (MMR) and FA-BRCA pathway in primary bladder carcinoma (BlCa) samples as well as in chemotolerant cell line. We analysed the genetic alterations of MLH1 and MSH2 (MMR-related genes) and after that we correlated it with the nuclear translocation of FANCD2 protein. Next, we evaluated this crosstalk in T24 BlCa cell line in response to doxorubicin treatment. In primary BlCa tumors, infrequent genetic deletion (17-20%) but frequent promoter methylation (28-55%) of MLH1 and MSH2 was observed, where MLH1 was significantly (p < 0.05) more methylated among the early staged samples (NMIBC). However, MSH2 was significantly more altered among the NMIBC samples, signifying the importance of MMR pathway during the early pathogenesis of the disease. Furthermore, BlCa samples with underexpressed MLH1/MSH2 protein possessed cytoplasmic FANCD2 protein; encouraging that inefficiency of MMR proteins might restrict FANCD2 nuclear translocation. Next, we analysed publicly available data in GEO2R tool where we observed that in response to chemotherapeutic drugs, expression of MLH1, MSH2 and FANCD2 were diminishing. Validating this result in doxorubicin tolerant T24 cells, we found that expression of MLH1 and MSH2 was gradually decreased with increasing dose of doxorubicin. Interestingly, FANCD2 mono-ubiquitination (L-form) was also reduced in chemotolerant T24 cells. The crosstalk between MMR and FA-BRCA pathway was substantiated in the primary BlCa tumors. Further, in response to doxorubicin, this crosstalk was found to be hampered due to under-expression of MLH1 and MSH2 gene, thereby rendering chemotolerance.
Assuntos
Carcinoma , Neoplasias da Bexiga Urinária , Humanos , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/genética , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/metabolismo , Proteína 2 Homóloga a MutS/genética , Proteína 2 Homóloga a MutS/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Bexiga Urinária/metabolismo , Reparo de Erro de Pareamento de DNA , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/genética , Doxorrubicina , Proteína 1 Homóloga a MutL/genética , Proteína 1 Homóloga a MutL/metabolismoRESUMO
Squamous cell carcinoma of the uterine cervix (CSCC) is one of the leading causes of death in Indian women. Protein tyrosine phosphatase receptor (PTPR) type J (also known as DEP1) is a recently reported tumour suppressor receptor phosphatase. Critical molecular analysis of PTPRJ/DEP1 (11p11.2) has not performed in CSCC to date. Here, we observed frequent downregulation of cancer samples (n=31) at the transcriptional level. Immunohistochemistry revealed concordant low expression of PTPRJ protein with a few samples showing intermediate expression. To probe for the cause of such downregulation of the gene in CSCC (n=155), we analysed the copy number and promoter methylation of PTPRJ. The genetic locus showed deletion (14.8%) and the promoter showed methylation (33.5%) of PTPRJ. To the best of our knowledge, for the first time we explored the molecular status of PTPRJ although we observed no statistically significant association with the prognosis of Indian CSCC patients (n=76). However, we observed enhanced expression of PTPRJ protein levels that contributes to effective cisplatin chemotherapy in the SiHa cell line. Thus, the present study paves the way for further research into the plausible mechanisms of downregulation of PTPRJ in cervical cancer.
Assuntos
Carcinoma de Células Escamosas , Neoplasias do Colo do Útero , Carcinoma de Células Escamosas/genética , Regulação para Baixo , Feminino , Humanos , Imuno-Histoquímica , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/genética , Neoplasias do Colo do Útero/genéticaRESUMO
BACKGROUND: Impulsivity (Imp), being one of the cardinal symptoms of Attention Deficit Hyperactivity Disorder (ADHD), often leads to inappropriate responses to stimuli. Since the dopaminergic system is the primary target for pharmaceutical intervention in ADHD, we investigated the association between ADHD-related Imp and functional gene variants of the dopamine transporter (SLC6A3) and catechol-O-methyltransferase involved in dopamine clearance. METHODS AND RESULTS: Indo-Caucasoid families with ADHD probands (N = 217) were recruited based on the Diagnostic and Statistical Manual of Mental Disorders (DSM). Imp of the probands was assessed using the Domain Specific Imp Scale for Children and DSM. Peripheral blood was collected after obtaining informed written consent for participation, genomic DNA was isolated, and target sites were genotyped by DNA sequencing. The association of genetic variants with Imp was examined by the Quantitative trait analysis (QTA) and Analysis of variance (ANOVA). Post-Hoc analysis following QTA and ANOVA showed significant associations of rs2254408, rs2981359, and rs2239393 with different domains of Imp (P < 0.05). Various haplotypic combinations also showed statistically significant associations with Imp (P < 0.05). Multifactor dimensionality reduction models revealed strong effects of the variants on Imp. ADHD probands harboring the risk alleles exhibited a deficit in performance during cognitive assessment. Longitudinal follow-up revealed a significant association of rs2254408 with trait persistence. CONCLUSION: The present study indicates the influence of the studied genetic variants on ADHD-associated imp, executive deficit, and disease persistence. Thus, these variants may be helpful as predictors for the success of individual therapeutic sessions during cognitive training.
Assuntos
Transtorno do Deficit de Atenção com Hiperatividade , Criança , Humanos , Transtorno do Deficit de Atenção com Hiperatividade/diagnóstico , Transtorno do Deficit de Atenção com Hiperatividade/genética , Catecol O-Metiltransferase/genética , Cognição , Dopamina , Comportamento Impulsivo , Índice de Gravidade de DoençaRESUMO
PURPOSE: The study was aimed to understand the importance of the hedgehog signaling pathway in development of head and neck squamous cell carcinoma (HNSCC). METHODS: The molecular profiles of the key regulatory genes of the pathway were analysed in the adjacent normal epithelium and tumor samples. The findings were validated in HNSCC cell line. RESULTS: In the bioinformatical analysis, severe reduction in the expression of HHIP was evident in the datasets. The protein and mRNA expression studies in our sample pool revealed interplay of various isoforms of PTCH1 gene (PTCH1-1 and 1B) together with high/medium expression of GLI, SHH, SMO and HHIP in the basal/parabasal layers of the normal epithelium. As the disease progressed, severe downregulation of HHIP coupled with upregulation of GLI1 and differential expression pattern of various PTCH1 gene isoform was evident. Promoter methylation analysis of PTCH1 gene revealed the involvement of more than one promoter of PTCH1 in regulating the expression of different isoform of this gene during tumorigenesis. Treating the FaDu cell line with the demethylating agent 5-aza-2'-deoxycytidine reversed the methylation effects of HHIP and PTCH1 and de-activated the pathway. Also, reduced expression of HHIP-AS1 was observed in our sample pool suggesting multiple ways of regulation of the HHIP gene. Lastly, the patients with under expression of HHIP, HHIP-AS1, high expression of GLI1 showed worse five-year over-all survival trend. CONCLUSION: Dynamic promoter switching of PTCH1 and frequent inactivation of HHIP are the key regulatory events of hedgehog pathway activation in HNSCC.
Assuntos
Proteínas de Transporte , Neoplasias de Cabeça e Pescoço , Glicoproteínas de Membrana , Receptor Patched-1 , Carcinoma de Células Escamosas de Cabeça e Pescoço , Proteínas de Transporte/genética , Regulação para Baixo , Neoplasias de Cabeça e Pescoço/genética , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Humanos , Glicoproteínas de Membrana/metabolismo , Receptor Patched-1/genética , Regiões Promotoras Genéticas , Transdução de Sinais , Carcinoma de Células Escamosas de Cabeça e Pescoço/genéticaRESUMO
Bladder cancer carries a poor prognosis and has proven resistance to chemotherapy. Pentacyclic Triterpenoid Acids (PTAs) are natural bioactive compounds that have a well-known impact on cancer research because of their cytotoxic and chemopreventive activities. This review focuses on bladder cancer which can no longer be successfully treated by DNA damaging drugs. Unlike most of the existing drugs against bladder cancer, PTAs are non-toxic to normal cells. Collecting findings from both in vitro and in vivo studies, it has been concluded that PTAs may serve as promising agents in future bladder cancer therapy. In this review, the roles of various PTAs in bladder cancer have been explored, and their mechanisms of action in the treatment of bladder cancer have been described. Specific PTAs have been shortlisted from each of the chief skeletons of pentacyclic triterpenoids, which could be effective against bladder cancer because of their mode of action. This review thereby throws light on the multi targets and mechanisms of PTAs, which are responsible for their selective anticancer effects and provides guidelines for further research and development of new natural antitumor compounds.
Assuntos
Antineoplásicos , Triterpenos , Neoplasias da Bexiga Urinária , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Humanos , Triterpenos/farmacologia , Triterpenos/uso terapêutico , Neoplasias da Bexiga Urinária/tratamento farmacológicoRESUMO
Amarogentin (active component of Chirata) was found to prevent CCl4/NDEA-induced liver carcinogenesis at mild dysplastic stage through modulation of cell cycle, apoptosis, self-renewal pathways. The cell cycle regulatory genes LIMD1, P16 and RBSP3 were found to be upregulated in restricted liver lesions. To understand the mechanism of upregulation during restriction of cacinogenesis, the effect of amarogentin on epigenetic modification was evaluated in this study. It was also validated in vitro. Hypermethylation of LIMD1 and P16 was seen in mouse hepatocellular carcinoma (30th week carcinogen control mice); however, hypomethylation of these genes was seen in amarogentin-treated liver. In the case of RBSP3, no such change was seen. DNMT1 expression (mRNA/protein) was significantly increased in later stages of carcinogenesis, whereas its expression was comparable to normal liver in the case of amarogentin treatment. No significant change in expression (mRNA/protein) of HDAC1/2 was observed irrespective of treatment. Amarogentin treatment upregulated the expression (mRNA/protein) of LIMD1, P16 and RBSP3 in the HepG2 cell line. Here also treated cells showed LIMD1 and P16 hypomethylation with DNMT1 downregulation. Increased expression of LIMD1, P16 and RBSP3 after treating cells with demethylating agent 5-aza-2-deoxycytidine indicated epigenetic modulation by amarogentin treatment.
Assuntos
Anticarcinógenos/farmacologia , Carcinogênese/efeitos dos fármacos , Carcinoma Hepatocelular/tratamento farmacológico , Iridoides/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Tetracloreto de Carbono/toxicidade , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Carcinoma Hepatocelular/induzido quimicamente , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/genética , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Decitabina/farmacologia , Dietilnitrosamina/toxicidade , Modelos Animais de Doenças , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Células Hep G2 , Histona Desacetilase 1/genética , Histona Desacetilase 1/metabolismo , Histona Desacetilase 2/genética , Histona Desacetilase 2/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Neoplasias Hepáticas/induzido quimicamente , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Camundongos , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , Transdução de SinaisRESUMO
Hyaluronic acid (HA)-CD44 pathway showed association with several malignancies. The natural polyphenols Plumbagin, Pongapin and Karanjin showed anti-cancer activities in different tumors including cervical carcinoma. To understand their mechanism of anti-cancer activity, the effect of the compounds on HA-CD44 pathway was analyzed in cervical cancer cell line HeLa. The mRNA expression of three different isoforms of CD44 i.e., CD44s, CD44v3, and CD44v6, was differentially downregulated by the compounds. This was validated by Western blot and immunocytochemical analysis of CD44s.The low molecular weight HA (LMW-HA) showed growth promoting activity in HeLa at low concentration, whereas high molecular weight HA (HMW-HA) had no such effect. The compounds could preferentially downregulate the LMW-HA level in HeLa, as evident in the cell as well as in the cell-free conditioned medium. Concentration-dependent upregulation of HA synthase-2 (HAS2) was seen in the cell by the compounds, whereas differential downregulation of hyalurinidases 1-4 (HYAL 1-4), predominantly HYAL1, were seen. The compounds could also downregulate the downstream target of the pathway p-AKT (T-308) in concentration-dependent manner. Thus, the compounds could attenuate the HA-CD44 pathway in HeLa cell to restrict the tumor growth.
Assuntos
Benzopiranos/farmacologia , Regulação para Baixo/efeitos dos fármacos , Flavonas/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Receptores de Hialuronatos/biossíntese , Ácido Hialurônico/metabolismo , Naftoquinonas/farmacologia , Proteínas de Neoplasias/biossíntese , Transdução de Sinais/efeitos dos fármacos , Neoplasias do Colo do Útero/metabolismo , Feminino , Células HeLa , Humanos , Receptores de Hialuronatos/genética , Ácido Hialurônico/genética , Proteínas de Neoplasias/genética , Transdução de Sinais/genética , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologiaRESUMO
PURPOSE: Our study was aimed to understand the importance of LIMD1-VHL-HIF1α pathway in development of bladder carcinoma (BlCa) in association with arsenic prevalence. METHODS: At first, the mRNA expression pattern of the genes of this pathway (LIMD1, VHL and HIF1α) was checked in GEO datasets and in our samples. Next, genetic and epigenetic profiling of LIMD1 and VHL was done in our sample pool, validated in T24 BlCa cell line. The results were next correlated with various clinico-pathological parameters. RESULTS: Differential under-expression of LIMD1 and VHL genes was found in muscle-invasive BlCa (MIBC) in comparison to non-muscle-invasive BlCa (NMIBC). However, HIF1α protein, but mRNA, was found to be overexpressed among the MIBC samples; depicting the probability of HIF1α protein stabilization. Analysis of genetic and epigenetic profiles of LIMD1 and VHL exposed a frequent promoter methylation of LIMD1 gene in MIBC samples. Further, in-depth look into the results unveiled that the high nuclear expression of HIF1α was significantly correlated with genetic alterations of LIMD1, alone or in combination with VHL. Moreover, treating the T24 cells with a de-methylating agent (5-aza-2'-deoxycytidine) re-expressed the methylated LIMD1 and VHL genes, which in turn, reduced the HIF1α protein level significantly. Additionally, patients with high arsenic content (> 112 ng/g, AsH) seemed to have recurrent promoter methylation in LIMD1, as well as co-methylation/alteration of LIMD1 and VHL gene. Lastly, high nuclear expression of HIF1α in association with co-alteration of VHL and LIMD1 showed the worst overall survival (OS) among the patients. CONCLUSION: To conclude, MIBC samples portrayed higher alterations in VHL and LIMD1, thereby, stabilizing HIF1α protein and lowering the OS of patients.
Assuntos
Carcinoma de Células de Transição/diagnóstico , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas com Domínio LIM/genética , Neoplasias da Bexiga Urinária/diagnóstico , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Intoxicação por Arsênico/diagnóstico , Intoxicação por Arsênico/epidemiologia , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma de Células de Transição/epidemiologia , Carcinoma de Células de Transição/genética , Carcinoma de Células de Transição/patologia , Linhagem Celular Tumoral , Núcleo Celular/genética , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Comorbidade , Metilação de DNA , Conjuntos de Dados como Assunto , Feminino , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/metabolismo , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Prevalência , Prognóstico , Análise de Sobrevida , Neoplasias da Bexiga Urinária/epidemiologia , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismoRESUMO
TNBC is the most aggressive and hormone receptor-negative subtype of breast cancer with molecular heterogeneity in bulk tumors hindering effective treatment. Toll-like receptors (TLRs) have the potential to ignite diverse immune responses in the tumor microenvironment (TME). This encouraged us to screen their transcript expression in the publically available TCGA datasets. Reported molecular subtypes of TNBC may represent different TMEs and we observed differentially expressed TLRs (DETs) i.e. TLR3/4/6/8/9 have unique expression pattern in the TNBC subtypes, particularly in Immunomodulatory (IM) TNBC subtype. We then dissected expression of the DETs in immune and other components of the TME. TLR4 and TLR8 showed significant (p-value ≤ 0.05) negative partial correlation with tumor purity compared to other DETs. Interestingly, TLR4 and TLR8 expression showed a significant (adjusted p-value ≤ 0.05) correlation with different subsets of immune infiltrating cells having the highest correlation with monocytes/macrophage/dendritic cell populations mediating both innate and adaptive response in TNBC. The co-expression network identified genes correlated with these immune cells. Further, GSEA analysis of co-expressed genes showed a significant association of TLR8 partners with 'Peptide ligand binding', 'Gά-signaling', and 'Cytokine-cytokine interaction' while TLR4 associated genes correlated with 'Adaptive immune system' and 'Systemic lupus erythematosus' interactome. Finally, the expression of TLR4 protein was validated in a panel of TNBC cell lines. TLR4 expression in chemoresponsive TNBC was also validated in TNBC cell lines upon Paclitaxel (PTX) treatment. Collectively, the present study identified specific DETs in TNBC and discovered a prospective role of TLR4 and TLR8 in the maintenance of tumor-immune-microenvironment.
Assuntos
Regulação Neoplásica da Expressão Gênica , Linfócitos do Interstício Tumoral/imunologia , Receptor 4 Toll-Like/genética , Receptor 8 Toll-Like/genética , Neoplasias de Mama Triplo Negativas/genética , Microambiente Tumoral/genética , Antineoplásicos Fitogênicos/uso terapêutico , Linhagem Celular Tumoral , Biologia Computacional/métodos , Bases de Dados Factuais , Feminino , Perfilação da Expressão Gênica , Ontologia Genética , Redes Reguladoras de Genes , Humanos , Linfócitos do Interstício Tumoral/classificação , Linfócitos do Interstício Tumoral/patologia , Anotação de Sequência Molecular , Proteínas de Neoplasias/classificação , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Paclitaxel/uso terapêutico , Transdução de Sinais , Receptor 4 Toll-Like/imunologia , Receptor 8 Toll-Like/imunologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/imunologia , Neoplasias de Mama Triplo Negativas/mortalidade , Microambiente Tumoral/imunologiaRESUMO
Oral squamous cell carcinoma of the gingivo-buccal region (OSCC-GB) accounts for the highest cancer morbidity and mortality among men in India. It has been observed that about one-third of individuals with oral leukoplakia, a dysplastic precancerous lesion in the oral cavity, progress to oral cancer. We aimed to identify systematic transcriptomic changes as a normal tissue in the oral cavity progresses to frank OSCC-GB. Seventy-two OSCC-GB patients, from multiple hospitals, were recruited, and transcriptome analysis of tumor and adjacent normal tissue (of all patients) and adjacent leukoplakia tissue (of a subset of 25 unselected patients with concomitant leukoplakia) was performed. We have identified many differences in the transcriptomic profiles between OSCC-GB and squamous cell carcinoma of the head and neck regions. Compared to the normal/precancerous tissue, significant enrichment of ECM-receptor interaction, PI3K-Akt signaling, cytokine-cytokine receptor interaction, focal adhesion, and cell cycle pathways were observed in OSCC-GB. Using gene set enrichment analysis, we identified a profound role of interferon receptor signaling in tumor growth by activating immune evasion mechanisms. The role of tumor-infiltrating immune cells further supported the growth and immunosuppressive mechanism of tumor tissues. Some immune evasion genes-CD274, CD80, and IDO1-were found to be activated even in the precancerous tissue. Taken together, our findings provide a clear insight into the sequential genetic dysregulation associated with progression to oral cancer. This insight provides a window to the development of predictive biomarkers and therapeutic targets for gingivo-buccal oral cancer.
RESUMO
Aims: To elucidate the impact of arsenic on progression and prognosis of bladder cancer. Patients & methods: Total arsenic in 145 tumors (80 non-muscle-invasive [NMIBC] and 65 muscle-invasive bladder cancer [MIBC]) was measured and associated with Ki67 expression, tumor-clinicopathological parameters and patient outcome. Results: Tumor arsenic concentration was higher in exposed than unexposed patients (256 µg/kg vs 77 µg/kg; p < 0.0001) and positively correlated (r = 0.65; p < 0.0001) with arsenic content of patient's drinking water. Arsenic concentration showed significant association with Ki67-overexpression (p = 0.001) and advanced tumor stages (NMIBC vs MIBC; p = 0.0009). In NMIBC, high tumor arsenic (>100 µg/kg) and Ki67 overexpression was established as predictors for recurrence (hazard ratio [HR]: 4.68; p = 0.005 and HR: 3.91; p = 0.018) and progression (HR: 6.04; p = 0.023 and HR: 6.87; p = 0.013). In MIBC, association of high arsenic remained significant with increased risk of recurrence (HR: 4.58; p = 0.04). Conclusion: In NMIBC, high arsenic and Ki67 overexpression and in MIBC, only high arsenic showed prognostic importance in predicting poor patient outcome.
Lay abstract Research work suggests arsenic as risk factor for bladder cancer. In developing countries such as India, arsenic contamination of underground drinking water is a major health problem. The present study aimed to evaluate impact of arsenic on parameters of bladder cancer aggressiveness and clinical outcome of patients from West Bengal, India. Our data showed accumulation of arsenic in bladder tumor of patients exposed mainly through contaminated drinking water. Arsenic content in tumor favored aggressive phenotypes in bladder cancer (higher cell proliferation and advanced tumor stages) and was found to be a potential predictor for cases of death and disease recurrence in patients after receiving primary treatment measures. Therefore, arsenic content in bladder tumor may be used to improve existing protocols for better prediction of patient outcomes in populations with a similar type of exposure.
Assuntos
Arsênio/metabolismo , Exposição Dietética/efeitos adversos , Neoplasias da Bexiga Urinária/metabolismo , Poluentes da Água/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Exposição Dietética/análise , Progressão da Doença , Feminino , Humanos , Índia , Antígeno Ki-67/metabolismo , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Prognóstico , Análise de Sobrevida , Neoplasias da Bexiga Urinária/patologiaRESUMO
A novel triterpenoid, holarol(1),3ß-lup-20(31)-en-3,29,30-triol along with one seco-triterpenoid, dihydrocanaric acid(2) and one known pentacyclic triterpenoid, betulin(3) have been isolated from Holarrhena antidysenterica (L.)Wall. (Family: Apocynaceae). The structures of the compounds were elucidated by extensive IR, 1D, 2D NMR and mass spectrometric analysis. The optimised geometry of (1) was calculated by density-functional theory (DFT) using M06-2X hybrid functional and 6-31 G(D) basis set. The compounds showed differential cytotoxic activities in the cell lines-HeLa, EAC, Raji and T24. Seco-triterpenoid (2) showed highest sensitivity (IC50: 1.710 µg/mL) against the bladder cancer cell line T24 followed by (1) (IC50 9.698 µg/mL) and (3) (IC50 11.769 µg/mL). Compound (1) showed highest reactive oxygen species (ROS) generation in T24 cell line followed by (3) and (2) resulting in induction of apoptosis through activation of caspase, cleavage of PARP and reduction of Bcl-2/Bax ratio. Thus compounds (1), (2) along with (3) could be potent anticancer agents.