Automated assessment of Ki-67 proliferation index in neuroendocrine tumors by deep learning

The Ki-67 proliferation index (PI) is a prognostic factor in neuroendocrine tumors (NETs) and defines tumor grade. Analysis of Ki-67 PI requires calculation of Ki-67-positive and Ki-67-negative tumor cells, which is highly subjective. To overcome this, we developed a deep learning-based Ki-67 PI algorithm (KAI) that objectively calculates Ki-67 PI. Our study material consisted of NETs divided into training (n = 39), testing (n = 124), and validation (n = 60) series. All slides were digitized and processed in the Aiforia(R) Create (Aiforia Technologies, Helsinki, Finland) platform. The ICC between the pathologists and the KAI was 0.89. In 46% of the tumors, the Ki-67 PIs calculated by the pathologists and the KAI were the same. In 12% of the tumors, the Ki-67 PI calculated by the KAI was 1% lower and in 42% of the tumors on average 3% higher. The DL-based Ki-67 PI algorithm yields results similar to human observers. While the algorithm cannot replace the pathologist, it can assist in the laborious Ki-67 PI assessment of NETs. In the future, this approach could be useful in, for example, multi-center clinical trials where objective estimation of Ki-67 PI is crucial.Peer reviewe

Artificial intelligence-based image analysis can predict outcome in high-grade serous carcinoma via histology alone

High-grade extrauterine serous carcinoma (HGSC) is an aggressive tumor with high rates of recurrence, frequent chemotherapy resistance, and overall 5-year survival of less than 50%. Beyond determining and confirming the diagnosis itself, pathologist review of histologic slides provides no prognostic or predictive information, which is in sharp contrast to almost all other carcinoma types. Deep-learning based image analysis has recently been able to predict outcome and/or identify morphology-based representations of underlying molecular alterations in other tumor types, such as colorectal carcinoma, lung carcinoma, breast carcinoma, and melanoma. Using a carefully stratified HGSC patient cohort consisting of women (n = 30) with similar presentations who experienced very different treatment responses (platinum free intervals of either = 18 months), we used whole slide images (WSI, n = 205) to train a convolutional neural network. The neural network was trained, in three steps, to identify morphologic regions (digital biomarkers) that are highly associating with one or the other treatment response group. We tested the classifier using a separate 22 slide test set, and 18/22 slides were correctly classified. We show that a neural network based approach can discriminate extremes in patient response to primary platinum-based chemotherapy with high sensitivity (73%) and specificity (91%). These proof-of-concept results are novel, because for the first time, prospective prognostic information is identified specifically within HGSC tumor morphology.Peer reviewe

Automated image analysis of keratin 7 staining can predict disease outcome in primary sclerosing cholangitis

Background and AimsPrimary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease that obstructs the bile ducts and causes liver cirrhosis and cholangiocarcinoma. Efficient surrogate markers are required to measure disease progression. The cytokeratin 7 (K7) load in a liver specimen is an independent prognostic indicator that can be measured from digitalized slides using artificial intelligence (AI)-based models. MethodsA K7-AI model 2.0 was built to measure the hepatocellular K7 load area of the parenchyma, portal tracts, and biliary epithelium. K7-stained PSC liver biopsy specimens (n = 295) were analyzed. A compound endpoint (liver transplantation, liver-related death, and cholangiocarcinoma) was applied in Kaplan-Meier survival analysis to measure AUC values and positive likelihood ratios for each histological variable detected by the model. ResultsThe K7-AI model 2.0 was a better prognostic tool than plasma alkaline phosphatase, the fibrosis stage evaluated by Nakanuma classification, or K7 score evaluated by a pathologist based on the AUC values of measured variables. A combination of parameters, such as portal tract volume and area of K7-positive hepatocytes analyzed by the model, produced an AUC of 0.81 for predicting the compound endpoint. Portal tract volume measured by the model correlated with the histological fibrosis stage. ConclusionsThe K7 staining of histological liver specimens in PSC provides significant information on disease outcomes through objective and reproducible data, including variables that cannot be measured by a human pathologist. The K7-AI model 2.0 could serve as a prognostic tool for clinical endpoints and as a surrogate marker in drug trials.Peer reviewe

Chronic cholestasis detection by a novel tool : automated analysis of cytokeratin 7-stained liver specimens

Background The objective was to build a novel method for automated image analysis to locate and quantify the number of cytokeratin 7 (K7)-positive hepatocytes reflecting cholestasis by applying deep learning neural networks (AI model) in a cohort of 210 liver specimens. We aimed to study the correlation between the AI model's results and disease progression. The cohort of liver biopsies which served as a model of chronic cholestatic liver disease comprised of patients diagnosed with primary sclerosing cholangitis (PSC). Methods In a cohort of patients with PSC identified from the PSC registry of the University Hospital of Helsinki, their K7-stained liver biopsy specimens were scored by a pathologist (human K7 score) and then digitally analyzed for K7-positive hepatocytes (K7%area). The digital analysis was by a K7-AI model created in an Aiforia Technologies cloud platform. For validation, values were human K7 score, stage of disease (Metavir and Nakunuma fibrosis score), and plasma liver enzymes indicating clinical cholestasis, all subjected to correlation analysis. Results The K7-AI model results (K7%area) correlated with the human K7 score (0.896; p < 2.2e(- 16)). In addition, K7%area correlated with stage of PSC (Metavir 0.446; p < 1.849e(- 10) and Nakanuma 0.424; p < 4.23e(- 10)) and with plasma alkaline phosphatase (P-ALP) levels (0.369, p < 5.749e(- 5)). Conclusions The accuracy of the AI-based analysis was comparable to that of the human K7 score. Automated quantitative image analysis correlated with stage of PSC and with P-ALP. Based on the results of the K7-AI model, we recommend K7 staining in the assessment of cholestasis by means of automated methods that provide fast (9.75 s/specimen) quantitative analysis.Peer reviewe

AI Model for Prostate Biopsies Predicts Cancer Survival

An artificial intelligence (AI) algorithm for prostate cancer detection and grading was developed for clinical diagnostics on biopsies. The study cohort included 4221 scanned slides from 872 biopsy sessions at the HUS Helsinki University Hospital during 2016–2017 and a subcohort of 126 patients treated by robot-assisted radical prostatectomy (RALP) during 2016–2019. In the validation cohort (n = 391), the model detected cancer with a sensitivity of 98% and specificity of 98% (weighted kappa 0.96 compared with the pathologist’s diagnosis). Algorithm-based detection of the grade area recapitulated the pathologist’s grade group. The area of AI-detected cancer was associated with extra-prostatic extension (G5 OR: 48.52; 95% CI 1.11–8.33), seminal vesicle invasion (cribriform G4 OR: 2.46; 95% CI 0.15–1.7; G5 OR: 5.58; 95% CI 0.45–3.42), and lymph node involvement (cribriform G4 OR: 2.66; 95% CI 0.2–1.8; G5 OR: 4.09; 95% CI 0.22–3). Algorithm-detected grade group 3–5 prostate cancer depicted increased risk for biochemical recurrence compared with grade groups 1–2 (HR: 5.91; 95% CI 1.96–17.83). This study showed that a deep learning model not only can find and grade prostate cancer on biopsies comparably with pathologists but also can predict adverse staging and probability for recurrence after surgical treatment

AI Model for Prostate Biopsies Predicts Cancer Survival

An artificial intelligence (AI) algorithm for prostate cancer detection and grading was developed for clinical diagnostics on biopsies. The study cohort included 4221 scanned slides from 872 biopsy sessions at the HUS Helsinki University Hospital during 2016–2017 and a subcohort of 126 patients treated by robot-assisted radical prostatectomy (RALP) during 2016–2019. In the validation cohort (n = 391), the model detected cancer with a sensitivity of 98% and specificity of 98% (weighted kappa 0.96 compared with the pathologist’s diagnosis). Algorithm-based detection of the grade area recapitulated the pathologist’s grade group. The area of AI-detected cancer was associated with extra-prostatic extension (G5 OR: 48.52; 95% CI 1.11–8.33), seminal vesicle invasion (cribriform G4 OR: 2.46; 95% CI 0.15–1.7; G5 OR: 5.58; 95% CI 0.45–3.42), and lymph node involvement (cribriform G4 OR: 2.66; 95% CI 0.2–1.8; G5 OR: 4.09; 95% CI 0.22–3). Algorithm-detected grade group 3–5 prostate cancer depicted increased risk for biochemical recurrence compared with grade groups 1–2 (HR: 5.91; 95% CI 1.96–17.83). This study showed that a deep learning model not only can find and grade prostate cancer on biopsies comparably with pathologists but also can predict adverse staging and probability for recurrence after surgical treatment

Systems pathology by multiplexed immunohistochemistry and whole-slide digital image analysis

The paradigm of molecular histopathology is shifting from a single-marker immunohistochemistry towards multiplexed detection of markers to better understand the complex pathological processes. However, there are no systems allowing multiplexed IHC (mIHC) with high-resolution whole-slide tissue imaging and analysis, yet providing feasible throughput for routine use. We present an mIHC platform combining fluorescent and chromogenic staining with automated whole-slide imaging and integrated whole-slide image analysis, enabling simultaneous detection of six protein markers and nuclei, and automatic quantification and classification of hundreds of thousands of cells in situ in formalin-fixed paraffin-embedded tissues. In the first proof-of-concept, we detected immune cells at cell-level resolution (n = 128,894 cells) in human prostate cancer, and analysed T cell subpopulations in different tumour compartments (epithelium vs. stroma). In the second proof-of-concept, we demonstrated an automatic classification of epithelial cell populations (n = 83,558) and glands (benign vs. cancer) in prostate cancer with simultaneous analysis of androgen receptor (AR) and alpha-methylacyl-CoA (AMACR) expression at cell-level resolution. We conclude that the open-source combination of 8-plex mIHC detection, whole-slide image acquisition and analysis provides a robust tool allowing quantitative, spatially resolved whole-slide tissue cytometry directly in formalin-fixed human tumour tissues for improved characterization of histology and the tumour microenvironment.Peer reviewe

Implementation of deep neural networks to count dopamine neurons in substantia nigra

Unbiased estimates of neuron numbers within substantia nigra are crucial for experimental Parkinson's disease models and gene-function studies. Unbiased stereological counting techniques with optical fractionation are successfully implemented, but are extremely laborious and time-consuming. The development of neural networks and deep learning has opened a new way to teach computers to count neurons. Implementation of a programming paradigm enables a computer to learn from the data and development of an automated cell counting method. The advantages of computerized counting are reproducibility, elimination of human error and fast high-capacity analysis. We implemented whole-slide digital imaging and deep convolutional neural networks (CNN) to count substantia nigra dopamine neurons. We compared the results of the developed method against independent manual counting by human observers and validated the CNN algorithm against previously published data in rats and mice, where tyrosine hydroxylase (TH)-immunoreactive neurons were counted using unbiased stereology. The developed CNN algorithm and fully cloud-embedded Aiforia (TM) platform provide robust and fast analysis of dopamine neurons in rat and mouse substantia nigra.Peer reviewe

Combined epithelial marker analysis of tumour budding in stage II colorectal cancer

Tumour budding predicts survival of stage II colorectal cancer (CRC) and has been suggested to be associated with epithelial-to-mesenchymal transition (EMT). However, the underlying molecular changes of tumour budding remain poorly understood. Here, we performed multiplex immunohistochemistry (mIHC) to phenotypically profile tumours using known EMT-associated markers: E-cadherin (adherence junctions), integrin beta 4 (ITGB4; basement membrane), ZO-1 (tight junctions), and pan-cytokeratin. A subpopulation of patients showed high ITGB4 expression in tumour buds, and this coincided with a switch of ITGB4 localisation from the basal membrane of intact epithelium to the cytoplasm of budding cells. Digital image analysis demonstrated that tumour budding with high ITGB4 expression in tissue microarray (TMA) cores correlated with tumour budding assessed from haematoxylin and eosin (H&E) whole sections and independently predicted poor disease-specific survival in two independent stage II CRC cohorts (hazard ratio [HR] = 4.50 (95% confidence interval [CI] = 1.50-13.5), n = 232; HR = 3.52 (95% CI = 1.30-9.53), n = 72). Furthermore, digitally obtained ITGB4-high bud count in random TMA cores was better associated with survival outcome than visual tumour bud count in corresponding H&E-stained samples. In summary, the mIHC-based phenotypic profiling of human tumour tissue shows strong potential for the molecular characterisation of tumour biology and for the discovery of novel prognostic biomarkers.Peer reviewe