Synthetical lethality of Werner helicase and mismatch repair deficiency is mediated by p53 and PUMA in colon cancer.

Synthetic lethality is a powerful approach for targeting oncogenic drivers in cancer. Recent studies revealed that cancer cells with microsatellite instability (MSI) require Werner (WRN) helicase for survival; however, the underlying mechanism remains unclear. In this study, we found that WRN depletion strongly induced p53 and its downstream apoptotic target PUMA in MSI colorectal cancer (CRC) cells. p53 or PUMA deletion abolished apoptosis induced by WRN depletion in MSI CRC cells. Importantly, correction of MSI abrogated the activation of p53/PUMA and cell killing, while induction of MSI led to sensitivity in isogenic CRC cells. Rare p53-mutant MSI CRC cells are resistant to WRN depletion due to lack of PUMA induction, which could be restored by wildtype (WT) p53 knock in or reconstitution. WRN depletion or treatment with the RecQ helicase inhibitor ML216 suppressed in vitro and in vivo growth of MSI CRCs in a p53/PUMA-dependent manner. ML216 treatment was efficacious in MSI CRC patient-derived xenografts. Interestingly, p53 gene remains WT in the majority of MSI CRCs. These results indicate a critical role of p53/PUMA-mediated apoptosis in the vulnerability of MSI CRCs to WRN loss, and support WRN as a promising therapeutic target in p53-WT MSI CRCs.

Immunotherapy efficacy on mismatch repair-deficient colorectal cancer: From bench to bedside.

Colorectal cancers (CRCs) with deficient mismatch repair (dMMR) or microsatellite instability-high (MSI-H) often have sustained responses to immune checkpoint inhibitors (ICIs) including selective monoclonal antibodies against Program Death 1 (PD-1), Programmed Death Ligand 1(PD-L1), and cytotoxic T lymphocyte associated antigen 4 (CTLA-4). However, a substantial fraction of dMMR CRCs do not respond or ultimately develop resistance to immunotherapy. The majority (~85%) of CRCs are MMR proficient (pMMR) or microsatellite stable (MSS) and lack response to ICIs. Understanding the biology and mechanisms underlying dMMR-associated immunogenicity is urgently needed for improving the therapeutic efficacy of immunotherapy on CRC. Compared to pMMR/MSS CRCs, dMMR/MSI CRCs typically have increased tumor mutational burden (TMB), lower response rate to 5-fluorouracil-based chemotherapy, distinctive immunological features such as high tumor-infiltrating lymphocytes (TILs), and better prognosis. Here, we review the current understanding of the clinical relevance of dMMR/MSI in CRCs, the molecular basis and rationales for targeting dMMR CRC with immunotherapy, and clinical approaches using ICIs as single agents or in combination with other therapies for MSI-H CRCs. Furthermore, we address the potential strategies to sensitize pMMR/MSS CRC to immunotherapy by converting an immunologically "cold" microenvironment into a "hot" one.

Untargeted Lipidomics Highlight the Depletion of Deoxyceramides during Therapy-Induced Senescence.

Therapy-induced senescence is a state of cell cycle arrest that occurs as a response to various chemotherapeutic reagents, especially ones that cause DNA damage. Senescent cells display resistance to cell death and can impair the efficacy of chemotherapeutic strategies. Since lipids can exhibit pro-survival activity, it is envisioned in this article that probing the lipidome could provide insights into novel lipids that are involved in senescence. Therefore, a tissue culture model system is established and the cellular lipidomes of senescent and proliferating cells are comparatively analyzed. Out of thousands of features detected, 17 species are identified that show significant changes in senescent cells. The majority of these species (11 out of 17) are atypical sphingolipids, 1-deoxyceramides/dihydroceramides, which are produced as a result of the utilization of alanine, instead of serine during sphingolipid biosynthesis. These lipids are depleted in senescent cells. Elevating the levels of deoxyceramides by supplementing the growth medium with metabolic precursors or by directly adding deoxyceramide result in decreased senescence, suggesting that these species might play a key role in this process.

An evolutionary transcriptomics approach links CD36 to membrane remodeling in replicative senescence.

Cellular senescence, the irreversible ceasing of cell division, has been associated with organismal aging, prevention of cancerogenesis, and developmental processes. As such, the evolutionary basis and biological features of cellular senescence remain a fascinating area of research. In this study, we conducted comparative RNAseq experiments to detect genes associated with replicative senescence in two different human fibroblast cell lines and at different time points. We identified 841 and 900 genes (core senescence-associated genes) that are significantly up- and downregulated in senescent cells, respectively, in both cell lines. Our functional enrichment analysis showed that downregulated core genes are primarily involved in cell cycle processes while upregulated core gene enrichment indicated various lipid-related processes. We further demonstrated that downregulated genes are significantly more conserved than upregulated genes. Using both transcriptomics and genetic variation data, we identified one of the upregulated, lipid metabolism genes, CD36, as an outlier. We found that overexpression of CD36 induces a senescence-like phenotype and, further, the media of CD36-overexpressing cells alone can induce a senescence-like phenotype in proliferating young cells. Moreover, we used a targeted lipidomics approach and showed that phosphatidylcholines accumulate during replicative senescence in these cells, suggesting that upregulation of CD36 could contribute to membrane remodeling during senescence. Overall, these results contribute to the understanding of evolution and biology of cellular senescence and identify several targets and questions for future studies.

Noncanonical Roles of Lipids in Different Cellular Fates.

Lipids are a diverse class of biomolecules. The biosynthesis and transport of these molecules are controlled by a considerable number of proteins, which facilitate spatiotemporal regulation of lipids during different fundamental cellular processes. Although lipids are traditionally considered as molecules for energy storage and as structural components of membranes, they are being increasingly recognized for their signaling roles. There is a growing appreciation of lipids' chemical diversity, which approaches that of proteins. In this Perspective, we discuss recent studies that suggest novel functions for distinct lipid species during different cellular processes. In particular, we discuss findings from our laboratory that illuminate the involvement of ceramides, polyunsaturated triacylglycerols, and very long chain fatty acids in different cellular fates. We also highlight recent innovative methods that have enabled the recognition of previously unknown lipid classes and/or roles of these molecules in different biological processes. We envision that advances in lipid identification, visualization, and perturbation will pave the way for broader investigations into this fascinating and influential class of biomolecules.

Regulation of lipids is central to replicative senescence.

Cellular replicative senescence, a state of permanent cell-cycle arrest, has been linked to organismal aging, tissue repair and tumorigenesis. In this study, we comparatively investigated the global lipid profiles and mRNA content of proliferating and senescent-state BJ fibroblasts. We found that both expression levels of lipid-regulating genes and the abundance of specific lipid families, are actively regulated. We further found that 19 specific polyunsaturated triacylglycerol species constituted the most prominent changes in lipid composition during replicative senescence. Based on the transcriptome analysis, we propose that the activation of CD36-mediated fatty acid uptake and diversion to glycerolipid biosynthesis could be responsible for the accumulation of triacylglycerols during replicative senescence. This, in turn, could be a cellular mechanism to prevent lipotoxicity under increased oxidative stress conditions observed in this process. Our results indicate that regulation of specific lipid species has a central role during replicative senescence.

Specific Triacylglycerols Accumulate via Increased Lipogenesis During 5-FU-Induced Apoptosis.

Lipids are emerging as key regulators of fundamental cellular processes including cell survival, division, and death. Apoptosis, a form of programmed cell death, is accompanied by numerous membrane-related phenotypic changes. However, we have an incomplete understanding of the involvement of specific lipid structures during this process. Here, we report that triacylglycerols are regulated at the molecular level during 5-fluorouracil-induced apoptosis in HCT-116. Mass-spectrometry-based global lipid profiling shows that specific triacylglycerols accumulate during apoptosis. Expression levels and activities of enzymes that are responsible for the biosynthesis and metabolic processing of triacylglycerols suggest that triacylglycerol biosynthesis is responsible for these accumulations. Based on our data, we propose that regulation of triacylglycerols at the molecular level happens downstream of p53 activation and potentially is a mechanism to prevent lipid oxidation during apoptosis.

Differential Regulation of Specific Sphingolipids in Colon Cancer Cells during Staurosporine-Induced Apoptosis.

Apoptosis is accompanied by distinct morphological changes at the plasma and organelle membrane level. Involvement of certain lipids in apoptosis has been established; however, we have limited understanding of the specific lipid structures that participate in this process. We used untargeted comparative lipidomics to study the changes in lipid composition during staurosporine-induced apoptosis in HCT-116. Our results revealed that ceramides, dihydroceramides, and sphingomyelins, with defined acyl chains, constitute the majority of changes in the lipidome. Expression levels and activities of enzymes responsible for the biosynthesis of lipids that change suggest that de novo synthesis causes these specific changes. Further analysis of the lipidome during apoptosis in other cancer and non-cancer cell lines suggested that accumulation of ceramides and dihydroceramides is specific to cancer cells. Taken together, our data propose that these molecules are regulated at the lipid-specific level during apoptosis and that this regulation differs between cancer and non-cancer cells.