Tumor-stroma ratio in preoperative biopsies and matched surgical specimens in oral squamous cell carcinoma: Concordance and impact on recurrence-free and overall survival.

Stroma-richness is commonly associated with decreased survival times as well as advanced tumor stages in various malignant tumors. A previous study on laryngeal squamous cell carcinomas showed very good agreement for tumor-stroma ratio assessment between pre-treatment biopsies and resection specimens. We therefore aimed to determine whether similar results could be shown for oral squamous cell carcinomas. 107 preoperative biopsies and matched surgical specimens were obtained from the histological archive, dating from 2011-2022. Tumor-stroma ratio was determined on all samples and cases were divided into stroma-rich (≥50% stroma) and stroma-poor (<50% stroma). Results were then correlated with recurrence-free and overall survival. Tumor-stroma ratio showed substantial agreement between preoperative biopsies and surgical specimens with a kappa correlation coefficient of 0.643. Concerning preoperative biopsies, 28 cases were stroma-rich (26.2%), in the group of tumor resections, 32 cases were stroma-rich (29.9%). No association with either recurrence-free or overall survival could be shown for both groups (p-values 0.158-0.495). Concordance between pre-treatment biopsies and resections was substantial in our study, however, as no association with survival times could be demonstrated, the prognostic significance in our cohort remains unclear. This might be attributable to the fact that almost 75% of our patients presented with early-stage tumors, which sometimes seem to show a less pronounced prognostic effect of the tumor-stroma ratio.

The expression of the adenosine pathway markers CD39 and CD73 in salivary gland carcinomas harbors the potential for novel immune checkpoint inhibition.

BACKGROUND: In salivary gland carcinomas (SGC), there is only a small fraction of entities that appears to profit from immune checkpoint inhibition (ICI). Recent findings connected the activation of adenosine-signaling with a tolerogenic microenvironment. Therefore, the inhibition of adenosine pathway markers (CD39 and/or CD73) can augment ICI and/or display a novel immunotherapeutic strategy beyond ICI. Here, we assessed the immuno-histochemical expression of CD39 and CD73 across a wide spectrum of SGCs. METHODS: In total, 114 patients with SGCs consecutively diagnosed between 2001 and 2021 were assessed for clinicopathological baseline characteristics and underwent confirmatory histopathological review. Immunohistochemical expression levels of CD39 and CD73 were assessed by applying the tumor proportion score (TPS) and the immune proportional score (IPS) comparable to PD-L1 expression analysis in routine clinical practice. Additionally, findings were correlated with PD-L1 expression levels. RESULTS: The median age was 60.6 and 51.8% patients were female. The cohort covered a spectrum of eight distinct entities. Advanced-stage disease (UICC/AJCC III/IVA-IVC) at initial diagnosis was present in the majority of patients (64/114). Immunohistochemical staining revealed positivity for CD39 and CD73 in 48.2% and 21.1% on tumor cells (TPS ≥ 1%) as well as 46.4% and 42.9% within the immune cell infiltrate (IPS ≥ 1%), respectively. Further comparative analyses revealed immune-cold entities such adenoid cystic carcinoma (AdCC), immune-hot tumors such as adenocarcinoma, not otherwise specified (AC (NOS)) and entities with intermediate immunologic features such as acinic cell carcinoma (ACC). CONCLUSION: Current results indicate entity-specific adenosine signaling signatures. These findings suggest that the adenosine pathway plays a decisive role in tumor immunity among the major spectrum of SGCs. Targeting the adenosine pathway might pose a promising therapeutic option for selected entities.

Tumor budding as a prognostic factor in pancreatic ductal adenocarcinoma.

In this retrospective study, we analyzed the association between tumor budding and perineural invasion as well as their prognostic role in pancreatic ductal adenocarcinoma. A total of N = 119 patients resected for pancreatic ductal carcinoma from 1996 to 2015 were included. Clinical and standard histopathological parameters were retrieved from the patient's records. One representative hematoxylin and eosin section from the tumor region was examined for perineural invasion and tumor budding using light microscopy. Tumor budding was assessed independently using two different methods: in the first approach, the number of buds was counted over three fields of 0.237 mm2 at 40-fold magnification; in the second approach, tumor budding was quantified according to the recommendation of the International Tumor Budding Consensus Conference (ITBCC) over a field of 0.785 mm2 at 20-fold magnification. Linear and logistic regression was applied to delineate association between perineural invasion, tumor budding, and other parameters; Kaplan-Meier and Cox regression were used in the survival analysis. Regardless of the quantification approach, high tumor budding was a significant negative prognostic factor in the univariable Cox regression (> 5 buds/0.237 mm2, hazard ratio (HR) 1.66, 95% confidence interval (CI) 1.06-2.61, p = 0.027; ≥ 10 buds/0.785 mm2, HR 1.68, 95% CI 1.07-2.64, p = 0.024). In the multivariable model adjusting for stage and standard histopathological parameters, lymph vessel invasion (HR = 2.43, 95% CI 1.47-4.03, p = 0.001) and tumor budding > 5 buds/0.237 mm2 (HR = 1.70, 95% CI 1.07-2.7, p = 0.026) were independent negative prognostic factors, while adjuvant therapy was a positive prognostic factor (HR = 0.54, 95% CI 0.33-0.86, p = 0.009). No significant prognostic value could be delineated for perineural invasion. In conclusion, tumor budding is an independent negative prognostic factor in pancreatic ductal adenocarcinoma associated with lymph node metastasis. The prognostic role of perineural invasion remains uncertain.

PD-L1 expression in carcinoma of the esophagogastric junction is positively correlated with T-cell infiltration and overall survival.

INTRODUCTION: In recent years antitumor immunity and inhibition of checkpoint molecules, such as PD-1 and PD-L1, have emerged as potential therapeutic strategies in advanced stages of various malignancies. We investigated PD-L1 expression in adenocarcinomas of the esophagogastric junction and correlated the results with densitiy of intratumoral T-lymphocytes. METHODS: Immunohistochemical staining for PD-L1 was carried out on 135 samples using a tissue microarray. Scoring was done according to the combined positivity score. RESULTS: 48.1% of tumors (65 cases) showed PD-L1 positivity with a score ≥ 1 while 51.9% were PD-L1 negative (70 cases). A positive correlation between PD-L1 negativity and mucinous and poorly cohesive carcinoma could be shown (p = 0.043), while no association existed for either gender, T-stage, N-stage, grading, surgical resection status, neoadjuvant therapy, distant metastases, lymphovascular or perineural invasion. No correlation of PD-L1 expression and overall survival could be detected (p = 0.497). Again, when stratified according to presence or absence of neoadjuvant therapy, no survival differences could be shown for either group (p = 0.540 and p = 0.736). When PD-L1 expression was correlated with density of tumor-infiltrating T-lymphocytes a positive correlation between PD-L1 positivity and denser T-cell infiltration could be shown (p = 0.001). Concerning overall survival, in PD-L1 negative cases, denser CD8-positive T-cell infiltrates were associated with prolonged survival times (p = 0.045). No differences could be shown for PD-L1 positive cases or CD103-positive T-cells. CONCLUSION: PD-L1 expression is frequent in esophagogastric adenocarcinoma and - when combined with dense CD8 infiltration - PD-L1 negativity correlates with prolonged overall survival.

Density of CD8-positive tumor-infiltrating T-lymphocytes is an independent prognostic factor in adenocarcinoma of the esophagogastric junction.

Tumor-infiltrating lymphocytes (TILs) have commonly been associated with markedly improved prognosis in a variety of human cancers, including carcinomas of the upper and lower gastrointestinal tract. Especially the presence of T-cells (cytotoxic as well as helper cells) seems to define a subgroup of patients with prolonged overall and event-free survival. The density of TILs was assessed via immunohistochemistry for CD8 and CD103 in a population of 228 adenocarcinomas of the esophagogastric junction. Density of CD8+ T-lymphocytes was inversely correlated with depth of tumor infiltration (p=0.013) while no correlation with any of the analyzed clinicopathologic factors could be established for CD103-density. High density of CD8-positive T-cells additionally showed significantly longer overall survival (OS) with a p-value of 0.024 while density of CD103+ cells was associated with prolonged tumor free survival (p-value 0.011). Independence could be demonstrated applying Cox proportional hazard analysis (Hazard Ratio 0.742; 95%-Confidence Interval 0.579-0.951; p=0.019). High density of CD8-positive T-lymphocytes identifies a patient subgroup with significantly prolonged overall survival, is correlated with tumor stage and might open up new therapeutic possibilities via immunomodulating drugs.

MicroRNAs as Potential Biomarkers for Chemoresistance in Adenocarcinomas of the Esophagogastric Junction.

Concerning adenocarcinomas of the esophagogastric junction, neoadjuvant chemotherapy is regularly implemented, but patients' response varies greatly, with some cases showing no therapeutic effect, being deemed as chemoresistant. Small, noncoding RNAs (miRNAs) have evolved as key players in biological processes, including malignant diseases, often promoting tumor growth and expansion. In addition, specific miRNAs have been implicated in the development of chemoresistance through evasion of apoptosis, cell cycle alterations, and drug target modification. We performed a retrospective study of 33 patients receiving neoadjuvant chemotherapy by measuring their miRNA expression profiles. Histologic tumor regression was evaluated using resection specimens, while miRNA profiles were prepared using preoperative biopsies without prior therapy. A preselected panel of 96 miRNAs, known to be of importance in various malignancies, was used to test for significant differences between responsive (chemosensitive) and nonresponsive (chemoresistant) cases. The cohort consisted of 12 nonresponsive and 21 responsive cases with the following 4 miRNAs differentially expressed between both the groups: hsa-let-7f-5p, hsa-miRNA-221-3p, hsa-miRNA-31-5p, and hsa-miRNA-191-5p. The former 3 showed upregulation in chemoresistant cases, while the latter showed upregulation in chemosensitive cases. In addition, significant correlation between high expression of hsa-miRNA-194-5p and prolonged survival could be demonstrated (p value <0.0001). In conclusion, we identified a panel of 3 miRNAs predicting chemoresistance and a single miRNA contributing to chemosensitivity. These miRNAs might function as prognostic biomarkers and enable clinicians to better predict the effect of one or more reliably select patients benefitting from (neoadjuvant) chemotherapy.

Impact of postoperative TNM stages after neoadjuvant therapy on prognosis of adenocarcinoma of the gastro-oesophageal junction tumours.

AIM: To compare prognostic relevance of postoperative tumour/node/metastasis (TMN) stages between patients with and without neoadjuvant treatment. METHODS: Data from patients with adenocarcinoma of the gastro-oesophageal junction (AEG) who had undergone surgical resection at a single German university centre were retrospectively analysed. Patients with or without neoadjuvant preoperative treatment were selected by exact matching based on preoperative staging. Standard assessment of preoperative (c)TNM stage was based on endoscopic ultrasound and computed tomography of the thorax and abdomen, according to the American Joint Committee on Cancer/Union for International Cancer Control classification system. Patients with cT1cN0cM0 and cT2cN0cM0 stages were excluded from the study, as these patients are generally not recommended for pretreatment. Long-term survival among the various postoperative TNM stages was compared between the groups of patients with or without neoadjuvant treatment. For statistical assessments, a P-value of ≤ 0.05 was considered significant. RESULTS: The study included a total of 174 patients. The group of patients who had received preoperative neoadjuvant treatment included more cases of AEG (Siewert) type 1 carcinoma (P < 0.001), and consequently oesophagectomy was performed more frequently among these patients (P < 0.001). The two groups (with or without preoperative neoadjuvant treatment) had comparable preoperative T stages, but the group of patients with preoperative neoadjuvant treatment presented a higher rate of preoperative N-positive disease (P = 0.020). Overall long-term survival was not different between the two groups of patients according to tumours of different AEG classifications, receipt of oesophagectomy or gastrectomy, nor between patients with similar postoperative TNM stage, resection margin and grading. However, an improvement of long-term survival was found for patients with nodal down-staging after neoadjuvant therapy (P = 0.053). CONCLUSION: The prognostic relevance of postoperative TNM stages is similar for AEG in patients with or without neoadjuvant preoperative treatment, but treatment-related nodal down-staging prognosticates longer-term survival.

MicroRNA analysis of gastroenteropancreatic neuroendocrine tumors and metastases.

The incidence of neuroendocrine neoplasias (NEN) continues to increase. Since the primary tumor cannot be diagnosed in some cases of metastatic disease, new biomarkers are clearly needed to find the most probable site of origin. Tissue samples from 79 patients were analyzed and microRNA profiles were generated from a total of 76 primary tumors, 31 lymph node and 14 solid organ metastases. NEN metastases were associated with elevated levels of miR-30a-5p, miR-210, miR-339-3p, miR-345 and miR-660. Three microRNAs showed a strong correlation between proliferation index and metastatic disease in general (miR-150, miR-21 and miR-660). Further, each anatomic location (primary or metastatic) had one or more site-specific microRNAs more highly expressed in these tissues. Comparison between primary tumors and metastases revealed an overlap only in pancreatic (miR-127) and ileal tumors (let-7g, miR-200a and miR-331). This thorough analysis of gastroenteropancreatic neuroendocrine tumors demonstrates site-specific microRNA profiles, correlation with proliferation indices as well as corresponding nodal and distant metastases. Using microRNA profiling might improve NEN diagnostics by linking metastases to a most probable site of origin.

ERG expression in multiple myeloma-A potential diagnostic pitfall.

INTRODUCTION: ERG expression has been described as a frequent event in prostate cancer indicating poor prognosis and promoting oncogenesis. It has also been demonstrated in Ewing's sarcoma, acute myeloid leukemia and acute T-lymphoblastic leukemia but could not be found in other epithelial tumors, Hodgkin's or Non-Hodgkin's lymphoma. We aimed to analyze ERG expression in multiple myeloma, following an index case of a patient with metastases of unknown origin in the spine strongly expressing ERG, which were thought to be of prostatic origin but turned out to be plasmacytic lesions. MATERIAL AND METHODS: We subsequently selected 12 formalin-fixed, paraffin-embedded tissue samples of multiple myeloma from our archives and performed immunohistochemical staining for ERG. RESULTS: All 12 analyzed cases showed strong nuclear expression of ERG in >90% of tumor cells (myeloma cells). CONCLUSIONS: This report highlights a potential and critical diagnostic pitfall in biopsy specimens where morphology is only of limited assistance in reaching the correct diagnosis. It urges pathologists to exercise caution in cases where strong ERG-positivity implicates the presence of a prostatic neoplasia and illustrates the need for further immunohistochemical examination.

Expression of cyclooxygenase-2 has no impact on survival in adenocarcinoma of the esophagogastric junction but is associated with favourable clinicopathologic features.

BACKGROUND: COX-2 expression induces carcinogenesis and is thought to be an adverse prognostic factor in gastric carcinomas while the prognostic value of DNA mismatch repair (MMR) is still controversial. Concerning adenocarcinomas of the esophagogastric junction, no comprehensive data regarding either factors are available as of yet. OBJECTIVE: We assessed expression of COX-2, MLH1 and MSH2 in adenocarcinoma of the esophagogastric junction in relation to patients' survival and various clinicopathologic features. DESIGN: Immunohistochemical studies (using antibodies against COX-2, MLH1 and MSH2) were performed in a study population of 228 tumours. Follow-up data was available for all patients with a mean follow-up time of 42.8 months. RESULTS: 78 (34.2%) tumours were COX-2 negative, 148 (64.9%) showed COX-2 positivity. Assessment of COX-2 expression and clinicopathologic features revealed an inverse correlation with depth of tumour invasion and number of metastatic lymph nodes (p=0,021 and p=0,004, respectively). No correlation with other features could be demonstrated. 62 cases (27.2%) showed loss of DNA repair enzymes MLH1 and/or MSH2. MMR differed significantly between COX-2 positive and negative cases (p=0,028). Kaplan-Meier survival analyses revealed no impact on patients' survival for COX-2 expression or MMR status (p=0.837 and p=0.972, respectively). CONCLUSIONS: Expression of COX-2 in adenocarcinomas of the esophagogastric junction seems to have no prognostic effect or impact on patients' survival but is associated with favourable clinicopathologic factors. MMR deficiency was more frequent in COX-2 negative tumours, but MMR status had no impact on survival and patients' outcome whatsoever.

High Density of Tumor-infiltrating B-Lymphocytes and Plasma Cells Signifies Prolonged Overall Survival in Adenocarcinoma of the Esophagogastric Junction.

BACKGROUND: Tumor-infiltrating lymphocytes (TILs) have been shown to be of prognostic significance in a variety of tumors. Not only T-cell, but also B-cell infiltration is commonly associated with improved survival. MATERIALS AND METHODS: We assessed the density of tumor-infiltrating B-cells, as well as that of plasma cells, in 210 adenocarcinomas of the esophagogastric junction through immunohistochemical analysis using antibodies against CD20 and CD138. RESULTS: No correlation between density of B-cells or plasma cells and various clinicopathologic features could be established. High density of tumor-infiltrating B-cells, as well as plasma cells, showed significantly better overall survival (OS) compared to patients with no infiltrates (p=0.047 and p=0.022, respectively). Cox proportional hazard analysis could verify B-cell infiltration as an independent prognostic factor (hazard ratio (HR)=0.683; 95% confidence interval (CI)=0.517-0.901; p=0.007). CONCLUSION: Plasma cell and B-cell infiltration correlates with prolonged OS and might identify a patient subset with favorable disease course.

Evaluation of general and coronary atherosclerosis and malignant disease demonstrates inverse correlations for specific cancer types as well as cancer in general.

CONTEXT: Historical data has shown that cancer in general has been associated with less atherosclerosis while recently several studies have demonstrated more heterogeneous relations. However, most investigations were carried out clinically or radiographically and updated exhaustive, comprehensive autopsy studies are lacking. In our study, we aimed to assess the relationship between malignancy and grade of atherosclerosis in different locations (general, coronary) in a large study population over the course of 14 years. METHODS: 2370 patients (autopsy reports) were analyzed retrospectively in regard to various parameters including demographics, organ weights, distribution and severity of atherosclerosis and presence or absence of malignancy. RESULTS: Our study shows that malignant disease is usually associated with less general and coronary atherosclerosis (p=0.002 and p<0.001). Especially breast, colorectal and pancreatic cancer as well as lymphomas/lymphoid leukaemia and sarcomas were associated with significantly less general and coronary AS (p-values<0.001-0.031). In addition, a positive correlation between coronary atherosclerosis and heart weight could be detected (ρ=0.302, p<0.001). As a common observation, male sex was associated with increased severity of general and coronary atherosclerosis in both cancer and non-cancer groups (p<0.001). CONCLUSIONS: We demonstrated that an inverse correlation between atherosclerosis and cancer in general is still sound today despite tremendous changes and advances in therapy strategies and diagnostics. Additionally, we could highlight that the effect was most pronounced in breast, colorectal and pancreatic cancer, sarcomas and lymphomas/lymphoid leukaemia.

Oncogenic mutations and chromosomal aberrations in primary extranodal diffuse large B-cell lymphomas of the thyroid--a study of 21 cases.

CONTEXT: Primary extranodal diffuse large B-cell lymphomas of the thyroid (ptDLBCL) constitute a rare entity, which until now was not fully explored. OBJECTIVE: Due to recently published data genetically linking ptDLBCL to a subset of thyroid carcinoma, we assessed the occurrence of oncogenic mutations and copy number alterations. DESIGN: A high-resolution array-based comparative genomic hybridization approach was applied to quantify genomic aberrations in a study population of 21 ptDLBCL patients. In addition, we investigated the frequency of mutations involving the BRAF, NRAS, and MYD88 genes in correlation with immunohistochemical data. RESULTS: Chromosomal gains were recurrently detected at 6p21.33-p21.31, 6p22.2, 12p13.31, 14q31.1, 14q32.33, 19p13.3, and 22q11.22; numeric losses were most frequently observed at 6p21.3-p21.31, 10q26.3, 19p13.3, 20q13.33, and 21q11.2. Aberrations affecting 6p22.2 and 14q32.33 as well as 22q11.22 differed slightly between germinal center B-cell (GCB) and non-GCB groups. Statistically significant deviations were detected at 20q13.33 and 21q11.2. These specific alterations do not seem to occur in thyroid carcinomas or other DLBCL, according to previously published literature. Analysis of BRAF and NRAS showed mutation frequencies of 4.8 and 9.5%, respectively. No MYD88 mutations could be detected in any of the analyzed cases. Fluorescence in situ hybridization demonstrated breakage events involving the BCL2, BCL6, and cMYC locus in 14.3, 9.5, and 9.5%, respectively. CONCLUSIONS: Our study revealed ptDLBCL to be predominantly composed of the GCB type, harboring no MYD88 mutations and showing infrequent mutations in the BRAF and NRAS genes. Additionally, array comparative genomic hybridization showed no overlapping alterations between ptDLBCL and thyroid carcinomas or other nodal or extranodal DLBCL.

Dysregulation of microRNAs in angioimmunoblastic T-cell lymphoma.

BACKGROUND: Angioimmunoblastic T-cell lymphomas (AITLs) are the second most frequent peripheral T-cell lymphomas in humans worldwide and histomorphologically well characterized. MicroRNAs are a group of small non-coding RNAs that can negatively regulate gene expression on a posttranscriptional level. Their dysregulation has been shown to be of importance in numerous tumour entities. MATERIALS AND METHODS: As a first step towards understanding the possible influence of microRNA-dysregulation in AITL, we analyzed the expression signatures of 760 microRNAs in 30 nodal AITLs in comparison to reactive lymphadenitis with T-zone hyperplasia. RESULTS: We found miR-34a, miR-146a and miR-193b to be up-regulated, as well as miR-140-3p, let-7g, miR-30b and miR-664 to be down-regulated in AITL to a significant level. CONCLUSION: The microRNA-signatures of AITL reveal some overlap to autoimmune diseases, virus-triggered lymphomas and angiogenic factors that, coupled with future studies, will potentially provide better understanding of this disease.

Granular cell tumour of the urinary bladder.

With only 16 cases reported in the literature, the mostly benign granular cell tumour of the urinary bladder is exceptionally rare. We present the case of a 68-year old patient with one of these lesions demonstrating our histological findings including several immunohistochemical stainings used to differentiate between other more common entities.