Mirghani H, Casiraghi O, Amen F, He M, Ma XJ, Saulnier P, Lacroix L, Drusch F, Ben Lakdhar A, Saint Guily JL, Badoual C, Scoazec JY, Vielh P.
PMID: 26403782 | DOI: 10.1038/modpathol.2015.113
Abstract
Accurate screening of HPV-driven head and neck squamous cell carcinoma is a critical issue. Although there are commercial direct and indirect assays for HPV-related head and neck squamous cell carcinoma, none are ideal. Recently, a novel RNA in situ hybridization test (the RNAscope HPV-test) has been developed for the detection of high-risk HPV E6/E7 mRNA in formalin-fixed paraffin-embedded tissue. However, validation of this assay against the 'gold standard' (identification of high-risk HPV E6/E7 mRNA in fresh-frozen tissue by quantitative real-time (qRT)-PCR) has only been reported by one team. Formalin-fixed paraffin-embedded samples from 50 patients with tonsil or tongue base carcinoma were tested using the RNAscope HPV-test, p16 immunohistochemistry, and chromogenic in situ hybridization for high-risk HPV-DNA. The results were compared with those of qRT-PCR on matched fresh-frozen samples. Compared with the reference test, the sensitivity, specificity, positive, and negative predictive values of the RNAscope HPV-test and of p16 immunohistochemistry were 93%, 94%, 96%, 88% and 96%, 93%, 96%, and 93%, respectively. Five cases were discrepant between the RNAscope HPV-test and p16-immunohistochemisrty. The RNAscope HPV-test demonstrated excellent analytical performance against the 'gold standard' and is easier to interpret than chromogenic in situ hybridization. p16-immunohistochemistry also performed very well, however its main weakness is that it is an indirect marker of the presence of HPV. These data suggest that the RNAscope HPV-test is a promising test that could be developed as a clinical standard for the precise identification of HPV-driven oropharyngeal squamous cell carcinoma.
Shi X, Wu S, Huo Z, Ling Q, Luo Y, Liang Z.
PMID: 26285694 | DOI: 10.1186/s13000-015-0376-z.
Abstract
BACKGROUNDS:
The aim of this study was to describe the clinicopathological characteristics and high-risk human papillomavirus (HPV) infection status in patients diagnosed with co-existing of adenoid cystic carcinoma (ACC) and invasive squamous cell carcinoma (SCC) of the uterine cervix.
METHODS:
Three patients were identified from the pathology databank of Peking Union Medical College Hospital from year 2000 to 2014. Immunohistochemistry and in situ hybridization (ISH) were employed in this study.
RESULTS:
The patients were aged 64, 77 and 63 years (average, 68 years old). All the patients were postmenopausal women who presented with bloody or watery vaginal discharge. The cervical cytology screening results were all suspicious for high-grade squamous intraepithelial lesion (HSIL). The subsequent cervical colposcopy biopsies all showed cervical intraepithelial neoplasia III (CINIII). One patient received only a cervical conization, whereas the other two patients underwent hysterectomy. The immunohistochemical results showed that the ACC compartments were positive for CK7 and CD117; the cases of SCC were negative for CK7 and CD117. P63 staining was strongly positive and diffuse throughout the SCC compartments, whereas only patchy positive areas were observed in the ACC. MYB exhibited strong nuclear staining in the ACC and SCC compartments but negative staining in the endocervical gland. In situ hybridization (ISH) signals for high-risk HPV DNA and mRNA were present in the two compartments of all three patients. The patients had no evidence of disease at an average follow-up time of 21.6 months.
CONCLUSION:
High-risk HPV was present in both the ACC and SCC compartments in all three patients.
Chuerduangphui J, Pientong C, Chaiyarit P, Patarapadungkit N, Chotiyano A, Kongyingyoes B, Promthet S, Swangphon P, Wongjampa W, Ekalaksananan T.
PMID: 27349249 | DOI: 10.1007/s12032-016-0800-6
Human papillomavirus (HPV) infection modulates several host cytokines contributing to cancer development. Oncostatin M (OSM), an IL-6 family cytokine, acts to promote cell senescence and inhibit growth. Its dysregulation promotes cell survival, cell proliferation and metastasis in various malignancies. The effect of HPV on OSM dysregulation has not been investigated. To elucidate this, immunohistochemistry was used on formalin-fixed, paraffin-embedded oral squamous cell carcinoma (OSCC) tissues: HPV-positive (50) and HPV-negative (50) cases. Immortalized human cervical keratinocytes expressing HPV16E6 (HCK1T, Tet-On system) were used to demonstrate the role of HPV16E6 in OSM expression. In addition, a vector containing HPV16E6/E7 was transiently transfected into oral cancer cell lines. Cell viability, cell-cycle progression and cell migration were evaluated using flow cytometry and a wound healing assay, respectively. The results showed various intensities of OSM expression in OSCC. Interestingly, the median percentages of strongly stained cells were significantly higher in HPV-positive OSCCs than in HPV-negative OSCCs. To explore the role of HPV oncoproteins on OSM expression, the expression of HPV16E6 in the HCK1T Tet-On condition was induced by doxycycline and HPV16E6 was found to significantly upregulate levels of OSM mRNA and protein, with concomitant upregulation of c-Myc. In addition, the levels of OSM mRNA and protein in E6/E7 transiently transfected oral cancer cells also gradually increased in a time-dependent manner and these transfected cells showed greater viability and higher migration rates and cell-cycle progression than controls. This result demonstrates that HPV16 oncoproteins upregulate OSM and play an important role to promote OSCC development.
Rao, X;Zheng, L;Wei, K;Li, M;Jiang, M;Qiu, J;Zhou, Y;Ke, R;Lin, C;
PMID: 36809088 | DOI: 10.1128/spectrum.03896-22
RNA plays a vital role in the physiological and pathological processes of cells and tissues. However, RNA in situ hybridization applications in clinical diagnostics are still limited to a few examples. In this study, we developed a novel in situ hybridization assay for human papillomavirus (HPV) E6/E7 mRNA by taking advantage of specific padlock probing and rolling circle amplification, combined with chromogenic readout. We designed padlock probes for 14 types of high-risk HPV and demonstrated that E6/E7 mRNA could be visualized in situ as discrete dot-like signals using bright-field microscopy. Overall, the results are consistent with the clinical diagnostics lab's hematoxylin and eosin (H&E) staining and p16 immunohistochemistry test results. Our work thus shows the potential applications of RNA in situ hybridization for clinical diagnostics using chromogenic single-molecule detection, offering an alternative technical option to the current commercially available kit based on branched DNA technology. IMPORTANCE In situ detection of viral mRNA expression in tissue samples is of great value for pathological diagnosis to access viral infection status. Unfortunately, conventional RNA in situ hybridization assays lack sensitivity and specificity for clinical diagnostic purposes. Currently, the commercially available branched DNA technology-based single-molecule RNA in situ detection method offers satisfactory results. Here, we present our padlock probe- and rolling circle amplification-based RNA in situ hybridization assay for detecting HPV E6/E7 mRNA expression in formalin-fixed paraffin-embedded tissue sections, providing an alternative yet robust method for viral RNA in situ visualization that is also applicable to different types of diseases.
Chahoud, J;Zacharias, NM;Pham, R;Qiao, W;Guo, M;Lu, X;Alaniz, A;Segarra, L;Martinez-Ferrer, M;Gleber-Netto, FO;Pickering, CR;Rao, P;Pettaway, CA;
PMID: 36551510 | DOI: 10.3390/cancers14246024
Penile Squamous Cell Carcinoma (PSCC) is associated with high-risk human papillomavirus (HR-HPV). The immunohistochemical (IHC) test for p16INK4a (p16) is highly correlated with HR-HPV expression in other SCCs. To investigate whether the expression of p16 IHC or HR-HPV is associated with survival in PSCC, we conducted a single institution analysis of 143 patients with a diagnosis of PSCC and, available tissue were tested for p16 IHC staining patterns, histological subtype, tumor grade, and lymphovascular invasion (LVI) by an experienced pathologist. HR-HPV status using the Cobas PCR Assay or the RNAScope high-risk HPV in situ hybridization kit were also assessed. Patient characteristics were summarized using descriptive statistics of clinico-pathologic variables. Kaplan-Meier was used to estimate median overall survival (OS), cancer specific survival (CSS) and correlated with HPV, p16, and other study variables. Patients with p16+ tumors had a significantly longer median CSS in comparison to the p16- group (p = 0.004), with respective 5-year CSS probability of 88% (95% CI; 0.84, 1) versus 58% (95% CI; 0.55, 0.76; p = 0.004). HPV status did not predict survival outcomes. Multivariable analysis with respect to OS and CSS, showed that p16+ status was associated with a lower risk of death (HR = 0.36, 95%CI; 0.20-0.67, p = 0.001), and improved CSS (HR = 0.20, 95% CI; 0.07-0.54, p = 0.002) after adjusting for covariates. In conclusion, tumor p16 status via IHC was an easy to perform independent prognostic factor for OS and CSS that correlates with HR-HPV expression.
Archives of pathology & laboratory medicine
Banet, N;Mao, Q;Chu, S;Ruhul Quddus, M;
PMID: 35738001 | DOI: 10.5858/arpa.2021-0426-OA
Human papillomavirus (HPV) in the postmenopausal age group is complex, with infected patients in this age group at increased risk of progressing to invasive disease and showing decreased clearance of the virus. Additionally, atrophic changes of the cervix can make histologic distinction of high-grade squamous intraepithelial lesions (HSILs) difficult.To determine morphologic and ancillary testing characteristics of atrophy and HSIL in postmenopausal patients.Files of patients at least 65 years of age were examined, with 81 patients (109 cases [53 benign, 56 HSIL]) included in the study. Results of morphology, immunostaining (p16 and Ki-67), and HPV RNA in situ hybridization (ISH) were noted on all cases with available material.Atrophy was present in 96 of 109 cases (88%) overall. Coarse nuclear chromatin was noted in none of the benign cases, in 19 of 30 HSIL biopsies (63%), and in 24 of 26 HSIL excisions (92%). All benign cases were negative for p16 and ISH. In the HSIL cases, 45 of 53 (89%) were positive for p16, and of cases with sufficient tissue for ISH, 44 of 45 (98%) were positive. Of the ISH/p16 discordant cases (n = 7), most were p16 negative/ISH positive (6 of 7; 86%), whereas 1 of 7 (14%) was p16 positive and ISH negative. A majority of HSIL cases showed near-full-thickness elevation of Ki-67 (45 of 54; 83%), whereas mitotic figures were less elevated.In postmenopausal patients with HSIL, mitotic activity is not reliably elevated, but Ki-67 is consistently high. ISH is a more direct method of HPV detection and should be considered in cases where morphology and immunolabeling show discordance.
Xing D, Zheng G, Schoolmeester JK, Li Z, Pallavajjala A, Haley L, Conner MG, Vang R, Hung CF, Wu TC, Ronnett BM.
PMID: 29505425 | DOI: 10.1097/PAS.0000000000001042
Small cell neuroendocrine carcinoma (SCNEC) of the uterine cervix is a rare but extremely aggressive tumor. While high-risk human papillomavirus (HPV) is involved at an early stage of oncogenesis in many tumors, additional driving events have been postulated to facilitate the progression of SCNECs. Identification of oncogenic drivers could guide targeted therapy of this neoplasm. Clinicopathologic features of 10 cervical SCNECs are reported. Analyses included immunohistochemical evaluation of p16, p53, synaptophysin, and chromogranin expression; in situ hybridizations and polymerase chain reaction for high-risk HPV and/or HPV 18; and next-generation sequencing based on a 637-gene panel. The patients ranged in age from 28 to 68 years (mean, 45.6 y; median, 40.5 y). All tumors had diffuse p16 and synaptophysin expression. All but 1 tumor was positive for chromogranin (extent of staining ranged from focal to diffuse). HPV 18 was detected in 6 tumors and HPV 35 in 1 tumor. At least 1 driver mutation was detected in 8 tumors. Four cases harbored TP53 somatic mutations, 3 of which correlated with an aberrant p53 staining pattern. Four PIK3CA mutations (p.G106A, p.N345T, p.E545K, and p.E545D) were detected in 3 tumors, 2 of which also harbored TP53 mutations. Oncogenic driver mutations involving KRAS, Erbb2, c-Myc, NOTCH1, BCL6, or NCOA3 were detected in 4 tumors. Mutations in caretaker tumor suppressors PTEN, RB1, BRCA1, BRCA2, and ARID1B were also identified in 4 tumors that commonly coharbored activating oncogenic mutations. Targeted next-generation gene sequencing identified genetic alterations involving the MAPK, PI3K/AKT/mTOR, and TP53/BRCA pathways in SCNECs. The presence of genetic alterations that are amenable to targeted therapy in SCNECs offers the potential for individualized management strategies for treatment of this aggressive tumor.
Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc
Sauer, MA;Yang, J;Isidro, RA;Dong, F;Patil, DT;Wee, JO;Agoston, AT;Deshpande, V;Zhao, L;
PMID: 35351978 | DOI: 10.1038/s41379-022-01060-4
We compared clinicopathologic and molecular features of esophageal squamous cell carcinoma (SCC) with basaloid features to conventional SCC using surgical resections of treatment naïve esophageal carcinomas and cases available from the TCGA database. Twenty-two cases of SCC with basaloid features were identified in the Mass General Brigham pathology archives, including 9 cases with pure basaloid morphology and 13 cases with mixed other features such as conventional well- or poorly differentiated areas or sarcomatoid areas. Thirty-eight cases of conventional SCC matched by tumor stage were used as controls. HPV infection status was tested by p16 immunohistochemistry and HPV mRNA ISH. Digital slides for 94 cases of esophageal SCC from TCGA found in the Genomic Data Commons (GDC) Data Portal were reviewed. Five cases of SCC with basaloid features were identified. Genomic profiles of SCC with basaloid features were compared to the rest of 89 SCCs without basaloid features. In addition, eight tumor sections from six patients selected from our cohort underwent in-house molecular profiling. Compared to conventional SCC, SCC with basaloid features were more frequently associated with diffuse or multifocal squamous dysplasia (p < 0.001). P16 IHC was positive in 2/13 cases, whereas HPV mRNA ISH was negative in 17/17 cases (including both p16-positive cases). SCC with basaloid features and conventional SCC from TCGA showed similar rates of TP53 mutations, CDKN2A/B deletions, and CCDN1 amplifications. TP53 variants were identified in all in-house samples that had sufficient coverage. Survival analyses between SCC with basaloid features versus conventional SCC (matched for tumor stage) did not reveal any statistically significant differences. In conclusion, esophageal SCC with basaloid features has similar survival and genomic alterations to those of conventional SCC, are more frequently associated with diffuse or multifocal dysplasia, and are not associated with HPV (high-risk strains) infection.
MEK/ERK signaling is a critical regulator of high-risk human papillomavirus oncogene expression revealing therapeutic targets for HPV-induced tumors
Luna, AJ;Sterk, RT;Griego-Fisher, AM;Chung, JY;Berggren, KL;Bondu, V;Barraza-Flores, P;Cowan, AT;Gan, GN;Yilmaz, E;Cho, H;Kim, JH;Hewitt, SM;Bauman, JE;Ozbun, MA;
PMID: 33481911 | DOI: 10.1371/journal.ppat.1009216
Intracellular pathogens have evolved to utilize normal cellular processes to complete their replicative cycles. Pathogens that interface with proliferative cell signaling pathways risk infections that can lead to cancers, but the factors that influence malignant outcomes are incompletely understood. Human papillomaviruses (HPVs) predominantly cause benign hyperplasia in stratifying epithelial tissues. However, a subset of carcinogenic or "high-risk" HPV (hr-HPV) genotypes are etiologically linked to nearly 5% of all human cancers. Progression of hr-HPV-induced lesions to malignancies is characterized by increased expression of the E6 and E7 oncogenes and the oncogenic functions of these viral proteins have been widely studied. Yet, the mechanisms that regulate hr-HPV oncogene transcription and suppress their expression in benign lesions remain poorly understood. Here, we demonstrate that EGFR/MEK/ERK signaling, influenced by epithelial contact inhibition and tissue differentiation cues, regulates hr-HPV oncogene expression. Using monolayer cells, epithelial organotypic tissue models, and neoplastic tissue biopsy materials, we show that cell-extrinsic activation of ERK overrides cellular control to promote HPV oncogene expression and the neoplastic phenotype. Our data suggest that HPVs are adapted to use the EGFR/MEK/ERK signaling pathway to regulate their productive replicative cycles. Mechanistic studies show that EGFR/MEK/ERK signaling influences AP-1 transcription factor activity and AP-1 factor knockdown reduces oncogene transcription. Furthermore, pharmacological inhibitors of EGFR, MEK, and ERK signaling quash HPV oncogene expression and the neoplastic phenotype, revealing a potential clinical strategy to suppress uncontrolled cell proliferation, reduce oncogene expression and treat HPV neoplasia.
JAAD Case Reports 1.4 (2015): 196-199.
Chen CH, Wu YY, Kuo KT, Liau JY, Liang CW.
PMID: http
Merkel cell carcinoma (MCC), an uncommon and highly aggressive cutaneous malignancy, usually occurs on the sun-damaged skin of the elderly and is characterized by coexpression of neuroendocrine markers and CK20, a discriminant from other types of visceral neuroendocrine neoplasias. Since the discovery of Merkel cell polyomavirus (MCV), many researchers have confirmed its presence in about 80% of cutaneous MCCs.1 Although some cutaneous MCCs were reported to be associated with squamous cell carcinomas (SCCs), such combined cases accounted for only a minor portion and the viral status appeared to be different from pure MCC.
The Journal of dermatology
Shimizu, A;Yamaguchi, R;Kuriyama, Y;
PMID: 36601717 | DOI: 10.1111/1346-8138.16697
More than 200 types of human papillomavirus (HPV) have been reported to date and have been associated with various dermatological diseases. Among dermatological diseases, viral verrucae are the most commonly reported to be associated with HPV. Epidermodysplasia verruciformis (EV) consists of three types: typical EV is an autosomal recessive genetic disorder with TMC6/TMC8 gene mutations, atypical EV develops due to various gene mutations that cause immunodeficiency, and acquired EV develops due to acquired immunodeficiency. Generalized verrucosis differs from EV in that it involves numerous verrucous nodules (mainly on the limbs), histopathologically no blue cells as seen in EV, and infection with cutaneous α-HPVs as well as β-HPVs. HPV-induced skin malignancies include squamous cell carcinoma (SCC) caused by β-HPV (especially HPV types 5 and 8) in EV patients, organ transplant recipients, and healthy individuals, and SCC of the vulva and nail unit caused by mucosal high-risk HPV infection. Carcinogenesis of β-HPV is associated with sunlight. Mucosal high-risk HPV-associated carcinomas may also be sexually transmitted. We focused on Bowen's disease of the nail, which has been the subject of our research for a long time and has recently come to the fore in the field of dermatology.
A Model of Impaired Langerhans Cell Maturation Associated With HPV Induced Epithelial Hyperplasia
Tuong, Z;Lukowski, S;Nguyen, Q;Chandra, J;Zhou, C;Gillinder, K;Bashaw, A;Ferdinand, J;Stewart, B;Teoh, S;Hanson, S;Devitt, K;Clatworthy, M;Powell, J;Frazer, I;
| DOI: 10.2139/ssrn.3889711
Langerhans cells (LC) are skin-resident antigen-presenting cells that regulate immune responses to epithelial commensal and pathogenic microorganisms. Infection of skin by human papillomavirus (HPV) is commonly persistant and can promote malignant epithelial transformation. As LCs are considered important in control of HPV infection, we compared the transcriptome of LCs from skin transformed by the oncogenic E7 protein of HPV16 and from healthy skin. We identified transcriptome heterogeneity at the single cell level amongst LCs in normal mouse skin, associated with cell ontogeny, cell cycle activity, and maturation. We identified a balanced co-existence of immune-stimulatory and immune-inhibitory LC cell states in normal skin that was signficantly disturbed in HPV16 E7-expressing hyperproliferative murine skin. Hyperplastic skin was depleted of immune-stimulatory LCs, and enriched for LCs with an immune-inhibitory gene signature, and LC-keratinocyte cross-talk was dysregulated. We identified reduced expression of the CSF1R ligand interleukin (IL)-34, a critical molecule for LC homeostasis. Enrichment of an immune-inhibitory LC gene signature and reduced levels of epithelial IL-34 were also found in human HPV-associated cervical epithelial cancers. HPV16 E7 transgenic hyperplastic murine skin thus provides a robust model to investigate interventions to overcome impaired LC maturation induced by epithelial proliferation in HPV associated cancer.