PACpAInt: a deep learning approach to identify molecular subtypes of pancreatic adenocarcinoma on histology slides

Pancreatic ductal adenocarcinoma (PAC) is a highly heterogeneous and plastic tumor with different transcriptomic molecular subtypes that hold great prognostic and theranostic values. We developed PACpAInt, a multistep approach using deep learning models to determine tumor cell type and their molecular phenotype on routine histological preparation at a resolution enabling to decipher complete intratumor heterogeneity on a massive scale never achieved before. PACpAInt effectively identified molecular subtypes at the slide level in three validation cohorts and had an independent prognostic value. It identified an interslide heterogeneity within a case in 39% of tumors that impacted survival. Diving at the cell level, PACpAInt identified pure classical and basal-like main subtypes as well as an intermediary phenotype and hybrid tumors that co-carried both classical and basal-like phenotypes. These novel artificial intelligence-based subtypes, together with the proportion of basal- like cells within a tumor had a strong prognostic impact.

Heterogenous Profiles Between Matched Primary Tumors and Brain Metastases Reveal Tumor Evolution

Background: Brain metastases (BMs) are the most common central nervous system (CNS) malignant tumors, with rapid disease progression and extremely poor prognosis. The heterogeneity between primary tumors and BMs leads to the divergent efficacy of the adjuvant therapy response to primary tumors and BMs. However, the extent of heterogeneity between primary tumors and BMs, and the evolutionary process remains little known. Methods: To deeply insight the extent of inter-tumor heterogeneity at single-patient level and the process of these evolutions, we retrospectively analyzed a total of 26 tumor samples from 11 patients with matched primary tumors and BMs. One patient underwent four times brain metastatic lesion surgery with diverse locations and one operation for the primary lesion. The genomic and immune heterogeneity between primary tumors and BMs was evaluated by utilizing the whole-exome sequencing (WESeq) and immunohistochemical analysis.Results: In addition to inheriting genomic phenotype and molecular phenotype from the primary tumors, massive unique genomic phenotype and molecular phenotype were also observed in BMs, which revealed unimaginable complexity of tumor evolution and extensive heterogeneity among lesions at single-patient level. Our study also verified that the expression level of immune checkpoints-related molecule Programmed Death-Ligand 1 (PD-L1) (P = 0.0013) and the density of tumor-infiltrating lymphocytes (TILs) (P = 0.0248) in BMs were significantly lower than that in paired primary tumors. Additionally, tumor microvascular density (MVD) and tumor invasiveness were also differed between primary tumors and paired BMs, indicating that temporal and spatial diversity profoundly contributes to the evolution of BMs heterogeneity.Conclusion: We verified the significance of temporal and spatial factors to the evolution of tumor heterogeneity by multi-dimensional analysis of matched primary tumors and BMs, which also provided novel insight for formulating individualized treatment strategies of BMs.

scONE-seq: A one-tube single-cell multi-omics method enables simultaneous dissection of molecular phenotype and genotype heterogeneity from frozen tumors

Genomic and transcriptomic heterogeneity both play important roles in normal cellular function as well as in disease development. To be able to characterize these different forms of cellular heterogeneity in diverse sample types, we developed scONE-seq, which enables simultaneous transcriptome and genome profiling in a one-tube reaction. Previous single-cell-whole-genome-RNA-sequencing (scWGS-RNA-seq) methods require physical separation of DNA and RNA, often by physical separation of the nucleus from the cytoplasm. Most of these methods are labor-intensive and technically demanding, time-consuming, or require special devices, and they are not applicable to frozen samples that cannot generate intact single-cell suspensions. scONE-seq is a one-tube reaction which eliminates loss due to transfer steps, and thus is highly scalable and compatible with frozen biobanked tissue, generating data that is superior in quality compared to other applicable methods. We benchmarked scONE-seq against existing methods using cell lines and lymphocytes from a healthy donor, and we applied it to a 2-year-frozen astrocytoma sample profiling over 1,200 nuclei, subsequently identifying a unique transcriptionally normal-like tumor clone. scONE-seq makes it possible to perform large-scale single-cell multi-omics interrogation with ease on the vast quantities of biobanked tissue, which could transform the scale of future multi-omics single-cell cancer profiling studies.

scONE-seq: A one-tube single-cell multi-omics method enables simultaneous dissection of molecular phenotype and genotype heterogeneity from frozen tumors

Genomic and transcriptomic heterogeneity both play important roles in normal cellular function as well as in disease development. To be able to characterize these different forms of cellular heterogeneity in diverse sample types, we developed scONE-seq, which enables simultaneous transcriptome and genome profiling in a one-tube reaction. Previous single-cell-whole-genome-RNA-sequencing (scWGS-RNA-seq) methods require physical separation of DNA and RNA, often by physical separation of the nucleus from the cytoplasm. These methods are labor-intensive and technically demanding, time-consuming, or require special devices, and they are not applicable to frozen samples that cannot generate intact single-cell suspensions. scONE-seq is a one-tube reaction, thus is highly scalable and is the first scWGS-RNA-seq method compatible with frozen biobanked tissue. We benchmarked scONE-seq against existing methods using cell lines and lymphocytes from a healthy donor, and we applied it to a 2-year-frozen astrocytoma sample profiling over 1,200 nuclei, subsequently identifying a unique transcriptionally normal-like tumor clone. scONE-seq makes it possible to perform large-scale single-cell multi-omics interrogation with ease on the vast quantities of biobanked tissue, which could transform the scale of future multi-omics single-cell cancer profiling studies.

Clinical and molecular features of acquired resistance to immunotherapy in non-small cell lung cancer

Although cancer immunotherapy with PD-(L)1 blockade is now routine treatment for patients with lung cancer, remarkably little is known about acquired resistance. We examined 1,201 patients with NSCLC treated with PD-(L)1 blockade to clinically characterize acquired resistance, finding it to be common (occurring in more than 60% of initial responders), with persistent but diminishing risk over time, and with distinct metastatic and survival patterns compared to primary resistance. To examine the molecular phenotype and potential mechanisms of acquired resistance, we performed whole transcriptome and exome tumor profiling in a subset of NSCLC patients (n=29) with acquired resistance. Systematic immunogenomic analysis revealed that tumors with acquired resistance generally had enriched signals of inflammation (including IFNγ signaling and inferred CD8+ T cells) and could be separated into IFNγ upregulated and stable subsets. IFNγ upregulated tumors had putative routes of resistance with signatures of dysfunctional interferon signaling and mutations in antigen presentation genes. Transcriptomic profiling of cancer cells from a murine model of acquired resistance to PD-(L)1 blockade also showed evidence of dysfunctional interferon signaling and acquired insensitivity to in vitro interferon gamma treatment. In summary, we characterized clinical and molecular features of acquired resistance to PD-(L)1 blockade in NSCLC and found evidence of ongoing but dysfunctional IFN response. The persistently inflamed, rather than excluded or deserted, tumor microenvironment of acquired resistance informs therapeutic strategies to effectively reprogram and reverse acquired resistance.

Case Report: Sellar Ependymomas: A Clinic-Pathological Study and Literature Review

Ependymomas are primary glial tumors arising from cells related to the ependymal lining of the ventricular system. They are classified into at least nine different molecular subtypes according to molecular phenotype, histological morphology, and tumor location. Primary sellar ependymoma is an extremely rare malignancy of the central nervous system, with only 12 known cases reported in humans. We herein report a case of ependymoma located at the pituitary region in a 44-year-old female patient and discuss the molecular subtype, natural history, clinical presentation, radiological findings, histological features, immunohistochemical characteristics, ultrastructural examinations, treatment, and prognosis of sellar ependymoma. This case report may serve as a helpful reference for clinicians and radiologists in clinical practice.

Neue Krankheiten mit Bluttranskriptomik entschlüsseln

The COVID-19 pandemic is leading to increasing numbers of patients all over the world. Reports on a dysregulated immune system in the severe cases calls for a better characterization of the ongoing changes. To dissect COVID-19-driven immune host responses, we profiled whole blood transcriptomes enabling a data-driven stratification based on molecular phenotype. This analysis allowed prediction of patient subgroup-specific drug candidates targeting the dysregulated systemic immune response of the host.