Prognostic factors in oral cancer surgery - results from a UK tertiary centre.

INTRODUCTION: Oral cancer surgery is complicated by the diverse nature of clinical and histopathological presentations that occur. Current National guidance recognises the significant role that surgical margin status plays in the overall survival of patients. Many other histopathological factors influence patient survival, the importance of which varies between the literature. MATERIALS AND METHODS: In this prospective longitudinal study, all patients diagnosed with squamous cell carcinoma who had primary surgical treatment under general anaesthesia were included. Surgery was performed by one surgical team within this tertiary referral centre. Patients were followed up for a maximum of 7 years following their surgery. RESULTS: A total of 250 patients were included from 2015 to 2022. Patients were 61.44 years old (SD 13.23) at diagnosis, and 56.4% were male (n = 141). Pathology was mainly pT1 (39.1%) and the most common sites were the border of tongue (31.2%) and floor of mouth (18.8%). 43.4% of patients had clear surgical margins, with overall survival being significantly associated with margin status (p = 0.0079). Extra-capsular spread was significantly associated with higher risk of death from metastatic head and neck cancer (p = 0.014), whereas presence of high-grade dysplasia at surgical margins and depth of invasion of tumour were not. CONCLUSION: This study has reinforced the importance of surgical margin clearance and as such the development of intra-operative techniques to ensure this is imperative. The significance of extra-capsular spread in survival has also been demonstrated. Discussion regarding the current deficiency in accurate pre-operative diagnostic methods for extra capsular spread is covered.

Necrotizing Fasciitis in the Immediate Post-Operative Period Following Resection and Free Flap Reconstruction for Oral Cancer.

Necrotising fasciitis (NF) is a rapidly spreading bacterial infection of the fascial planes and can be fatal if is not treated urgently. Here, we present the case of a 65-year-old female, with oral squamous cell carcinoma, treated surgically with curative intent. On the second post-operative day from a mandibulectomy, selective neck dissection and reconstruction with a fibula free flap, she developed rapidly progressing NF, at the surgical site.

It's not all about the thyroid! Extrinsic and unusual pathology affecting the thyroid gland: A pictorial review.

There is a growing epidemic of thyroid nodules, commonly detected as incidental imaging findings. The vast majority of nodules are benign and of primary thyroidal origin. However, a multitude of non-native, extrinsic or systemic conditions may affect the thyroid and mimic primary thyroid nodules. Contributing factors may include the glands' location in an anatomically dense area, rich vascular and lymphatic network, and embryological origin. In this article we describe a variety of extrinsic and unusual pathology which can affect the thyroid gland. Conditions are classified into benign congenital, benign acquired, cancers which secondarily involve the thyroid gland and unusual cancers arising from within the gland itself. The imaging findings, primarily on high-resolution ultrasound, are reviewed and illustrated with examples. Where possible, imaging features which suggest a specific pathological category or entity are highlighted. It is important that those performing ultrasound examination of the thyroid gland are aware that thyroid nodules may not exclusively represent pathology native or intrinsic to the gland itself.

Homozygosity for the pathogenic RET hotspot variant p.Cys634Trp: A consanguineous family with MEN2A.

Multiple endocrine neoplasia type 2 (MEN2) is a dominantly inherited condition with defined correlations between the genetic variant and clinical presentations. The location of pathogenic variants in the RET gene is a significant determinant of disease presentation and is associated with variable gene activation. Heterozygous pathogenic variants in codon 634 result in earlier onset of medullary thyroid carcinoma and higher incidence of phaeochromocytoma. Here we describe a consanguineous family with MEN2A that includes two children homozygous for the established pathogenic variant p. Cys634Trp. Both parents and a sibling were confirmed to being heterozygotes. Previous reports of biallelic or multiple RET variants have been limited to weakly activating variants. We present the first report of individuals homozygous for the highly activating RET p. Cys634Trp pathogenic variant and discuss disease severity and onset in this rare occurrence.

The genomic road to invasion-examining the similarities and differences in the genomes of associated oral pre-cancer and cancer samples.

BACKGROUND: It is frequently assumed that pre-invasive lesions are simpler precursors of cancer and will contain a limited subset of the genomic changes seen in their associated invasive disease. Driver mutations are thought to occur early, but it is not known how many of these are present in pre-invasive lesions. These assumptions need to be tested with the increasing focus on both personalised cancer treatments and early detection methodologies. METHODS: We examined genomic copy number changes in 256 pre-invasive and invasive samples from 69 oral cancer patients. Forty-eight samples from 16 patients were further examined using exome sequencing. RESULTS: Evidence of a shared ancestor of both dysplasia and carcinoma was seen in all but one patient. One-third of dysplasias showed independent copy number events. The remainder had a copy number pattern that was similar to or simpler than that of the carcinoma. All dysplasias examined contained somatic mutations absent in the related carcinoma. Previously observed copy number changes and TP53 mutations were very frequently observed, and almost always shared between dysplasia and carcinoma. Other gene changes were more sporadic. Pathway analysis confirmed that each patient's disease developed in a different way. Examining the numbers of shared mutations and the rate of accumulation of mutations showed evidence that all samples contain a population of sub-clones, with little evidence of selective advantage of a subset of these. CONCLUSIONS: These findings suggest that most of the genomic changes driving oral cancer occur in the pre-cancerous state by way of gradual random accumulation rather than a dramatic single event.

Elucidating drivers of oral epithelial dysplasia formation and malignant transformation to cancer using RNAseq.

Oral squamous cell carcinoma (OSCC) is a prevalent cancer with poor prognosis. Most OSCC progresses via a non-malignant stage called dysplasia. Effective treatment of dysplasia prior to potential malignant transformation is an unmet clinical need. To identify markers of early disease, we performed RNA sequencing of 19 matched HPV negative patient trios: normal oral mucosa, dysplasia and associated OSCC. We performed differential gene expression, principal component and correlated gene network analysis using these data. We found differences in the immune cell signatures present at different disease stages and were able to distinguish early events in pathogenesis, such as upregulation of many HOX genes, from later events, such as down-regulation of adherens junctions. We herein highlight novel coding and non-coding candidates for involvement in oral dysplasia development and malignant transformation, and speculate on how our findings may guide further translational research into the treatment of oral dysplasia.

The clonal relationships between pre-cancer and cancer revealed by ultra-deep sequencing.

The study of the relationships between pre-cancer and cancer and identification of early driver mutations is becoming increasingly important as the value of molecular markers of early disease and personalised drug targets is recognized, especially now the extent of clonal heterogeneity in fully invasive disease is being realized. It has been assumed that pre-cancerous lesions exhibit a fairly passive progression to invasive disease; the degree to which they, too, are heterogeneous is unknown. We performed ultra-deep sequencing of thousands of selected mutations, together with copy number analysis, from multiple, matched pre-invasive lesions, primary tumours and metastases from five patients with oral cancer, some with multiple primary tumours presenting either synchronously or metachronously, totalling 75 samples. This allowed the clonal relationships between the samples to be observed for each patient. We expose for the first time the unexpected variety and complexity of the relationships between this group of oral dysplasias and their associated carcinomas and, ultimately, the diversity of processes by which tumours are initiated, spread and metastasize. Instead of a series of genomic precursors of their adjacent invasive disease, we have shown dysplasia to be a distinct dynamic entity, refuting the belief that pre-cancer and invasive tumours with a close spatial relationship always have linearly related genomes. We show that oral pre-cancer exhibits considerable subclonal heterogeneity in its own right, that mutational changes in pre-cancer do not predict the onset of invasion, and that the genomic pathway to invasion is neither unified nor predictable. Sequence data from this study have been deposited in the European Nucleotide Archive, Accession No. PRJEB6588.

A novel genomic signature reclassifies an oral cancer subtype.

Verrucous carcinoma of the oral cavity (OVC) is considered a subtype of classical oral squamous cell carcinoma (OSCC). Diagnosis is problematic, and additional biomarkers are needed to better stratify patients. To investigate their molecular signature, we performed low-coverage copy number (CN) sequencing on 57 OVC and exome and RNA sequencing on a subset of these and compared the data to the same OSCC parameters. CN results showed that OVC lacked any of the classical OSCC patterns such as gain of 3q and loss of 3p and demonstrated considerably fewer genomic rearrangements compared to the OSCC cohort. OVC and OSCC samples could be clearly differentiated. Exome sequencing showed that OVC samples lacked mutations in genes commonly associated with OSCC (TP53, NOTCH1, NOTCH2, CDKN2A and FAT1). RNA sequencing identified genes that were differentially expressed between the groups. In silico functional analysis showed that the mutated and differentially expressed genes in OVC samples were involved in cell adhesion and keratinocyte proliferation, while those in the OSCC cohort were enriched for cell death and apoptosis pathways. This is the largest and most detailed genomic and transcriptomic analysis yet performed on this tumour type, which, as an example of non-metastatic cancer, may shed light on the nature of metastases. These three independent investigations consistently show substantial differences between the cohorts. Taken together, they lead to the conclusion that OVC is not a subtype of OSCC, but should be classified as a distinct entity.

A rare case of gingival metastases from papillary thyroid carcinoma.

Metastatic oral malignancy accounts for 1% of all oral cancers. Oral soft tissue involvement is rare and accounts for less than 0.1% of all oral tumours with the attached gingiva being the commonest site affected. We present the first reported case of a papillary thyroid carcinoma (PTC) with sarcomatoid transformation giving rise to gingival metastasis. A 71 year old man with a history of PTC presented with an asymptomatic gingival swelling adjacent to his lower right lateral incisor. Subsequent biopsy of the lesion confirmed PTC metastasis with aggressive sarcomatoid features. We present a clinical photograph of the gingival swelling and the pathology images demonstrating both the papillary and sarcomatoid features of the gingival biopsy. The prognosis of PTC is usually excellent but some histological variants of PTC behave more aggressively. The histology in our case demonstrated solid areas and sarcomatoid transformation and behaved far more aggressively than typical PTC. Sarcomatoid transformation in PTC has not been previously described and indicates a poor prognosis and the need for planning urgent palliation. These lesions can present a diagnostic challenge to both pathologists and clinicians in identifying the lesion as metastatic and locating the primary cancer. This case demonstrates the need for vigilance amongst health professionals when presented with an oral soft tissue mass in patients with a known primary malignancy. This may be the first evidence of disseminated disease and emphasises a low threshold to biopsy oral soft tissue lesions in patients with a history of malignant disease.

Next-generation sequencing analysis for detecting human papillomavirus in oral verrucous carcinoma.

OBJECTIVE: The etiology of oral verrucous carcinoma is unknown, and human papillomavirus 'involvement' remains contentious. The uncertainty can be attributed to varied detection procedures and difficulties in defining 'gold-standard' histologic criteria for diagnosing 'verrucous' lesions. Their paucity also hampers investigation. We aimed to analyze oral verrucous lesions for human papillomavirus (HPV) subtype genomes. STUDY DESIGN: We used next-generation sequencing for the detection of papillomavirus sequences, identifying subtypes and computing viral loads. We identified a total of 78 oral verrucous cases (62 carcinomas and 16 hyperplasias). DNA was extracted from all and sequenced at a coverage between 2.5% and 13%. RESULTS: An HPV-16 sequence was detected in 1 carcinoma and 1 hyperplasia, and an HPV-2 sequence was detected in 1 carcinoma out of the 78 cases, with viral loads of 2.24, 8.16, and 0.33 viral genomes per cell, respectively. CONCLUSIONS: Our results indicate no conclusive human papillomavirus involvement in oral verrucous carcinoma or hyperplasia.

Next-generation sequencing for simultaneous determination of human papillomavirus load, subtype, and associated genomic copy number changes in tumors.

Human papillomavirus (HPV) infection in cases of squamous cell carcinoma of the oropharynx is a powerful predictive and prognostic biomarker. We describe how the use of next-generation sequencing can provide a novel method for the detection of HPV in DNA isolated from formalin-fixed paraffin-embedded tissues. Using this methodology in a cohort of 44 head and neck tumors, we identified the samples that contained HPV sequences, the viral subtype involved, and a direct readout of viral load. Specificity of HPV detection by sequencing compared to traditional detection methods using either PCR or p16 immunohistochemistry was 100%. Sensitivity was 50% when either compared to PCR [confidence interval (CI) = 29% to 71%] or 75% when compared to p16 (CI = 47% to 91%). In addition, we demonstrate the ability of next-generation sequencing to detect other HPV subtypes that would not have been detected by traditional methods, and we demonstrated the ability to apply this method to any tumor and any virus in a panel of eight human cancer cell lines. This methodology also provides a tumor genomic copy number karyogram, and in the samples analyzed here, a lower level of chromosome instability was detected in HPV-positive tumors compared to HPV-negative tumors, as observed in previous studies. Thus, the use of next-generation sequencing for the detection of HPV provides a multiplicity of data with clinical significance in a single test.