Beatrice S. Knudsen

Shikha Dubey, Tushar Kataria, Beatrice Knudsen et al.

With the advent of digital scanners and deep learning, diagnostic operations may move from a microscope to a desktop. Hematoxylin and Eosin (H&E) staining is one of the most frequently used stains for disease analysis, diagnosis, and grading, but pathologists do need different immunohistochemical (IHC) stains to analyze specific structures or cells. Obtaining all of these stains (H&E and different IHCs) on a single specimen is a tedious and time-consuming task. Consequently, virtual staining has emerged as an essential research direction. Here, we propose a novel generative model, Structural Cycle-GAN (SC-GAN), for synthesizing IHC stains from H&E images, and vice versa. Our method expressly incorporates structural information in the form of edges (in addition to color data) and employs attention modules exclusively in the decoder of the proposed generator model. This integration enhances feature localization and preserves contextual information during the generation process. In addition, a structural loss is incorporated to ensure accurate structure alignment between the generated and input markers. To demonstrate the efficacy of the proposed model, experiments are conducted with two IHC markers emphasizing distinct structures of glands in the colon: the nucleus of epithelial cells (CDX2) and the cytoplasm (CK818). Quantitative metrics such as FID and SSIM are frequently used for the analysis of generative models, but they do not correlate explicitly with higher-quality virtual staining results. Therefore, we propose two new quantitative metrics that correlate directly with the virtual staining specificity of IHC markers.

Tushar Kataria, Beatrice Knudsen, Shireen Elhabian

Annotating medical imaging datasets is costly, so fine-tuning (or transfer learning) is the most effective method for digital pathology vision applications such as disease classification and semantic segmentation. However, due to texture bias in models trained on real-world images, transfer learning for histopathology applications might result in underperforming models, which necessitates the need for using unlabeled histopathology data and self-supervised methods to discover domain-specific characteristics. Here, we tested the premise that histopathology-specific pretrained models provide better initializations for pathology vision tasks, i.e., gland and cell segmentation. In this study, we compare the performance of gland and cell segmentation tasks with histopathology domain-specific and non-domain-specific (real-world images) pretrained weights. Moreover, we investigate the dataset size at which domain-specific pretraining produces significant gains in performance. In addition, we investigated whether domain-specific initialization improves the effectiveness of out-of-distribution testing on distinct datasets but the same task. The results indicate that performance gain using domain-specific pretrained weights depends on both the task and the size of the training dataset. In instances with limited dataset sizes, a significant improvement in gland segmentation performance was also observed, whereas models trained on cell segmentation datasets exhibit no improvement.

Bodong Zhang, Beatrice Knudsen, Deepika Sirohi et al.

We propose a novel semi-supervised learning approach for classification of histopathology images. We employ strong supervision with patch-level annotations combined with a novel co-training loss to create a semi-supervised learning framework. Co-training relies on multiple conditionally independent and sufficient views of the data. We separate the hematoxylin and eosin channels in pathology images using color deconvolution to create two views of each slide that can partially fulfill these requirements. Two separate CNNs are used to embed the two views into a joint feature space. We use a contrastive loss between the views in this feature space to implement co-training. We evaluate our approach in clear cell renal cell and prostate carcinomas, and demonstrate improvement over state-of-the-art semi-supervised learning methods.

Xiaoya Tang, Bodong Zhang, Beatrice S. Knudsen et al.

We here propose a novel hierarchical transformer model that adeptly integrates the feature extraction capabilities of Convolutional Neural Networks (CNNs) with the advanced representational potential of Vision Transformers (ViTs). Addressing the lack of inductive biases and dependence on extensive training datasets in ViTs, our model employs a CNN backbone to generate hierarchical visual representations. These representations are then adapted for transformer input through an innovative patch tokenization. We also introduce a 'scale attention' mechanism that captures cross-scale dependencies, complementing patch attention to enhance spatial understanding and preserve global perception. Our approach significantly outperforms baseline models on small and medium-sized medical datasets, demonstrating its efficiency and generalizability. The components are designed as plug-and-play for different CNN architectures and can be adapted for multiple applications. The code is available at https://github.com/xiaoyatang/DuoFormer.git.

Shikha Dubey, Yosep Chong, Beatrice Knudsen et al.

This paper introduces a Virtual Immunohistochemistry Multiplex staining (VIMs) model designed to generate multiple immunohistochemistry (IHC) stains from a single hematoxylin and eosin (H&E) stained tissue section. IHC stains are crucial in pathology practice for resolving complex diagnostic questions and guiding patient treatment decisions. While commercial laboratories offer a wide array of up to 400 different antibody-based IHC stains, small biopsies often lack sufficient tissue for multiple stains while preserving material for subsequent molecular testing. This highlights the need for virtual IHC staining. Notably, VIMs is the first model to address this need, leveraging a large vision-language single-step diffusion model for virtual IHC multiplexing through text prompts for each IHC marker. VIMs is trained on uniplex paired H&E and IHC images, employing an adversarial training module. Testing of VIMs includes both paired and unpaired image sets. To enhance computational efficiency, VIMs utilizes a pre-trained large latent diffusion model fine-tuned with small, trainable weights through the Low-Rank Adapter (LoRA) approach. Experiments on nuclear and cytoplasmic IHC markers demonstrate that VIMs outperforms the base diffusion model and achieves performance comparable to Pix2Pix, a standard generative model for paired image translation. Multiple evaluation methods, including assessments by two pathologists, are used to determine the performance of VIMs. Additionally, experiments with different prompts highlight the impact of text conditioning. This paper represents the first attempt to accelerate histopathology research by demonstrating the generation of multiple IHC stains from a single H&E input using a single model trained solely on uniplex data.

Tushar Kataria, Shikha Dubey, Mary Bronner et al.

Deep learning models can generate virtual immunohistochemistry (IHC) stains from hematoxylin and eosin (H&E) images, offering a scalable and low-cost alternative to laboratory IHC. However, reliable evaluation of image quality remains a challenge as current texture- and distribution-based metrics quantify image fidelity rather than the accuracy of IHC staining. Here, we introduce an automated and accuracy grounded framework to determine image quality across sixteen paired or unpaired image translation models. Using color deconvolution, we generate masks of pixels stained brown (i.e., IHC-positive) as predicted by each virtual IHC model. We use the segmented masks of real and virtual IHC to compute stain accuracy metrics (Dice, IoU, Hausdorff distance) that directly quantify correct pixel - level labeling without needing expert manual annotations. Our results demonstrate that conventional image fidelity metrics, including Frechet Inception Distance (FID), peak signal-to-noise ratio (PSNR), and structural similarity (SSIM), correlate poorly with stain accuracy and pathologist assessment. Paired models such as PyramidPix2Pix and AdaptiveNCE achieve the highest stain accuracy, whereas unpaired diffusion- and GAN-based models are less reliable in providing accurate IHC positive pixel labels. Moreover, whole-slide images (WSI) reveal performance declines that are invisible in patch-based evaluations, emphasizing the need for WSI-level benchmarks. Together, this framework defines a reproducible approach for assessing the quality of virtual IHC models, a critical step to accelerate translation towards routine use by pathologists.

Bodong Zhang, Hamid Manoochehri, Man Minh Ho et al.

Histopathological image classification is an important task in medical image analysis. Recent approaches generally rely on weakly supervised learning due to the ease of acquiring case-level labels from pathology reports. However, patch-level classification is preferable in applications where only a limited number of cases are available or when local prediction accuracy is critical. On the other hand, acquiring extensive datasets with localized labels for training is not feasible. In this paper, we propose a semi-supervised patch-level histopathological image classification model, named CLASS-M, that does not require extensively labeled datasets. CLASS-M is formed by two main parts: a contrastive learning module that uses separated Hematoxylin and Eosin images generated through an adaptive stain separation process, and a module with pseudo-labels using MixUp. We compare our model with other state-of-the-art models on two clear cell renal cell carcinoma datasets. We demonstrate that our CLASS-M model has the best performance on both datasets. Our code is available at github.com/BzhangURU/Paper_CLASS-M/tree/main

Bodong Zhang, Hamid Manoochehri, Xiwen Li et al.

Weakly supervised multiple instance learning (MIL) is a challenging task given that only bag-level labels are provided, while each bag typically contains multiple instances. This topic has been extensively studied in histopathological image analysis, where labels are usually available only at the whole slide image (WSI) level, while each WSI could be divided into thousands of small image patches for training. The dominant MIL approaches focus on feature aggregation and take fixed patch features as inputs. However, weakly supervised feature representation learning in MIL settings is always neglected. Those features used to be generated by self-supervised learning methods that do not utilize weak labels, or by foundation encoders pre-trained on other large datasets. In this paper, we propose a novel weakly supervised feature representation learning method called Weakly Supervised Contrastive Learning (WeakSupCon) that utilizes bag-level labels. In our method, we employ multi-task learning and define distinct contrastive losses for samples with different bag labels. Our experiments demonstrate that the features generated using WeakSupCon with limited computing resources significantly enhance MIL classification performance compared to self-supervised approaches across three datasets. Our WeakSupCon code is available at github.com/BzhangURU/Paper_WeakSupCon

Xiaoya Tang, Bodong Zhang, Man Minh Ho et al.

Despite the widespread adoption of transformers in medical applications, the exploration of multi-scale learning through transformers remains limited, while hierarchical representations are considered advantageous for computer-aided medical diagnosis. We propose a novel hierarchical transformer model that adeptly integrates the feature extraction capabilities of Convolutional Neural Networks (CNNs) with the advanced representational potential of Vision Transformers (ViTs). Addressing the lack of inductive biases and dependence on extensive training datasets in ViTs, our model employs a CNN backbone to generate hierarchical visual representations. These representations are adapted for transformer input through an innovative patch tokenization process, preserving the inherited multi-scale inductive biases. We also introduce a scale-wise attention mechanism that directly captures intra-scale and inter-scale associations. This mechanism complements patch-wise attention by enhancing spatial understanding and preserving global perception, which we refer to as local and global attention, respectively. Our model significantly outperforms baseline models in terms of classification accuracy, demonstrating its efficiency in bridging the gap between Convolutional Neural Networks (CNNs) and Vision Transformers (ViTs). The components are designed as plug-and-play for different CNN architectures and can be adapted for multiple applications. The code is available at https://github.com/xiaoyatang/DuoFormer.git.

Tushar Kataria, Beatrice Knudsen, Shireen Y. Elhabian

Hematoxylin and Eosin (H&E) staining is widely regarded as the standard in pathology for diagnosing diseases and tracking tumor recurrence. While H&E staining shows tissue structures, it lacks the ability to reveal specific proteins that are associated with disease severity and treatment response. Immunohistochemical (IHC) stains use antibodies to highlight the expression of these proteins on their respective cell types, improving diagnostic accuracy, and assisting with drug selection for treatment. Despite their value, IHC stains require additional time and resources, limiting their utilization in some clinical settings. Recent advances in deep learning have positioned Image-to-Image (I2I) translation as a computational, cost-effective alternative for IHC. I2I generates high fidelity stain transformations digitally, potentially replacing manual staining in IHC. Diffusion models, the current state of the art in image generation and conditional tasks, are particularly well suited for virtual IHC due to their ability to produce high quality images and resilience to mode collapse. However, these models require extensive and diverse datasets (often millions of samples) to achieve a robust performance, a challenge in virtual staining applications where only thousands of samples are typically available. Inspired by the success of multitask deep learning models in scenarios with limited data, we introduce STAINDIFFUSER, a novel multitask diffusion architecture tailored to virtual staining that achieves convergence with smaller datasets. STAINDIFFUSER simultaneously trains two diffusion processes: (a) generating cell specific IHC stains from H&E images and (b) performing H&E based cell segmentation, utilizing coarse segmentation labels exclusively during training. STAINDIFFUSER generates high-quality virtual stains for two markers, outperforming over twenty I2I baselines.

Man M. Ho, Shikha Dubey, Yosep Chong et al.

The Frozen Section (FS) technique is a rapid and efficient method, taking only 15-30 minutes to prepare slides for pathologists' evaluation during surgery, enabling immediate decisions on further surgical interventions. However, FS process often introduces artifacts and distortions like folds and ice-crystal effects. In contrast, these artifacts and distortions are absent in the higher-quality formalin-fixed paraffin-embedded (FFPE) slides, which require 2-3 days to prepare. While Generative Adversarial Network (GAN)-based methods have been used to translate FS to FFPE images (F2F), they may leave morphological inaccuracies with remaining FS artifacts or introduce new artifacts, reducing the quality of these translations for clinical assessments. In this study, we benchmark recent generative models, focusing on GANs and Latent Diffusion Models (LDMs), to overcome these limitations. We introduce a novel approach that combines LDMs with Histopathology Pre-Trained Embeddings to enhance restoration of FS images. Our framework leverages LDMs conditioned by both text and pre-trained embeddings to learn meaningful features of FS and FFPE histopathology images. Through diffusion and denoising techniques, our approach not only preserves essential diagnostic attributes like color staining and tissue morphology but also proposes an embedding translation mechanism to better predict the targeted FFPE representation of input FS images. As a result, this work achieves a significant improvement in classification performance, with the Area Under the Curve rising from 81.99% to 94.64%, accompanied by an advantageous CaseFD. This work establishes a new benchmark for FS to FFPE image translation quality, promising enhanced reliability and accuracy in histopathology FS image analysis. Our work is available at https://minhmanho.github.io/f2f_ldm/.

Man M. Ho, Elham Ghelichkhan, Yosep Chong et al.

Latent Diffusion Models (LDMs) can generate high-fidelity images from noise, offering a promising approach for augmenting histopathology images for training cancer grading models. While previous works successfully generated high-fidelity histopathology images using LDMs, the generation of image tiles to improve prostate cancer grading has not yet been explored. Additionally, LDMs face challenges in accurately generating admixtures of multiple cancer grades in a tile when conditioned by a tile mask. In this study, we train specific LDMs to generate synthetic tiles that contain multiple Gleason Grades (GGs) by leveraging pixel-wise annotations in input tiles. We introduce a novel framework named Self-Distillation from Separated Conditions (DISC) that generates GG patterns guided by GG masks. Finally, we deploy a training framework for pixel-level and slide-level prostate cancer grading, where synthetic tiles are effectively utilized to improve the cancer grading performance of existing models. As a result, this work surpasses previous works in two domains: 1) our LDMs enhanced with DISC produce more accurate tiles in terms of GG patterns, and 2) our training scheme, incorporating synthetic data, significantly improves the generalization of the baseline model for prostate cancer grading, particularly in challenging cases of rare GG5, demonstrating the potential of generative models to enhance cancer grading when data is limited.

Souradeep Chakraborty, Ruoyu Xue, Rajarsi Gupta et al.

The ability to predict the attention of expert pathologists could lead to decision support systems for better pathology training. We developed methods to predict the spatio-temporal (where and when) movements of pathologists' attention as they grade whole slide images (WSIs) of prostate cancer. We characterize a pathologist's attention trajectory by their x, y, and m (magnification) movements of a viewport as they navigate WSIs using a digital microscope. This information was obtained from 43 pathologists across 123 WSIs, and we consider the task of predicting the pathologist attention scanpaths constructed from the viewport centers. We introduce a fixation extraction algorithm that simplifies an attention trajectory by extracting fixations in the pathologist's viewing while preserving semantic information, and we use these pre-processed data to train and test a two-stage model to predict the dynamic (scanpath) allocation of attention during WSI reading via intermediate attention heatmap prediction. In the first stage, a transformer-based sub-network predicts the attention heatmaps (static attention) across different magnifications. In the second stage, we predict the attention scanpath by sequentially modeling the next fixation points in an autoregressive manner using a transformer-based approach, starting at the WSI center and leveraging multi-magnification feature representations from the first stage. Experimental results show that our scanpath prediction model outperforms chance and baseline models. Tools developed from this model could assist pathology trainees in learning to allocate their attention during WSI reading like an expert.

Tushar Kataria, Beatrice Knudsen, Shireen Y. Elhabian

Hematoxylin and eosin (H&E) staining is a gold standard for microscopic diagnosis in pathology. However, H&E staining does not capture all the diagnostic information that may be needed. To obtain additional molecular information, immunohistochemical (IHC) stains highlight proteins that mark specific cell types, such as CD3 for T-cells or CK8/18 for epithelial cells. While IHC stains are vital for prognosis and treatment guidance, they are typically only available at specialized centers and time consuming to acquire, leading to treatment delays for patients. Virtual staining, enabled by deep learning-based image translation models, provides a promising alternative by computationally generating IHC stains from H&E stained images. Although many GAN and diffusion based image to image (I2I) translation methods have been used for virtual staining, these models treat image patches as independent data points, which results in increased and more diverse data requirements for effective generation. We present ImplicitStainer, a novel approach that leverages local implicit functions to improve image translation, specifically virtual staining performance, by focusing on pixel-level predictions. This method enhances robustness to variations in dataset sizes, delivering high-quality results even with limited data. We validate our approach on two datasets using a comprehensive set of metrics and benchmark it against over fifteen state-of-the-art GAN- and diffusion based models. Full Code and models trained will be released publicly via Github upon acceptance.

Hamid Manoochehri, Bodong Zhang, Beatrice S. Knudsen et al.

Self-supervised learning has become a cornerstone in various areas, particularly histopathological image analysis. Image augmentation plays a crucial role in self-supervised learning, as it generates variations in image samples. However, traditional image augmentation techniques often overlook the unique characteristics of histopathological images. In this paper, we propose a new histopathology-specific image augmentation method called stain reconstruction augmentation (SRA). We integrate our SRA with MoCo v3, a leading model in self-supervised contrastive learning, along with our additional contrastive loss terms, and call the new model SRA-MoCo v3. We demonstrate that our SRA-MoCo v3 always outperforms the standard MoCo v3 across various downstream tasks and achieves comparable or superior performance to other foundation models pre-trained on significantly larger histopathology datasets.

Tushar Kataria, Beatrice Knudsen, Shireen Elhabian

Semantic segmentation is a critical step in automated image interpretation and analysis where pixels are classified into one or more predefined semantically meaningful classes. Deep learning approaches for semantic segmentation rely on harnessing the power of annotated images to learn features indicative of these semantic classes. Nonetheless, they often fail to generalize when there is a significant domain (i.e., distributional) shift between the training (i.e., source) data and the dataset(s) encountered when deployed (i.e., target), necessitating manual annotations for the target data to achieve acceptable performance. This is especially important in medical imaging because different image modalities have significant intra- and inter-site variations due to protocol and vendor variability. Current techniques are sensitive to hyperparameter tuning and target dataset size. This paper presents an unsupervised domain adaptation approach for semantic segmentation that alleviates the need for annotating target data. Using kernel density estimation, we match the target data distribution to the source in the feature space, particularly when the number of target samples is limited (3% of the target dataset size). We demonstrate the efficacy of our proposed approach on 2 datasets, multisite prostate MRI and histopathology images.

Souradeep Chakraborty, Ke Ma, Rajarsi Gupta et al.

We study the attention of pathologists as they examine whole-slide images (WSIs) of prostate cancer tissue using a digital microscope. To the best of our knowledge, our study is the first to report in detail how pathologists navigate WSIs of prostate cancer as they accumulate information for their diagnoses. We collected slide navigation data (i.e., viewport location, magnification level, and time) from 13 pathologists in 2 groups (5 genitourinary (GU) specialists and 8 general pathologists) and generated visual attention heatmaps and scanpaths. Each pathologist examined five WSIs from the TCGA PRAD dataset, which were selected by a GU pathology specialist. We examined and analyzed the distributions of visual attention for each group of pathologists after each WSI was examined. To quantify the relationship between a pathologist's attention and evidence for cancer in the WSI, we obtained tumor annotations from a genitourinary specialist. We used these annotations to compute the overlap between the distribution of visual attention and annotated tumor region to identify strong correlations. Motivated by this analysis, we trained a deep learning model to predict visual attention on unseen WSIs. We find that the attention heatmaps predicted by our model correlate quite well with the ground truth attention heatmap and tumor annotations on a test set of 17 WSIs by using various spatial and temporal evaluation metrics.

Jiayun Li, Wenyuan Li, Arkadiusz Gertych et al.

Histology review is often used as the `gold standard' for disease diagnosis. Computer aided diagnosis tools can potentially help improve current pathology workflows by reducing examination time and interobserver variability. Previous work in cancer grading has focused mainly on classifying pre-defined regions of interest (ROIs), or relied on large amounts of fine-grained labels. In this paper, we propose a two-stage attention-based multiple instance learning model for slide-level cancer grading and weakly-supervised ROI detection and demonstrate its use in prostate cancer. Compared with existing Gleason classification models, our model goes a step further by utilizing visualized saliency maps to select informative tiles for fine-grained grade classification. The model was primarily developed on a large-scale whole slide dataset consisting of 3,521 prostate biopsy slides with only slide-level labels from 718 patients. The model achieved state-of-the-art performance for prostate cancer grading with an accuracy of 85.11\% for classifying benign, low-grade (Gleason grade 3+3 or 3+4), and high-grade (Gleason grade 4+3 or higher) slides on an independent test set.