Immunoreactivity of LMO7 and other molecular markers as potential prognostic factors in oropharyngeal squamous cell carcinoma.

BACKGROUND: Despite the better prognosis associated with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC), some patients experience relapse and succumb to the disease; thus, there is a need for biomarkers identifying these patients for intensified treatment. Leucine-rich repeats and immunoglobulin-like domain (LRIG) protein 1 is a negative regulator of receptor tyrosine kinase signaling and a positive prognostic factor in OPSCC. Studies indicate that LRIG1 interacts with the LIM domain 7 protein (LMO7), a stabilizer of adherence junctions. Its role in OPSCC has not been studied before. METHODS: A total of 145 patients diagnosed with OPSCC were enrolled. Immunohistochemical LMO7 expression and staining intensity were evaluated in the tumors and correlated with known clinical and pathological prognostic factors, such as HPV status and LRIG1, CD44, Ki67, and p53 expression. RESULTS: Our results show that high LMO7 expression is associated with significantly longer overall survival (OS) (p = 0.044). LMO7 was a positive prognostic factor for OS in univariate analysis (HR 0.515, 95% CI: 0.267-0.994, p = 0.048) but not in multivariate analysis. The LMO7 expression correlated with LRIG1 expression (p = 0.048), consistent with previous findings. Interestingly, strong LRIG1 staining intensity was an independent negative prognostic factor in the HPV-driven group of tumors (HR 2.847, 95% Cl: 1.036-7.825, p = 0.043). CONCLUSIONS: We show for the first time that high LMO7 expression is a positive prognostic factor in OPSCC, and we propose that LMO7 should be further explored as a biomarker. In contrast to previous reports, LRIG1 expression was shown to be an independent negative prognostic factor in HPV-driven OPSCC.

Association of Human Papillomavirus Infection with Tonsillar Cancers: A Systematic Review.

Vaccinations have shown a decrease in human papillomavirus (HPV) infection-related cervical cancer in women, but there has been a sharp rise in the HPV infection-related oropharyngeal cancer cases over the past few decades. Recent studies have suggested the association of HPV infections with tonsillar cancers as well and suggestions regarding preventive tonsillectomies in order to achieve a decrease in HPV infection-related oropharyngeal or tonsillar cancer have arisen. However, there is limited cumulative evidence validated at a global level to support the endorsement of this strategy. This research revolves around the concept of burden of tonsillar carcinomas due to oropharyngeal HPV infection. Thus, a systematic review and meta-analysis of existing studies was undertaken to estimate the pooled prevalence of tonsillar cancer associated with oropharyngeal HPV infection. Published articles on tonsillar cancer with and without HPV infection from PubMed, Embase, Scopus and Web of Science were systematically searched from inception until 23 December 2021. A random-effects model was used to estimate the pooled prevalence forest plots. The systematic review revealed that 50% of the reported cases of tonsillar cancer had an oropharyngeal HPV infection, questioning the preventive nature of an early tonsillectomy which is essentially an invasive surgical procedure. Large heterogeneity was reported in the included studies, and there was insufficient data for sub-group analysis. Future research and representative studies are required to thoroughly explore the correlation between HPV infection and tonsillar cancer. Supplementary Information: The online version contains supplementary material available at 10.1007/s12070-023-04140-2.

Association between tonsillectomy and oropharyngeal cancer risk: a retrospective cohort study.

BACKGROUND: The purpose of this retrospective cohort study is to describe the association between the history of tonsillectomy and the risk of oropharyngeal squamous cell carcinoma (OPSSC), using a large cohort of patients. MATERIALS AND METHODS: We performed a retrospective cohort study with 3620 patients diagnosed with OPٍSCC from 2010 to 2021. We utilized the University of Florida patients' registry i2b2 system. Three subsets of OPSSC were defined, base of tongue(BOT) cancer, tonsillar cancer, and other OPSSC. Tumor demographics and history of tonsillectomy were collected. Odds ratio for OPSSC were assessed utilizing a logistic regression model with adjusting for gender, race, and age. P < 0.05 was deemed significant. RESULTS: Of the 3620 OPSSC patients were BOT cancer (N = 964), tonsillar cancer (N = 995), and other OPSSC (N = 1661). There was a statistically significant reduction in tonsillar cancer and BOT cancer odds ratio in patients with a history of tonsillectomy vs. patients without tonsillectomy (0.086 and 0.117), respectively, with a P value < .0001. The odds ratio of OPSSC in patients with a history of tonsillectomy vs. patients without tonsillectomy is 1.031. CONCLUSION: This study showed that the OPSSC and previous history of tonsillectomy are associated. Our results showed a significant reduction in BOT and tonsillar cancer risk in patients with a history of tonsillectomy and an insignificant decrease in other OPSSC. This study could emphasize the importance of the development of future clinical trials to investigate the role of prophylactic tonsillectomy as a secondary preventive strategy to reduce OPSSC.

Neoadjuvant chemotherapy followed by surgery for HPV-associated tonsillar cancer: Does imaging reflect the pathological response?

OBJECTIVES: The purpose of this study was to present our evaluation of the outcome of tonsillar cancer managed with neoadjuvant chemotherapy followed by surgery as definitive treatment. MATERIALS AND METHODS: Thirty-eight patients with human papilloma virus (HPV)-associated tonsillar cancer were treated with neoadjuvant chemotherapy followed by surgery. The volume reduction response of the tumor to neoadjuvant chemotherapy was evaluated and verified by histopathology. RESULTS: The complete pathologic response (pCR) rates at primary and nodal sites were 60 % and 45 %. Tumor volume reduction ≥78.8 % following neoadjuvant chemotherapy predicted pCR of the cervical node. In addition, the optimal cut-off value to predict pCR at the primary tumor site was 83.4 % volume reduction but was not a significant result. For pCR, neoadjuvant chemotherapy decreased the pathological adverse features, significantly reducing the need for adjuvant therapy. The overall survival of the adjuvant group was 79.2 %, and that of the non-adjuvant group was 87.5 %, with disease-free survival of 65.9 % and 54.2 %. There was no significant difference between the two groups. CONCLUSION: Neoadjuvant chemotherapy followed by surgery proved to be a good therapeutic option for management of HPV-associated tonsillar cancer. A greater reduction in tumor volume in post-neoadjuvant chemotherapy imaging predicts a complete pathologic response.

Targeted spatial proteomic analysis of CD8+ T- and myeloid cells in tonsillar cancer.

Background: Tonsillar cancer is caused by high-risk human papillomavirus (HPV), tobacco smoking, and alcohol abuse. Aspects of the patient's immune response to this disease have arisen as prognostic factors and treatment targets, reflecting differences in the type and protein expression profile of immune cells. Because tonsillar cancers are heterogenous lesions such data need to be spatially resolved. Methods: In this study, we aim to explore inter-patient and intra-tumoral sources of variation in tonsillar cancer using immunofluorescence and targeted spatial proteomics to interrogate a cohort of 105 patients. Furthermore, we assess prognostic factors and elucidate molecular targets. We have used CD8, CD11c, and Pan-cytokeratin (PanCK) to quantify and locate immune cells driving antigen-specific cellular immunity. Guided by immunofluorescence information, we selected 355 CD8+, CD11c+, or PanCK+ areas inside and outside (i.e., stroma) cancer-cell islets, to quantify 43 immune-related proteins using digital spatial profiling. Results: Quantitative analysis of immunofluorescence in combination with clinical data revealed that the abundance of total CD8+ cells and CD8+ cells infiltrating cancer-cell islets, respectively, were associated with higher 5-year disease-free survival and overall survival, independently of HPV-status and clinical stage. Comparison of CD8+ cells inside and outside cancer-cell islets revealed an upregulation of effector CD8+ T-cell and immune checkpoint molecules in the former. Among these, the expression of PD-L1 by CD8+ T-cells was associated with lower all-cause mortality in a univariate proportional hazards model. Similarly, a comparison of tumor boundary and stroma CD11c+ cells showed upregulation of both co-stimulatory and immune checkpoint molecules with proximity to tumor cell islets. Conclusion: Our findings highlight the relevance of analyzing aspects of tumor micro-architecture in the search of prognostic markers and molecular targets for tonsillar cancer. The abundance of intra-tumoral CD8+ T-cells can be considered a positive predictive marker for tonsillar cancer, while the significance of PD-L1 expression by intra-tumoral CD8+ T-cells warrants further evaluation. Location-based differences in CD8+ and CD11c+ cells suggest an immune cell-altering effect on the tumor microenvironment, and grant new insight into which cells that can be targeted by novel therapeutic agents.

High Levels of FGF11 Correlate with Poor Survival in Patients with Human Papillomavirus (HPV)-Positive Oropharyngeal Squamous Cell Carcinoma.

Human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) is associated with a favourable prognosis. It has therefore been suggested that treatment should be individualized and separated by HPV status. However, additional prognostic markers are still needed before treatment can be individualized for this patient group. For this purpose, all patients diagnosed with HPV and p16-positive OPSCC in Stockholm 2000-2009, identified as having a partial/nonresponse to treatment and having viable tumour cells in their neck specimen with material available were categorized as cases. These were matched to controls (complete responders), and the differences in the gene expression were analysed. Two separate verification cohorts were identified including patients with HPV- and p16-positive OPSCC, and the data from the case-control study were verified by qPCR and immunohistochemistry (IHC) in the respective cohorts. A separation of gene expression in correlation with survival was observed in the case-control study, and FGF11 expression was identified as significantly differently expressed between the two groups. The prognostic role of FGF11 was validated in the two cohorts on the RNA and protein levels, respectively. Taken together, our findings suggest that FGF11 may indicate a poor prognosis in HPV-positive OPSCC and may serve as a prognostic biomarker.

Targeted Therapy of HPV Positive and Negative Tonsillar Squamous Cell Carcinoma Cell Lines Reveals Synergy between CDK4/6, PI3K and Sometimes FGFR Inhibitors, but Rarely between PARP and WEE1 Inhibitors.

Human papillomavirus positive (HPV+) tonsillar and base of tongue squamous cell carcinoma (TSCC/BOTSCC) have a favorable outcome, but upon relapse, survival is poor and new therapeutical options are needed. Recently, we found synergistic effects by combining the food and drug administration approved (FDA) phosphoinositide 3-kinase (PI3K) and fibroblast-growth-factor-receptor (FGFR) inhibitors BYL719 and JNJ-42756493 on TSCC cell lines. Here this approach was extended and Cyclin-Dependent-Kinase-4/6 (CDK4/6) and Poly-ADP-ribose-polymerase (PARP) and WEE1 inhibitors PD-0332991, and MK-1775 respectively were also examined. HPV+ CU-OP-2, -3, -20, and HPV- CU-OP-17 TSCC cell lines were treated with either BYL719 and JNJ-42756493, PD-0332991 BMN-673 and MK-1775 alone or in different combinations. Viability, proliferation, and cytotoxicity were followed by WST-1 assays and the IncuCyte S3 Live® Cell Analysis System. All inhibitors presented dose-dependent inhibitory effects on tested TSCC lines. Synergy was frequently obtained when combining CDK4/6 with PI3K inhibitors, but only sometimes or rarely when combining CDK4/6 with FGFR inhibitors or PARP with WEE1 inhibitors. To conclude, using CDK4/6 with PI3K or FGFR inhibitors, especially PD-0332991 with BYL719 presented synergy and enhanced the decrease of viability considerably, while although dose dependent responses were obtained with PARP and WEE1 inhibitors (BMN-673 and MK-1775 resp.), synergy was rarely disclosed.

Spontaneous regression of advanced-stage oropharyngeal squamous cell carcinoma.

BACKGROUND: Spontaneous regression is defined as the partial or complete disappearance of a malignant tumour proven by microscopic examination in the absence of any substantial treatment. This paper presents the case of an older woman whose advanced-stage tonsillar squamous cell carcinoma was noted to have spontaneously regressed at seven months. CASE REPORT: A 66-year-old woman presented with a 4-month history of dysphagia and odynophagia in September 2020. An exophytic tumour was seen on the right tonsil; this was diagnosed radiologically and histologically as a squamous cell carcinoma of the tonsils, with tumour-node-metastasis staging of T4aN0M0. The patient received best supportive care. Seven months later, the oropharyngeal lesion had disappeared, with no treatment. Subsequent computed tomography imaging showed radiological resolution of the previously noted right-sided oropharyngeal lesion. CONCLUSION: Several mechanisms of spontaneous regression are discussed. Further studies should review this case in conjunction with other reports of spontaneous tumour regressions, to elucidate underlying mechanisms.

Single-cell analysis of myeloid cells in HPV+ tonsillar cancer.

The incidence of human papillomavirus-positive (HPV+) tonsillar cancer has been sharply rising during the last decades. Myeloid cells represent an appropriate therapeutic target due to their proximity to virus-infected tumor cells, and their ability to orchestrate antigen-specific immunity, within the tonsil. However, the interrelationship of steady-state and inflammatory myeloid cell subsets, and their impact on patient survival remains unexplored. Here, we used single-cell RNA-sequencing to map the myeloid compartment in HPV+ tonsillar cancer. We observed an expansion of the myeloid compartment in HPV+ tonsillar cancer, accompanied by interferon-induced cellular responses both in dendritic cells (DCs) and monocyte-macrophages. Our analysis unveiled the existence of four DC lineages, two macrophage polarization processes, and their sequential maturation profiles. Within the DC lineages, we described a balance shift in the frequency of progenitor and mature cDC favoring the cDC1 lineage in detriment of cDC2s. Furthermore, we observed that all DC lineages apart from DC5s matured into a common activated DC transcriptional program involving upregulation of interferon-inducible genes. In turn, the monocyte-macrophage lineage was subjected to early monocyte polarization events, which give rise to either interferon-activated or CXCL-producing macrophages, the latter enriched in advanced tumor stages. We validated the existence of most of the single-cell RNA-seq clusters using 26-plex flow cytometry, and described a positive impact of cDC1 and interferon-activated DCs and macrophages on patient survival using gene signature scoring. The current study contributes to the understanding of myeloid ontogeny and dynamics in HPV-driven tonsillar cancer, and highlights myeloid biomarkers that can be used to assess patient prognosis.

Targeted Therapy with PI3K, PARP, and WEE1 Inhibitors and Radiotherapy in HPV Positive and Negative Tonsillar Squamous Cell Carcinoma Cell Lines Reveals Synergy while Effects with APR-246 Are Limited.

Human papillomavirus positive (HPV+) tonsillar and base of tongue cancer (TSCC/BOTSCC) is rising in incidence, but chemoradiotherapy is not curative for all. Therefore, targeted therapy with PI3K (BYL719), PARP (BMN-673), and WEE1 (MK-1775) inhibitors alone or combined was pursued with or without 10 Gy and their effects were analyzed by viability, proliferation, and cytotoxicity assays on the TSCC/BOTSCC cell lines HPV+ UPCI-SCC-154 and HPV- UT-SCC-60A. Effective single drug/10 Gy combinations were validated on additional TSCC lines. Finally, APR-246 was assessed on several TSCC/BOTSCC cell lines. BYL719, BMN-673, and MK-1775 treatments induced dose dependent responses in HPV+ UPCI-SCC-154 and HPV- UT-SCC-60A and when combined with 10 Gy, synergistic effects were disclosed, as was also the case upon validation. Using BYL719/BMN-673, BYL719/MK-1775, or BMN-673/MK-1775 combinations on HPV+ UPCI-SCC-154 and HPV- UT-SCC-60A also induced synergy compared to single drug administrations, but adding 10 Gy to these synergistic drug combinations had no further major effects. Low APR-246 concentrations had limited usefulness. To conclude, synergistic effects were disclosed when complementing single BYL719 BMN-673 and MK-1775 administrations with 10 Gy or when combining the inhibitors, while adding 10 Gy to the latter did not further enhance their already additive/synergistic effects. APR-246 was suboptimal in the present context.

Tonsillar Cancer with High CD8+ T-Cell Infiltration Features Increased Levels of Dendritic Cells and Transcriptional Regulation Associated with an Inflamed Tumor Microenvironment.

Human papillomavirus (HPV) is the main causal agent of tonsillar cancer (TC) and HPV+ TC has a favorable prognosis compared to HPV- disease. In this study, we examined aspects of the tumor microenvironment of TC, focusing on T-cells, dendritic cells (DC), and macrophages. Fresh biopsies of TC and the contralateral healthy tonsil (HT) were obtained from 20 patients, analyzed by multiparameter flow cytometry, and assessed against a detailed HPV-status. Additionally, RNA-sequencing data from 38 TC samples available in the public database, The Cancer Genome Atlas (TCGA), were explored, focusing on the same leukocyte populations. HPV+ TC featured increased levels of CD8+ T-cells and antigen-presenting cells (cf. HPV- TC and HT, respectively). In HPV+ TC, CD8+ T-cell frequencies correlated to DC levels independently of tumor stage, HPV 16 copy number, and E7 oncogene expression as well as frequencies of other leukocytes. Similarly, RNA sequencing data were explored by dividing the HPV+ TCs according to predefined CD8+ T-cell scores in silico. Higher levels of genes expressed by antigen-presenting cells and effector T-cells, such as immune checkpoints and cytokines, were detected in the CD8HIGH HPV+ TC samples (cf. CD8LOW HPV+ TC). In conclusion, CD8HIGH HPV+ TC displays a unique inflammatory profile associated with increased effector T-cell functions and the presence of antigen-presenting cells in the tumor microenvironment. Further studies are warranted to assess if this information can be used on an individual basis to aid in prognosis and treatment decisions.

Long-Term Survival and Recurrence in Oropharyngeal Squamous Cell Carcinoma in Relation to Subsites, HPV, and p16-Status.

Long-term survival data in relation to sub-sites, human papillomavirus (HPV), and p16INK4a (p16) for patients with oropharyngeal squamous cell carcinoma (OPSCC) is still sparse. Furthermore, reports have indicated atypical and late recurrences for patients with HPV and p16 positive OPSCC. Therefore, we assessed long-term survival and recurrence in relation to oropharyngeal subsite and HPV/p16 status. A total of 529 patients with OPSCC, diagnosed in the period 2000-2010, with known HPVDNA and p16-status, were included. HPV/p16 status and sub-sites were correlated to disease-free and overall survival (DFS and OS respectively). The overexpression of p16 (p16+) is associated with significantly better long-term OS and DFS in tonsillar and base of tongue carcinomas (TSCC/BOTSCC), but not in patients with other OPSCC. Patients with HPVDNA+/p16+ TSCC/BOTSCC presented better OS and DFS compared to those with HPVDNA-/p16- tumors, while those with HPVDNA-/p16+ cancer had an intermediate survival. Late recurrences were rare, and significantly more frequent in patients with p16- tumors, while the prognosis after relapse was poor independent of HPVDNA+/-/p16+/- status. In conclusion, patients with p16+ OPSCC do not have more late recurrences than p16-, and a clear prognostic value of p16+ was only observed in TSCC/BOTSCC. Finally, the combination of HPVDNA and p16 provided superior prognostic information compared to p16 alone in TSCC/BOTSCC.

Targeted Therapy With PI3K and FGFR Inhibitors on Human Papillomavirus Positive and Negative Tonsillar and Base of Tongue Cancer Lines With and Without Corresponding Mutations.

OBJECTIVES: Human papillomavirus positive (HPV+) tonsillar and base of tongue squamous cell carcinoma (TSCC/BOTSCC), the major subsites of oropharyngeal squamous cell carcinoma (OPSCC) have favorable outcome, but upon relapse, outcome is poor and new therapies needed. Since, phosphatidyl-inositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA) and fibroblast-growth-factor-receptor-3 (FGFR3) mutations often occur in such tumors, here, we tested targeted therapy directed to such genes in TSCC/BOTSCC cell lines. We also combined the two types of inhibitors with each other, and cisplatin or docetaxel that are used clinically. METHODS: The HPV+ CU-OP-2, -3, -20, UPCI-SCC-154, and HPV- CU-OP-17 and UT-SCC-60A cell lines were first tested for common PIK3CA/FGFR3 mutations by competitive-allele-specific TaqMan-PCR. They were then treated with the food and drug administration (FDA) approved drugs, alpelisib (BYL719) and erdafitinib (JNJ-42756493) alone and in combination with cisplatin or docetaxel. Viability, proliferation, apoptosis and cytotoxicity responses were thereafter followed by WST-1 assays and the IncuCyte S3 Live® Cell Analysis System. RESULTS: HPV+ CU-OP-2 had a pS249C-FGFR3, and like CU-OP-20, a pE545K-PIK3CA mutation, while no other lines had such mutations. Irrespectively, dose dependent responses to all PI3K/FGFR inhibitors were obtained, and upon combining the inhibitors, positive effects were observed. Cisplatin and docetaxel also induced dose dependent responses, and upon combination with the inhibitors, both positive and neutral effects were found. CONCLUSIONS: The data suggest that FDA approved drugs alpelisib and erdafitinib efficiently inhibit TSCC/BOTSCC cell line growth, especially when combined irrespective of presence of corresponding mutations and should be further explored, for use upon recurrent disease.

History of tonsillectomy and risk of oropharyngeal cancer.

OBJECTIVE: To investigate whether palatine tonsillectomy in youth influences the risk of oropharyngeal cancers (OPC) by assessing the association between history of tonsillectomy and risk of tonsillar, base of tongue (BOT) cancer, and other head and neck cancers (HNC). MATERIALS AND METHODS: RACKAM was a case-case study comparing frequency of tonsillectomy history in individuals diagnosed with HNC from 2013 to 2018 in 15 centers across France. History of tonsillectomy was defined using combined assessment of patients' recollections and surgeons' visualizations of tonsil area. OPC subsite-specific odds ratios (OR) of tonsillectomy were calculated using multinomial logistic regression with non-oropharyngeal HNC as reference. RESULTS: 1045 patients were included in the study. Frequency of tonsillectomy was 19.5% in patients with tonsillar cancer (N = 85), 49.3% in BOT (N = 76), 33.8% in other oropharyngeal cancers (N = 202) and 38.0% in non-oropharyngeal HNC (N = 682). History of tonsillectomy was inversely associated with tonsillar cancer (adjusted OR 0.4; 95% CI 0.2-0.8), and positively associated with BOT cancer (adjusted OR 1.8; 95% CI 1.1-3.1), but was not associated with all OPC combined (adjusted OR 1.1; 95% CI 0.8-1.4). Sensitivity analyses considering only patients' or surgeons' assessments of tonsillectomy provided comparable results. CONCLUSION: We confirm the long-term protective effect of tonsillectomy performed in youth on future risk of tonsillar cancer, and our study is the second to report a concurrent increased risk of BOT cancer. Our data suggest that tonsillectomy in youth shifts the site of the first diagnosed oropharyngeal tumor and has a limited impact on overall risk of OPC.

Transoral robotic submandibular gland transposition to reconstruct radical tonsillar resection defects.

Significant dysphagia, pain, and risk of bleeding occur after transoral robotic surgery (TORS) radical tonsillectomy. We present a novel surgical technique utilizing robotically assisted submandibular gland transposition (SMGT) to reconstruct the radical tonsillar defect. A 48-year-old male with p16+ tonsillar squamous cell carcinoma underwent deep TORS radical tonsillectomy, contralateral tonsillectomy, ipsilateral neck dissection, and TORS-assisted reconstruction of the radical defect with ipsilateral SMGT. Postoperatively, the patient experienced minimal pain and was discharged on postoperative day (POD) 3 tolerating a soft diet. There were no episodes of postoperative bleeding. This procedure was performed in five other cases as well. Transoral robotic SMGT can be used successfully to repair deep TORS radical tonsillectomy defects and may theoretically reduce dysphagia, pain, and the risk of hemorrhage.

Whole-Exome Sequencing of HPV Positive Tonsillar and Base of Tongue Squamous Cell Carcinomas Reveals a Global Mutational Pattern along with Relapse-Specific Somatic Variants.

To identify predictive/targetable markers in human papillomavirus positive (HPV+) tonsillar and base of tongue cancer (TSCC/BOTSCC), whole-exome sequencing (WES) of tumours of patients with/without recurrence was performed. Forty primary tumours and adjacent normal tissue were separated by micro-dissection from formalin-fixed paraffin-embedded tissue from patients treated with curative intent 2000-2014 at Karolinska University Hospital. Successful sequencing was obtained in primary tumours of 18 patients without and primaries of 17 with local or distant recurrence, as well as in 10 corresponding recurrences (i.e., five local relapses and five distant metastases) from these 17 patients. One variant-a high-impact deletion in the CDC27 gene-was observed only in primaries of 5/17 patients that had a recurrence after full treatment but in none of those without recurrence. In addition, 3 variants and 26 mutated genes, including CDC27, BCLAF1 and AQP7, were present in at least 30% of all primary tumours independent of prognosis. To conclude, a CDC27 deletion was specific and found in ~30% of samples from patients with a local relapse/distant metastasis and could, therefore, potentially be a prospective marker to predict prognosis. Commonly mutated genes, such as BCLAF1, should be further studied in the context of targeted therapy.

Human papilloma virus (HPV) prevalence upon HPV vaccination in Swedish youth: a review based on our findings 2008-2018, and perspectives on cancer prevention.

PURPOSE: Three human papillomavirus (HPV) vaccines are available against up to nine HPV types. In Sweden, from 2012, Gardasil was offered to 10-12 year old girls through the school-based vaccination program, and as catchup vaccination for women up to 26 years. To obtain a baseline, and follow HPV vaccination effects, during 2008-2018, cervical and oral HPV prevalence were followed at a youth clinic in Stockholm, and in 2013 for comparison oral HPV prevalence was examined in high-school youth in a middle-sized county in Sweden. METHODS: In this review, we discuss all our data with cervical and oral mouthwash samples that were collected and tested for 24-27 HPV types by a bead-based multiplex assay from 2008. RESULTS: Compared with 2008-2011, with ~ 35% HPV16 and > 60% high risk (HR) HPV cervical prevalence at the youth clinic, a decrease of vaccine HPV types was observed between 2013 and 2018, with e.g., HPV16 falling to 5% in catchup vaccinated women and 15-18% in nonvaccinated women. Most common cervical HR-HPV types were HPV39, 51, 52, 56, and 59 together accounting for ~ 10% of cervical cancer, and where only HPV52 is included in Gardasil-9. At baseline 2009-2011, oral HPV prevalence was ~ 10% in unvaccinated youth at the youth clinic, but after 2013 it dropped to < 2% at the youth clinic and high schools. CONCLUSION: To conclude, Gardasil HPV types have decreased, but it is still important to follow remaining HR-HPV types and cancer development, since there is an ongoing increase in the incidence of HPV-associated tonsillar and base of tongue cancer, and cervical cancer in Sweden.

A global epidemic increase of an HPV-induced tonsil and tongue base cancer - potential benefit from a pan-gender use of HPV vaccine.

In 2007, human papillomavirus (HPV) type 16 was finally recognized as a risk factor, besides smoking and alcohol, for oropharyngeal squamous cell carcinoma (OPSCC), including tonsillar squamous cell carcinoma (TSCC), by the International Agency for Research against Cancer. Just before, in 2006, the Food and Drug Administration had approved Gardasil, the first vaccine against HPV16, 18, 6 and 11, for preventive vaccination women against cervical cancer. Concurrently, some Western countries, where smoking was decreasing, disclosed an epidemic increase in the incidence of OPSCC, especially of TSCC and base of tongue cancer (BOTSCC), together accounting for 80-90% of all OPSCCs, and mainly affecting men. The epidemic was later revealed to be due to a rise in HPV-positive cases, and scientists in the field suggested HPV vaccination also of boys. Globally, there are roughly 96 000 incident OPSCC cases/year of which 20-24% are caused by HPV, thereby accounting for around 22 000 OPSCC cases annually. Of these cases, 80-90% are due to HPV16 infection and would be prevented with the presently registered HPV vaccines. In Western countries, such as Sweden (with almost 400 TSCC and BOTSCC cases per year) and the United States, HPV prevalence in OPSCC is higher and around 70%. HPV vaccination of girls has been initiated in many countries, and the vaccines have been efficient and their side effects limited. HPV vaccination of boys has, however, been the exception, but should definitely not be delayed any further. It would benefit both girls and boys directly, and result in better and more robust herd immunity. Today, we have the possibility to eliminate several high-risk HPV types in the younger generations and avoid more than 600 000 cancer cases annually worldwide, and this possibility should be embraced by offering global pan-gender HPV vaccination.

In vitro antitumor effects of FGFR and PI3K inhibitors on human papillomavirus positive and negative tonsillar and base of tongue cancer cell lines.

Human papillomavirus positive (HPV+) tonsillar and base of tongue squamous cell carcinoma (TSCC/BOTSCC) have better outcomes than corresponding HPV- negative (HPV-) cancer cases. Our previous study demonstrated that fibroblast growth factor receptor 3 (FGFR3) and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit a (PIK3CA) are often mutated in HPV+ cancer. To investigate whether targeted therapy is an option for TSCC/BOTSCC, two HPV+ and one HPV- TSCC/BOTSCC cell lines were tested for their sensitivity towards FGFR and PI3K inhibitors. The HPV+ cell lines UM-SCC-47 and UPCI-SCC-154, and the HPV- cell line UT-SSC-60A were tested by competitive allele-specific TaqMan-PCR for presence/absence of frequently occurring FGFR3 and PIK3CA mutations. All cells were then treated with FGFR inhibitor AZD4547 and PI3K inhibitors BEZ235 and BKM120 alone, or with AZD4547 and BEZ235 in combination. Viability was analyzed using a WST-1 assay, cytotoxicity tested by a CellTox Green cytotoxicity assay, apoptosis analyzed by a Caspase Glo 3/7 assay and proliferation examined with the xCELLigence system. HPV+ UM-SCC-47 and UPCI-SCC-154 cells, and HPV- UT-SSC-60A cells, did not exhibit any common FGFR3 or PIK3CA mutations, but were all sensitive to FGFR inhibitor AZD4547 and PI3K inhibitors BEZ235 and BKM120. Notably, HPV+ UPCI-SCC-154 cells were more sensitive than the other two cell lines. Furthermore, when AZD4547 and BEZ235 treatment was combined in HPV+ UPCI-SCC-154 and HPV- UT-SSC-60A cells, potentiated combination effects were observed. HPV+ UM-SCC-47 and UPCI-SCC-154 cells, and HPV- UT-SSC-60A cells had no common FGFR3 or PIK3CA mutations, but were sensitive to FGFR inhibitor AZD4547, and PI3K inhibitors BEZ235 and BKM120. Furthermore, the latter two cell lines were particularly sensitive to combinations of AZD4547 and BEZ235.

Differences in gene expression between high-grade dysplasia and invasive HPV+ and HPV- tonsillar and base of tongue cancer.

BACKGROUND: Human papillomavirus (HPV) is a causative agent for tonsillar and base of tongue squamous cell carcinoma (TSCC/BOTSCC), as well as for cervical cancer. Premalignant stages in cervical cancer have been studied extensively, while little is known about premalignant stages in TSCC/BOTSCC and the role of HPV. Here we analyzed differences in gene and protein expression between high-grade dysplasia and invasive cancer in both HPV-positive (HPV+ ) and HPV-negative (HPV- ) TSCC/BOTSCC. METHODS: High-grade dysplasia and invasive carcinoma were laser microdissected from HPV+ and HPV- TSCC/BOTSCC tumor sections. Differential gene expression was studied utilizing nanoString RNA-panels and genes of interest were validated on the protein level by immunohistochemistry. RESULTS: Forty genes in the HPV+ tumors showed significantly different expression between high-grade dysplasia and invasive cancer and 33 genes in the HPV- tumors. Five out of the nine most significant pathways showed similar increased activity in invasive cancer as compared to high-grade dysplasia in both HPV+ and HPV- tumors. Lastly, significant differences in protein expression was confirmed for SPARC, psoriasin, type I collagen and galectin-1 in both HPV+ and HPV- tumors. CONCLUSIONS: This is to our knowledge the first study disclosing differences and similarities in gene expression between dysplastic and invasive HPV+ and HPV- TSCC/BOTSCC.