Lewis, JS;Smith, MH;Wang, X;Tong, F;Mehrad, M;Lang-Kuhs, KA;
PMID: 35802245 | DOI: 10.1007/s12105-022-01467-0
HPV-associated oral cavity squamous cell carcinoma (SCC) is not well-characterized in the literature, and also has a clinical significance that is poorly understood.We gathered a cohort of oral cavity (OC) SCC with nonkeratinizing morphology, either in the invasive or in situ carcinoma (or both), tested for p16 by immunohistochemistry and high risk HPV E6/E7 mRNA by RTPCR (reference standard for transcriptionally-active high risk HPV) and gathered detailed morphologic and clinicopathologic data.Thirteen patients from two institutions were proven to be HPV-associated by combined p16 and high risk HPV mRNA positivity. All 13 patients (100%) were males, all were heavy smokers (average 57 pack/year), and most were active drinkers (9/11 or 81.8%). All 13 (100%) involved the tongue and/or floor of mouth. All had nonkeratinizing features, but maturing squamous differentiation varied widely (0-90%; mean 37.3%). Nonkeratinizing areas had high N:C ratios and larger nests, frequently with pushing borders, and minimal (or no) stromal desmoplasia. The carcinoma in situ, when present, was Bowenoid/nonkeratinizing with cells with high N:C ratios, full thickness loss of maturation, and abundant apoptosis and mitosis. HPV was type 16 in 11 patients (84.6%) and type 33 in two (15.4%). Nine patients had treatment data available. These underwent primary surgical resection with tumors ranging from 1.6 to 5.2 cm. Most had bone invasion (6/9-66.7% were T4a tumors), and most (6/9-66.7%) had extensive SCC in situ with all 6 of these patients having final margins positive for in situ carcinoma.HPV-associated OCSCC is an uncommon entity that shows certain distinct clinical and pathologic features. Recognition of these features may help pathologic diagnosis and could potentially help guide clinical management.
Rajendra S, Yang T, Xuan W, Sharma P, Pavey D, Soon Lee C, Le S, Collins J, Wang B.
PMID: 28722212 | DOI: 10.1002/ijc.30896
We have previously demonstrated that transcriptionally active high-risk HPV (hr-HPV) is strongly incriminated in Barrett's dysplasia (BD) and oesophageal adenocarcinoma (OAC) using mainly fresh frozen tissue. This study aimed to identify biomarkers of active HPV infection in Barrett's metaplasia, (BM)/BD/OAC by immunohistochemical staining (IHC) of formalin-fixed paraffin embedded (FFPE) tissue for aberrations of p53 and the retinoblastoma (pRb) pathway which are targets for the viral oncoproteins, E6/E7 respectively. Prospectively, BM(n=81)/BD(n=72)/OAC(n=65) FFPE specimens were subjected to IHC staining for pRb, p16INK4A , cyclin D1 , p53 and RNA in-situ hybridization (ISH) for E6/E7 transcripts. HPV DNA was determined via PCR in fresh frozen specimens. Viral load measurement (real-time PCR) and Next Generation Sequencing of TP53 was also performed. Of 218 patients, 56 were HPV DNA positive [HPV16 (n=42), 18 (n=13), 6 (n=1)]. Viral load was low. Transcriptionally active HPV (DNA+ /RNA+ ) was only found in the dysplastic and adenocarcinoma group (n=21). The majority of HPV DNA+ /RNA+ BD/OAC were characterized by p16INK4Ahigh (14/21, 66.7%), pRblow (15/21, 71.4%) and p53low (20/21, 95%) and was significantly different to controls [combination of HPV DNA- /RNA- (n=94) and HPV DNA+ /RNA- cohorts (n=22)]. p53low had the strongest association with DNA+ /RNA+ oesophageal lesions (OR=23.5, 95% CI=2.94-187.8, p=0.0029). Seventeen HPV DNA+ /RNA+BD/OAC identified as p53low, were sequenced and all but one exhibited wild-type status. pRblow /p53low provided the best balance of strength of association (OR=8.0, 95% CI=2.6-25.0, p=0.0003) and sensitivity (71.4%)/specificity (71.6%) for DNA+ /RNA+ BD/OAC. Active HPV involvement in BD/OAC is characterized by wild-type p53 and aberrations of the retinoblastoma protein pathway.
D'Souza G, Westra WH, Wang SJ, van Zante A, Wentz A, Kluz N, Rettig E, Ryan WR, Ha PK, Kang H, Bishop J, Quon H, Kiess AP, Richmon JD, Eisele DW, Fakhry C.
PMID: 27930766 | DOI: 10.1001/jamaoncol.2016.3067
Abstract
IMPORTANCE:
Human papillomavirus (HPV) causes an increasing proportion of oropharyngeal squamous cell carcinomas (OPSCCs), particularly in white men. The prevalence of HPV among other demographic groups and other anatomic sites of HNSCC is unclear.
OBJECTIVE:
To explore the role of HPV tumor status among women and nonwhites with OPSCC and patients with nonoropharyngeal head and neck squamous cell carcinoma (non-OP HNSCC).
DESIGN, SETTING, AND PARTICIPANTS:
Retrospective cohort study at 2 tertiary academic centers including cases diagnosed 1995 through 2012, oversampled for minorities and females. A stratified random sample of 863 patients with newly diagnosed SCC of the oral cavity, oropharynx, larynx, or nasopharynx was used.
MAIN OUTCOMES AND MEASURES:
Outcomes were HPV status as measured by p16 immunohistochemical analysis, HPV16 DNA in situ hybridization (ISH), and high-risk HPV E6/E7 mRNA ISH.
RESULTS:
Of 863 patients, 551 (63.9%) were male and median age was 58 years (interquartile range, 51-68 years). Among 240 OPSCCs, 144 (60%) were p16 positive (p16+), 115 (48%) were HPV16 DNA ISH positive (ISH16+), and 134 (56%) were positive for any oncogenic HPV type (ISH+). From 1995 to 2012, the proportion of p16+ OPSCC increased significantly among women (from 29% to 77%; P = .005 for trend) and men (36% to 72%; P < .001 for trend), as well as among whites (39% to 86%; P < .001 for trend) and nonwhites (32% to 62%; P = .02 for trend). Similar results were observed for ISH+ OPSCC (P ≤ .01 for all). Among 623 non-OP HNSCCs, a higher proportion were p16+ compared with ISH positive (62 [10%] vs 30 [5%]; P = .001). A high proportion (26 of 62 [42%]) of these p16+ non-OP HNSCCs were found in sites adjacent to the oropharynx. The proportion of p16+ and ISH+ non-OP HNSCCs were similar by sex. Over time, the proportion of non-OP HNSCCs that were p16+ (or ISH+) increased among whites (P = .04 for trend) but not among nonwhites (each P > .51 for trend). Among OPSCCs, p16 had high sensitivity (100%), specificity (91%), and positive (93%) and negative predictive value (100%) for ISH positivity. In non-OP HNSCCs, p16 had lower sensitivity (83%) and positive predictive value (40%) but high specificity (94%) and negative predictive value (99%) for ISH positivity.
CONCLUSIONS AND RELEVANCE:
During 1995 through 2012, the proportion of OPSCCs caused by HPV has increased significantly. This increase was not restricted to white men but was a consistent trend for women and men, as well as for white and nonwhite racial groups. Few non-OP HNSCCs were HPV related. P16 positivity was a good surrogate for ISH+ tumor status among OPSCC, but not a good surrogate for non-OP HNSCC.
Chinese journal of pathology
Zhao YH, Bai YP, Mao ML, Zhang H, Zhao XL, Yang DM, Wan HF, Liu HG.
PMID: 30695865 | DOI: 10.3760/cma.j.issn.0529-5807.2019.02.010
Objective: To observe the clinicopathologic features of oropharyngeal squamous cell carcinoma associated with human papilloma virus (OPSCC-HPV) and discuss the role and value of different in situ hybridization (ISH) detection methods for HPV in pathologic diagnosis. Methods: Fifteen cases of OPSCC-HPV were collected from Department of Pathology, Beijing Tongren Hospital, Capital Medical University from January 2016 to August 2018. These cases were diagnosed in accordance with the WHO classification of head and neck tumors. The histopathologic features and the clinicopathologic data were retrospectively analyzed. Immunohistochemistry (two-step EnVision method) was done to evaluate the expression of p16, Ki-67 and p53. ISH was used to detect HPV DNA (6/11 and 16/18). RNAscope technology was used to evaluate the presence of HPV mRNAs (16 and 18). Results: The mean age for the 15 patients (8 males, 7 females) was 47 years (range from 30 to 69 years). OPSCC-HPV typically presentedat an advanced clinical stage, six patients had cervical lymphadenopathy (large and cystic), seven had tonsillar swelling, one had tumor at base of tongue, and one had odynophagia. Microscopically the tumors exhibited distinctive non-keratinizing squamous cell carcinoma morphology. Cervical nodal metastases were large and cystic, with thickening of lymph node capsules. OPSCC-HPV raised from crypt epithelium and extended beneath the tonsillar surface epithelial lining as nests and lobules, often with central necrosis. Tumor cells displayed a high N: C ratio, and high mitotic and apoptotic rates. Tumor nests are often embedded within lymphoid stroma, and may be infiltrated by lymphoid cells.Fifteen cases (15/15) were strongly positive for p16; Ki-67 index were 60%-90%; they were focally positive or negative for p53. Ten cases (10/10) were negative for HPV 6/11 DNA, and one case(1/10) was focally positive for HPV16/18 DNA. Eleven cases (11/11) were strongly positive for HPV16 mRNA, one case was focally positive for HPV18 mRNA. Conclusions: OPSCC-HPV is a pathologically and clinically distinct form of head and neck squamous cell carcinoma. OPSCC-HPV is associated with high-risk HPV (type 16) in all cases. Detection of high-risk HPV16 mRNA by RNAscope is of great significance in the final diagnosis and pathogen identification.
Myers, JE;Schaal, DL;Nkadi, EH;Ward, BJH;Bienkowska-Haba, M;Sapp, M;Bodily, JM;Scott, RS;
PMID: 36719239 | DOI: 10.1128/jvi.01032-22
Coinfection of human papillomavirus (HPV) and Epstein-Barr virus (EBV) has been detected in oropharyngeal squamous cell carcinoma. Although HPV and EBV replicate in differentiated epithelial cells, we previously reported that HPV epithelial immortalization reduces EBV replication within organotypic raft culture and that the HPV16 oncoprotein E7 was sufficient to inhibit EBV replication. A well-established function of HPV E7 is the degradation of the retinoblastoma (Rb) family of pocket proteins (pRb, p107, and p130). Here, we show that pRb knockdown in differentiated epithelia and EBV-positive Burkitt lymphoma (BL) reduces EBV lytic replication following de novo infection and reactivation, respectively. In differentiated epithelia, EBV immediate early (IE) transactivators were expressed, but loss of pRb blocked expression of the early gene product, EA-D. Although no alterations were observed in markers of epithelial differentiation, DNA damage, and p16, increased markers of S-phase progression and altered p107 and p130 levels were observed in suprabasal keratinocytes after pRb knockdown. In contrast, pRb interference in Akata BX1 Burkitt lymphoma cells showed a distinct phenotype from differentiated epithelia with no significant effect on EBV IE or EA-D expression. Instead, pRb knockdown reduced the levels of the plasmablast differentiation marker PRDM1/Blimp1 and increased the abundance of c-Myc protein in reactivated Akata BL with pRb knockdown. c-Myc RNA levels also increased following the loss of pRb in epithelial rafts. These results suggest that pRb is required to suppress c-Myc for efficient EBV replication in BL cells and identifies a mechanism for how HPV immortalization, through degradation of the retinoblastoma pocket proteins, interferes with EBV replication in coinfected epithelia. IMPORTANCE Terminally differentiated epithelium is known to support EBV genome amplification and virion morphogenesis following infection. The contribution of the cell cycle in differentiated tissues to efficient EBV replication is not understood. Using organotypic epithelial raft cultures and genetic interference, we can identify factors required for EBV replication in quiescent cells. Here, we phenocopied HPV16 E7 inhibition of EBV replication through knockdown of pRb. Loss of pRb was found to reduce EBV early gene expression and viral replication. Interruption of the viral life cycle was accompanied by increased S-phase gene expression in postmitotic keratinocytes, a process also observed in E7-positive epithelia, and deregulation of other pocket proteins. Together, these findings provide evidence of a global requirement for pRb in EBV lytic replication and provide a mechanistic framework for how HPV E7 may facilitate a latent EBV infection through its mediated degradation of pRb in copositive epithelia.
Annals of oncology : official journal of the European Society for Medical Oncology
Rischin, D;Mehanna, H;Young, RJ;Bressel, M;Dunn, J;Corry, J;Soni, P;Fulton-Lieuw, T;Iqbal, G;Kenny, L;Porceddu, S;Wratten, C;Robinson, M;Solomon, BJ;Trans-Tasman Radiation Oncology Group and the De-ESCALaTE HPV Trial Group, ;
PMID: 35525376 | DOI: 10.1016/j.annonc.2022.04.074
High CD103+ intratumoral immune cell (ITIC) abundance is associated with better prognosis in unselected patients with human papilloma virus associated oropharyngeal squamous cell carcinoma(HPV-associated OPSCC) treated with cisplatin and radiotherapy(CIS/RT). Substituting cetuximab(CETUX) for CIS with RT in HPV-associated OPSCC resulted in inferior efficacy. Our aim was to determine if quantification of ITIC CD103 could be used to identify a population of HPV-associated OPSCC with superior prognosis.We pooled data from the TROG 12.01 and De-ESCALaTE randomised trials that compared CETUX/70GyRT with CIS/70GyRT in low risk HPV-associated OPSCC: AJCC 7th Stage III (excluding T1-2N1) or stage IV (excluding N2b-c if smoking history >10 pack years and/or distant metastases), including all patients with available tumor samples. The primary endpoint was failure-free survival (FFS) in patients receiving CETUX/ RT comparing CD103+ ITIC high (>30%) versus low (<30%). High/low CD103 were compared using Cox regression adjusting for age, stage and trial.Tumor samples were available in 159/182 patients on TROG 12.01 and 145/334 on De-ESCALaTE. CD103+ ITIC abundance was high in 27% of patients. The median follow-up was 3.2 years. The 3-year FFS in patients treated with CETUX/RT were 93% (95% CI: 79-98%) in high CD103 and 74% (95% CI: 63-81%) in low CD103, adjusted HR 0.22 (95% CI: 0.12-0.41); p<0.001. The 3-year overall survival in patients treated with CETUX/RT was 100% in high CD103 and 86% (95% CI: 76-92%) in low CD103, p<0.001. In patients treated with CIS/RT there was no significant difference in FFS.CD103+ ITIC expression separates CETUX/RT treated low risk HPV-associated OPSCC into excellent and poor prognosis subgroups. The high CD103 population is a rational target for de-intensification trials.
Virchows Archiv (2015): 1-9.
Hauck F, Oliveira-Silva M, Dreyer JH, Ferreira Perrusi VJ, Arcuri RA, Hassan R, Bonvicino CR, Barros MHM, Niedobitek G.
PMID: 25820374 | DOI: 10.1007/s00428-015-1761-4
Rising prevalence rates of high-risk human papillomaviruses (hrHPV) infection in oropharyngeal carcinoma (up to 80 %) have been reported in North America and Scandinavia. We have analysed 424 German and 163 Brazilian head and neck squamous cell carcinomas (HNSCC) from the oral cavity (OSCC), oropharynx (OPSCC) and hypopharynx (HPSCC) using p16 immunohistochemistry, HPV DNA PCR and sequencing, hrHPV DNA in situ hybridisation (ISH) and hrHPV E6/E7 RNA ISH. In the German series, 52/424 cases (12.3 %) were p16-positive/hrHPV-positive (OSCC 3.8 % [10/265], OPSCC 34.4 % [42/122], HPSCC 0 % [0/37]). In addition, there were 9 cases that were p16-positive/hrHPV-negative (5 OPSCC and 4 OSCC). In the Brazilian series, the overall hrHPV DNA prevalence by PCR was 11.0 % ([18/163]; OSCC 6 % [5/83], OPSCC 15.5 % [11/71], HPSCC 22.2 % [2/9]). Ten of these cases were hrHPV-positive/p16-positive. The remaining 8 hrHPV-positive/p16-negative cases were also negative in both ISH assays. Furthermore, 5 p16-positive/hrHPV-negative cases (2 OPSCC and 3 OSCC) were identified. In both series, HPV16 was by far the most common HPV type detected. We confirm that regardless of geographical origin, the highest hrHPV prevalence in HNSCC is observed in oropharyngeal carcinomas. The proportion of HPV-associated OPSCC was substantially higher in the German cohort than in the Brazilian series (34.4 vs. 15.5 %), and in both groups, the prevalence of hrHPV in OPSCC was much lower than in recent reports from North America and Scandinavia. We suggest, therefore, that it may be possible to define areas with high (e.g. USA, Canada, Scandinavia), intermediate (e.g. Germany) and low (e.g. Brazil) prevalences of HPV infection in OPSCC.
ER-positive endocervical adenocarcinoma mimicking endometrioid adenocarcinoma in morphology and immunohistochemical profile: A case report of application of HPV RNAscope detection
Chen, R;Qin, P;Luo, Q;Yang, W;Tan, X;Cai, T;Jiang, Q;Chen, H;
PMID: 33787580 | DOI: 10.1097/MD.0000000000024927
Usual-type endocervical adenocarcinoma (ECA), high-risk HPV associated, is the most common type of glandular carcinoma in the endocervix. Mucin-depleted usual-type ECA is 1 end of morphological lineage of usual-type ECA and morphologically may show endometrioid features, which could cause diagnostic challenge with uterine endometrioid adenocarcinoma (EEC) and primary endometrioid ECA, especially in the setting of small biopsy and endocervical curettage (ECC). A 37-year-old women presented with dyspareunia for 1 year, showing atypical glandular cell on a liquid-based Pap TCT examination and positive for HPV16 detection. ECC showed EEC in another hospital based on its "endometrioid" morphology and immunohistochemical profiles (ER/PR/PAX8 strongly positive, though p16 also strongly positive). The specimen of hysterectomy in our hospital displayed a lesion confined to the uterine cervix showing the same morphology and immunohistochemical profiles as ECC. Finally, we successfully performed HPV RNAscope and detected high-risk human papilloma virus (HPV) E6/E7 mRNA particles in tumor cells in situ, which warranted usual-type ECA with mucin-depleted feature, a rare deviation of usual-type of ECA. The patient underwent total hysterectomy with lymph node dissection. To date, 14 months after surgery, the patient is well without recurrence or distant metastasis, and undergoes regular reexamination. We report a rare case of mucin-depleted usual-type ECA showing overlapping morphological and immunohistochemical profiles with EEC. The pathological diagnosis was confirmed by high-risk HPV RNAscope detection which is superior than immunohistochemistry to identify usual-type ECA, warranting an important role in assisting the diagnosis of morphological vague cases.
Virchows Archiv : an international journal of pathology
Asaka, S;Nakajima, T;Ida, K;Asaka, R;Kobayashi, C;Ito, M;Miyamoto, T;Uehara, T;Ota, H;
PMID: 34581850 | DOI: 10.1007/s00428-021-03207-1
In 2020, the WHO published a new system for classifying invasive endocervical adenocarcinoma based on histological features and high-risk human papillomavirus (HPV) infection. However, immunophenotypes of each histological subtype require further investigation. We immunohistochemically analyzed 66 invasive endocervical adenocarcinomas using three cell-lineage-specific markers: claudin 18 (CLDN18) for gastric, cadherin 17 (CDH17) for intestinal, and PAX8 for Müllerian epithelial cells. We identified five immunophenotypes of endocervical adenocarcinoma: gastric (21%); intestinal (14%); gastrointestinal (11%); Müllerian (35%); and not otherwise specified (NOS) (20%). Adenocarcinomas with gastric immunophenotype, characterized by aging (p = 0.0050), infrequent HPV infection (p < 0.0001), concurrent lobular endocervical glandular hyperplasia (p = 0.0060), lymphovascular invasion (p = 0.0073), advanced clinical stage (p = 0.0001), and the poorest progression-free (p < 0.0001) and overall (p = 0.0023) survivals, were morphologically compatible with gastric-type adenocarcinoma of the WHO 2020 classification. Conversely, most adenocarcinomas with Müllerian (91%) and intestinal (89%) immunophenotypes were HPV associated and morphologically compatible with usual- or intestinal-type adenocarcinomas of the WHO 2020 classification. The morphology of adenocarcinomas with gastrointestinal immunophenotype was intermediate or mixed between those of gastric and intestinal immunophenotypes; 57% were HPV associated. Adenocarcinomas with NOS immunophenotype were mainly HPV associated (85%) and histologically poorly differentiated. Multivariate analysis revealed that gastric (p = 0.008), intestinal + gastrointestinal (p = 0.0103), and NOS (p = 0.009) immunophenotypes were independent predictors of progression-free survival. Immunophenotypes characterized by CLDN18, CDH17, and PAX8 exhibited clinicopathological relevance and may improve the diagnostic accuracy and prognostic value of conventional histological classification.
Role of HPV status and PD-L1 expression in prognosis of laryngeal squamous cell carcinoma
International journal of clinical and experimental pathology
Yang, SM;Wu, M;Han, FY;Sun, YM;Yang, JQ;Liu, HX;
PMID: 33532028
Human papillomavirus (HPV) infection has been recognized as a cause of head and neck squamous cell carcinomas (HNSCC). Laryngeal squamous cell carcinoma (LSCC) is one of the most common pathologic types of HNSCC. Clinical trials show that there are differences in response to immunotherapy according to HPV status. It was reported that a high level of programmed cell death-ligand 1 (PD-L1) is correlated with better survival in HPV-positive head and neck cancer. In this study, we investigated the expression of PD-L1 in HPV-positive and HPV-negative LSCC to determine its prevalence and prognostic value. 52 cases of LSCC were collected from Tangshan Head and Neck Disease Pathology Research Base. PCR-reverse dot blot hybridization and RNAscope in situ hybridization were used to detect HPV status. PD-L1 expression was evaluated by immunohistochemistry and all cases were followed up for survival. SPSS24.0 was used for data entry and statistical analysis. Kaplan-Meier method and Log-rank time series analysis were used for single factor analysis. Multivariate analysis was performed using Cox proportional hazard regression model, and HR and 95% CI were calculated. Of the 52 LSCC patients, 32.7% (17/52) were HPV-positive by RNAscope in situ hybridization, and 51.9% (27/52) of patients were positive for PD-L1 expression by immunohistochemistry. Regression analysis showed that with a median follow-up period of 69 months, smoking and late stage were associated with poor overall survival (OS), whereas HPV positivity and PD-L1 expression showed a better overall survival outcome. Smoking status, tumor stage, HPV status, and PD-L1 expression in tumor cells may represent useful prognostic biomarkers in patients with LSCC. IJCEP
Cancer Cytopathol. 2015 Aug 4.
Jalaly JB, Lewis JS Jr, Collins BT, Wu X, Ma XJ, Luo Y, Bernadt CT.
PMID: 26242494 | DOI: 10.1002/cncy.21600.
Abstract BACKGROUND: Human papillomavirus (HPV)-related oropharyngeal squamous cell carcinoma (SCC) is a unique form of carcinoma that is important to identify for prognosis and treatment. Immunohistochemistry (IHC) for p16 (also known as cyclin-dependent kinase inhibitor 2A, multiple tumor suppressor 1) is used as a surrogate marker for transcriptionally active, high-risk HPV. The primary objective of this study was to correlate p16 IHC of cell blocks from fine-needle aspirations (FNAs) with surgical pathology specimens of HPV-related oropharyngeal SCC. METHODS: In total, 48 patients who had a diagnosis of oropharyngeal or nonoropharyngeal SCC and also had an FNA that demonstrated metastatic SCC with available cell block material were identified. IHC for p16 was evaluated on both FNA cell blocks and surgical pathology specimens. In situ hybridization for high-risk HPV messenger RNA was performed on 31 of the FNA cell blocks. RESULTS: Although partial p16 staining was observed in the majority of cell blocks, there was concordance in 47 of 48 FNAs (98%) with surgical pathology specimens when strong positive p16 staining of at least 15% of tumor cells in FNA cell block material was present. In addition, high-risk HPV RNA in situ hybridization demonstrated a high correlation with p16 staining in surgical pathology specimens (96%) and FNAs (93%). CONCLUSIONS: There was excellent correlation between p16 IHC of FNA cell blocks and surgical pathology specimens using a cutoff of at least 15% positive staining in cell blocks. The recommended threshold (70% positive staining) for surgical pathology specimens may yield a high rate of false-negative results if applied to FNA cell blocks.
Mod Pathol. 2012 Sep;25(9):1212-20.
Lewis JS Jr1, Chernock RD, Ma XJ, Flanagan JJ, Luo Y, Gao G, Wang X, El-Mofty SK (2012)
PMID: 22596101doi
Human papillomavirus (HPV)-related oropharyngeal squamous cell carcinoma has unique biology and better outcomes. p16 immunostaining is used as a surrogate marker for transcriptionally active HPV. Although diffuse staining is generally accepted as positive, the significance of partial staining has not been established, nor has the cutoff for extent of p16 staining that should be used to identify a tumor as HPV-related. From three other large studies utilizing p16 immunohistochemistry, we identified all cases with partial positive staining. The p16-stained slides were reviewed by three study pathologists for staining (nuclear and cytoplasmic) extent (in quartiles), and also for percentage that was confluent (ie, back-to-back cell staining). Tumors were histologically typed (keratinizing, non-keratinizing, or non-keratinizing with maturation) and tested for high-risk HPV by RNA in-situ hybridization and reverse-transcriptase PCR. For the 16 cases, there were two 4+(13%), five 3+(31%), six 2+(38%), and three 1+(19%) p16 staining tumors. Extent of staining ranged from 5 to 90% of cells positive with 25% or more confluent staining in 4/16 (25%). Of the 16 (31%) cases, 5 were HPV-related on the basis of RNA in-situ hybridization and reverse-transcriptase PCR. All of these cases had >50% p16 staining, 4/5 (80%) had more than 25% confluent staining, and 4/7 (57%) had non-keratinizing histological features. Only one of the p16 1+/2+ tumors was HPV RNA-positive (by reverse-transcriptase PCR only and low level). All 1+/2+ cases were keratinizing type or undifferentiated. By sensitive detection methods, most partial p16-positive squamous cell carcinoma cases with >50% staining harbor transcriptionally active HPV, and most HPV+ tumors have significant amounts of confluent staining. Cases with <50% p16 staining and lacking significant confluent staining rarely harbor HPV. These results support that greater than 75% p16 staining or, alternatively, >50% staining combined with >25% confluent areas, are suitable cutoffs for defining positivity.
