Refining the ITBCC tumor budding scoring system with a "zero-budding" category in colorectal cancer.

Tumor budding scoring guidelines from the International Tumor Budding Consensus Conference (ITBCC) for colorectal cancer propose three groups: BD1 (0-4 buds/0.785 mm2), BD2 (5-9 buds/0.785 mm2), and BD3 (10 or more buds/0.785 mm2). Here, we investigate whether a fourth scoring category, namely zero buds, may have additional clinical relevance. The number of tumor buds/0.785 mm2 was scored in 959 cases. Those with zero tumor buds were considered BD0, while a new BD1 category of 1-4 buds was proposed. Associations of both scoring approaches with clinicopathological features were analyzed. Conventional ITBCC scoring showed expected associations with unfavorable histopathological prognostic factors. In total, 111/959 (11.6%) were BD0. A significant difference was found when BD0 was compared statistically to BD1 (1-4 buds) for pT, TNM, tumor grade, and lymphatic, venous, and perineural invasion (p < 0.01, all). Tumors with BD0 occur relatively frequently and contribute additional information on tumor behavior. BD0 should be considered for subsequent ITBCC guidelines.

Classification of colorectal tissue images from high throughput tissue microarrays by ensemble deep learning methods.

Tissue microarray (TMA) core images are a treasure trove for artificial intelligence applications. However, a common problem of TMAs is multiple sectioning, which can change the content of the intended tissue core and requires re-labelling. Here, we investigate different ensemble methods for colorectal tissue classification using high-throughput TMAs. Hematoxylin and Eosin (H&E) core images of 0.6 mm or 1.0 mm diameter from three international cohorts were extracted from 54 digital slides (n = 15,150 cores). After TMA core extraction and color enhancement, five different flows of independent and ensemble deep learning were applied. Training and testing data with 2144 and 13,006 cores included three classes: tumor, normal or "other" tissue. Ground-truth data were collected from 30 ngTMA slides (n = 8689 cores). A test augmentation is applied to reduce the uncertain prediction. Predictive accuracy of the best method, namely Soft Voting Ensemble of one VGG and one CapsNet models was 0.982, 0.947 and 0.939 for normal, "other" and tumor, which outperformed to independent or ensemble learning with one base-estimator. Our high-accuracy algorithm for colorectal tissue classification in high-throughput TMAs is amenable to images from different institutions, core sizes and stain intensity. It helps to reduce error in TMA core evaluations with previously given labels.

Are tumour grade and tumour budding equivalent in colorectal cancer? A retrospective analysis of 771 patients.

BACKGROUND: Tumour grade is traditionally considered in the management of patients with colorectal cancer. However, a large body of literature suggests that a related feature, namely tumour budding, may have a more important clinical impact. The aim of our study is to determine the correlation between tumour grade and tumour budding and their impact on patient outcome. METHODS: A retrospective collective of 771 patients with colorectal cancer were included in the study. Clinicopathological information included tumour grade (World Health Organisation 2010; G1, G2 and G3) and tumour budding evaluated as BD1, BD2 and BD3 and representing 0-4 buds, 5-9 buds and 10 or more buds per 0.785 mm2, respectively. RESULTS: Tumour grade and tumour budding were correlated (p < 0.0001, percent concordance: 33.8%). Of the BD1 cases, 18.1% were of G3. Only two BD3 cases were G1. Both high tumour grade and tumour budding were associated with higher pT, lymph node metastasis, distant metastasis and lymphatic and venous vessel invasion (p < 0.01, all), but only tumour grade was additionally associated with right-sided tumour location and mucinous histology. Higher tumour budding led to worse overall (p = 0.0286) and disease-free survival (p = 0.001), but tumour grade did not. Budding was independent of tumour grade in multivariate analysis. DISCUSSION: Tumour grade and tumour budding are distinct features, as recognised by their different clinicopathological associations, reflecting different underlying biological processes. Nonetheless, tumour budding seems to outperform tumour grade in terms of predicting disease-free survival.

Tumour budding is associated with the mesenchymal colon cancer subtype and RAS/RAF mutations: a study of 1320 colorectal cancers with Consensus Molecular Subgroup (CMS) data.

BACKGROUND: Tumour budding is an important prognostic factor in colorectal cancer (CRC). Molecular profiling of tumour buds suggests (partial) epithelial-mesenchymal transition and cancer stem-cell phenotype, similarly described in the "mesenchymal" Consensus Molecular Subtype 4 (CMS4), which identifies a particularly poor prognostic subgroup. Here, we determine the association of tumour budding with CMS classification, prognosis, and response to therapy. METHODS: AMC-AJCCII-90 cohort (n = 76, stage II) was evaluated for peritumoural budding on H&E slides. LUMC (n = 270, stage I-IV), CAIRO (n = 504, metastatic CRC) and CAIRO2 (n = 472, metastatic CRC) cohorts were investigated for intratumoural budding using pan-cytokeratin-stained tissue microarrays. Budding was scored as count/area, then classified as <5 or ≥5 buds. For all cohorts, CMS classifications were available (gene-expression/immunohistochemistry-based classifiers). RESULTS: High (≥5) budding predicted a worse outcome in multivariate analysis in AMC-AJCCII-90 (p = 0.018), LUMC (p < 0.0001), and CAIRO (p = 0.03), and in CAIRO2 (continuous variable, p = 0.02). Tumour budding counts were higher in CMS4 compared to epithelial CMS2/3 cancers (p < 0.01, all), and associated with KRAS/BRAF mutations (p < 0.01, AMC-AJCCII-90, CAIRO, CAIRO2). CONCLUSION: Tumour budding is an adverse prognostic factor across all CRC stages and is associated with the mesenchymal CMS4 phenotype. KRAS/BRAF mutations are strongly correlated with tumour budding suggesting their involvement in the regulation of this process.

Comprehensive assessment of tumour budding by cytokeratin staining in colorectal cancer.

AIMS: Tumour budding in colorectal cancer (CRC) is a recognized prognostic parameter. The aim of this study was to address the use of cytokeratin immunostaining for the visualization and scoring of tumour buds. METHODS AND RESULTS: Ten hotspots (0.238 mm2 ) of peritumoural budding (PTB) and intratumoural budding (ITB) were evaluated in surgical resections from 215 patients. The budding counts in the 10 densest regions anywhere in the tumour were combined into an overall tumour budding (OTB) score. The PTB, ITB and OTB hotspot with the maximum budding count was then evaluated. Finally, continuous and cut-off values of 10 buds per high-power field (HPF) (PTB10HPF ), five buds per HPF (ITB10HPF ) and eight buds per HPF (OTB10HPF ) were used to categorize budding counts into low-grade and high-grade scores. All budding scores were highly correlated. PTB and ITB counts were associated with many clinicopathological features, including tumour stage, lymph node and distant metastasis, venous and lymphovascular invasion, and disease-free survival (DFS) (all P < 0.05). Analyses of OTB counts recapitulated these associations, including a lower DFS with a greater number of tumour buds (P = 0.0309; hazard ratio 1.0332, 95% confidence interval 1.003-1.062). One OTB hotspot performed similarly to 10 OTB hotspots in terms of relationship with outcome. These statistical significances were largely lost when cut-offs were applied to PTB, ITB or OTB counts. CONCLUSIONS: An OTB count in a single hotspot on cytokeratin-stained CRC tissue sections is a fast and reliable prognostic scoring system for the assessment of tumour budding. This approach should be considered in future studies.

VE1 immunohistochemistry predicts BRAF V600E mutation status and clinical outcome in colorectal cancer.

AIM: VE1 is a monoclonal antibody detecting mutant BRAFV(600E) protein by immunohistochemistry. Here we aim to determine the inter-observer agreement and concordance of VE1 with mutational status, investigate heterogeneity in colorectal cancers and metastases and determine the prognostic effect of VE1 in colorectal cancer patients. METHODS: Concordance of VE1 with mutational status and inter-observer agreement were tested on a pilot cohort of colorectal cancers (n = 34), melanomas (n = 23) and thyroid cancers (n = 8). Two prognostic cohorts were evaluated (n = 259, Cohort 1 and n = 226, Cohort 2) by multiple-punch tissue microarrays. VE1 staining on preoperative biopsies (n = 118 patients) was compared to expression in resections. Primary tumors and metastases from 13 patients were tested for VE1 heterogeneity using a tissue microarray generated from all available blocks (n = 100 blocks). RESULTS: Inter-observer agreement was 100% (kappa = 1.0). Concordance between VE1 and V600E mutation was 98.5%. Cohort 1: VE1 positivity (seen in 13.5%) was associated with older age (p = 0.0175) and MLH1 deficiency (p < 0.0001). Cohort 2: VE1 positivity (seen in 12.8%) was associated with female gender (p = 0.0016), right-sided tumor location (p < 0.0001), higher tumor grade (p < 0.0001) and mismatch repair (MMR)-deficiency (p < 0.0001). In survival analysis, MMR status and postoperative therapy were identified as possible confounding factors. Adjusting for these features, VE1 was an unfavorable prognostic factor. Preoperative biopsy staining matched resections in all cases except one. No heterogeneity was found across any primary/metastatic tumor blocks. CONCLUSION: VE1 is highly concordant for V600E and homogeneously expressed suggesting staining can be analysed on resection specimens, preoperative biopsies, metastatic lesions and tissue microarrays.

Fostering disability-inclusive HIV/AIDS programs in northeast India: a participatory study.

BACKGROUND: Manipur and Nagaland in northeast India are among the Indian states with the highest prevalence of HIV. Most prevention and care programs focus on identified "high risk" groups, but recent data suggest the epidemic is increasing among the general population, primarily through heterosexual sex. People with disability (PWD) in India are more likely than the general population to be illiterate, unemployed and impoverished, but little is known of their HIV risk. METHODS: This project aimed to enable HIV programs in Manipur and Nagaland to be more disability-inclusive. The objectives were to: explore HIV risk and risk perception in relation to PWD among HIV and disability programmers, and PWD themselves; identify HIV-related education and service needs and preferences of PWD; and utilise findings and stakeholder consultation to draft practical guidelines for inclusion of disability into HIV programming. Data were collected through a survey and several qualitative tools. RESULTS: The findings revealed that participants believe PWD in these states are potentially vulnerable to HIV transmission due to social exclusion and poverty, lack of knowledge, gender norms and obstacles to accessing HIV programs. Neither HIV nor disability organisations currently address the risks, needs and preferences of PWD. CONCLUSION: The Guidelines produced in the project and disseminated to stakeholders emphasise opportunities for taking action with minimal cost and resources, such as using the networks and expertise of both HIV and disability sectors, producing HIV material in a variety of formats, and promoting accessibility to mainstream HIV education and services. The human rights obligations and public health benefits of modifying national and state policies and programs to assist this highly disadvantaged population are also highlighted.