The impact of the tumor microenvironment on the survival of penile cancer patients.

PeCa is a rare entity with rising incidence rates due to increased infections with human papillomaviruses (HPV). The distinct subtypes of PeCa with an individual pathogenesis demand biomarkers for a precise patient risk assessment regarding disease progression and therapeutic susceptibility. We recently identified promising candidates associated with an HPV-instructed tumor microenvironment (TME) using HPV-positive PeCa cell lines and tissue microarrays (TMA). The capacity of HPV + p63 + PeCa cells to release neutrophil-attracting CXCL-8 provided a molecular link explaining the infiltration of CD15 + myeloid cells in PeCa specimens. The candidate biomarkers HPV, p63, CD15, DKK1, and CD147 linked a tumor-promoting TME with a higher TNM classification reflecting more aggressive and metastasizing cancers. Based on immune-reactive scores (IRS) from TMA staining for these biomarkers, we calculated correlations and conducted association analyses to assess the degree of relationship between all biomarkers. We then conducted Kaplan-Meier survival estimates and Cox regression analyses to delineate the impact on PeCa patient survival. There is a notable predictive potential regarding the survival of patients with biomarker profiles beyond the potency of the individual biomarker. From all candidate biomarkers and biomarker profiles, the combination of CD147 and infiltrating CD15 + cells linked to an active HPV-driven transformation displayed cancer-immune dynamics with dismal prognosis for patients. After deciphering relevant interdependencies, the HPV + CD147 + CD15 + status was the most potent profile predicting metastasis-free survival of PeCa patients. The results of this report underscore the need for analysis of the TME and the development of multi-parameter composite scores that reflect fundamental cancer-immune relationships to tailor therapeutic interventions based on actual cancer immune dynamics.

Comparative Analysis of Inhibitory and Activating Immune Checkpoints PD-1, PD-L1, CD28, and CD86 in Non-Melanoma Skin Cancer.

The establishment of immunotherapy applying immune checkpoint inhibitors (ICI) has provided an important new option for the treatment of solid malignant diseases. However, different tumor entities show dramatically different responses to this therapy. BCC responds worse to anti-PD-1 ICIs as compared to cSCC. Differential immune checkpoint expression could explain this discrepancy and, therefore, the aim of this study was to analyze activating and inhibitory immune checkpoints in cSCC and BCC tissues. Tissue microarrays of the invasive front as well as the tumor core of BCC and cSCC samples were used to evaluate PD-1, PD-L1, CD28, and CD86 expression and their topographic distribution profiles by chromogenic immunohistochemistry. QuPath was used to determine the labeling index. The expression of PD-1, PD-L1, and CD28 was significantly higher in both the tumor core and the invasive front of cSCC samples as compared to BCC (p < 0.001). In addition, the ratios of PD-L1/CD86 (p < 0.001) and CD28/CD86 (p < 0.001) were significantly higher in cSCC. The invasive front of both tumor entities showed higher expression levels of all immune markers compared to the tumor core in both tumor entities. The significantly higher expression of PD-1, PD-L1, and CD28 in cSCC, along with the predominance of the inhibitory ligand PD-L1 as compared to the activating CD86 in cSCC, provide a potential explanation for the better objective response rates to anti-PD-1 immunotherapy as compared to BCC. Furthermore, the predominant site of interaction between the immune system and the tumor was within the invasive front in both tumor types.

Evaluation of Prognostic Parameters to Identify Aggressive Penile Carcinomas.

BACKGROUND: Advanced penile carcinoma is characterized by poor prognosis. Most data on prognostic factors are based on small study cohorts, and even meta-analyses are limited in patient numbers. Therefore, there is still a lack of evidence for clinical decisions. In addition, the most recent TNM classification is questionable; in line with previous studies, we found that it has not improved prognosis estimation. METHODS: We evaluated 297 patients from Germany, Russia, and Portugal. Tissue samples from 233 patients were re-analyzed by two experienced pathologists. HPV status, p16, and histopathological parameters were evaluated for all patients. RESULTS: Advanced lymph node metastases (N2, N3) were highly significantly associated with reductions in metastasis-free (MFS), cancer-specific (CS), and overall survival (OS) rates (p = <0.001), while lymphovascular invasion was a significant parameter for reduced CS and OS (p = 0.005; p = 0.007). Concerning the primary tumor stage, a significant difference in MFS was found only between pT1b and pT1a (p = 0.017), whereas CS and OS did not significantly differ between T categories. In patients without lymph node metastasis at the time of primary diagnosis, lymphovascular invasion was a significant prognostic parameter for lower MFS (p = 0.032). Histological subtypes differed in prognosis, with the worst outcome in basaloid carcinomas, but without statistical significance. HPV status was not associated with prognosis, either in the total cohort or in the usual type alone. CONCLUSION: Lymphatic involvement has the highest impact on prognosis in penile cancer, whereas HPV status alone is not suitable as a prognostic parameter. The pT1b stage, which includes grading, as well as lymphovascular and perineural invasion in the T stage, seems questionable; a revision of the TNM classification is therefore required.

Cytology and HPV Co-Testing for Detection of Vaginal Intraepithelial Neoplasia: A Retrospective Study.

(1) Background: Vaginal intraepithelial neoplasia (VaIN) is a rare premalignant disease caused by persistent human papillomavirus (HPV) infection. Diagnosing VaIN is challenging; abnormal cytology and positive HPV tests are usually the first signs, but published data on their accuracy for detecting it are rare and contradictory. The aim of this study is to compare the results of hrHPV and cytology co-testing with the histological findings of the vagina. (2) Methods: In the certified Dysplasia Unit at Erlangen University Hospital, cytology and HPV samples from the uterine cervix or vaginal wall after hysterectomy were obtained between 2015 and 2023 and correlated with histological findings in biopsies from the vaginal wall. Women without vaginal biopsy findings or concomitant cervical disease were excluded. (3) Results: In all, 279 colposcopies in 209 women were included. The histological results were: benign (n = 86), VaIN I/vLSIL (n = 116), VaIN II/vHSIL (n = 41), VaIN III/vHSIL (n = 33), and carcinoma (n = 3). Accuracy for detecting VaIN was higher in women with previous hysterectomies. Positive HPV testing during colposcopy increased the likelihood for VaIN II/III/vHSIL threefold. The detection rate for VaIN III/vHSIL was 50% after hysterectomy and 36.4% without hysterectomy. (4) Conclusions: Women with risk factors for VaIN, including HPV-16 infection or prior HPV-related disease, need careful work-up of the entire vaginal wall. Hysterectomy for HPV-related disease and a history of cervical intraepithelial neoplasia (CIN) also increased the risk for VaIN II/III/vHSIL.

A20 as a Potential New Tool in Predicting Recurrence and Patient's Survival in Oral Squamous Cell Carcinoma.

A20, known as a potent inhibitor of NF-κB signaling, has been characterized in numerous clinical as well as preclinical studies. Recently, especially in various malignant diseases, the prognostic and therapeutic relevance of A20 was investigated. In oral squamous cell carcinoma (OSCC) however, the characterization of A20 is uncharted territory. We analyzed a tissue microarray (TMA) of 229 surgically-treated OSCC patients (2003-2013). Immunohistochemical (IHC) stainings were performed for A20 and CD3; additionally, standard haematoxylin-eosin staining was applied. IHC findings were correlated with a comprehensive dataset, comprising clinical and pathohistological information. A20 expression was analyzed in tumor cells as well as in tumor infiltrating lymphocytes (TILs) and correlated with the overall survival (OS) and recurrence-free survival (RFS) using uni- and multivariable Cox regression. The median follow-up time was 10.9 years and the A20 expression was significantly decreased in CD3+ TILs compared to mucosa-infiltrating lymphocytes (MILs). In the Kaplan-Meier analyses, higher A20 expression in TILs was correlated with better OS (p = 0.017) and RFS (p = 0.020). In the multivariable survival analysis, A20 overexpression correlated with improved OS (HR: 0.582; 95% CI 0.388-0.873, p = 0.009) and RFS (HR 0.605; 95% CI 0.411-0.889, p = 0.011). Our results indicate a novel prognostic role for A20 in OSCC. Due to its elevated expression in TILs, further research is highly desirable, which therefore could offer new therapeutic opportunities for patients suffering from OSCC.

Proposal of a T3 Subclassification for Colon Carcinoma.

The TNM classification system is one of the most important factors determining prognosis for cancer patients. In colorectal cancer, the T category reflects the depth of tumor invasion. T3 is defined by a tumor that invades through the muscularis propria into pericolorectal tissues. The data of 1047 patients with complete mesocolic excision were analyzed. The depth of invasion beyond the outer border of the muscularis propria into the subserosa or into nonperitonealized pericolic tissue was measured and categorized in 655 pT3 patients: pT3a (≤1 mm), pT3b,c (>1−15 mm) and pT3d (>15 mm). The prognosis of these categories was compared. Five-year distant metastasis increased significantly from pT3a (5.7%) over pT3b,c (17.7%) to pT3d (37.2%; p = 0.001). There was no difference between pT2 (5.3%) and pT3a or between pT3d and pT4a (42.1%) or pT4b (33.7%). The 5-year disease-free survival decreased significantly from pT3a (77.4%) over pT3b,c (65.4%) to pT3d (50.1%; p = 0.015). No significant difference was found between pT2 (80.5%) and pT3a or between pT3d and pT4a (43.9%; p = 0.296) or pT4b (53.4%). The prognostic inhomogeneity in pT3 colon carcinoma has been demonstrated. A three-level subdivision of T3 for colon carcinoma in the TNM system into T3a (≤1 mm), T3b (>1−15 mm), and T3c (>15 mm) is recommended.

Differences and Similarities in the Pattern of Early Metabolic and Morphologic Response after Induction Chemo-Immunotherapy versus Induction Chemotherapy Alone in Locally Advanced Squamous Cell Head and Neck Cancer.

Background: In head and neck cancer patients, parameters of metabolic and morphologic response of the tumor to single-cycle induction chemotherapy (IC) with docetaxel, cis- or carboplatin are used to decide the further course of treatment. This study investigated the effect of adding a double immune checkpoint blockade (DICB) of tremelimumab and durvalumab to IC on imaging parameters and their significance with regard to tumor cell remission. Methods: Response variables of 53 patients treated with IC+DICB (ICIT) were compared with those of 104 who received IC alone. Three weeks after one cycle, pathologic and, in some cases, clinical and endoscopic primary tumor responses were evaluated and correlated with a change in 18F-FDG PET and CT/MRI-based maximum-standardized uptake values (SUVmax) before (SUVmaxpre), after treatment (SUVmaxpost) and residually (resSUVmax in % of SUVmaxpre), and in maximum tumor diameter (Dmax) before (Dmaxpre) and after treatment (Dmaxpost) and residually (resD). Results: Reduction of SUVmax and Dmax occurred in both groups; values were SUVmaxpre: 14.4, SUVmaxpost: 6.6, Dmaxpre: 30 mm and Dmaxpost: 23 mm for ICIT versus SUVmaxpre: 16.5, SUVmaxpost: 6.4, Dmaxpre: 21 mm, and Dmaxpost: 16 mm for IC alone (all p < 0.05). ResSUVmax was the best predictor of complete response (IC: AUC: 0.77; ICIT: AUC: 0.76). Metabolic responders with resSUVmax ≤ 40% tended to have a higher rate of CR to ICIT (88%; n = 15/17) than to IC (65%; n = 30/46; p = 0.11). Of the metabolic nonresponders (resSUVmax > 80%), 33% (n = 5/15) achieved a clinical CR to ICIT versus 6% (n = 1/15) to IC (p = 0.01). Conclusions: ICIT and IC quickly induce a response and 18F-FDG PET is the more accurate modality for identifying complete remission. The rate of discrepant response, i.e., pCR with metabolic nonresponse after ICIT was >30%.

The prominent role of the S100A8/S100A9-CD147 axis in the progression of penile cancer.

Currently, no established biomarkers are recommended for the routine diagnosis of penile carcinoma (PeCa). The rising incidence of this human papillomavirus (HPV)-related cancer entity highlights the need for promising candidates. The Calprotectin subunits S100A8 and S100A9 mark myeloid-derived suppressor cells in other HPV-related entities while their receptor CD147 was discussed to identify patients with PeCa at a higher risk for poor prognoses and treatment failure. We thus examined their expression using immunohistochemistry staining of PeCa specimens from 74 patients on tissue microarrays of the tumor center, the invasion front, and lymph node metastases. Notably, whereas the tumor center was significantly more intensively stained than the invasion front, lymph node metastases were thoroughly positive for both S100 subunits. An HPV-positive status combined with an S100A8+S100A9+ profile was related with an elevated risk for metastases. We observed several PeCa specimens with S100A8+S100A9+-infiltrating immune cells overlapping with CD15 marking neutrophils. The S100A8+S100A9+CD15+ profile was associated with dedifferentiated and metastasizing PeCa, predominantly of HPV-associated subtype. These data suggest a contribution of neutrophil-derived suppressor cells to the progression of HPV-driven penile carcinogenesis. CD147 was elevated, expressed in PeCa specimens, prominently at the tumor center and in HPV-positive PeCa cell lines. CD147+HPV+ PeCa specimens were with the higher-frequency metastasizing cancers. Moreover, an elevated expression of CD147 of HPV-positive PeCa cell lines correlated negatively with the susceptibility to IgA-based neutrophil-mediated tumor cell killing. Finally, stratifying patients regarding their HPV/S100A8/S100A9/CD15/CD147 profile may help identify patients with progressing cancer and tailor immunotherapeutic treatment strategies.

Vedolizumab blocks α4ß7 integrin-mediated T cell adhesion to MAdCAM-1 in microscopic colitis.

Background: In Crohn's disease and ulcerative colitis, the anti-α4ß7 integrin antibody vedolizumab has demonstrated efficacy in phase III trials and has been successfully used under real-world conditions. Occasionally, it has also been used in other forms of inflammatory bowel disease (IBD) such as microscopic colitis (MC). However, the mechanisms of vedolizumab in MC have not been studied to date. Therefore, we aimed to investigate the expression and functional role of gut-homing integrins and in particular α4ß7 integrin in a cohort study in MC. Methods: We studied the expression of gut homing integrins on T cells from patients with MC and healthy controls by flow cytometry. To investigate the function of α4ß7 integrin in MC and the potential of vedolizumab to block it, we used dynamic adhesion assays and transmigrations assays. Moreover, we describe two clinical cases of MC patients treated with vedolizumab. Results: A specific profile of gut homing markers can be found on T cells from patients with MC. α4ß7 integrin functionally leads to firm adhesion to MAdCAM-1 and supports transmigration. Vedolizumab is able to block both processes. In two cases of MC, we observed reduced clinical symptoms and histologic improvement upon therapy with vedolizumab. Conclusion: Our data suggest that α4ß7 mediates gut homing of T cells also in MC and that, on single cell level, vedolizumab blocks the function of α4ß7 in MC. Thus, we provide mechanistic evidence supporting vedolizumab as promising therapeutic option for MC.

IL-9 Producing Tumor-Infiltrating Lymphocytes and Treg Subsets Drive Immune Escape of Tumor Cells in Non-Small Cell Lung Cancer.

Although lung cancer is the leading cause of cancer deaths worldwide, the mechanisms how lung cancer cells evade the immune system remain incompletely understood. Here, we discovered IL-9-dependent signaling mechanisms that drive immune evasion in non-small cell lung cancer (NSCLC). We found increased IL-9 and IL-21 production by T cells in the tumoral region of the lung of patients with NSCLC, suggesting the presence of Th9 cells in the lung tumor microenvironment. Moreover, we noted IL-9 producing Tregs in NSCLC. IL-9 target cells in NSCLC consisted of IL-9R+ tumor cells and tumor-infiltrating lymphocytes. In two murine experimental models of NSCLC, and in vitro, IL-9 prevented cell death and controlled growth of lung adenocarcinoma cells. Targeted deletion of IL-9 resulted in successful lung tumor rejection in vivo associated with an induction of IL-21 and reduction of Treg cells. Finally, anti-IL-9 antibody immunotherapy resulted in suppression of tumor development even in established experimental NSCLC and was associated with reduced IL-10 production in the lung. In conclusion, our findings indicate that IL-9 drives immune escape of lung tumor cells via effects on tumor cell survival and tumor infiltrating T cells. Thus, strategies blocking IL-9 emerge as a new approach for clinical therapy of lung cancer.

The comparison of the morphology of the mid-palatal suture between edentulous individuals and dentate jaws shows morphological differences.

BACKGROUND: A profound understanding of the evolution and anatomy of the viscero- and neurocranium is quintessentially important for orthodontists. This particularly alludes to structures, which are directly targeted by orthodontic therapy such as the maxilla and the mid-palatal suture. The anatomy of the mid-palatal suture of toothed individuals is well described, whereas little is known about sutures' morphological changes after tooth loss. Therefore, the aim of this study was to evaluate the edentulous mid-palatal suture by means of histologic and histomorphometric analysis. METHODS: Ten mid-palatal sutures of edentulous donors as well as six age- and sex matched dentulous controls were examined. For the histological and histomorphometric analysis (sutural width, obliteration, vascularization and interdigitation) conventional staining protocols (HE, Movat-Pentachrome, Sirius Red) and immunofluorescence (vWF, TRAP) were performed. Histomorphometric analysis was carried out using NIS-elements imaging software. RESULTS: When compared to dentulous controls, the edentulous investigation group showed a decreased vascularization and sutural width as well as an increased sutural obliteration. Notably, a high variability and inhomogeneity within regard the histomorphometric parameters was seen in edentulous samples. CONCLUSIONS: The mid-palatal suture of edentulous individuals showed significant morphological differences compared to individuals with toothed jaws. The loss of teeth and thereby functional loading seems to have a considerable impact on sutures' morphology.

Macrophage and T-Cell Infiltration and Topographic Immune Cell Distribution in Non-Melanoma Skin Cancer of the Head and Neck.

Non-melanoma skin cancer (NMSC) is a heterogeneous tumor entity that is vastly determined by age and UV-light exposure leading to a great mutational burden in cancer cells. However, the success of immune checkpoint blockade in advanced NMSC and the incidence and disease control rates of NMSC in organ transplant recipients compared to immunologically uncompromised patients point toward the emerging importance of the immunologic activity of NMSC. To gain first insight into the role of T-cell and macrophage infiltration in NMSC of the head and neck and capture their different immunogenic profiles, which appear to be highly relevant for the response to immunotherapy, we conducted a whole slide analysis of 107 basal cell carcinoma (BCC) samples and 117 cutaneous squamous cell carcinoma (cSCC) samples. The CD8+ and CD68+ immune cell expression in both cancer types was evaluated by immunohistochemistry and a topographic distribution profile, and the proportion of both cell populations within the two tumor entities was assessed. The results show highly significant differences in terms of CD8+ T-cell and CD68+ macrophage infiltration in BCC and cSCC and indicate cSCC as a highly immunogenic tumor. Yet, BCC presents less immune cell infiltration; the relation between the immune cells compared to cSCC does not show any significant difference. These findings help explain disparities in local aggressiveness, distant metastasis, and eligibility for immune checkpoint blockade in both tumor entities and encourage further research.

Fast whole-slide cartography in colon cancer histology using superpixels and CNN classification.

Purpose: Automatic outlining of different tissue types in digitized histological specimen provides a basis for follow-up analyses and can potentially guide subsequent medical decisions. The immense size of whole-slide-images (WSIs), however, poses a challenge in terms of computation time. In this regard, the analysis of nonoverlapping patches outperforms pixelwise segmentation approaches but still leaves room for optimization. Furthermore, the division into patches, regardless of the biological structures they contain, is a drawback due to the loss of local dependencies. Approach: We propose to subdivide the WSI into coherent regions prior to classification by grouping visually similar adjacent pixels into superpixels. Afterward, only a random subset of patches per superpixel is classified and patch labels are combined into a superpixel label. We propose a metric for identifying superpixels with an uncertain classification and evaluate two medical applications, namely tumor area and invasive margin estimation and tumor composition analysis. Results: The algorithm has been developed on 159 hand-annotated WSIs of colon resections and its performance is compared with an analysis without prior segmentation. The algorithm shows an average speed-up of 41% and an increase in accuracy from 93.8% to 95.7%. By assigning a rejection label to uncertain superpixels, we further increase the accuracy by 0.4%. While tumor area estimation shows high concordance to the annotated area, the analysis of tumor composition highlights limitations of our approach. Conclusion: By combining superpixel segmentation and patch classification, we designed a fast and accurate framework for whole-slide cartography that is AI-model agnostic and provides the basis for various medical endpoints.

DKK1 inhibits canonical Wnt signaling in human papillomavirus-positive penile cancer cells.

Penile squamous cell cancer (PSCC) is the most frequent penile malignant disease. Infections with human papillomaviruses (HPV) are a major etiologic driver of PSCC. However, the molecular details of the underlying carcinogenesis are understudied because of rare clinical specimens and missing cell lines. Here, we investigated if the expression of high-risk HPV16 oncogenes causes an augmentation of the Wnt pathway using unique HPV-positive penile cancer (PeCa) cell lines in monolayer and organotypic 3D raft cultures as well as tissue micro arrays containing clinical tissue specimens. The HPV oncoproteins enhanced the expression of Leucine-rich repeat-containing G-protein coupled receptor 6 (LGR6) and the HPV-positive PeCa cells expressed a signature of Wnt target and stemness-associated genes. However, the notable lack of nuclear ß-catenin in vitro and in situ raised the question if the enhanced expression of Wnt pathway factors is tantamount to an active Wnt signaling. Subsequent TOP-flash reporter assays revealed Wnt signaling as absent and not inducible by respective Wnt ligands in PeCa cell lines. The HPV-positive PeCa cells and especially HPV-positive PeCa specimens of the tumor core expressed the Wnt antagonist and negative feedback-regulator Dickkopf1 (DKK1). Subsequent neutralization experiments using PeCa cell line-conditioned media demonstrated that DKK1 is capable to impair ligand-induced Wnt signaling. While gene expression analyses suggested an augmented and active canonical Wnt pathway, the respective signaling was inhibited due to the endogenous expression of the antagonist DKK1. Subsequent TMA stainings indicated Dkk1 as linked with HPV-positivity and metastatic disease progression in PeCa suggesting potential as a prognostic marker.

α4ß7 integrin-dependent adhesion of T cells to MAdCAM-1 is blocked by vedolizumab in patients with chronic refractory pouchitis.

BACKGROUND: The anti-α4ß7 integrin antibody vedolizumab is an established therapeutic option for the treatment of inflammatory bowel disease (IBD). It has also been successfully used in patients with chronic antibiotic-refractory pouchitis following proctocolectomey with ileal pouch-anal anastomosis. However, the expression and function of gut-homing markers as well as strategies to predict the response to vedolizumab in pouchitis are understudied so far. METHODS: We used flow cytometry and dynamic adhesion assays to study the expression and function of gut-homing integrins on T cells from patients with pouchitis and controls as well as longitudinally during therapy of pouchitis with vedolizumab. Moreover, we describe clinical effects of vedolizumab in a cohort of patients with pouchitis. RESULTS: T cells from patients with pouchitis express a specific profile of gut-homing integrins. Integrin α4ß7 on T cells from patients with pouchitis mediates adhesion to mucosal addressin cell adhesion molecule (MAdCAM)-1, which can be blocked by vedolizumab in vitro. Vedolizumab efficiently treats pouchitis in a portion of patients and response correlates with dynamic adhesion profiles to MAdCAM-1. CONCLUSION: Our data suggest that T cell trafficking seems to be important for the pathogenesis of pouchitis and support the therapeutic use of vedolizumab. Integrin function might serve as a biomarker to predict response to vedolizumab.

Implementation of Double Immune Checkpoint Blockade Increases Response Rate to Induction Chemotherapy in Head and Neck Cancer.

To determine whether a single dose of double immune checkpoint blockade (induction chemoimmunotherapy (ICIT)) adds benefit to induction single-cycle platinum doublet (induction chemotherapy (IC)) in locally advanced head and neck squamous cell carcinoma (HNSCC), patients treated with cisplatin 30 mg/m2 d1-3 and docetaxel 75 mg/m2 d1 combined with durvalumab 1500 mg fixed dose d5 and tremelimumab 75 mg fixed dose d5 (ICIT) within the CheckRad-CD8 trial were compared with a retrospective cohort receiving the same chemotherapy (IC) without immunotherapy. The endpoint of this analysis was the complete response rate (CR). A total of 53 patients were treated with ICIT and 104 patients with IC only. CR rates were 60.3% for ICIT and 40.3% for IC (p = 0.018). In the total population (n = 157), the most important predictor to achieve a CR was treatment type (OR: 2.21 for ICIT vs. IC; p = 0.038, multivariate analysis). The most diverse effects in CR rates between ICIT and IC were observed in younger (age ≤ 60) patients with HPV-positive OPSCCs (82% vs. 33%, p = 0.176), while there was no difference in older patients without HPV-positive OPSCCs (53% vs. 48%). The analysis provides initial evidence that ICIT could result in higher CR rates than IC. Young patients with HPV-positive OPSCCs may have the greatest benefit from additional immune checkpoint inhibitors.

Lineage-associated connexin 43 expression in bisphosphonate-exposed rat bones.

Expression of signaling proteins in bone cells depends on their embryological mesoderm-derived (e.g. tibia) or cranial neural crest (CNC)-derived (e.g. jaw) origin. Connexin 43 (Cx43) is a gap junction protein that plays an essential role in the mode of action of bisphosphonates (BP). This study aimed to investigate Cx43 expression and the influence of BP application on mesoderm- and CNC-derived bone. Using a rat model, molar extraction and tibia osteotomy with (Group 4) or without (Group 3) previous BP application was performed. Untreated (Group 1) and animals selectively treated with BPs (Group 2) served as controls. Cx43 expression was immunohistochemically determined 12 and 16 weeks postoperatively via a labeling index. Cx43 expression in CNC-derived bone was significantly higher compared with mesodermal bone. BP application decreased Cx43 expression; however, detected expression levels were still higher in jawbone (Group 2 tibia vs jaw: 5.83 ± 5.06 vs 23.52 ± 6.42; p = 0.007). During bone healing after surgical intervention (Group 3) there were no expression differences between tibia and jawbone. BP treatment prior to surgery resulted in significantly lower Cx43 expression in CNC-derived compared with tibia bone (Group 4 tibia vs jaw: 56.84 ± 15.57 vs 16.40 ± 5.66; p < 0.01). Increased Cx43 expression in jaw compared with tibia bone is in line with their embryological origins. A significant Cx43 suppression in jawbone after BP application and surgery might contribute to the selectively altered osseous turnover and development of MRONJ in CNC-derived bone.

The HPV and p63 Status in Penile Cancer Are Linked with the Infiltration and Therapeutic Availability of Neutrophils.

Squamous penile cancer displays a rare human papillomavirus (HPV)-associated tumor entity. Investigations on the molecular pathogenesis of HPV-driven penile cancer are impaired by the rareness of clinical specimens and, in particular, are missing relevant cell culture models. Here, we identified in HPV-positive penile cancer cell lines that HPV16 oncoproteins control TP63 expression by modulating critical regulators, while integration into the TP63 open reading frame facilitates oncogene expression. The resulting feed-forward loop leads to elevated p63 levels that in turn enhance the release of the neutrophil-recruiting chemokine CXCL8. Remarkably, elevated CXCL8 amounts lead to the increased surface exposition of the Fc receptor of human IgA antibodies, FcαRI, on neutrophils and correlated with a higher susceptibility to antibody-dependent neutrophil-mediated cytotoxicity (ADCC) using an EGFR-specific IgA2 antibody. IHC staining of tissue microarrays proved that elevated expression of p63 together with neutrophil infiltration were significantly more frequent in HPV-positive penile cancer displaying a higher tumor grade. In summary, we identified a promising marker profile of patients with penile cancer at higher risk for worse prognosis. However, these patients may benefit from immunotherapeutic approaches efficiently engaging neutrophils for tumor cell killing.

Seizure-induced neuronal apoptosis is related to dysregulation of the RNA-edited GluR2 subunit in the developing mouse brain.

Ca2+-permeable AMPA receptors (AMPAR) which crucially modify maturational programs of the developing brain are involved in seizure-induced glutamate excitotoxicity and apoptosis. Regulatory effects on AMPAR subunit composition and RNA-editing in the developing brain and their significance as therapeutic targets are not well understood. Here, we analyzed acute effects of recurrent pilocarpine-induced neonatal seizures on age- and region-specific expression of AMPAR subunits and adenosine deaminases (ADAR) in the developing mouse brain (P10). After recurrent seizure activity and regeneration periods of 6-72 h cerebral mRNA levels of GluR (glutamate receptor subunit) 1, GluR2, GluR3, and GluR4 were unaffected compared to controls. However, ratio of GluR2 and GluR4 to pooled GluR1-4 mRNA concentration significantly decreased in seizure-exposed brains in comparison to controls. After a regeneration period of 24-72 h ADAR1 and ADAR2 mRNA expression was significantly lower in seizure-exposed brains than in those of controls. This was confirmed at the protein level in the hippocampal CA3 region. We observed a regionally increased apoptosis (TUNEL+ and CC3+ cells) in the hippocampus, parietal cortex and subventricular zone of seizure-exposed brains in comparison to controls. Together, present in vivo data demonstrate the maturational age-specific, functional role of RNA-edited GluR2 in seizure-induced excitotoxicity in the developing mouse brain. In response to recurrent seizure activity, we observed reduced expression of GluR2 and the GluR2 mRNA-editing enzymes ADAR1 and ADAR2 accompanied by increased apoptosis in a region-specific manner. Thus, AMPA receptor subtype-specific mRNA editing is assessed as a promising target of novel neuroprotective treatment strategies in consideration of age-related developmental mechanisms.