Hallmarks of a genomically distinct subclass of head and neck cancer.

Cancer is caused by an accumulation of somatic mutations and copy number alterations (CNAs). Besides mutations, these copy number changes are key characteristics of cancer development. Nonetheless, some tumors show hardly any CNAs, a remarkable phenomenon in oncogenesis. Head and neck squamous cell carcinomas (HNSCCs) arise by either exposure to carcinogens, or infection with the human papillomavirus (HPV). HPV-negative HNSCCs are generally characterized by many CNAs and frequent mutations in CDKN2A, TP53, FAT1, and NOTCH1. Here, we present the hallmarks of the distinct subgroup of HPV-negative HNSCC with no or few CNAs (CNA-quiet) by genetic profiling of 802 oral cavity squamous cell carcinomas (OCSCCs). In total, 73 OCSCC (9.1%) are classified as CNA-quiet and 729 as CNA-other. The CNA-quiet group is characterized by wild-type TP53, frequent CASP8 and HRAS mutations, and a less immunosuppressed tumor immune microenvironment with lower density of regulatory T cells. Patients with CNA-quiet OCSCC are older, more often women, less frequently current smokers, and have a better 5-year overall survival compared to CNA-other OCSCC. This study demonstrates that CNA-quiet OCSCC should be considered as a distinct, clinically relevant subclass. Given the clinical characteristics, the patient group with these tumors will rapidly increase in the aging population.

Immune cell topography of head and neck cancer.

BACKGROUND: Approximately 50% of head and neck squamous cell carcinomas (HNSCC) recur after treatment with curative intent. Immune checkpoint inhibitors are treatment options for recurrent/metastatic HNSCC; however, less than 20% of patients respond. To increase this response rate, it is fundamental to increase our understanding of the spatial tumor immune microenvironment (TIME). METHODS: In total, 53 HNSCC specimens were included. Using a seven-color multiplex immunohistochemistry panel we identified tumor cells, CD163+macrophages, B cells, CD8+T cells, CD4+T helper cells and regulatory T cells (Tregs) in treatment-naive surgical resection specimens (n=29) and biopsies (n=18). To further characterize tumor-infiltrating CD8+T cells, we stained surgical resection specimens (n=12) with a five-color tumor-resident panel including CD103, Ki67, CD8 and pan-cytokeratin. Secretome analysis was performed on matched tumor suspensions (n=11) to measure protein levels. RESULTS: Based on CD8+T cell infiltrates, we identified four different immunotypes: fully infiltrated, stroma-restricted, immune-excluded, and immune-desert. We found higher cytokine levels in fully infiltrated tumors compared with other immunotypes. While the highest immune infiltrates were observed in the invasive margin for all immune cells, CD163+macrophages and Tregs had the highest tendency to infiltrate the tumor center. Within the tumor center, especially B cells stayed at the tumor stroma, whereas CD163+macrophages, followed by T cells, were more often localized within tumor fields. Also, B cells were found further away from other cells and often formed aggregates while T cells and CD163+macrophages tended to be more closely located to each other. Across resection specimens from various anatomical sites within the head and neck, oral cavity tumors exhibited the highest densities of Tregs. Moreover, the distance from B cells and T cells to tumor cells was shortest in oral cavity squamous cell carcinoma (OCSCC), suggesting more interaction between lymphocytes and tumor cells. Also, the fraction of T cells within 10 µm of CD163+macrophages was lowest in OCSCC, indicating fewer myeloid/T-cell suppressive interactions in OCSCC. CONCLUSIONS: We comprehensively described the TIME of HNSCC using a unique data set of resection specimens. We discovered that the composition, as well as the relative localization of immune cells in the TIME, differed in distinct anatomical sites of the head and neck.

Circulating T cell status and molecular imaging may predict clinical benefit of neoadjuvant PD-1 blockade in oral cancer.

BACKGROUND: Addition of neoadjuvant immune checkpoint inhibition to standard-of-care interventions for locally advanced oral cancer could improve clinical outcome. METHODS: In this study, 16 evaluable patients with stage III/IV oral cancer were treated with one dose of 480 mg nivolumab 3 weeks prior to surgery. Primary objectives were safety, feasibility, and suitability of programmed death receptor ligand-1 positron emission tomography (PD-L1 PET) as a biomarker for response. Imaging included 18F-BMS-986192 (PD-L1) PET and 18F-fluorodeoxyglucose (FDG) PET before and after nivolumab treatment. Secondary objectives included clinical and pathological response, and immune profiling of peripheral blood mononuclear cells (PBMCs) for response prediction. Baseline tumor biopsies and postnivolumab resection specimens were evaluated by histopathology. RESULTS: Grade III or higher adverse events were not observed and treatment was not delayed in relation to nivolumab administration and other study procedures. Six patients (38%) had a pathological response, of whom three (19%) had a major (≥90%) pathological response (MPR). Tumor PD-L1 PET uptake (quantified using standard uptake value) was not statistically different in patients with or without MPR (median 5.3 vs 3.4). All major responders showed a significantly postnivolumab decreased signal on FDG PET. PBMC immune phenotyping showed higher levels of CD8+ T cell activation in MPR patients, evidenced by higher baseline expression levels of PD-1, TIGIT, IFNγ and lower levels of PD-L1. CONCLUSION: Together these data support that neoadjuvant treatment of advanced-stage oral cancers with nivolumab was safe and induced an MPR in a promising 19% of patients. Response was associated with decreased FDG PET uptake as well as activation status of peripheral T cell populations.

Architectural dysplasia in surgical margins and the risk of local relapse in oral cancer.

BACKGROUND: A major challenge in the clinical management of oral cavity squamous cell carcinoma is local relapse. Even when surgical margins are tumor-free, local relapses occur frequently, and relapse prediction by histology remains suboptimal. In leukoplakia, an oral potentially malignant disorder, the presence of architectural dysplasia is a critical risk factor for malignant transformation. This study aimed to investigate whether the presence of architectural dysplasia in oral cavity squamous cell carcinoma surgical margins is a risk factor for local relapse. METHODS: Hematoxylin and eosin-stained slides of resection margins from a consecutive cohort of surgically treated patients diagnosed with stage I-IV oral cavity squamous cell carcinoma between 2008 and 2014 were assessed for the presence of architectural dysplasia (N = 311). Five-year local relapse-free survival rates of oral cavity squamous cell carcinoma with architectural dysplasia were compared to those of oral cavity squamous cell carcinoma without architectural dysplasia. RESULTS: In total, 92 of 311 (29.6%) of oral cavity squamous cell carcinoma displayed architectural dysplasia in the margins. The presence of architectural dysplasia was associated with higher patient age, female sex, less pack years, lower cT-stage, and a cohesive tumor growth pattern. In oral cavity squamous cell carcinomas with architectural dysplasia, postoperative (chemo)radiotherapy was less often indicated compared with oral cavity squamous cell carcinoma without architectural dysplasia (19.5% vs. 36.1%, p = 0.009). Five-year local relapse-free survival was significantly lower in oral cavity squamous cell carcinoma with architectural dysplasia than in oral cavity squamous cell carcinoma without architectural dysplasia (83.1% vs. 94.9%, p = 0.017). CONCLUSIONS: Oral cavity squamous cell carcinoma arising in the background of architectural dysplasia displays relatively favorable clinical and histopathological characteristics. Nonetheless, the presence of architectural dysplasia in oral cavity squamous cell carcinoma surgical margins is associated with a higher risk of local relapse, indicating its clinical relevance.

Design and evaluation of an immunology and pathology course that is tailored to today's dentistry students.

Curricular reform provides new opportunities to renovate important pillars of the dentistry curriculum, such as immunology and pathology, with novel approaches that appeal to new generations of students. When redesigning a course that integrates both immunology and pathology at the level that provides dentistry students with sustainable knowledge that is useful for their entire career, several challenges must be met. The objective of the present study was to describe the considerations involved in the design phase of such a new course. First, the course should be compatible with the new view on the incorporation of more active learning and teaching methods. Practically, this means that the course design should contain fewer lectures and more seminars and tutorials, where the students have fewer contact hours and actively engage in using recently acquired knowledge within a contextual background. A mandatory session of team-based learning provides opportunities to apply knowledge in combination with academic reasoning skills, teamwork, and communication. Second, for a 4-week course, choices must be made: students will not become immunologists nor pathologists in such a short period. A governing principle for this course's design is that it should be based on understanding the basic principles of immunology and pathology. The ultimate goal for the students is to make the course immuno-logical and patho-logical, challenging them to reach a next level but clearly without oversimplification. Part of the course design should allow room for students to further study an immunological topic of their own choice, thereby contributing to their immunological curiosity and to their academic development. Third, to make it tailored to a new generation of dentists, examples from the field of dentistry are actively integrated in all aspects of the course. Finally, the era of ChatGPT provides novel opportunities to use generative artificial intelligence (AI) tools in the learning process, but it demands critical thinking of how to use it in a newly designed course. A mid-course evaluation revealed that students acknowledged that immunology and pathology were presented as an integrated course. The final course evaluation endorsed the use of these various educational methods. These methods proved to be appropriate and logical choices for reaching the learning goals of the course.

Genomic Engineering of Oral Keratinocytes to Establish In Vitro Oral Potentially Malignant Disease Models as a Platform for Treatment Investigation.

Precancerous cells in the oral cavity may appear as oral potentially malignant disorders, but they may also present as dysplasia without visual manifestation in tumor-adjacent tissue. As it is currently not possible to prevent the malignant transformation of these oral precancers, new treatments are urgently awaited. Here, we generated precancer culture models using a previously established method for the generation of oral keratinocyte cultures and incorporated CRISPR/Cas9 editing. The generated cell lines were used to investigate the efficacy of a set of small molecule inhibitors. Tumor-adjacent mucosa and oral leukoplakia biopsies were cultured and genetically characterized. Mutations were introduced in CDKN2A and TP53 using CRISPR/Cas9 and combined with the ectopic activation of telomerase to generate cell lines with prolonged proliferation. The method was tested in normal oral keratinocytes and tumor-adjacent biopsies and subsequently applied to a large set of oral leukoplakia biopsies. Finally, a subset of the immortalized cell lines was used to assess the efficacy of a set of small molecule inhibitors. Culturing and genomic engineering was highly efficient for normal and tumor-adjacent oral keratinocytes, but success rates in oral leukoplakia were remarkably low. Knock-out of CDKN2A in combination with either the activation of telomerase or knock-out of TP53 seemed a prerequisite for immortalization. Prolonged culturing was accompanied by additional genetic aberrations in these cultures. The generated cell lines were more sensitive than normal keratinocytes to small molecule inhibitors of previously identified targets. In conclusion, while very effective for normal keratinocytes and tumor-adjacent biopsies, the success rate of oral leukoplakia cell culturing methods was very low. Genomic engineering enabled the prolonged culturing of OL-derived keratinocytes but was associated with acquired genetic changes. Further studies are required to assess to what extent the immortalized cultures faithfully represent characteristics of the cells in vivo.

An Antibody-Drug Conjugate Directed to Tissue Factor Shows Preclinical Antitumor Activity in Head and Neck Cancer as a Single Agent and in Combination with Chemoradiotherapy.

Head and neck squamous cell carcinoma (HNSCC) is a solid tumor type that arises in the squamous epithelial cells lining the mucosal surfaces of the upper aerodigestive tract. Long-term survival of patients with advanced disease stage remains disappointing with current treatment options. We show that tissue factor is abundantly expressed on patient-derived HNSCC cell lines, xenograft tumor material, and tumor biopsies from patients with HNSCC. Tisotumab vedotin (TV) is an antibody-drug conjugate (ADC) directed to tissue factor, a protein expressed in many solid tumors. HNSCC cells and xenograft tumors were efficiently eliminated in vitro and in vivo with TV-monotherapy compared with treatment with a control antibody conjugated to monomethyl auristatin E (MMAE). Antitumor activity of TV was also tested in vivo in combination with chemoradiotherapy, standard of care for patients with advanced stage HNSCC tumors outside the oral cavity. Preclinical studies showed that by adding TV to chemoradiotherapy, survival was markedly improved, and TV, not radiotherapy or chemotherapy, was the main driver of antitumor activity. Interestingly, TV-induced cell death in xenograft tumors showed an influx of macrophages indicative of a potential immune-mediated mode-of-action. In conclusion, on the basis of these preclinical data, TV may be a novel treatment modality for patients suffering from head and neck cancer and is hypothesized to improve efficacy of chemoradiotherapy. SIGNIFICANCE: This work shows preclinical in vitro and in vivo antitumor activity of the antibody-drug conjugate Tisotumab vedotin in head and neck cancer models, and enhanced activity in combination with chemoradiotherapy, supporting further clinical development for this cancer type.

The value of regular follow-up of oral leukoplakia for early detection of malignant transformation.

OBJECTIVES: Evaluate whether regular follow-up of oral leukoplakia (OL) resulted in early detection of malignant transformation (MT). METHOD: Two hundred and twenty-two consecutive patients with OL (147 females, 75 males); median follow-up period of 64 months (range: 12-300). Three groups were distinguished: group A (n = 92) follow-up at the hospital; group B (n = 84) follow-up by their dentist; group C (n = 46) lost to follow-up. RESULTS: OLs in group B compared to group A, were smaller in size (<2 cm; p < 0.001), showed more hyperkeratosis (p < 0.001) and less moderate/severe dysplasia (p < 0.001). MT occurred in 45 (20%) patients: 32 (35%) in group A, five (6%) in group B and eight (17%) in group C. There was no significant difference in clinical tumour size between group A (median: 15 mm, range: 1-40) and group B (median: 10 mm, range: 3-25; p = 0.496). Tumour size was smaller for patients in groups A and B (median: 10 mm, range 1-40) compared to group C (median: 33 mm, range: 3-100; p = 0.003). There was a positive correlation between tumour size and interval between the last visit in all patients (p = 0.022). CONCLUSION: Regular follow-up of OL resulted in early detection of MT. If properly selected, follow-up of OL performed by the dentist seems feasible.

The role of differentiated dysplasia in the prediction of malignant transformation of oral leukoplakia.

OBJECTIVE: Oral leukoplakia is the most common oral potentially malignant disorder. Malignant transformation of oral leukoplakia occurs at an annual rate of 1%-7%. WHO-defined classic epithelial dysplasia is an important predictor of malignant transformation of oral leukoplakia, but we have previously shown in a proof of concept study that prediction improves by incorporation of an architectural pattern of dysplasia, also coined as differentiated dysplasia. We aimed to analyze this finding in a larger cohort of patients. METHOD: For this retrospective study 176 oral leukoplakia patients were included. Biopsies for all patients were assessed for the presence of dysplasia and analyzed for cytokeratin 13 and 17 expression. Moreover, the inter-observer agreement for the diagnosis of differentiated dysplasia was determined. RESULTS: In total, 33 of 176 patients developed oral squamous cell carcinoma during follow-up. Presence of classic epithelial dysplasia increased cancer risk two-fold (HR = 2.18, p = 0.026). Lesions without classic epithelial dysplasia could be further risk-stratified by the presence of differentiated dysplasia (HR = 7.36, p < 0.001). Combined classic epithelial and differentiated dysplasia imparted a seven-fold increased risk of malignant transformation (7.34, p = 0.001). Inter-observer agreement for the diagnosis of dysplasia, including differentiated dysplasia, was moderate (κ = 0.56, p < 0.001). DISCUSSION: This study emphasizes the importance of the recognition of the architectural pattern of differentiated dysplasia as a separate entity for risk prediction of malignant transformation of oral leukoplakia. Presence of any pattern of dysplasia results in accurate prediction of malignant transformation risk of oral leukoplakia.

Elucidating the Genetic Landscape of Oral Leukoplakia to Predict Malignant Transformation.

PURPOSE: Oral leukoplakia is the most common oral potentially malignant disorder with an annual malignant transformation rate of 1% to 5%. Consequently, oral leukoplakia patients have a 30% to 50% lifetime risk to develop oral squamous cell carcinoma. Although risk factors for malignant transformation of oral leukoplakia have been investigated, no definitive risk stratification model has been proposed. Next-generation sequencing can elucidate the genetic landscape of oral leukoplakia, which may be used to predict the risk for malignant transformation. EXPERIMENTAL DESIGN: We investigated a retrospective cohort of 89 oral leukoplakia patients, and analyzed their oral leukoplakia lesions for the presence of genomic copy-number alterations and mutations in genes associated with oral squamous cell carcinoma. RESULTS: In 25 of 89 (28%) patients, oral squamous cell carcinoma developed during follow-up. Seventy-nine of 89 (89%) oral leukoplakias harbored at least one genetic event. Copy-number alterations were present in 61 of 89 (69%) oral leukoplakias, most commonly gains of chromosome regions 8q24 (46%) and 20p11 (20%) and loss of 13q12 (19%). Mutations were present in 59 of 89 (66%) oral leukoplakias, most commonly in TP53 (28%), FAT1 (20%), and NOTCH1 (13%). Genetic data were combined with the presence of dysplasia to generate a prediction model, identifying three groups with a distinct risk for malignant transformation. CONCLUSIONS: We provide an extensive description of genetic alterations in oral leukoplakia and its relation to malignant transformation. On the basis of our data we provide a model for the prediction of malignant transformation of oral leukoplakia using dysplasia and genetic markers.

Associations between clinical and histopathological characteristics in oral leukoplakia.

OBJECTIVES: To identify possible associations between patients' demographics and habits and the clinical aspects and histopathological characteristics of oral leukoplakia (OL) at patients' first visit. METHOD: A total of 140 consecutive patients with OL at a single institute between 1997 and 2019. All biopsies were microscopically examined for classic dysplasia (CD) (WHO definition oral epithelial dysplasia) and differentiated dysplasia (DD) known from differentiated vulvar intraepithelial neoplasia. Clinical characteristics were correlated to histopathological diagnosis and odds ratios (OR) were calculated. RESULTS: A total of 96 females and 44 males, mean age 58 years, were presented. OLs were found mainly on the tongue (41%) and floor of mouth (FOM) (18%). Homogeneous OLs (58%) were associated with smoking, FOM and size <2cm and non-homogeneous OLs (42%) with non-smokers. No dysplasia was present in 40% and any dysplasia (AD) in 60%. Tongue OLs were correlated with AD (OR:6.0) and CD (OR:5.7). FOM OLs were correlated with CD (OR:4.5). DD was correlated with non-homogeneous OLs (OR:2.6). CONCLUSIONS: CD was most frequently observed in tongue and FOM OLs, while DD was associated with non-homogeneous OLs. In this series of patients, there was no consistent reliable association between the clinical and histopathological features and clinical characteristics can therefore not substitute microscopic examination of biopsies.

Oral cancer prediction by noninvasive genetic screening.

Oral squamous cell carcinomas (OSCCs) develop in genetically altered epithelium in the mucosal lining, also coined as fields, which are mostly not visible but occasionally present as white oral leukoplakia (OL) lesions. We developed a noninvasive genetic assay using next-generation sequencing (NGS) on brushed cells to detect the presence of genetically altered fields, including those that are not macroscopically visible. The assay demonstrated high accuracy in OL patients when brush samples were compared with biopsies as gold standard. In a cohort of Fanconi anemia patients, detection of mutations in prospectively collected oral brushes predicted oral cancer also when visible abnormalities were absent. We further provide insight in the molecular landscape of OL with frequent changes of TP53, FAT1 and NOTCH1. NGS analysis of noninvasively collected samples offers a highly accurate method to detect genetically altered fields in the oral cavity, and predicts development of OSCC in high-risk individuals. Noninvasive genetic screening can be employed to screen high-risk populations for cancer and precancer, map the extension of OL lesions beyond what is visible, map the oral cavity for precancerous changes even when visible abnormalities are absent, test accuracy of promising imaging modalities, monitor interventions and determine genetic progression as well as the natural history of the disease in the human patient.

Oral leukoplakia classification and staging system with incorporation of differentiated dysplasia.

OBJECTIVES: A classification and staging system for oral leukoplakia (OL) was introduced to promote uniform reporting. In this system, size and the histopathologic diagnosis are assessed and combined in a staging system. The various stages could be predictive for malignant transformation of OL. Differentiated dysplasia (DD) was recently recognized as an important architectural pattern of dysplasia and is highly associated with malignant transformation (MT) of OL. In the present study, DD was incorporated in the OL-system. The aim of the present study was to test the adapted system on a cohort of patients with OL. PATIENT AND METHODS: The group consisted of 140 patients. The size, absence or presence and degree of classic dysplasia (CD) and DD were incorporated into the OL-system. RESULTS: In 31/140 patients, MT occurred. Size was not statistically significant with MT (p = 0.422). The presence of dysplasia was predictive for MT (p = 0.003), whereby severe CD and DD were highly statistically significant for MT (p = 0.008). Stage IV was statistically significant for MT (p = 0.011). CONCLUSIONS: The present study emphasizes the value of the slightly modified OL-system with incorporation of DD in uniform reporting of OL and the value in predicting MT.

Irradiation Alters the Expression of MUC1, CD44 and Hyaluronan in Oral Mucosal Epithelium.

Purpose: It is well established that cancer cells exploit aberrant synthesis of mucin 1 (MUC1) and hyaluronan (HA) synthesis along with HA's physiological cell surface receptor CD44. However, their role in irradiated oral tissue has not been reported previously. We, therefore, aimed to study MUC1, CD44 and HA immunohistochemically in irradiated oral mucosa and their role in the long-term effects after radiotherapy. Materials and Methods: Oral mucosal biopsies were obtained from healthy subjects as controls and from patients after radiotherapy for head and neck cancer (irradiated group) during dental implant surgery.The presence of MUC1, CD44, and HA in oral mucosa was studied by immunohistochemical methods. The differences in the localization and intensity in the oral epithelium between control and irradiated tissue were analyzed. Results: The staining intensity of MUC1 was confined to the superficial epithelial layer, whereas HA and CD44 were found in the cell membranes in the epithelial basal and intermediate layers of control specimens. In irradiated epithelium, MUC1 staining was distributed throughout all the layers of the oral epithelium, with significant staining in the basal and intermediate layers. Accordingly, HA and CD44 staining extended to involve the superficial cells of the irradiated epithelium. The staining pattern of MUC1 and CD44 showed significant changes in irradiated samples. Conclusions: Our results showed that the staining intensities of MUC1, CD44, and HA were significantly elevated in irradiated tissue compared to controls. MUC1, CD44, and HA are important markers and take part in long-term changes in the oral mucosa after radiotherapy.

Effect of smoking on MUC1 expression in oral epithelial dysplasia, oral cancer, and irradiated oral epithelium.

OBJECTIVES: The aim of this study was to assess the MUC1 expression in the oral epithelium of normal, oral epithelial dysplasia (OED), oral squamous cell carcinoma (OSCC), and irradiated oral epithelium (IROE) and its association with smoking habits in non-smokers and smokers. DESIGN: Oral mucosal biopsies from controls, OED, OSCC, and IROE groups were obtained and categorized based on the smoking history as non-smokers, smoker I (25 pack-years), and smoker II (>25 pack-years). Immunohistochemical staining of MUC1 using human milk fat globule 1 (HMFG 1) antibody was performed, and the MUC1 score was calculated. The relation between MUC1 expression and clinicopathological findings was examined. RESULTS: MUC1 staining of superficial oral epithelial cells with mild MUC1 score was detected in all control samples. The MUC1 staining extended from superficial to basal cell layer of oral epithelium with the increase in MUC1 score from moderate to strong in OED, OSCC, and IROE, and the difference was significant (p < 0.004, p < 0.002 and p < 0.004, respectively) compared to controls. A positive association between smoking and MUC1 score was observed within groups (p < 0.05). CONCLUSION: The depolarization of MUC1 protein expression is associated with smoking habits in OED and OSCC. In the IROE, the radiation causes subcellular and molecular changes, observed as altered MUC1 expression and accelerated by smoking, furthermore, complicating the oral mucosal adaptation and progress to radiation-induced lesions as a delayed effect.

Osteocyte Apoptosis, Bone Marrow Adiposity, and Fibrosis in the Irradiated Human Mandible.

Purpose: To assess the effect of radiation therapy on osteocyte apoptosis, osteocyte death, and bone marrow adipocytes in the human mandible and its contribution to the pathophysiology of radiation damage to the mandibular bone. Methods and Materials: Mandibular cancellous bone biopsies were taken from irradiated patients and nonirradiated controls. Immunohistochemical detection of cleaved caspase-3 was performed to visualize apoptotic osteocytes. The number of apoptotic osteocytes per bone area and per total amount of osteocytes, osteocytes per bone area, and empty lacunae per bone area were counted manually. The percentage fibrotic tissue and adipose tissue per bone marrow area, the percentage bone marrow of total area, and the mean adipocyte diameter (µm) was determined digitally from adjacent Goldner stained sections. Results: Biopsies of 15 irradiated patients (12 men and 3 women) and 7 nonirradiated controls (5 men and 2 women) were assessed. In the study group a significant increase was seen in the number of empty lacunae, the percentage of adipose tissue of bone marrow area, and the adipocyte diameter. There was no significant difference in bone marrow fibrosis nor apoptotic osteocytes between the irradiated group and the controls. Conclusions: Irradiation alone does not seem to induce excessive bone marrow fibrosis. The damage to bone mesenchymal stem cells leads to increased marrow adipogenesis and decreased osteoblastogenic potential. Early osteocyte death resulting in avital persisting bone matrix with severely impaired regenerative potential may contribute to the vulnerability of irradiated bone to infection and necrosis.

The impact of a pathologist's personality on the interobserver variability and diagnostic accuracy of predictive PD-L1 immunohistochemistry in lung cancer.

OBJECTIVES: Programmed death-ligand 1 (PD-L1) is the only approved predictive biomarker for immunotherapy in non-small cell lung cancer (NSCLC). However, predictive PD-L1 immunohistochemistry is subject to interobserver variability. We hypothesized that a pathologist's personality influences the interobserver variability and diagnostic accuracy of PD-L1 immunoscoring. MATERIALS AND METHODS: Seventeen pathologists performed PD-L1 immunoscoring on 50 resected NSCLC tumors in three categories (<1%;1-49%;≥50%). Also, the pathologists completed a certified personality test (NEO-PI-r), assessing five personality traits: neuroticism, extraversion, openness, altruism and conscientiousness. RESULTS: The overall agreement among pathologists for a series of 47 tumors was substantial (kappa = 0.63). Of these, 23/47 (49%) tumors were entirely negative or largely positive, resulting in a kappa value of 0.93. The remaining 24/47 (51%) tumors had a PD-L1 score around the cutoff value, generating a kappa value of 0.32. Pathologists with high scores for conscientiousness (careful, diligent) had the least interobserver variability (r = 0.6, p = 0.009). Also, they showed a trend towards higher sensitivity (74% vs. 68%, p = 0.4), specificity (86% vs. 82%, p = 0.3) and percent agreement (83% vs. 79%, p = 0.3), although not significant. In contrast, pathologists with high scores for neuroticism (sensitive, anxious) had significantly lower specificity (80% vs. 87%, p = 0.03) and percent agreement (78% vs. 85%, p = 0.03). Also, a trend towards high interobserver variability (r = -0.3, p = 0.2) and lower sensitivity (68% vs. 74%, p = 0.3) was observed, although not significant. Pathologists with relatively high scores for conscientiousness scored fewer tumors PD-L1 positive at the ≥ 1% cut-off (r = -0.5, p = 0.03). In contrast, pathologists with relatively high scores for neuroticism score more tumors PD-L1 positive at ≥ 1% (r = 0.6, p = 0.017) and ≥ 50% cut-offs (r = 0.6, p = 0.009). CONCLUSIONS: This study is the first to demonstrate the impact of a pathologist's personality on the interobserver variability and diagnostic accuracy of immunostaining, in the context of PD-L1 in NSCLC. Larger studies are needed for validation of these findings.

Prognostic factors in acinic cell carcinoma of the head and neck: The Amsterdam experience.

INTRODUCTION: The aim was to analyse prognosticators in acinic cell carcinoma (AciCC) in two head and neck referral centers in Amsterdam, the Netherlands. MATERIALS AND METHODS: Eighty- nine cases of AciCC treated between 1979 and 2016 were retrospectively reviewed. Five, - 10 -and 20- year estimates of survival were executed as well as univariate analysis of prognosticators. RESULTS: The majority of AciCC were T1-T2; 89%. Two percent had nodal disease (2%). The most affected organ was the parotid gland (84%) with a female preponderance (67%). Mean age was 52 years with most cases diagnosed in the fourth to sixth decade. The majority of patients received adjuvant radiotherapy. Elective neck dissection (END) in the N0 neck showed no metastases. High grade transformation (HGT) was found in 21% of cases. Median follow up was 101.9 months. Median time to recurrence was 26 months. Nine patients developed distant metastases (DM) of whom 6 had HGT-AciCC. Median survival with DM was 7 months. Five,- ten -and twenty- year estimates were 84%, 81% and 81% for recurrence free survival respectively. Negative clinical features were advanced stage disease and tumour size > 2.6 cm. Negative histological features were a high mitotic rate, HGT, close and involved surgical margins and necrosis. CONCLUSION: AciCC- HGT excluded- of the head and neck has an excellent prognosis and shows acceptable long term results. END can be considered as part of the standard treatment due to the relative high incidence of HGT- AciCC and low accuracy of cytology.

Fewer head and neck cancer diagnoses and faster treatment initiation during COVID-19 in 2020: A nationwide population-based analysis.

BACKGROUND: Inevitably, the emergence of COVID-19 has impacted non-COVID care. Because timely diagnosis and treatment are essential, especially for patients with head and neck cancer (HNC) with fast-growing tumours in a functionally and aesthetically important area, we wished to quantify the impact of the COVID-19 pandemic on HNC care in the Netherlands. MATERIAL AND METHODS: This population-based study covered all, in total 8468, newly diagnosed primary HNC cases in the Netherlands in 2018, 2019 and 2020. We compared incidence, patient and tumour characteristics, primary treatment characteristics, and time-to-treatment in the first COVID-19 year 2020 with corresponding periods in 2018 and 2019 (i.e. pre-COVID). RESULTS: The incidence of HNC was nearly 25% less during the first wave (n = 433) than in 2019 (n = 595) and 2018 (n = 598). In April and May 2020, the incidence of oral cavity and laryngeal carcinomas was significantly lower than in pre-COVID years. There were no shifts in tumour stage or alterations in initial treatment modalities. Regardless of the first treatment modality and specific period, the median number of days between first visit to a HNC centre and start of treatment was significantly shorter during the COVID-19 year (26-28 days) than pre-COVID (31-32 days, p < 0.001). CONCLUSION: The incidence of HNC during the Netherlands' first COVID-19 wave was significantly lower than expected. The expected increase in incidence during the remainder of 2020 was not observed. Despite the overloaded healthcare system, the standard treatment for HNC patients could be delivered within a shorter time interval.