LANGUAGE= "English" Objectives: DKK-1, a negative regulator of ?-catenin in the Wnt pathway is commonly upregulated in cancer cells but its expression and association to chemoresistance have not been ...investigated extensively. Our aim was to evaluate the expression of DKK-1 and ?-catenin as well as the effect of the Wnt pathway modulators iCRT-14 (iCRT) and WAY626611 (WAY) alone or in combination with Nigericin (NIG) on the viability of cancer cells grown under routine conditions (RC) or as floating spheroids (FSs). Methods: RC or FSs from human prostate (PC3 and LnCAP) and breast (MCF-7 and MDA-MB231) cells were treated for 72 hs with WAY or iCRT. Cell viability was determined by the MTT or the CCK-8 assays and protein expression was determined by western blot analysis. Results: DKK-1 was expressed at different basal levels in RC but was undetectable in FSs. ?-catenin expression was in?creased in FSs. RC of PC3 and MCF-7 were sensitive to both iCRT and WAY but FSs generated from these cell lines were resistant to both drugs. NIG potentiated the effect of both Wnt inhibitors in FSs. Conclusion: Marked changes in the expression of Wnt pathway proteins, DKK-1 and ?-catenin in FSs are associated with resistance to WAY and iCRT.
Although it is well established that all brain regions contain various neuronal subtypes with different functions, astrocytes have traditionally been thought to be homogenous. However, recent ...evidence has shown that astrocytes in the mammalian CNS display distinct inter- and intra-regional features, as well as functional diversity. In the CNS, astrocyte processes fill the local environment in non-overlapping domains. Therefore, a potential advantage of region-specified astrocytes might be their capacity to regulate local development or optimize local neural circuit function. An overview of the regional heterogeneity of neuron-astrocyte interactions indicates novel ways in which they could regulate normal neurological function and shows how they might become dysregulated in disease.
Isolated brain capillaries are essential for analyzing the changes of protein expressions at the blood–brain barrier (BBB) under pathological conditions. The standard brain capillary isolation ...methods require the use of at least five mouse brains in order to obtain a sufficient amount and purity of brain capillaries. The purpose of this study was to establish a brain capillary isolation method from a single mouse brain for protein expression analysis. We successfully isolated brain capillaries from a single frozen mouse brain by using a bead homogenizer in the brain homogenization step and combination of cell strainers and glass beads in the purification step. Western blot and proteomic analysis showed that proteins expressed at the BBB in mouse brain capillaries isolated by the developed method were more enriched than those isolated from a pool of five mouse brains by the standard method. By using the developed method, we further verified the changes in expression of BBB proteins in Glut1-deficient mouse. The developed method is useful for the analysis of various mice models with low numbers and enables us to understand, in more detail, the physiology and pathology of BBB.
The adoptive transfer of T lymphocytes reprogrammed to target tumour cells has demonstrated potential for treatment of various cancers
. However, little is known about the long-term potential and ...clonal stability of the infused cells. Here we studied long-lasting CD19-redirected chimeric antigen receptor (CAR) T cells in two patients with chronic lymphocytic leukaemia
who achieved a complete remission in 2010. CAR T cells remained detectable more than ten years after infusion, with sustained remission in both patients. Notably, a highly activated CD4
population emerged in both patients, dominating the CAR T cell population at the later time points. This transition was reflected in the stabilization of the clonal make-up of CAR T cells with a repertoire dominated by a small number of clones. Single-cell profiling demonstrated that these long-persisting CD4
CAR T cells exhibited cytotoxic characteristics along with ongoing functional activation and proliferation. In addition, longitudinal profiling revealed a population of gamma delta CAR T cells that prominently expanded in one patient concomitant with CD8
CAR T cells during the initial response phase. Our identification and characterization of these unexpected CAR T cell populations provide novel insight into the CAR T cell characteristics associated with anti-cancer response and long-term remission in leukaemia.
Methods for profiling RNA and protein expression in a spatially resolved manner are rapidly evolving, making it possible to comprehensively characterize cells and tissues in health and disease. To ...maximize the biological insights obtained using these techniques, it is critical to both clearly articulate the key biological questions in spatial analysis of tissues and develop the requisite computational tools to address them. Developers of analytical tools need to decide on the intrinsic molecular features of each cell that need to be considered, and how cell shape and morphological features are incorporated into the analysis. Also, optimal ways to compare different tissue samples at various length scales are still being sought. Grouping these biological problems and related computational algorithms into classes across length scales, thus characterizing common issues that need to be addressed, will facilitate further progress in spatial transcriptomics and proteomics.
The most common type of lung cancer is adenocarcinoma (ADC), comprising around 40% of all lung cancer cases. In spite of achievements in understanding the pathogenesis of this disease and the ...development of new approaches in its treatment, unfortunately, lung ADC is still one of the most aggressive and rapidly fatal tumor types with overall survival less than 5 years. Lung ADC is often diagnosed at advanced stages involving disseminated metastatic tumors. This is particularly important for the successful development of new approaches in cancer therapy. The high resistance of lung ADC to conventional radiotherapies and chemotherapies represents a major challenge for treatment effectiveness. Here we discuss recent advances in understanding the molecular pathways driving tumor progression and related targeted therapies in lung ADCs. In addition, the cell death mechanisms induced by different treatment strategies and their contribution to therapy resistance are analyzed. The focus is on approaches to overcoming drug resistance in order to improve future treatment decisions.
About 25% of pancreatic cancers harbour actionable molecular alterations, defined as molecular alterations for which there is clinical or strong preclinical evidence of a predictive benefit from a ...specific therapy. The Know Your Tumor (KYT) programme includes US patients with pancreatic cancer and enables patients to undergo commercially available multi-omic profiling to provide molecularly tailored therapy options and clinical trial recommendations. We sought to determine whether patients with pancreatic cancer whose tumours harboured such actionable molecular alterations and who received molecularly matched therapy had a longer median overall survival than similar patients who did not receive molecularly matched therapy.
In this retrospective analysis, treatment history and longitudinal survival outcomes were analysed in patients aged 18 years or older with biopsy-confirmed pancreatic cancer of any stage, enrolled in the KYT programme and who received molecular testing results. Since the timing of KYT enrolment varied for each patient, the primary outcome measurement of median overall survival was calculated from the initial diagnosis of advanced disease until death. We compared median overall survival in patients with actionable mutations who were treated with a matched therapy versus those who were not treated with a matched therapy.
Of 1856 patients with pancreatic cancer who were referred to the KYT programme between June 16, 2014, and March 31, 2019, 1082 (58%) patients received personalised reports based on their molecular testing results. Actionable molecular alterations were identified in 282 (26%) of 1082 samples. Among 677 patients for whom outcomes were available, 189 had actionable molecular alterations. With a median follow-up of 383 days (IQR 214–588), those patients with actionable molecular alterations who received a matched therapy (n=46) had significantly longer median overall survival than did those patients who only received unmatched therapies (n=143; 2·58 years 95% CI 2·39 to not reached vs 1·51 years 1·33–1·87; hazard ratio 0·42 95% CI 0·26–0·68, p=0·0004). The 46 patients who received a matched therapy also had significantly longer overall survival than the 488 patients who did not have an actionable molecular alteration (2·58 years 95% CI 2·39 to not reached vs 1·32 years 1·25–1·47; HR 0·34 95% CI 0·22–0·53, p<0·0001). However, median overall survival did not differ between the patients who received unmatched therapy and those without an actionable molecular alteration (HR 0·82 95% CI 0·64–1·04, p=0·10).
These real-world outcomes suggest that the adoption of precision medicine can have a substantial effect on survival in patients with pancreatic cancer, and that molecularly guided treatments targeting oncogenic drivers and the DNA damage response and repair pathway warrant further prospective evaluation.
Pancreatic Cancer Action Network and Perthera.
There are numerous cell types with scarcely understood functions, whose interactions with the immune system are not well characterized. To facilitate their study, we generated a mouse bearing ...enhanced green fluorescent protein (EGFP)-specific CD8
T cells. Transfer of the T cells into EGFP reporter animals can be used to kill EGFP-expressing cells, allowing selective depletion of desired cell types, or to interrogate T-cell interactions with specific populations. Using this system, we eliminate a rare EGFP-expressing cell type in the heart and demonstrate its role in cardiac function. We also show that naive T cells are recruited into the mouse brain by antigen-expressing microglia, providing evidence of an immune surveillance pathway in the central nervous system. The just EGFP death-inducing (Jedi) T cells enable visualization of a T-cell antigen. They also make it possible to utilize hundreds of existing EGFP-expressing mice, tumors, pathogens and other tools, to study T-cell interactions with many different cell types, to model disease states and to determine the functions of poorly characterized cell populations.
The emergency use authorizations (EUAs) of two mRNA-based severe acute respiratory syndrome coronavirus (SARS-CoV)-2 vaccines approximately 11 months after publication of the viral sequence ...highlights the transformative potential of this nucleic acid technology. Most clinical applications of mRNA to date have focused on vaccines for infectious disease and cancer for which low doses, low protein expression and local delivery can be effective because of the inherent immunostimulatory properties of some mRNA species and formulations. In addition, work on mRNA-encoded protein or cellular immunotherapies has also begun, for which minimal immune stimulation, high protein expression in target cells and tissues, and the need for repeated administration have led to additional manufacturing and formulation challenges for clinical translation. Building on this momentum, the past year has seen clinical progress with second-generation coronavirus disease 2019 (COVID-19) vaccines, Omicron-specific boosters and vaccines against seasonal influenza, Epstein-Barr virus, human immunodeficiency virus (HIV) and cancer. Here we review the clinical progress of mRNA therapy as well as provide an overview and future outlook of the transformative technology behind these mRNA-based drugs.
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