Chemoproteomics reveals immunogenic and tumor-associated cell surface substrates of ectokinase CK2α.

New epitopes for immune recognition provide the basis of anticancer immunity. Due to the high concentration of extracellular adenosine triphosphate in the tumor microenvironment, we hypothesized that extracellular kinases (ectokinases) could have dysregulated activity and introduce aberrant phosphorylation sites on cell surface proteins. We engineered a cell-tethered version of the extracellular kinase CK2α, demonstrated it was active on cells under tumor-relevant conditions, and profiled its substrate scope using a chemoproteomic workflow. We then demonstrated that mice developed polyreactive antisera in response to syngeneic tumor cells that had been subjected to surface hyperphosphorylation with CK2α. Interestingly, these mice developed B cell and CD4+ T cell responses in response to these antigens but failed to develop a CD8+ T cell response. This work provides a workflow for probing the extracellular phosphoproteome and demonstrates that extracellular phosphoproteins are immunogenic even in a syngeneic system.

Allosteric SHP2 inhibition increases apoptotic dependency on BCL2 and synergizes with venetoclax in FLT3- and KIT-mutant AML.

Mutations in the receptor tyrosine kinases (RTKs) FLT3 and KIT are frequent and associated with poor outcomes in acute myeloid leukemia (AML). Although selective FLT3 inhibitors (FLT3i) are clinically effective, remissions are short-lived due to secondary resistance characterized by acquired mutations constitutively activating the RAS/MAPK pathway. Hereby, we report the pre-clinical efficacy of co-targeting SHP2, a critical node in MAPK signaling, and BCL2 in RTK-driven AML. The allosteric SHP2 inhibitor RMC-4550 suppresses proliferation of AML cell lines with FLT3 and KIT mutations, including cell lines with acquired resistance to FLT3i. We demonstrate that pharmacologic SHP2 inhibition unveils an Achilles' heel of RTK-driven AML, increasing apoptotic dependency on BCL2 via MAPK-dependent mechanisms, including upregulation of BMF and downregulation of MCL1. Consequently, RMC-4550 and venetoclax are synergistically lethal in AML cell lines and in clinically relevant xenograft models. Our results provide mechanistic rationale and pre-clinical evidence for co-targeting SHP2 and BCL2 in RTK-driven AML.

As above, so below: Deposition, modification, and reutilization of human remains at Marmoles cave (Cueva de los Marmoles: Southern Spain, 4000-1000 cal. BCE).

The deposition and manipulation of human remains in natural caves are well known for the Neolithic of Southern Iberia. The cultural meaning of these practices is however still largely unclear. Cueva de los Marmoles (CM, Priego-Córdoba) is one of the most important cave contexts from Southern Spain, which returned a large number of commingled skeletal remains suggesting its funerary use from the Neolithic to the Late Bronze Age. Here we discuss CM from a chronological and cultural perspective based on new radiocarbon, anthropological, and taphonomic analyses. These include the estimation of the minimum number of individuals, the exploration of fragmentation patterns characterizing different skeletal regions, and the macroscopic and microscopic analysis of modifications to the remains of possible anthropic origin. Radiocarbon data point to a funerary use of CM between the 5th -2nd millennium cal. BCE. MNI estimates reveal the presence of at least 12 individuals (seven adults and five nonadults). The low representation of elements from hands and feet suggests that individuals were placed in the cave while partially decomposed. Anthropic traces on the remains (e.g. fresh fractures, marrow canal modifications, and scraping marks) hint at their intentional fragmentation, cleaning from residual soft tissues, and in some cases reutilization. These practices are well-exemplified by the recovery of one "skull cup" and of two long bones used as tools. These data align with those from other cave contexts from the same geographic region, suggesting the presence, especially during the Neolithic period, of shared ideologies centered on the human body.

Ultrasound Guided Surgery as a Refinement Tool in Oncology Research.

Refinement is one of the ethical pillars of the use of animals in research. Ultrasonography is currently used in human medicine as a surgical tool for guided biopsies and this idea can be applied to preclinical research thanks to the development of specific instruments. This will eliminate the necessity of a surgical opening for implanting cells in specific organs or taking samples from tissues. The approach for the injection will depend on the target but most of the case is going to be lateral, with the probe in a ventral position and the needle going into from the lateral. This is the situation for the thyroid gland, heart, liver, spleen, kidney, pancreas, uterus, and testicles. Other approaches, such as the dorsal, can be used in the spleen or kidney. The maximum injected volume will depend on the size of the structure. For biopsies, the technical protocol is similar to the injection knowing that in big organs such as the liver, spleen, or kidney we can take several samples moving slightly the needle inside the structure. In all cases, animals must be anesthetized and minimum pain management is required after the intervention.