Current management and future challenges in salivary glands cancer.

Salivary gland cancers (SGCs) are rare, accounting for less than 5% of all malignancies of the head and neck region, and are morphologically heterogeneous. The diagnosis is mainly based on histology, with the complementary aid of molecular profiling, which is helpful in recognizing some poorly differentiated, borderline, or atypical lesions. Instrumental imaging defines the diagnosis, representing a remarkable tool in the treatment plan. Ultrasound and magnetic resonance are the most common procedures used to describe the primary tumour. The treatment of SGCs is multimodal and consists of surgery, radiotherapy, and systemic therapy; each treatment plan is, however, featured on the patient and disease's characteristics. On 24 June 2022, in the meeting "Current management and future challenges in salivary gland cancers" many experts in this field discussed the state of the art of SGCs research, the future challenges and developments. After the meeting, the same pool of experts maintained close contact to keep these data further updated in the conference proceedings presented here. This review collects the insights and suggestions that emerged from the discussion during and after the meeting per se.

Lung mesenchymal cells from patients with COVID-19 driven lung fibrosis: Several features with CTD-ILD derived cells but with higher response to fibrogenic signals and might be more pro-inflammatory.

A subset of severe COVID19 patients develop pulmonary fibrosis, but the pathophysiology of this complication is still unclear. We previously described the possibility to isolate lung mesenchymal cells (LMC) by culturing broncho-alveolar lavage (BAL) cells from patients with pulmonary fibrosis or chronic lung allograft dysfunction. Aim of this study was to investigate the possibility to isolate and characterize LMC from BAL of patients that, two months after discharge for severe COVID19, show CT signs of post-COVID19 fibrosis (Post-COVID) and in some cases has been considered transplant indication. Results were compared with those from BAL of patients with collagen tissue disease-associated interstitial fibrosis (CTD-ILD). BAL fluid levels of TGFß, VEGF, TIMP2, RANTES, IL6, IL8, and PAI1 were assessed. LMC were cultured and expanded, phenotyped by flow cytometry, and tested for osteogenic and adipogenic differentiation. Finally, we tested immunomodulatory and proliferative capabilities, collagen I production + /- TGF-beta stimulation. BAL cytokine and growth factor levels were comparable in the two groups. Efficiency of isolation from BAL was 100% in post-COVID compared to 63% in CTD-ILD. LMC from post-COVID were positive for CD105, CD73, CD90, and negative for CD45, CD34, CD19 and HLA-DR as in CTD-ILD samples. Post-COVID LMC displayed higher collagen production with respect to CTD-ILD LMC. Immunomodulatory capacity towards lymphocytes was very low, while Post-COVID LMC significantly upregulated pro-inflammatory cytokine production by healthy PBMCs. Our preliminary data suggest that LMC from post-COVID19 fibrosis patients share several features with CTD-ILD ones but might have a higher response to fibrogenic signals and pro-inflammatory profile.

Effect of Radio-Chemotherapy on PD-L1 Immunohistochemical Expression in Head and Neck Squamous Cell Carcinoma.

BACKGROUND: Programmed death-ligand 1 (PD-L1) checkpoint inhibitors represent a mainstay of therapy in head and neck squamous cell cancer (HNSCC). However, little is known about the influence of combined therapy on PD-L1 expression. The study aims to gather evidence on this topic. METHODS: A systematic search was carried out in electronic databases Pubmed-MEDLINE and Embase to retrieve studies on the comparison of PD-L1 expression before and after conventional therapy. Data were extracted and a quantitative analysis with pooled odds ratios (ORs) was performed when applicable. RESULTS: Of 5688 items, 15 were finally included. Only a minority of studies assessed PD-L1 with the recommended combined positive score (CPS). The results are highly heterogeneous, with some studies reporting an increase in PD-L1 expression and others reporting a decrease. Three studies allowed for quantitative analysis and showed a pooled OR of 0.49 (CI 0.27-0.90). CONCLUSIONS: From the present evidence, a clear conclusion towards an increase or decrease in PD-L1 expression after combined therapy cannot be drawn, but even with few studies available, a trend towards an increase in expression in tumor cells at a cutoff of 1% can be noted in patients undergoing platinum-based therapy. Future studies will provide more robust data on the effect of combined therapy on PD-L1 expression.

Impaired intratumoral natural killer cell function in head and neck carcinoma.

Natural killer (NK) cells are emerging as unique players in the immune response against cancer; however, only limited data are available on tumor infiltrating NK cells in head and neck squamous cell carcinoma (HNSCC), one of the most common cancer. Occurrence of HNSCC is closely related to the immune microenvironment, and immunotherapy is increasingly being applied to this setting. However, the limited success of this type of treatment in this tumor calls for further investigation in the field. Surgical HNSSC specimens of 32 consecutive patients were mechanically and enzymatically dissociated. Tumor cells were separated from infiltrating cells by short centrifugation and infiltrating NK cells were phenotypically and functionally characterized by multiple antibody staining and flow cytometry. Tumor infiltrating NK cells in HNSCC showed a peculiar phenotype predominantly characterized by increased NKG2A and reduced Siglec-7, NKG2D, NKp30 and CD16 expression. This phenotype was associated with a decreased ability to perform antibody-dependent cellular cytotoxicity (ADCC). However, NK, CD4 and CD8 shared an increment of glucocorticoid-induced tumor necrosis factor-related (GITR) costimulatory receptor which could be exploited for immunotherapy with agonistic anti-GITR antibodies combined with checkpoint inhibitors.

Feasibility and safety of extended pleurectomy/decortication for malignant pleural mesothelioma. A single group experience.

Surgery is part of a multimodal therapeutic approach to malignant pleural mesothelioma (MPM) although its real beneficial effect is still controversial. The optimal precise sequence of treatments within the trimodality is unclear, and should be decided upon a multidisciplinary consensus for each individual patient. Here, we analyzed the perioperative data of 19 MPM patients who underwent extended pleurectomy/decortication (EPD) with curative intent. The mean age at diagnosis was 67 years; 11 males and eight females. Ten patients were diagnosed with MPM via medical thoracoscopy (MT), and nine via video-assisted thoracoscopic surgery (VATS). The vast majority of cases harbored epitheliod forms. We compared neoadjuvant chemotherapy (NCT) followed by surgery (11 cases) versus surgery followed by adjuvant chemotherapy (ACT, 8 cases) within a 3-year period. All patients had extended pleurectomy/decortication and none had an extended pneumonectomy. Analysis of survival curves suggested that the short-term outcomes are better with upfront EDP followed by ACT if compared to EDP preceded by NCT. Although limited, the data highlighted the safety and feasibility of EPD, with manageable postoperative complications and no major burden for the patients.

TITF1 Screening in Human Congenital Diaphragmatic Hernia (CDH).

TITF1 (Thyroid Transcription Factor-1) is a homeodomain-containing transcription factor. Previous studies showed that Titf1 null mice are characterized by failure of tracheo-oesophageal separation and impaired lung morphogenesis resulting in Pulmonary Hypoplasia (PH). In this study, we aim to evaluate the role of TITF1 in the pathogenesis of congenital diaphragmatic hernia (CDH) in humans. We investigated TITF1 expression in human trachea and lungs and performed direct mutation analysis in a CDH population. We studied 13 human fetuses at 14 to 24 weeks of gestation. Five µm sections were fixed in paraformaldehyde and incubated with anti-TITF1 primary antibody. Positive staining was visualized by biotinylated secondary antibody. We also performed TITF1 screening on genomic DNA extracted from peripheral blood of 16 patients affected by CDH and different degrees of PH, searching for mutations, insertions, and/or deletions, by sequencing the exonic regions of the gene. Histochemical studies showed positive brown staining of fetal follicular thyroid epithelium, normal fetal trachea, and normal fetal lung bronchial epithelium. Fetal esophageal wall was immunohistochemically negative. Molecular genetic analysis showed complete identity between the sequences obtained and the Wild Type (WT) form of the gene in all cases. No mutation, insertion and/or deletion was detected. Although TITF1 is expressed in the human fetal lung and has been considered to have a role in the pathogenesis of PH in CDH, the results of our study do not support the hypothesis that TITF1 mutations play a key role in the etiopathogenesis of CDH.

Post-acute coronavirus disease 2019 (COVID 19) syndrome: HLH and cholangiopathy in a lung transplant recipient.

Background: Sars-CoV2 can cause severe pneumonia and acute respiratory distress syndrome (ARDS). In COVID-19-associated respiratory failure, lung transplantation might be an option (Bharat A). Case report: A previously healthy 63-year-old man with a nasopharyngeal swab positive for SarsCoV2 and radiological evidence of interstitial lung consolidations developed acute respiratory distress that required intubation and veno-venous extracorporeal membrane oxygenation support (VV ECMO). Because of no recovery of his lung function, he underwent a bilateral lung transplant. ICU stay was complicated by several episodes of bacterial superinfections and an increase of liver function tests (LFTs). Afterward, he faced a progressive clinical worsening associated to severe anemia, further rise of indices of cholestasis, hypertriglyceridemia and hyperferritinemia. Bone marrow smear showed a picture compatible with haemophagocytic lymphohistocytosis (HLH) and first and second line therapy were started. In addition, a transjugular hepatic biopsy was performed with histopathological evidence of portal and periportal fibrosis, compatible with Covid 19-related cholangiopathy. During the hospital stay, he developed several MDR opportunistic infections. The patient died few months later from multiorgan failure secondary to septic shock. A post-mortem confirmed a diagnosis of cholangiopathy, and medullary erythro-haemophagocytosis. Conclusion: Post Covid19 syndrome is a clinical entity that includes novel and old sequelae following recovery from Sars-CoV2 infections. Early identification of these diseases is crucial for adequate management and might influence the long term prognosis of these patients.

PD-L1 in oral squamous cell carcinoma: A key biomarker from the laboratory to the bedside.

OBJECTIVES AND BACKGROUND: Oral squamous cell carcinoma (OSCC) is a highly malignant disease with an increasing incidence. The need to improve therapeutic strategies for patients affected by OSCC is an urgent challenge. Currently, the advent of immunotherapy represents an important step toward this aim. Programmed cell death-ligand 1 (PD-L1), a membrane protein that can be expressed on tumor and inflammatory cells is a key biomarker whose expression is determined by means of immunohistochemistry and is necessary for selecting patients for immunotherapy. METHODS: In this study, we review the methods of PD-L1 assessment and outcomes achieved with immunotherapy in the treatment of OSCC patients. RESULTS: Based on a meta-analysis we demonstrate a lack of prognostic significance of PD-L1 in OSCC. CONCLUSIONS: We also highlight unresolved issues including difficulties in standardizing PD-L1 evaluation and discuss future opportunities such as leveraging digital pathology.

Concordance between Three PD-L1 Immunohistochemical Assays in Head and Neck Squamous Cell Carcinoma (HNSCC) in a Multicenter Study.

The introduction of immunotherapy targeting the programmed death-1 (PD-1)/programmed death-ligand-1 (PD-L1) axis has represented a turning point in the treatment of HNSCC. Harmonization studies comparing the different antibodies and immunohistochemistry platforms available for the evaluation of PD-L1 expression with Combined Positive Score (CPS) in HNSCC are strongly required. Tissue microarrays (TMA) constructed from formalin-fixed, paraffin-embedded (FFPE) tissue blocks of HNSCC tumor were stained with two commercial in-vitro diagnostic (IVD) PD-L1 immunohistochemical assays (22C3 pharmDx on Autostainer Link48 and Omnis platforms, and SP263) and were reviewed by seven trained pathologists to assess CPS. We found a very similar distribution for PD-L1 expression between 22C3 pharmDx assay with both platforms and SP263 assay and a strong significant correlation between the two assays in different platforms (p < 0.0001). The interobserver reliability among pathologists for the continuous scores of CPS with intraclass correlation coefficient (ICC) and the correlation between the two assays were both good. Moreover, the agreement rate between assays was high at all cut-offs, while the kappa values were from substantial to almost perfect. These data suggest the interchangeability of the two antibodies and of the different immunohistochemical platforms in the selection of patients with HNSCC for immunotherapy.

Neuropathological findings from COVID-19 patients with neurological symptoms argue against a direct brain invasion of SARS-CoV-2: A critical systematic review.

BACKGROUND AND PURPOSE: Neuropathological studies can elucidate the mechanisms of nervous system damage associated with SARS-CoV-2 infection. Despite literature on this topic is rapidly expanding, correlations between neurological symptoms and brain pathology findings in COVID-19 patients remain largely unknown. METHODS: We performed a systematic literature review on neuropathological studies in COVID-19, including 438 patients from 45 articles published by April 22, 2021. We retrieved quantitative data regarding demographic, clinical, and neuropathological findings. We carried out a Wilcoxon rank sum test or χ2 test to compare patients' subgroups based on different clinical and brain pathology features. RESULTS: Neuropathological findings in COVID-19 patients were microgliosis (52.5%), astrogliosis (45.6%), inflammatory infiltrates (44.0%), hypoxic-ischemic lesions (40.8%), edema (25.3%), and hemorrhagic lesions (20.5%). SARS-CoV-2 RNA and proteins were identified in brain specimens of 41.9% and 28.3% of subjects, respectively. Detailed clinical information was available from 245 patients (55.9%), and among them, 96 subjects (39.2%) had presented with neurological symptoms in association with typical COVID-19 manifestations. We found that: (i) the detection rate of SARS-CoV-2 RNA and proteins in brain specimens did not differ between patients with versus those without neurological symptoms; (ii) brain edema, hypoxic-ischemic lesions, and inflammatory infiltrates were more frequent in subjects with neurological impairment; (iii) neurological symptoms were more common among older individuals. CONCLUSIONS: Our systematic revision of clinical correlates in COVID-19 highlights the pathogenic relevance of brain inflammatory reaction and hypoxic-ischemic damage rather than neuronal viral load. This analysis indicates that a more focused study design is needed, especially in the perspective of potential therapeutic trials.

Integrating data from multidisciplinary Management of Malignant Pleural Mesothelioma: a cohort study.

BACKGROUND: Malignant pleural mesothelioma (MPM) is a rare and aggressive malignancy that most commonly affects the pleural layers. MPM has a strong association with asbestos, mainly caused by exposure to its biopersistent fibers in at least 80% of cases. Individuals with a chronic exposure to asbestos might develop disease with a 20-40-year latency with few or no symptoms. Such has been the case in the Italian regions of Piedmont and Lombardy, where industrial production of materials laden with asbestos, mainly cements, has been responsible for the onset of a large epidemic. Since 2018, a multidisciplinary team at San Matteo hospital in Pavia has been collecting data on over 100 patients with MPM. The main goal of this project is to define and describe an integrated profile for each MPM case at diagnosis by using data mining and partition analysis. METHODS: Here we bring together exhaustive epidemiologic, histologic and radiologic data of 88 MPM patients that came to our observation and draw correlations with predictive and prognostic significance. RESULTS: The median overall survival (OS) was 15.6 months. Most patients presented with pleural effusion, irrespective of disease stage. Quite unexpectedly, no statistically significant association was demonstrated between OS and TNM disease stage at diagnosis. Although average OS is similar in male and female patients, partition analysis of data underlined a significant differential hierarchy of predictor categories based on patient gender. In females with no smoking history, full chemotherapeutic regimens are associated with better outcomes. Moreover, concerning second line treatments, vinorelbine emerged as the most advantageous choice for female patients, whereas in the male subgroup no statistically significant difference resulted between gemcitabine and vinorelbine. CONCLUSION: A multidisciplinary approach to MPM is mandatory to define better therapeutic approaches, personalize the management and improve patient outcomes.

Acute-onset diplopia from intracranial hypertension due to torcular herophili obstruction by an hemorrhagic intradiploic epidermoid cyst.

BACKGROUND: Epidermoid cysts are benign slow-growing congenital lesions, constituting approximately 1% of all cranial tumors. Most of these lesions are located intradurally, while about 10-25% of them are located within the diploic spaces. Intradiploic epidermoid cysts are usually discovered incidentally and may remain asymptomatic for many years, but in rare instances, they may grow intracranially and produce brain compression. Sometimes, intradiploic epidermoid cysts may occlude the main cranial venous sinuses causing intracranial hypertension. CASE DESCRIPTION: We present the case of a 24-year-old male harboring a paramedian right occipital intradiploic cyst with erosion of both outer and inner bony tables, which occluded the torcular herophili producing a worsening symptomatology with acute-onset diplopia from right sixth cranial nerve palsy; the patient also presented bilateral papilledema, but only reported mild headache and dizziness. Neuroradiological studies evidentiated a lesion compatible with intradiploic epidermoid cyst with intralesional hemorrhagic component, overlying and almost completely occluding the torcular herophili. Considering the fast worsening of symptomatology and the evidence of intracranial hypertension, the patient was operated on immediately after completion of clinical and radiological assessment. The lesion was radically removed with almost immediate reversal of signs and symptoms. Histopathology confirmed the diagnosis of epidermoid cyst with intralesional hemorrhagic components. CONCLUSION: Intradiploic epidermoid cysts may cause intracranial hypertension by occlusion of main cranial venous sinuses; intralesional hemorrhage may act as precipitating factor in occlusion of the torcular herophili, producing rapidly worsening intracranial hypertension, which requires prompt surgical treatment to reverse symptomatology. Radical surgical resection is necessary to avoid recurrence.

miRNAs Potentially Involved in Post Lung Transplant-Obliterative Bronchiolitis: The Role of miR-21-5p.

Epigenetic changes, including miRNAs deregulation, have been suggested to play a significant role in development of obliterative bronchiolitis (OB) in transplanted lungs. Many studies have tried to identify ideal candidate miRNAs and the downstream pathways implicated in the bronchiolar fibro-obliterative process. Several candidate miRNAs, previously indicated as possibly being associated with OB, were analyzed by combining the quantitative real time-polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH) of lung tissues of OB affected patients. Disease and OB-lesion-specific expression of miR-21-5p was confirmed and by computational analysis we were able to identify the network of genes most probably associated miR-21-5p in the context of OB fibrogenesis. Among all potentially associated genes, STAT3 had a very high probability score. Immunohistochemistry showed that STAT3/miR-21-5p were co-over expressed in OB lesions, thus, suggesting miR-21-5p could regulate STAT3 expression. However, miR-21-5p inhibition in cultures of bronchiolitis obliterans syndrome (BOS) derived myofibroblasts did not significantly affect STAT3 mRNA and protein expression levels. This study demonstrates the specificity of miR-21-5p over-expression in OB lesions and contributes to existing knowledge on the miR-21-5p downstream pathway. Activation of STAT3 is associated with miR-21-5p upregulation, however, STAT-3 network activation is most likely complex and miR-21-5p is not the sole regulator of STAT3.

Recent advancement on PD-L1 expression quantification: the radiologist perspective on CT-guided FNAC.

PURPOSE: We aimed to evaluate the feasibility, accuracy, and safety of Programmed Death-1/ Programmed Death-Ligand 1 (PD-1/ PD-L1) expression quantification in cytology cell-block samples obtained through transthoracic CT-guided fine-needle aspiration cytology (FNAC) from the interventional radiologist's perspective. METHODS: We performed a consecutive unselected series of 361 CT-guided biopsies of pulmonary nodules and masses which came to our observation from June 2017 to October 2018. For each case, exhaustive clinical, morphologic, molecular and tomographic data were available. All the material obtained was fixed in formalin to obtain a cell-block for the pathologist, who performed immunohistochemical analysis to detect PD-L1 expression levels on each sample. RESULTS: Of all the analyzed samples, 93.6% (338/361) were defined to be diagnostic, including neoplastic (72%, 260/361) and non-neoplastic lesions (21.6%, 78/361); only 6.4% (23/361) of them resulted in nondiagnostic specimens. Non-small cell lung cancer (NSCLC) accounted for 73.8% of neoplastic lesions (192/260): most of them were adenocarcinoma (83%, 160/192), followed by squamous carcinoma (14%, 27/192) and poorly differentiated carcinoma (3%, 5/192). In 96% of NSCLC (184/192), the diagnosis was reached either in the absence of complications or with early minor complications. PD-L1 expression was evaluated in all 192 NSCLC cytology specimens: 180 immunostainings were found to be adequate for PD-L1 testing. In 76% of cases, PD-L1 expression level was lower than 50%. CONCLUSION: The findings of our study indicate that PD-L1 quantification using a cell-block approach on CT-guided FNAC is a feasible and safe technique and should be taken into account alongside with core biopsy approach, especially in case of advanced disease and/or fragile and older patients.

PD-L1 Testing and Squamous Cell Carcinoma of the Head and Neck: A Multicenter Study on the Diagnostic Reproducibility of Different Protocols.

Immune checkpoint inhibitors for blocking the programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) axis are now available for squamous cell carcinoma of the head and neck (HNSCC) in relapsing and/or metastatic settings. In this work, we compared the resulting combined positive score (CPS) of PD-L1 using alternative methods adopted in routine clinical practice and determined the level of diagnostic agreement and inter-observer reliability in this setting. The study applied 5 different protocols on 40 tissue microarrays from HNSCC. The error rate of the individual protocols ranged from a minimum of 7% to a maximum of 21%, the sensitivity from 79% to 96%, and the specificity from 50% to 100%. In the intermediate group (1 ≤ CPS < 20), the majority of errors consisted of an underestimation of PD-L1 expression. In strong expressors, 5 out of 14 samples (36%) were correctly evaluated by all the protocols, but no protocol was able to correctly identify all the "strong expressors". The overall inter-observer agreement in PD-L1 CPS reached 87%. The inter-observer reliability was moderate, with an ICC of 0.774 (95% CI (0.651; 0.871)). In conclusion, our study showed moderate interobserver reliability among different protocols. In order to improve the performances, adequate specific training to evaluate PD-L1 by CPS in the HNSCC setting should be coordinated.

Fibrotic progression and radiologic correlation in matched lung samples from COVID-19 post-mortems.

Data on the pathology of COVID-19 are scarce; available studies show diffuse alveolar damage; however, there is scarce information on the chronologic evolution of COVID-19 lung lesions. The primary aim of the study is to describe the chronology of lung pathologic changes in COVID-19 by using a post-mortem transbronchial lung cryobiopsy approach. Our secondary aim is to correlate the histologic findings with computed tomography patterns. SARS-CoV-2-positive patients, who died while intubated and mechanically ventilated, were enrolled. The procedure was performed 30 min after death, and all lung lobes sampled. Histopathologic analysis was performed on thirty-nine adequate samples from eight patients: two patients (illness duration < 14 days) showed early/exudative phase diffuse alveolar damage, while the remaining 6 patients (median illness duration-32 days) showed progressive histologic patterns (3 with mid/proliferative phase; 3 with late/fibrotic phase diffuse alveolar damage, one of which with honeycombing). Immunohistochemistry for SARS-CoV-2 nucleocapsid protein was positive predominantly in early-phase lesions. Histologic patterns and tomography categories were correlated: early/exudative phase was associated with ground-glass opacity, mid/proliferative lesions with crazy paving, while late/fibrous phase correlated with the consolidation pattern, more frequently seen in the lower/middle lobes. This study uses an innovative cryobiopsy approach for the post-mortem sampling of lung tissues from COVID-19 patients demonstrating the progression of fibrosis in time and correlation with computed tomography features. These findings may prove to be useful in the correct staging of disease, and this could have implications for treatment and patient follow-up.