RESUMEN
BACKGROUND: Cervical cancer, primarily caused by HPV infection, remains a global health concern. Current treatments face challenges including drug resistance and toxicity. This study investigates combining E5-siRNA with chemotherapy drugs, Oxaliplatin and Ifosfamide, to enhance treatment efficacy in HPV-16 positive cervical cancer cells, targeting E5 oncoprotein to overcome limitations of existing therapies. METHODS: The CaSki cervical cancer cell line was transfected with E5-siRNA, and subsequently treated with Oxaliplatin/Ifosfamide. Quantitative real-time PCR was employed to assess the expression of related genes including p53, MMP2, Nanog, and Caspases. Cell apoptosis, cell cycle progression, and cell viability were evaluated using Annexin V/PI staining, DAPI staining, and MTT test, respectively. Furthermore, stemness ability was determined through a colony formation assay, and cell motility was assessed by wound healing assay. RESULTS: E5-siRNA transfection significantly reduced E5 mRNA expression in CaSki cells compared to the control group. The MTT assay revealed that monotherapy with E5-siRNA, Oxaliplatin, or Ifosfamide had moderate effects on cell viability. However, combination therapy showed synergistic effects, reducing the IC50 of Oxaliplatin from 11.42 × 10-8 M (45.36 µg/ml) to 6.71 × 10-8 M (26.66 µg/ml) and Ifosfamide from 12.52 × 10-5 M (32.7 µg/ml) to 8.206 × 10-5 M (21.43 µg/ml). Flow cytometry analysis demonstrated a significant increase in apoptosis for combination treatments, with apoptosis rates rising from 11.02 % (Oxaliplatin alone) and 16.98 % (Ifosfamide alone) to 24.8 % (Oxaliplatin + E5-siRNA) and 34.9 % (Ifosfamide + E5-siRNA). The sub-G1 cell population increased from 15.7 % (Oxaliplatin alone) and 18 % (Ifosfamide alone) to 21.9 % (Oxaliplatin + E5-siRNA) and 27.1 % (Ifosfamide + E5-siRNA), indicating cell cycle arrest. The colony formation assay revealed a substantial decrease in the number of colonies following combination treatment. qRT-PCR analysis showed decreased expression of stemness-related genes CD44 and Nanog, and migration-related genes MMP2 and CXCL8 in the combination groups. Apoptosis-related genes Casp-3, Casp-9, and pP53 showed increased expression following combination therapy, while BAX expression increased and BCL2 expression decreased relative to the control. CONCLUSION: The study demonstrates that combining E5-siRNA with Oxaliplatin or Ifosfamide enhances the efficacy of chemotherapy in HPV-16 positive cervical cancer cells. This synergistic approach effectively targets multiple aspects of cancer cell behavior, including proliferation, apoptosis, migration, and stemness. The findings suggest that this combination strategy could potentially allow for lower chemotherapy doses, thereby reducing toxicity while maintaining therapeutic efficacy. This research provides valuable insights into targeting HPV E5 as a complementary approach to existing therapies focused on E6 and E7 oncoproteins, opening new avenues for combination therapies in cervical cancer treatment.
Asunto(s)
Apoptosis , Papillomavirus Humano 16 , Ifosfamida , Oxaliplatino , ARN Interferente Pequeño , Neoplasias del Cuello Uterino , Humanos , Neoplasias del Cuello Uterino/tratamiento farmacológico , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/virología , Oxaliplatino/farmacología , Femenino , ARN Interferente Pequeño/genética , Línea Celular Tumoral , Ifosfamida/farmacología , Apoptosis/efectos de los fármacos , Papillomavirus Humano 16/genética , Infecciones por Papillomavirus/tratamiento farmacológico , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/virología , Supervivencia Celular/efectos de los fármacos , Proteínas Oncogénicas Virales/genética , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Antineoplásicos/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacosRESUMEN
Human papillomaviruses (HPVs) and herpesviruses are detected in patients with epithelial ovarian cancer (EOC). We sought to analyze the prevalence of HPV's 16 and 18, cytomegalovirus (CMV), and Epstein-Barr virus (EBV) DNA in peripheral blood, ovarian, and fallopian tube (FT) tissue samples collected from 97 EOC patients, including 71 cases of high-grade serous ovarian carcinoma (HGSOC), and from 60 women with other tumors or non-neoplastic gynecological diseases. DNA isolates were analyzed by PCR methods, including droplet digital PCR. The results demonstrate that (1) HPV16 DNA has been detected in one-third of the FT and tumor samples from EOCs; (2) the prevalence and quantity of HPV16 DNA were significantly higher in FT samples from HGSOCs, non-HGSOCs, and ovarian metastases than in those from non-neoplastic diseases; (3) CMV and EBV have been detected in approximately one-seventh of EOC samples. The results suggest that HPV16 might be a potential risk factor for EOC development.
Asunto(s)
Carcinoma Epitelial de Ovario , Trompas Uterinas , Papillomavirus Humano 16 , Neoplasias Ováricas , Infecciones por Papillomavirus , Humanos , Femenino , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/complicaciones , Factores de Riesgo , Carcinoma Epitelial de Ovario/virología , Carcinoma Epitelial de Ovario/patología , Persona de Mediana Edad , Trompas Uterinas/virología , Trompas Uterinas/patología , Adulto , Anciano , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Neoplasias Ováricas/virología , Neoplasias Ováricas/patología , Herpesvirus Humano 4/aislamiento & purificación , Herpesvirus Humano 4/genética , Citomegalovirus/aislamiento & purificación , Citomegalovirus/genética , ADN Viral/genéticaRESUMEN
Invasive cervical cancers (ICC), caused by HPV infections, have a heterogeneous molecular landscape. We investigate the detection, timing, and HPV type specificity of somatic mutations in 3929 HPV-positive exfoliated cervical cell samples from individuals undergoing cervical screening in the U.S. using deep targeted sequencing in ICC cases, precancers, and HPV-positive controls. We discover a subset of hotspot mutations rare in controls (2.6%) but significantly more prevalent in precancers, particularly glandular precancer lesions (10.2%), and cancers (25.7%), supporting their involvement in ICC carcinogenesis. Hotspot mutations differ by HPV type, and HPV18/45-positive ICC are more likely to have multiple hotspot mutations compared to HPV16-positive ICC. The proportion of cells containing hotspot mutations is higher (i.e., higher variant allele fraction) in ICC and mutations are detectable up to 6 years prior to cancer diagnosis. Our findings demonstrate the feasibility of using exfoliated cervical cells for detection of somatic mutations as potential diagnostic biomarkers.
Asunto(s)
Cuello del Útero , Mutación , Infecciones por Papillomavirus , Neoplasias del Cuello Uterino , Humanos , Femenino , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patología , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/genética , Cuello del Útero/virología , Cuello del Útero/patología , Adulto , Persona de Mediana Edad , Papillomaviridae/genética , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Secuenciación de Nucleótidos de Alto Rendimiento , Displasia del Cuello del Útero/virología , Displasia del Cuello del Útero/genética , Displasia del Cuello del Útero/patologíaRESUMEN
The natural history of cervical cancer is closely linked to that of high-risk human papillomaviruses (HPV) infection. It is recognized that upon HPV DNA integration, partial or complete loss of the E2 open reading frame precludes expression of the corresponding protein, resulting in upregulation of the E6 and E7 viral oncoproteins. To better characterize HPV16 infection at the cervical level, viral load, viral DNA integration, and viral early transcript expression (E2, E5, and E6) were analyzed in a series of 158 cervical specimens representative of the full spectrum of cervical disease. Overall, the frequency of early transcript detection varied from 45% to 90% and tended to increase with lesion severity. In addition, the levels of E2, E5, and E6 transcript expression were slightly higher in high-grade lesions than in cervical specimens without abnormalities. Notably, early transcript expression was clearly associated with viral load, and no inverse correlation was found between the expression of E2 and E6 transcripts. No clear association was found between early transcript expression and HPV16 DNA integration, with the exception that samples with a fully integrated HPV16 genome did not harbor E2 or E5 transcripts. In conclusion, early HPV16 transcript expression appears to be associated with viral load rather than lesion grade. From a practical point of view, quantification of HPV16 early transcripts is difficult to translate into a relevant biomarker for cervical cancer screening.
Asunto(s)
Papillomavirus Humano 16 , Proteínas Oncogénicas Virales , Infecciones por Papillomavirus , Neoplasias del Cuello Uterino , Carga Viral , Humanos , Femenino , Papillomavirus Humano 16/genética , Infecciones por Papillomavirus/virología , Proteínas Oncogénicas Virales/genética , Neoplasias del Cuello Uterino/virología , Integración Viral , Adulto , Persona de Mediana Edad , ADN Viral/genética , Anciano , Cuello del Útero/virología , Cuello del Útero/patologíaRESUMEN
Human papillomavirus (HPV) drives cervical cancer (CaCx) pathogenesis and viral oncoproteins jeopardize global gene expression in such cancers. In this study, our aim was to identify differentially expressed coding (DEcGs) and long noncoding RNA genes (DElncGs) specifically sense intronic and Natural Antisense Transcripts as they are located in the genic regions and may have a direct influence on the expression pattern of their neighbouring coding genes. We compared HPV16-positive CaCx patients (N = 44) with HPV-negative normal individuals (N = 34) by employing strand-specific RNA-seq and determined the relationships between DEcGs and DElncGs and their clinical implications. By performing Gene set enrichment and protein-protein interaction (PPI) analyses of DEcGs, we identified enrichment of processes crucial for abortive virus life cycle and cancer progression. The DEcGs formed 16 gene clusters which we identified through Molecular Complex Detection (MCODE) plugin of Cytoscape. All the gene clusters portrayed cancer-related functions. We recorded significantly correlated expression levels of 79 DElncGs with DEcGs at proximal genomic loci based on Pearson's Correlation coefficients. Of these gene pairs, 24 pairs portrayed significantly altered correlation coefficients among patients, compared to normal individuals. Of these, 6 DEcGs of 6 such gene pairs, belonged to 5 of the identified gene clusters, one of which was survival-associated. Out of the 24 correlated DEcG: DElncG pairs, we identified 3 pairs, where expression of both members was significantly associated with patient overall survival. The findings justify the cooperative roles of these gene pairs, in patient prognostication, thereby bearing immense potential for translation. Thus, elucidation of correlative strengths between paired DElncGs and DEcGs in patient and normal samples, could serve as a foundation for identification of therapeutic and prognostic targets of HPV16-positive CaCx.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Papillomavirus Humano 16 , Infecciones por Papillomavirus , ARN Largo no Codificante , Neoplasias del Cuello Uterino , Humanos , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/patología , Femenino , ARN Largo no Codificante/genética , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/patogenicidad , Regulación Neoplásica de la Expresión Génica/genética , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/virología , Persona de Mediana Edad , Familia de Multigenes/genética , Adulto , Relevancia ClínicaRESUMEN
BACKGROUND: To investigate the technical feasibility of RT-PCR and direct sequencing to quantify HPV DNA in the saliva of patients with Human-Papilloma-Virus related oropharyngeal cancer (HPV-OPC), the level of which is known to predict prognosis after treatment. METHODS: Nine patients with locally advanced HPV-OPC treated with definitive radiotherapy with chemotherapy or cetuximab, or radiotherapy alone between April 2016 and September 2017, were enrolled, two of whom also received induction chemotherapy. Saliva was collected before (baseline), during (mid-RT) and after (post-RT) radiotherapy. HPV-16 DNAs (E6 and E7) in saliva were quantified by RT-PCR and sequencing, the latter using a custom cancer panel. Correlations between HPV DNA levels and clinical outcomes were assessed. RESULTS: Compared to the baseline, the relative cycle threshold (Ct) value of E6 and E7 reduced at the point of mid-RT in the majority of the patients (100% and 75% for E6 and E7, respectively). Similarly, the relative Ct value from the baseline to post-RT reduced in 86% and 100% of the patients for E6 and E7, respectively. During the follow-up period, three patients (33%) experienced disease progression. The relative baseline Ct values of these three patients were in the top 4 of all the patients. The sequences of HPV DNA were detected in five (83%) of six samples of the baseline saliva that underwent DNA sequencing, along with several gene mutations, such as TP53,CDKN2A and PIK3CA. CONCLUSIONS: This study demonstrates that, in addition to detection and quantification of HPV DNA by RT-PCR, detection by sequencing of HPV-DNA using a customized cancer panel is technically possible.
Asunto(s)
ADN Viral , Neoplasias Orofaríngeas , Infecciones por Papillomavirus , Saliva , Humanos , Neoplasias Orofaríngeas/virología , Neoplasias Orofaríngeas/radioterapia , Saliva/virología , ADN Viral/análisis , Persona de Mediana Edad , Masculino , Femenino , Anciano , Infecciones por Papillomavirus/virología , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificaciónRESUMEN
BACKGROUND: An increasing number of studies have focused on the association between Human papillomavirus (HPV) infection and systemic lupus erythematosus (SLE). However, current evidence is largely based on retrospective studies, which are susceptible to confounding factors and cannot establish causation. METHODS: A bidirectional two-sample Mendelian randomization (MR) design was used to evaluate the causal relationship between HPV and SLE. Mononucleoside polymers (SNPS) with strong evidence for genome-wide association studies (GWAS) were selected from the HPV exposure dataset and used as an instrumental variable (IV) for this study. For the MR Analysis results, the MR-Egger intercept P test, MR-Presso global test, CochranQ test and leave-one test were used for sensitivity analysis. RESULTS: Based on the evidence of MR Analysis, this study finally determined that there was no causal association between HPV16 and HPV18 and SLE. CONCLUSIONS: Possible regulation of HPV infection is not significantly associated with regulation of SLE. These findings provide new insights into the underlying mechanisms of HPV and SLE and need to be validated by further studies.
Asunto(s)
Estudio de Asociación del Genoma Completo , Lupus Eritematoso Sistémico , Análisis de la Aleatorización Mendeliana , Infecciones por Papillomavirus , Polimorfismo de Nucleótido Simple , Humanos , Lupus Eritematoso Sistémico/genética , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/complicaciones , Papillomavirus Humano 16/genética , Papillomavirus Humano 18/genética , FemeninoRESUMEN
We have demonstrated that SAMHD1 (sterile alpha motif and histidine-aspartic domain HD-containing protein 1) is a restriction factor for the human papillomavirus 16 (HPV16) life cycle. Here, we demonstrate that in HPV-negative cervical cancer C33a cells and human foreskin keratinocytes immortalized by HPV16 (HFK+HPV16), SAMHD1 is recruited to E1-E2 replicating DNA. Homologous recombination (HR) factors are required for HPV16 replication, and viral replication promotes phosphorylation of SAMHD1, which converts it from a dNTPase to an HR factor independent from E6/E7 expression. A SAMHD1 phospho-mimic (SAMHD1 T592D) reduces E1-E2-mediated DNA replication in C33a cells and has enhanced recruitment to the replicating DNA. In HFK+HPV16 cells, SAMHD1 T592D is recruited to the viral DNA and attenuates cellular growth, but does not attenuate growth in isogenic HFK cells immortalized by E6/E7 alone. SAMHD1 T592D also attenuates the development of viral replication foci following keratinocyte differentiation. The results indicated that enhanced SAMHD1 phosphorylation could be therapeutically beneficial in cells with HPV16 replicating genomes. Protein phosphatase 2A (PP2A) can dephosphorylate SAMHD1, and PP2A function can be inhibited by endothall. We demonstrate that endothall reduces E1-E2 replication and promotes SAMHD1 recruitment to E1-E2 replicating DNA, mimicking the SAMHD1 T592D phenotypes. Finally, we demonstrate that in head and neck cancer cell lines with HPV16 episomal genomes, endothall attenuates their growth and promotes recruitment of SAMHD1 to the viral genome. The results suggest that targeting cellular phosphatases has therapeutic potential for the treatment of HPV infections and cancers. IMPORTANCE: Human papillomaviruses (HPVs) are causative agents in around 5% of all human cancers. The development of anti-viral therapeutics depends upon an increased understanding of the viral life cycle. Here, we demonstrate that HPV16 replication converts sterile alpha motif and histidine-aspartic domain HD-containing protein 1 (SAMHD1) into a homologous recombination (HR) factor via phosphorylation. This phosphorylation promotes recruitment of SAMHD1 to viral DNA to assist with replication. A SAMHD1 mutant that mimics phosphorylation is hyper-recruited to viral DNA and attenuates viral replication. Expression of this mutant in HPV16-immortalized cells attenuates the growth of these cells, but not cells immortalized by the viral oncogenes E6/E7 alone. Finally, we demonstrate that the phosphatase inhibitor endothall promotes hyper-recruitment of endogenous SAMHD1 to HPV16 replicating DNA and can attenuate the growth of both HPV16-immortalized human foreskin keratinocytes (HFKs) and HPV16-positive head and neck cancer cell lines. We propose that phosphatase inhibitors represent a novel tool for combating HPV infections and disease.
Asunto(s)
ADN Viral , Papillomavirus Humano 16 , Queratinocitos , Proteína 1 que Contiene Dominios SAM y HD , Replicación Viral , Proteína 1 que Contiene Dominios SAM y HD/metabolismo , Proteína 1 que Contiene Dominios SAM y HD/genética , Humanos , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/metabolismo , Papillomavirus Humano 16/fisiología , ADN Viral/genética , ADN Viral/metabolismo , Queratinocitos/virología , Queratinocitos/metabolismo , Fosforilación , Línea Celular Tumoral , Recombinación Homóloga , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/metabolismo , Infecciones por Papillomavirus/genética , Replicación del ADNRESUMEN
BACKGROUND: Human papillomavirus 16 (HPV16) is an oncogenic virus responsible for the majority of invasive cervical cancer cases worldwide. Due to genetic modifications, some variants are more oncogenic than others. We analysed the HPV16 phylogeny in HPV16-positive cervical Desoxyribonucleic Acid (DNA) samples collected from South African and Mozambican women to detect the circulating lineages. METHODS: Polymerase chain reaction (PCR) amplification of the long control region (LCR) and 300 nucleotides of the E6 region was performed using HPV16-specific primers on HPV16-positive cervical samples collected in women from South Africa and Mozambique. HPV16 sequences were obtained through Next Generation Sequencing (NGS) methods. Geneious prime and MEGA 11 software were used to align the sequences to 16 HPV16 reference sequences, gathering the A, B, C, and D lineages and generating the phylogenetic tree. Single nucleotide polymorphisms (SNPs) in the LCR and E6 regions were analysed and the phylogenetic tree was generated using Geneious Prime software. RESULTS: Fifty-eight sequences were analysed. Of these sequences, 79% (46/58) were from women who had abnormal cervical cytology. Fifteen SNPs in the LCR and eight in the E6 region were found to be the most common in all sequences. The phylogenetic analysis determined that 45% of the isolates belonged to the A1 sublineage (European variant), 34% belonged to the C1 sublineage (African 1 variant), 16% belonged to the B1 and B2 sublineage (African 2 variant), two isolates belonged to the D1-3 sublineages (Asian-American variant), and one to the North American variant. CONCLUSIONS: The African and European HPV16 variants were the most common circulating lineages in South African and Mozambican women. A high-grade squamous intraepithelial lesion (HSIL) was the most common cervical abnormality observed and linked to European and African lineages. These findings may contribute to understanding molecular HPV16 epidemiology in South Africa and Mozambique.
Asunto(s)
Papillomavirus Humano 16 , Infecciones por Papillomavirus , Filogenia , Neoplasias del Cuello Uterino , Humanos , Femenino , Mozambique/epidemiología , Sudáfrica/epidemiología , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Papillomavirus Humano 16/clasificación , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/epidemiología , Neoplasias del Cuello Uterino/virología , Adulto , Cuello del Útero/virología , Cuello del Útero/patología , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple , ADN Viral/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Proteínas Oncogénicas Virales/genética , Adulto Joven , CitologíaRESUMEN
BACKGROUND: Nitric oxide (NO) may contribute to the persistence of high-risk human papillomavirus (hrHPV) infection, which has been linked to the development of premalignant lesions and cervical cancer. Our study aimed to examine the relationship between cervical NO metabolite (NOx) levels, hrHPV infection, and cytopathological findings. Additionally, we assessed cervical NOx levels as a biomarker for predicting hrHPV infection and epithelial atypia. METHODS: The study involved 74 women who attended the Gynecology and Obstetrics outpatient clinics at Cairo University Hospitals between November 2021 and August 2022. Cervical samples were subjected to Pap testing, assessment of NOx levels by the Griess method, and detection of hrHPV DNA by real-time polymerase chain reaction. RESULTS: High-risk HPV was detected in 37.8% of women. EA was found in 17.1% of cases, with a higher percentage among hrHPV-positive than negative cases (35.7% vs. 4.3%, p = 0.001). The most prevalent hrHPV genotype was HPV 16 (89.3%). The cervical NOx level in hrHPV-positive cases was significantly higher (37.4 µmol/mL, IQR: 34.5-45.8) compared to negative cases (2.3 µmol/mL, IQR: 1.2-9.8) (p = < 0.001). Patients with high-grade atypia showed significantly higher NOx levels (38.0 µmol/mL, IQR: 24.6-94.7) in comparison to NILM and low-grade atypia cases (5.0 µmol/mL, IQR: 1.6-33.3 and 34.5 µmol/mL, IQR: 11.7-61.7, respectively) (p = 0.006). Although the NOx levels among hrHPV-positive cases with low-grade atypia (40.4 µmol/mL, IQR: 33.3â61.8) were higher than those with NILM (36.2 µmol/mL, IQR: 35.7â44.0) and high-grade atypia (38.0 µmol/mL, IQR: 24.6â94.7), the difference was not significant (p = 0.771). ROC curve analysis indicated that the cervical NOx cut-off values of > 23.61 µmol/mL and > 11.35 µmol/mL exhibited good diagnostic accuracy for the prediction of hrHPV infection and EA, respectively. CONCLUSIONS: The high prevalence of hrHPV infection, particularly HPV 16, in our hospital warrants targeted treatment and comprehensive screening. Elevated cervical NOx levels are associated with hrHPV infection and high-grade atypia, suggesting their potential use as biomarkers for predicting the presence of hrHPV and abnormal cytological changes.
Asunto(s)
Cuello del Útero , Óxido Nítrico , Infecciones por Papillomavirus , Humanos , Femenino , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/diagnóstico , Infecciones por Papillomavirus/patología , Óxido Nítrico/análisis , Óxido Nítrico/metabolismo , Adulto , Cuello del Útero/virología , Cuello del Útero/patología , Persona de Mediana Edad , Papillomaviridae/genética , Papillomaviridae/aislamiento & purificación , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/patología , Neoplasias del Cuello Uterino/diagnóstico , Adulto Joven , ADN Viral/genética , Displasia del Cuello del Útero/virología , Displasia del Cuello del Útero/patología , Displasia del Cuello del Útero/diagnóstico , Biomarcadores/análisis , Genotipo , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Frotis Vaginal , Prueba de Papanicolaou , CitologíaRESUMEN
Human papillomavirus (HPV) 11/16 E6/E7 proteins have been recognized to be pivotal in viral pathogenesis. This study sought to uncover the potential mechanisms of how HPV11/16 E6/E7-transfected keratinocytes inhibit cytokine secretion in peripheral blood mononuclear cells (PBMC). Upon co-culturing HPV11/16 E6/E7-transfected keratinocytes with PBMC in a non-contact manner, we observed a marked decrease in various cytokines secreted by PBMC. To determine if this suppression was mediated by specific common secreted factors, we conducted transcriptomic sequencing on these transfected cells. This analysis identified 53 common differentially secreted genes in all four HPV-transfected cells. Bioinformatics analysis demonstrated these genes were predominantly involved in immune regulation. Results from quantitative PCR (qPCR) and an extensive literature review suggested the downregulation of 12 genes (ACE2, BMP3, BPIFB1, CLU, CST6, CTF1, HMGB2, MMP12, PDGFA, RNASE7, SULF2, TGM2), and upregulation of 7 genes (CCL17, CCL22, FBLN1, PLAU, S100A7, S100A8, S100A9), may be crucial in modulating tumor immunity and combating pathogenic infections, with genes S100A8 and S100A9, and IL-17 signaling pathway being particularly noteworthy. Thus, HPV11/16 E6/E7 proteins may inhibit cytokine secretion of immune cells by altering the expression of host-secreted genes. Further exploration of these genes may yield new insights into the complex dynamics of HPV infection.
Asunto(s)
Citocinas , Leucocitos Mononucleares , Proteínas Oncogénicas Virales , Humanos , Citocinas/metabolismo , Citocinas/genética , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/virología , Leucocitos Mononucleares/metabolismo , Proteínas Oncogénicas Virales/genética , Proteínas Oncogénicas Virales/metabolismo , Proteínas Oncogénicas Virales/inmunología , Queratinocitos/virología , Queratinocitos/inmunología , Queratinocitos/metabolismo , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/inmunología , Papillomavirus Humano 11/genética , Papillomavirus Humano 11/inmunología , Perfilación de la Expresión Génica , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/inmunología , Infecciones por Papillomavirus/genética , Proteínas E7 de Papillomavirus/genética , Proteínas E7 de Papillomavirus/metabolismo , Proteínas E7 de Papillomavirus/inmunología , Técnicas de Cocultivo , Interacciones Huésped-Patógeno/inmunología , Interacciones Huésped-Patógeno/genéticaRESUMEN
Cervical cancer is among the most common malignant tumors in women. The development of rapid screening techniques plays an important role in early screening for cancer treatment. We have developed an HPV screening method, which effectively combines the high-efficiency nucleic acid enrichment of chitosan-modified filter paper and the rapid visual detectability of colorimetric LAMP, along with the enhancement of the tolerance ability of the pH-sensitive LAMP reagent to acidic original samples, making the detection of HPV 16/18 easy to carry out and reliable, which is helpful for the epidemiological prevention and control strategies of HPV-induced cancer. This technique can simultaneously exhibit the "in situ amplification" capability of chitosan-modified filter paper and the nontemperature cycle dependence of visual LAMP detection. Therefore, DNA extraction and amplification can be performed efficiently and quickly within a single reaction where all DNA is concentrated in the QF paper disc. By embedding amino-modified filter paper into the plastic chip, a simple and reliable disposable chip was prepared for rapid HPV16 and HPV18 detection from clinical endometrial samples, and the results were 100% consistent with clinical diagnosis. More importantly, even after the sample was diluted 100-fold, HPV16/18-infected cells could be accurately identified, showing the advantages of the system in early cancer screening. Moreover, for endometrial samples containing plenty of cells, the filter paper could be used to enrich cells by filtration, preventing the acidic fluid from impacting pH-induced colorimetric LAMP detection and realizing direct amplification for HPV identification without nucleic acid extraction. This easy-to-operate system that can analyze a wide range of samples will be suitable for routine on-site HPV screening, dramatically extending the applications and utility for rapid, near-patient nucleic acid testing.
Asunto(s)
Colorimetría , Papillomavirus Humano 16 , Papillomavirus Humano 18 , Técnicas de Amplificación de Ácido Nucleico , Papel , Humanos , Colorimetría/métodos , Papillomavirus Humano 18/genética , Papillomavirus Humano 18/aislamiento & purificación , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Técnicas de Amplificación de Ácido Nucleico/métodos , Femenino , ADN Viral/análisis , ADN Viral/genética , Quitosano/química , Virus del Papiloma HumanoRESUMEN
BACKGROUND: Esophageal carcinoma is a growing concern in regions that have a high incidence of human papillomavirus (HPV) infection such as East Africa. HPV, particularly the high-risk genotypes, is increasingly recognized as a risk factor for esophageal carcinoma. We set out to investigate the prevalence and associated factors of high-risk HPV in formalin-fixed paraffin-embedded (FFPE) tissue blocks with esophageal carcinoma at Bugando Medical Center, a tertiary referral hospital in Mwanza, Tanzania, East Africa. METHODS: A total of 118 esophageal carcinoma FFPE tissue blocks, collected from January 2021 to December 2022, were analyzed. Genomic DNA was extracted from these tissues, and multiplex polymerase chain reaction (PCR) was performed to detect HPV using degenerate primers for the L1 region and type-specific primers for detecting HPV16, HPV18, and other high-risk HPV genotypes. Data were collected using questionnaires and factors associated with high-risk HPV genotypes were analyzed using STATA version 15 software. RESULTS: Of the 118 patients' samples investigated, the mean age was 58.3 ± 13.4 years with a range of 29-88 years. The majority of the tissue blocks were from male patients 81/118 (68.7%), and most of them were from patients residing in Mwanza region 44/118 (37.3%). Esophageal Squamous Cell Carcinoma (ESCC) was the predominant histological type 107/118 (91.0%). Almost half of the tissue blocks 63/118 (53.3%) tested positive for high-risk HPV. Among these, HPV genotype 16 (HPV16) was the most common 41/63 (65.1%), followed by HPV genotype 18 (HPV18) 15/63 (23.8%), and the rest were other high-risk HPV genotypes detected by the degenerate primers 7/63 (11.1%). The factors associated with high-risk HPV genotypes were cigarette smoking (p-value < 0.001) and alcohol consumption (p-value < 0.001). CONCLUSION: A substantial number of esophageal carcinomas from Bugando Medical Center in Tanzania tested positive for HPV, with HPV genotype 16 being the most prevalent. This study also revealed a significant association between HPV status and cigarette smoking and alcohol consumption. These findings provide important insights into the role of high-risk HPV in esophageal carcinoma in this region.
Asunto(s)
Neoplasias Esofágicas , Genotipo , Virus del Papiloma Humano , Infecciones por Papillomavirus , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Esofágicas/virología , Neoplasias Esofágicas/epidemiología , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Virus del Papiloma Humano/genética , Virus del Papiloma Humano/aislamiento & purificación , Papillomaviridae/genética , Papillomaviridae/aislamiento & purificación , Infecciones por Papillomavirus/epidemiología , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/complicaciones , Prevalencia , Factores de Riesgo , Tanzanía/epidemiologíaRESUMEN
BACKGROUND: To investigate the causal relationship between human papillomavirus (HPV) and lung cancer, we conducted a study using the two-sample Mendelian randomization (TSMR). METHOD: Data from genome-wide association studies (GWAS) were analyzed with HPV E7 Type 16 and HPV E7 Type 18 as exposure factors. The outcome variables included lung cancer, small cell lung cancer, adenocarcinoma and squamous cell lung cancer. Causality was estimated using inverse variance weighted (IVW), MR-Egger and weighted median methods. Heterogeneity testing, sensitivity analysis, and multiple validity analysis were also performed.. RESULTS: The results showed that HPV E7 Type 16 infection was associated with a higher risk of squamous cell lung cancer (OR = 7.69; 95% CI:1.98-29.85; p = 0.0149). HPV E7 Type 18 infection significantly increased the risk of lung adenocarcinoma (OR = 0.71; 95% CI: 0.38-1.31; p = 0.0079) and lung cancer (OR = 7.69; 95% CI:1.98-29.85; p = 0.0292). No significant causal relationship was found between HPV E7 Type 16 and lung adenocarcinoma, lung cancer, or small cell lung carcinoma, and between HPV E7 Type 18 and squamous cell lung cancer or small cell lung carcinoma. CONCLUSIONS: This study has revealed a causal relationship between HPV and lung cancers. Our findings provide valuable insights for further mechanistic and clinical studies on HPV-mediated cancer.
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Estudio de Asociación del Genoma Completo , Neoplasias Pulmonares , Análisis de la Aleatorización Mendeliana , Infecciones por Papillomavirus , Humanos , Neoplasias Pulmonares/virología , Neoplasias Pulmonares/genética , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/complicaciones , Polimorfismo de Nucleótido Simple , Papillomavirus Humano 16/genética , Papillomaviridae/genética , Virus del Papiloma HumanoRESUMEN
BACKGROUND: There is a high incidence of cervical cancer in Xinjiang. Genetic variation in human papillomavirus may increase its ability to invade, spread, and escape host immune response. METHODS: HPV16 genome was sequenced for 90 positive samples of HPV16 infection. Sequences of the E4, E5 and L2 genes were analysed to reveal sequence variation of HPV16 in Xinjiang and the distribution of variation among the positive samples of HPV16 infection. RESULTS: Eighty-one of the 90 samples of HPV16 infection showed variation in HPV16 E4 gene with 18 nucleotide variation sites, of which 8 sites were synonymous variations and 11 missense variations. 90 samples of HPV16 infection showed variation in HPV16 E5 and L2 genes with 16 nucleotide variation sites (6 synonymous, 11 missense variations) in the E5 gene and 100 nucleotide variation sites in L2 gene (37 synonymous, 67 missense variations). The frequency of HPV16 L2 gene missense variations G3377A, G3599A, G3703A, and G3757A was higher in the case groups than in the control groups. CONCLUSIONS: Phylogenetic tree analysis showed that 87 samples were European strains, 3 cases were Asian strains, there were no other variations, and G4181A was related to Asian strains. HPV16 L2 gene missense variations G3377A, G3599A, G3703A, and G3757A were significantly more frequent in the case groups than in the control groups.
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Variación Genética , Papillomavirus Humano 16 , Proteínas Oncogénicas Virales , Infecciones por Papillomavirus , Filogenia , Humanos , Femenino , China , Papillomavirus Humano 16/genética , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/genética , Proteínas Oncogénicas Virales/genética , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/genética , Adulto , Persona de Mediana Edad , Mutación MissenseRESUMEN
The microbial community has a profound effect on the host microenvironment by altering metabolites. Persistent high-risk human papillomavirus (HRHPV) infection has been implicated as contributors to the initiation and progression of cervical cancer, but the involved mechanisms are unknown. Assessing the metabolic profile of the cervicovaginal microenvironment has the potential to reveal the functional interactions among the host, metabolites and microbes in HRHPV persistence infection and progression to cancer. The vaginal swabs of women were collected and divided into three groups according to the HPV HybridenPture DNA test (HC2). The participants, include 9 who were categorized as HPV-negative, 8 as positive for HPV16, and 9 as positive for HPV18. 16S rRNA gene sequencing and metabolomics analyses were applied to determine the influence of the vaginal microbiota and host metabolism on the link between HPV and cervicovaginal microenvironment. These findings revealed that HRHPV groups have unique metabolic fingerprints that distinguish them from heathy controls. We showed that HRHPV affects changes in microbial metabolic function, which has important implications for the host. Our study further demonstrated metabolite-driven complex host-microbe interactions and assist in understanding the alterations in the HRHPV-induced cervicovaginal microenvironment.
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Metaboloma , Microbiota , Infecciones por Papillomavirus , ARN Ribosómico 16S , Vagina , Femenino , Humanos , Vagina/microbiología , Vagina/virología , Vagina/metabolismo , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/metabolismo , ARN Ribosómico 16S/genética , Adulto , Cuello del Útero/microbiología , Cuello del Útero/virología , Cuello del Útero/metabolismo , Interacciones Microbiota-Huesped , Papillomavirus Humano 18/genética , Papillomavirus Humano 18/metabolismo , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/metabolismo , Metabolómica , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/microbiología , Neoplasias del Cuello Uterino/metabolismo , Papillomaviridae/genética , Virus del Papiloma HumanoRESUMEN
The present study was aimed at showing the importance of HPV DNA status and the clinical history of the patients required by the cytologist for accurate reporting. A total of 1250 symptomatic women who attended the gynaecology outpatient department of the Mahavir Cancer Sansthan and Nalanda Medical College, Patna, for pap smear examinations were screened and recruited for the study. Due to highly clinical symptoms out of the negative with inflammatory smears reported, one hundred and ten patients were randomly advised for biopsy and HPV 16/18 DNA analysis by a gynaecologist to correlate negative smears included in the study. Pap smear reports revealed that 1178 (94.24%) were negative for intraepithelial lesions (NILM) with inflammatory smears, 23 (1.84%) smears showed low-grade squamous intraepithelial lesions (LSIL), 12 (0.96%) smears showed high-grade squamous intraepithelial lesions, and 37 (2.96%) smears showed an atypical squamous cell of undetermined significance (ASC-US). A biopsy of 110 out of 1178 (NILM) patients revealed that 15 (13.63%) women had cervical cancer, 29 women had CIN I, 17 women had CIN II + CIN III, 35 women had benign cervical changes, and 14 women had haemorrhages. On the other hand, HPV 16/18 DNA was detected as positive in 87 out of 110. The high positivity of HPV in biopsied cases where frank cervical cancer and at-risk cancer were also observed in the negative smear-screened patients reveals that the HPV status and clinical history of the patients will be quite helpful to the cytologist for accurate reporting, and suggests that a negative HPV DNA result may be a stronger predictor of cervical cancer risk than a negative Pap test.
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ADN Viral , Prueba de Papanicolaou , Infecciones por Papillomavirus , Neoplasias del Cuello Uterino , Humanos , Femenino , Adulto , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/diagnóstico , Neoplasias del Cuello Uterino/patología , ADN Viral/análisis , ADN Viral/genética , Persona de Mediana Edad , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/diagnóstico , Displasia del Cuello del Útero/virología , Displasia del Cuello del Útero/diagnóstico , Displasia del Cuello del Útero/patología , Frotis Vaginal , Papillomavirus Humano 18/genética , Papillomavirus Humano 18/aislamiento & purificación , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/aislamiento & purificación , Adulto Joven , Anciano , BiopsiaRESUMEN
Human papillomaviruses (HPVs) cause most cervical cancers and an increasing number of anogenital and oral carcinomas, with most cases caused by HPV16 or HPV18. HPV hijacks host signalling pathways to promote carcinogenesis. Understanding these interactions could permit identification of much-needed therapeutics for HPV-driven malignancies. The Hippo signalling pathway is important in HPV+ cancers, with the downstream effector YAP playing a pro-oncogenic role. In contrast, the significance of its paralogue TAZ remains largely uncharacterised in these cancers. We demonstrate that TAZ is dysregulated in a HPV-type dependent manner by a distinct mechanism to that of YAP and controls proliferation via alternative cellular targets. Analysis of cervical cancer cell lines and patient biopsies revealed that TAZ expression was only significantly increased in HPV18+ and HPV18-like cells and TAZ knockdown reduced proliferation, migration and invasion only in HPV18+ cells. RNA-sequencing of HPV18+ cervical cells revealed that YAP and TAZ have distinct targets, suggesting they promote carcinogenesis by different mechanisms. Thus, in HPV18+ cancers, YAP and TAZ play non-redundant roles. This analysis identified TOGARAM2 as a previously uncharacterised TAZ target and demonstrates its role as a key effector of TAZ-mediated proliferation, migration and invasion in HPV18+ cancers.
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Proteínas Adaptadoras Transductoras de Señales , Proliferación Celular , Vía de Señalización Hippo , Papillomavirus Humano 18 , Infecciones por Papillomavirus , Proteínas Serina-Treonina Quinasas , Transducción de Señal , Factores de Transcripción , Neoplasias del Cuello Uterino , Proteínas Señalizadoras YAP , Femenino , Humanos , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Carcinogénesis/genética , Línea Celular Tumoral , Movimiento Celular , Regulación Neoplásica de la Expresión Génica , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/metabolismo , Papillomavirus Humano 18/genética , Papillomavirus Humano 18/metabolismo , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/metabolismo , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Transactivadores/metabolismo , Transactivadores/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZ/metabolismo , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/metabolismo , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patología , Proteínas Señalizadoras YAP/metabolismoRESUMEN
This study was designed to investigate the expression of HPV16 L1-protein in biopsies of oral mucosa samples. The expression of HPV16 L1 protein was investigated in biopsies taken from oral mucosa from patients who required pathological diagnosis of oral lesions. Seventy-two samples were incubated with anti-L1 protein monoclonal antibodies and protein detection was revealed with diaminobenzidine. Expression of L1 protein was performed by a pathologist blinded for tissue diagnosis under light microscopy. Most of the lesions of oral mucosa were present in lining mucosa (75 %) and the most frequent lesion were mucocele (n = 17, 23.6 %), epithelial hyperplasia (n = 6, 8.33 %), fibroma (n = 5, 6.9 %) and inflammatory hyperplasia (n = 5, 6.9 %). L1 protein expression was observed only in five (6.9 %) samples (two squamous cell carcinomas, two epithelial hyperplasia, and one gingival hyperplasia). We concluded that L1 expression in oral biopsies presented a low frequency in oral mucosal biopsies samples.
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Proteínas de la Cápside , Mucosa Bucal , Proteínas Oncogénicas Virales , Infecciones por Papillomavirus , Humanos , Biopsia , Femenino , Mucosa Bucal/virología , Mucosa Bucal/patología , Infecciones por Papillomavirus/diagnóstico , Infecciones por Papillomavirus/virología , Adulto , Masculino , Proteínas Oncogénicas Virales/genética , Persona de Mediana Edad , Ecuador/epidemiología , Proteínas de la Cápside/genética , Proteínas de la Cápside/inmunología , Adulto Joven , Adolescente , Anciano , Prevalencia , Enfermedades de la Boca/virología , Enfermedades de la Boca/patología , Enfermedades de la Boca/epidemiología , Papillomavirus Humano 16/genética , Inmunohistoquímica , Neoplasias de la Boca/virología , Neoplasias de la Boca/patología , Neoplasias de la Boca/diagnósticoRESUMEN
Cervical cancer (CC) screening in women comprises human papillomavirus (HPV) testing followed by cytology triage of positive cases. Drawbacks, including cytology's low reproducibility and requirement for short screening intervals, raise the need for alternative triage methods. Here we used an innovative triage technique, the WID-qCIN test, to assess the DNA methylation of human genes DPP6, RALYL and GSX1 in a real-life cohort of 28,017 women aged ≥30 years who attended CC screening in Stockholm between January and March 2017. In the analysis of all 2,377 HPV-positive samples, a combination of WID-qCIN (with a predefined threshold) and HPV16 and/or HPV18 (HPV16/18) detected 93.4% of cervical intraepithelial neoplasia grade 3 and 100% of invasive CCs. The WID-qCIN/HPV16/18 combination predicted 69.4% of incident cervical intraepithelial neoplasia grade 2 or worse compared with 18.2% predicted by cytology. Cytology or WID-qCIN/HPV16/18 triage would require 4.1 and 2.4 colposcopy referrals to detect one cervical intraepithelial neoplasia grade 2 or worse, respectively, during the 6 year period. These findings support the use of WID-qCIN/HPV16/18 as an improved triage strategy for HPV-positive women.