Highlights d PDAC regional heterogeneity stems from sub-tumor microenvironments (subTMEs) d SubTMEs exhibit distinct immune phenotypes and CAF differentiation states d SubTMEs execute distinct tumor-promoting and chemoprotective functions d Intratumoral subTME co-occurrence links stromal heterogeneity to patient outcome

Individuals with acute myeloid leukemia (AML) harboring an internal tandem duplication (ITD) in the gene encoding Fms-related tyrosine kinase 3 (FLT3) who relapse after allogeneic hematopoietic cell transplantation (allo-HCT) have a 1-year survival rate below 20%. We observed that sorafenib, a multitargeted tyrosine kinase inhibitor, increased IL-15 production by FLT3-ITD leukemia cells. This synergized with the allogeneic CD8 T cell response, leading to long-term survival in six mouse models of FLT3-ITD AML. Sorafenib-related IL-15 production caused an increase in CD8CD107aIFN-γ T cells with features of longevity (high levels of Bcl-2 and reduced PD-1 levels), which eradicated leukemia in secondary recipients. Mechanistically, sorafenib reduced expression of the transcription factor ATF4, thereby blocking negative regulation of interferon regulatory factor 7 (IRF7) activation, which enhanced IL-15 transcription. Both IRF7 knockdown and ATF4 overexpression in leukemia cells antagonized sorafenib-induced IL-15 production in vitro. Human FLT3-ITD AML cells obtained from sorafenib responders following sorafenib therapy showed increased levels of IL-15, phosphorylated IRF7, and a transcriptionally active IRF7 chromatin state. The mitochondrial spare respiratory capacity and glycolytic capacity of CD8 T cells increased upon sorafenib treatment in sorafenib responders but not in nonresponders. Our findings indicate that the synergism of T cells and sorafenib is mediated via reduced ATF4 expression, causing activation of the IRF7-IL-15 axis in leukemia cells and thereby leading to metabolic reprogramming of leukemia-reactive T cells in humans. Therefore, sorafenib treatment has the potential to contribute to an immune-mediated cure of FLT3-ITD-mutant AML relapse, an otherwise fatal complication after allo-HCT.

No abstract

Extrapulmonary manifestations of COVID-19 have gained attention due to their links to clinical outcomes and their potential long-term sequelae1. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) displays tropism towards several organs, including the heart and kidney. Whether it also directly affects the liver has been debated2,3. Here we provide clinical, histopathological, molecular and bioinformatic evidence for the hepatic tropism of SARS-CoV-2. We find that liver injury, indicated by a high frequency of abnormal liver function tests, is a common clinical feature of COVID-19 in two independent cohorts of patients with COVID-19 requiring hospitalization. Using autopsy samples obtained from a third patient cohort, we provide multiple levels of evidence for SARS-CoV-2 liver tropism, including viral RNA detection in 69% of autopsy liver specimens, and successful isolation of infectious SARS-CoV-2 from liver tissue postmortem. Furthermore, we identify transcription-, proteomic- and transcription factor-based activity profiles in hepatic autopsy samples, revealing similarities to the signatures associated with multiple other viral infections of the human liver. Together, we provide a comprehensive multimodal analysis of SARS-CoV-2 liver tropism, which increases our understanding of the molecular consequences of severe COVID-19 and could be useful for the identification of organ-specific pharmacological targets.

Wiskott-Aldrich syndrome (WAS) is an X-linked primary immunodeficiency with severe platelet abnormalities and complex immunodeficiency. Although clinical gene therapy approaches using lentiviral vectors have produced encouraging results, full immune and platelet reconstitution is not always achieved. Here we show that a CRISPR/Cas9-based genome editing strategy allows the precise correction of WAS mutations in up to 60% of human hematopoietic stem and progenitor cells (HSPCs), without impairing cell viability and differentiation potential. Delivery of the editing reagents to WAS HSPCs led to full rescue of WASp expression and correction of functional defects in myeloid and lymphoid cells. Primary and secondary transplantation of corrected WAS HSPCs into immunodeficient mice showed persistence of edited cells for up to 26 weeks and efficient targeting of long-term repopulating stem cells. Finally, no major genotoxicity was associated with the gene editing process, paving the way for an alternative, yet highly efficient and safe therapy.

Oncogenic Ras mutations occur in various leukemias. It was unclear if, besides the direct transforming effect via constant RAS/MEK/ERK signaling, an inflammation-related effect of KRAS contributes to the disease. Here, we identify a functional link between oncogenic Kras G12D and NLRP3 inflammasome activation in murine and human cells. Mice expressing active Kras G12D in the hematopoietic system developed myeloproliferation and cytopenia, which is reversed in Kras G12D mice lacking NLRP3 in the hematopoietic system. Therapeutic IL-1-receptor blockade or NLRP3-inhibition reduces myeloproliferation and improves hematopoiesis. Mechanistically, Kras G12D-RAC1 activation induces reactive oxygen species (ROS) production causing NLRP3 inflammasome-activation. In agreement with our observations in mice, patient-derived myeloid leukemia cells exhibit KRAS/RAC1/ROS/NLRP3/IL-1β axis activity. Our findings indicate that oncogenic KRAS not only act via its canonical oncogenic driver function, but also enhances the activation of the pro-inflammatory RAC1/ROS/NLRP3/ IL-1β axis. This paves the way for a therapeutic approach based on immune modulation via NLRP3 blockade in KRAS-mutant myeloid malignancies.

Organoid cultures derived from colorectal cancer (CRC) samples are increasingly used as preclinical models for studying tumor biology and the effects of targeted therapies under conditions capturing in vitro the genetic make-up of heterogeneous and even individual neoplasms. While 3D cultures are initiated from surgical specimens comprising multiple cell populations, the impact of tumor heterogeneity on drug effects in organoid cultures has not been addressed systematically. Here we have used a cohort of well-characterized CRC organoids to study the influence of tumor heterogeneity on the activity of the KRAS/MAPK-signaling pathway and the consequences of treatment by inhibitors targeting EGFR and downstream effectors. MAPK signaling, analyzed by targeted proteomics, shows unexpected heterogeneity irrespective of RAS mutations and is associated with variable responses to EGFR inhibition. In addition, we obtained evidence for intratumoral heterogeneity in drug response among parallel “sibling” 3D cultures established from a single KRAS -mutant CRC. Our results imply that separate testing of drug effects in multiple subpopulations may help to elucidate molecular correlates of tumor heterogeneity and to improve therapy response prediction in patients.

Intracellular mechanisms of copper cytotoxicity.