Real-world data on the prevalence of BRCA1/2 and HRR gene mutations in patients with primary and metastatic castration resistant prostate cancer.

PURPOSE: This study seeks to contribute real-world data on the prevalence of BRCA1/2 and HRR gene mutations in prostate cancer. METHODS: We compiled sequencing data of 197 cases of primary and metastatic prostate cancer, in which HRR mutation analysis was performed upon clinical request within the last 5 years. All cases were analyzed using a targeted NGS BRCAness multigene panel, including 8 HRR genes (ATM, BRCA1, BRCA2, CDK12, CHEK2, FANCA, HDAC2, PALB2). RESULTS: Our findings reveal a prevalence of potentially targetable mutations based on FDA criteria of 20.8%, which is comparable to the literature. However, the frequency of targetable BRCA2 mutations within our cohort was lower than reported for mCRPC and ATM and CHEK2 mutations were more prevalent instead. Thus, while 20.8% (n = 38) of the cases meet the criteria for olaparib treatment per FDA approval, only 4.9% (n = 9) align with the eligibility criteria according to the EMA approval. CONCLUSION: This study offers valuable real-world insights into the landscape of BRCA1/2 and HRR gene mutations and the practical clinical management of HRR gene testing in prostate cancer, contributing to a better understanding of patient eligibility for PARPi treatment.

Pathology skills lab: use of macroscopic tumor models in pathology teaching.

BACKGROUND: The shortage of pathologists in Germany, coupled with an aging workforce, requires innovative approaches to attract medical students to the field. Medical education must address different learning styles to ensure that all students are successful. METHODS: The pilot project "Practical Pathology" aims to enhance students' understanding of pathology by providing hands-on experience in macroscopic gross analysis through the use of tumor dummies built from scratch. RESULTS: An evaluation survey, completed by 63 participating students provided positive feedback on the course methodology, its relevance to understanding the pathology workflow, and its improvement over traditional teaching methods. The majority of students recognized the importance of hands-on training in medical education. Students with previous work experience rated the impact of the course on knowledge acquisition even more positively. CONCLUSION: The course improved students' understanding of pathological processes and potential sources of clinical-pathological misunderstanding. An increase in motivation for a potential career in the field of pathology was observed in a minority of students, although this exceeded the percentage of pathologists in the total medical workforce.

The Different Immune Profiles of Normal Colonic Mucosa in Cancer-Free Lynch Syndrome Carriers and Lynch Syndrome Colorectal Cancer Patients.

BACKGROUND & AIMS: Owing to the high load of immunogenic frameshift neoantigens, tumors arising in individuals with Lynch syndrome (LS), the most common inherited colorectal cancer (CRC) syndrome, are characterized by a pronounced immune infiltration. However, the immune status of normal colorectal mucosa in LS is not well characterized. We assessed the immune infiltrate in tumor-distant normal colorectal mucosa from LS CRC patients, sporadic microsatellite-unstable (MSI) and microsatellite-stable (MSS) CRC patients, and cancer-free LS carriers. METHODS: CD3-positive, FOXP3-positive, and CD8-positive T cells were quantified in, respectively, 219, 233, and 201 formalin-fixed paraffin-embedded (FFPE) normal colonic mucosa tissue sections from CRC patients and cancer-free LS carriers and 26, 22, and 19 LS CRCs. CD3-positive T cells were also quantified in an independent cohort of 97 FFPE normal rectal mucosa tissue sections from LS carriers enrolled in the CAPP2 clinical trial. The expression of 770 immune-relevant genes was analyzed in a subset of samples with the use of the NanoString nCounter platform. RESULTS: LS normal mucosa specimens showed significantly elevated CD3-, FOXP3-, and CD8-positive T-cell densities compared with non-LS control specimens. Gene expression profiling and cluster analysis revealed distinct immune profiles in LS carrier mucosa with and without cancer manifestation. Long-term follow-up of LS carriers within the CAPP2 trial found a correlation between mucosal T-cell infiltrate and time to subsequent tumor occurrence. CONCLUSIONS: LS carriers show elevated mucosal T-cell infiltration even in the absence of cancer. The normal mucosa immune profile may be a temporary or permanent tumor risk modifier in LS carriers.

[Molecular pathology of urogenital tumors : Recommendations from the 2019 International Society of Urological Pathology (ISUP) Consensus Conference]. / Molekularpathologie bei urologischen Tumoren : Empfehlungen der Konsenskonferenz der Internationalen Gesellschaft für Uropathologie (ISUP) 2019.

Comprehensive understanding of molecular principles in cancer and the diversification of oncological therapy promise individual therapeutic concepts, which have not yet found their way into urogenital cancer therapy. In March 2019 the International Society of Urogenital Pathology (ISUP) therefore held a consensus conference on recommendations for molecular diagnostics of genitourinary tumors, which were published in five separate manuscripts and are summarized in this article.In preparation for the conference, a comprehensive survey of current practices for molecular testing of urogenital tumors was carried out by members of the ISUP. At the conference, the results and the corresponding background information were presented by five working groups and recommendations for action for diagnostics were developed. An agreement between 66% of the conference participants was defined as consensus.

Prospective selective embedding of radical prostatectomy specimens is not inferior to full embedding regarding established and new prognostic parameters.

The histopathological examination of radical prostatectomy specimens is essential for assessing critical tumor characteristics, including stage, grade, and margins, all of which impact patient prognosis. However, the extent of embedding the prostate has long been a subject of debate, with some advocating partial/selective embedding and others favoring complete embedding. This study establishes a standardized and time-efficient protocol for processing radical prostatectomy specimens with limited embedding while maintaining diagnostic accuracy. Two hundred twenty-six prostatectomy specimens were analyzed, and the results of a highly standardized selective embedding protocol, systematically embedding the apex, the base, the transition to the seminal vesicles, and selected horizontal sections, were compared with full embedding as the gold standard. Non-inferiority testing was conducted by one-sided binomial tests and Pearson-Clopper confidence intervals. Selective embedding provided consistent and accurate diagnostic information with up to 90-98% concordance in pT, margins, ISUP-grade groups, and presence of IDC-P and cribriform tumor growth. In summary, this study establishes an economical standardized protocol for selective embedding of radical prostatectomy specimens with only minimal loss of information.

Peer-to-peer validation of Ki-67 scoring in a pathology quality circle as a tool to assess interobserver variability: are we better than we thought?

Ki-67, a nuclear protein expressed in all stages of cellular proliferation, is a valuable tool to assess tumor proliferation and has been linked to more aggressive tumor behavior. However, interlaboratory staining heterogeneity and inter-observer variability challenge its reproducibility. Round Robin tests are a suitable tool to standardize and harmonize immunohistochemical and molecular analyses in histopathology. The study investigates the interrater and interlaboratory reproducibility of Ki-67-scoring using both manual and automated approaches. Unstained TMA slides comprising diverse tumor types (breast cancer, neuroendocrine tumors, lymphomas, and head and neck squamous cell carcinoma) were distributed to six pathology laboratories, each employing their routine staining protocols. Manual and automated scoring methods were applied, and interrater and interlaboratory agreement assessed using intraclass correlation coefficients (ICC). The results highlight good-to-excellent reliability overall, with automated scoring demonstrating higher consistency (ICC 0.955) than manual scoring (ICC 0.871). Results were more variable when looking at the individual entities. Reliability remained good for lymphomas (ICC 0.878) and breast cancer (ICC 0.784) and was poor in well-differentiated neuroendocrine tumors (ICC 0.354). This study clearly advocates standardized practices and training to ensure consistency in Ki-67-assessment, and it demonstrates that this can be achieved in a peer-to-peer approach in local quality-circles.

Clinical implications of AGR2 in primary prostate cancer: Results from a large-scale study.

Human anterior gradient-2 (AGR2) has been implicated in carcinogenesis of various solid tumours, but the expression data in prostate cancer are contradictory regarding its prognostic value. The objective of this study is to evaluate the expression of AGR2 in a large prostate cancer cohort and to correlate it with clinicopathological data. AGR2 protein expression was analysed immunohistochemically in 1023 well-characterized prostate cancer samples with a validated antibody. AGR2 expression levels in carcinomas were compared with matched tissue samples of adjacent normal glands. AGR2 expression levels were dichotomized and tested for statistical significance. Increased AGR2 expression was found in 93.5% of prostate cancer cases. AGR2 levels were significantly higher in prostate cancer compared with normal prostate tissue. A gradual loss of AGR2 expression was associated with increasing tumour grade (ISUP), and AGR2 expression is inversely related to patient survival, however, multivariable significance is not achieved. AGR2 is clearly upregulated in the majority of prostate cancer cases, yet a true diagnostic value appears unlikely. In spite of the negative correlation of AGR2 expression with increasing tumour grade, no independent prognostic significance was found in this large-scale study.

Real-time use of artificial intelligence (CADEYE) in colorectal cancer surveillance of patients with Lynch syndrome-A randomized controlled pilot trial (CADLY).

BACKGROUND: Lynch syndrome (LS), an autosomal dominant disorder caused by pathogenic germline variants in DNA mismatch repair (MMR) genes, represents the most common hereditary colorectal cancer (CRC) syndrome. Lynch syndrome patients are at high risk of CRC despite regular endoscopic surveillance. OBJECTIVE: Our aim was to investigate the diagnostic performance of artificial intelligence (AI)-assisted colonoscopy in comparison to High-Definition white-light endoscopy (HD-WLE) for the first time. METHODS: Patients ≥18 years with LS, with a pathogenic germline variant (MLH1, MHS2, MSH6), and at least one previous colonoscopy (interval 10-36 months) were eligible. Patients were stratified by previous CRC and affected MMR gene with a 1:1 allocation ratio (AI-assisted vs. HD white-light endoscopy) in this exploratory pilot trial. RESULTS: Between Dec-2021 and Dec-2022, 101 LS patients were randomised and 96 patients were finally analyzed after exclusion of 5 patients due to insufficient bowel preparation. In the HD-WLE arm, adenomas were detected in 12/46 patients compared to 18/50 in the AI arm (26.1% [95% CI 14.3-41.1] vs. 36.0% [22.9-50.8]; p = 0.379). The use of AI-assisted colonoscopy especially increased detection of flat adenomas (Paris classification 0-IIb) (examinations with detected flat adenomas: 3/46 [6.5%] vs. 10/50 [20%]; p = 0.07; numbers of detected flat adenomas: 4/20 vs. 17/30, p = 0.018). The median withdrawal time did not differ significantly between HD-WLE and AI (14 vs. 15 min; p = 0.170). CONCLUSION: We here present first data suggesting that real-time AI-assisted colonoscopy is a promising approach to optimize endoscopic surveillance in LS patients, in particular to improve the detection of flat adenomas.

CDCP1 expression is frequently increased in aggressive urothelial carcinoma and promotes urothelial tumor progression.

The prognosis of patients with advanced urothelial carcinoma (UC) remains poor and improving treatment continues to be a major medical need. CUB domain containing protein 1 (CDCP1) is a known oncogene in various types of solid cancers and its overexpression is associated with impaired prognosis. However, its role in UC remains undetermined. Here we assessed the clinical relevance of CDCP1 in two cohorts of UC at different stages of the disease. Immunohistochemistry showed that CDCP1 is highly expressed in advanced UC, which significantly correlates with shorter overall survival. Importantly, the basal/squamous UC subtype showed significantly enriched CDCP1 at the mRNA and protein levels. The functional role of CDCP1 overexpression was assessed taking advantage of ex vivo organoids derived from the CDCP1pcLSL/+ transgenic mouse model. Furthermore, CDCP1 knockout UC cell lines were generated using CRISPR/Cas9 technology. Interestingly, CDCP1 overexpression significantly induced the activation of MAPK/ERK pathways in ex vivo organoids and increased their proliferation. Similarly, CDCP1 knockout in UC cell lines reduced their proliferation and migration, concomitant with MAPK/ERK pathway activity reduction. Our results highlight the relevance of CDCP1 in advanced UC and demonstrate its oncogenic role, suggesting that targeting CDCP1 could be a rational therapeutic strategy for the treatment of advanced UC.

Identification of Embryonic Neural Plate Border Stem Cells and Their Generation by Direct Reprogramming from Adult Human Blood Cells.

We report the direct reprogramming of both adult human fibroblasts and blood cells into induced neural plate border stem cells (iNBSCs) by ectopic expression of four neural transcription factors. Self-renewing, clonal iNBSCs can be robustly expanded in defined media while retaining multilineage differentiation potential. They generate functional cell types of neural crest and CNS lineages and could be used to model a human pain syndrome via gene editing of SCN9A in iNBSCs. NBSCs can also be derived from human pluripotent stem cells and share functional and molecular features with NBSCs isolated from embryonic day 8.5 (E8.5) mouse neural folds. Single-cell RNA sequencing identified the anterior hindbrain as the origin of mouse NBSCs, with human iNBSCs sharing a similar regional identity. In summary, we identify embryonic NBSCs and report their generation by direct reprogramming in human, which may facilitate insights into neural development and provide a neural stem cell source for applications in regenerative medicine.

Characterization of a novel cell penetrating peptide derived from human Oct4.

BACKGROUND: Oct4 is a transcription factor that plays a major role for the preservation of the pluripotent state in embryonic stem cells as well as for efficient reprogramming of somatic cells to induced pluripotent stem cells (iPSC) or other progenitors. Protein-based reprogramming methods mainly rely on the addition of a fused cell penetrating peptide. This study describes that Oct4 inherently carries a protein transduction domain, which can translocate into human and mouse cells. RESULTS: A 16 amino acid peptide representing the third helix of the human Oct4 homeodomain, referred to as Oct4 protein transduction domain (Oct4-PTD), can internalize in mammalian cells upon conjugation to a fluorescence moiety thereby acting as a cell penetrating peptide (CPP). The cellular distribution of Oct4-PTD shows diffuse cytosolic and nuclear staining, whereas penetratin is strictly localized to a punctuate pattern in the cytoplasm. By using a Cre/loxP-based reporter system, we show that this peptide also drives translocation of a functionally active Oct4-PTD-Cre-fusion protein. We further provide evidence for translocation of full length Oct4 into human and mouse cell lines without the addition of any kind of cationic fusion tag. Finally, physico-chemical properties of the novel CPP are characterized, showing that in contrast to penetratin a helical structure of Oct4-PTD is only observed if the FITC label is present on the N-terminus of the peptide. CONCLUSIONS: Oct4 is a key transcription factor in stem cell research and cellular reprogramming. Since it has been shown that recombinant Oct4 fused to a cationic fusion tag can drive generation of iPSCs, our finding might contribute to further development of protein-based methods to generate iPSCs. Moreover, our data support the idea that transcription factors might be part of an alternative paracrine signalling pathway, where the proteins are transferred to neighbouring cells thereby actively changing the behaviour of the recipient cell.