Protein lipoylation: mitochondria, cuproptosis, and beyond.

Protein lipoylation, a crucial post-translational modification (PTM), plays a pivotal role in mitochondrial function and emerges as a key player in cell death through cuproptosis. This novel copper-driven cell death pathway is activated by excessive copper ions binding to lipoylated mitochondrial proteins, disrupting energy production and causing lethal protein aggregation and cell death. The intricate relationship among protein lipoylation, cellular energy metabolism, and cuproptosis offers a promising avenue for regulating essential cellular functions. This review focuses on the mechanisms of lipoylation and its significant impact on cell metabolism and cuproptosis, emphasizing the key genes involved and their implications for human diseases. It offers valuable insights into targeting dysregulated cellular metabolism for therapeutic purposes.

Nardilysin-regulated scission mechanism activates polo-like kinase 3 to suppress the development of pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) develops through step-wise genetic and molecular alterations including Kras mutation and inactivation of various apoptotic pathways. Here, we find that development of apoptotic resistance and metastasis of KrasG12D-driven PDAC in mice is accelerated by deleting Plk3, explaining the often-reduced Plk3 expression in human PDAC. Importantly, a 41-kDa Plk3 (p41Plk3) that contains the entire kinase domain at the N-terminus (1-353 aa) is activated by scission of the precursor p72Plk3 at Arg354 by metalloendopeptidase nardilysin (NRDC), and the resulting p32Plk3 C-terminal Polo-box domain (PBD) is removed by proteasome degradation, preventing the inhibition of p41Plk3 by PBD. We find that p41Plk3 is the activated form of Plk3 that regulates a feed-forward mechanism to promote apoptosis and suppress PDAC and metastasis. p41Plk3 phosphorylates c-Fos on Thr164, which in turn induces expression of Plk3 and pro-apoptotic genes. These findings uncover an NRDC-regulated post-translational mechanism that activates Plk3, establishing a prototypic regulation by scission mechanism.

Unlocking c-MET: A comprehensive journey into targeted therapies for breast cancer.

Breast cancer is the most common malignancy among women, posing a formidable health challenge worldwide. In this complex landscape, the c-MET (cellular-mesenchymal epithelial transition factor) receptor tyrosine kinase (RTK), also recognized as the hepatocyte growth factor (HGF) receptor (HGFR), emerges as a prominent protagonist, displaying overexpression in nearly 50% of breast cancer cases. Activation of c-MET by its ligand, HGF, secreted by neighboring mesenchymal cells, contributes to a cascade of tumorigenic processes, including cell proliferation, metastasis, angiogenesis, and immunosuppression. While c-MET inhibitors such as crizotinib, capmatinib, tepotinib and cabozantinib have garnered FDA approval for non-small cell lung cancer (NSCLC), their potential within breast cancer therapy is still undetermined. This comprehensive review embarks on a journey through structural biology, multifaceted functions, and intricate signaling pathways orchestrated by c-MET across cancer types. Furthermore, we highlight the pivotal role of c-MET-targeted therapies in breast cancer, offering a clinical perspective on this promising avenue of intervention. In this pursuit, we strive to unravel the potential of c-MET as a beacon of hope in the fight against breast cancer, unveiling new horizons for therapeutic innovation.

BRCA1-mediated dual regulation of ferroptosis exposes a vulnerability to GPX4 and PARP co-inhibition in BRCA1-deficient cancers.

Resistance to poly (ADP-ribose) polymerase inhibitors (PARPi) limits the therapeutic efficacy of PARP inhibition in treating breast cancer susceptibility gene 1 (BRCA1)-deficient cancers. Here we reveal that BRCA1 has a dual role in regulating ferroptosis. BRCA1 promotes the transcription of voltage-dependent anion channel 3 (VDAC3) and glutathione peroxidase 4 (GPX4); consequently, BRCA1 deficiency promotes cellular resistance to erastin-induced ferroptosis but sensitizes cancer cells to ferroptosis induced by GPX4 inhibitors (GPX4i). In addition, nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy and defective GPX4 induction unleash potent ferroptosis in BRCA1-deficient cancer cells upon PARPi and GPX4i co-treatment. Finally, we show that xenograft tumors derived from BRCA1-mutant breast cancer patients with PARPi resistance exhibit decreased GPX4 expression and high sensitivity to PARP and GPX4 co-inhibition. Our results show that BRCA1 deficiency induces a ferroptosis vulnerability to PARP and GPX4 co-inhibition and inform a therapeutic strategy for overcoming PARPi resistance in BRCA1-deficient cancers.

DDX3 regulates cancer immune surveillance via 3' UTR-mediated cell-surface expression of PD-L1.

Programmed death-1 (PD-1)/PD ligand-1 (PD-L1)-mediated immune escape contributes to cancer development and has been targeted as an anti-cancer strategy. Here, we show that inhibition of the RNA helicase DDX3 increased CD8+ T cell infiltration in syngeneic oral squamous cell carcinoma tumors. DDX3 knockdown compromised interferon-γ-induced PD-L1 expression and, in particular, reduced the level of cell-surface PD-L1. DDX3 promoted surface PD-L1 expression by recruiting the adaptor protein 2 (AP2) complex to the 3' UTR of PD-L1 mRNA. DDX3 depletion or 3' UTR truncation increased the binding of the coatomer protein complexes to PD-L1, leading to its intracellular accumulation. Therefore, this 3' UTR-dependent mechanism may counteract cellular negative effects on surface trafficking of PD-L1. Finally, pharmaceutic disruption of DDX3's interaction with AP2 reduced surface PD-L1 expression, supporting that the DDX3-AP2 pathway routes PD-L1 to the cell surface. Targeting DDX3 to modulate surface trafficking of immune checkpoint proteins may provide a potential strategy for cancer immunotherapy.

Targeting ALK averts ribonuclease 1-induced immunosuppression and enhances antitumor immunity in hepatocellular carcinoma.

Tumor-secreted factors contribute to the development of a microenvironment that facilitates the escape of cancer cells from immunotherapy. In this study, we conduct a retrospective comparison of the proteins secreted by hepatocellular carcinoma (HCC) cells in responders and non-responders among a cohort of ten patients who received Nivolumab (anti-PD-1 antibody). Our findings indicate that non-responders have a high abundance of secreted RNase1, which is associated with a poor prognosis in various cancer types. Furthermore, mice implanted with HCC cells that overexpress RNase1 exhibit immunosuppressive tumor microenvironments and diminished response to anti-PD-1 therapy. RNase1 induces the polarization of macrophages towards a tumor growth-promoting phenotype through activation of the anaplastic lymphoma kinase (ALK) signaling pathway. Targeting the RNase1/ALK axis reprograms the macrophage polarization, with increased CD8+ T- and Th1- cell recruitment. Moreover, simultaneous targeting of the checkpoint protein PD-1 unleashes cytotoxic CD8+ T-cell responses. Treatment utilizing both an ALK inhibitor and an anti-PD-1 antibody exhibits enhanced tumor regression and facilitates long-term immunity. Our study elucidates the role of RNase1 in mediating tumor resistance to immunotherapy and reveals an RNase1-mediated immunosuppressive tumor microenvironment, highlighting the potential of targeting RNase1 as a promising strategy for cancer immunotherapy in HCC.

CRKL dictates anti-PD-1 resistance by mediating tumor-associated neutrophil infiltration in hepatocellular carcinoma.

BACKGROUND & AIMS: The effectiveness of immune checkpoint inhibitor (ICI) therapy for hepatocellular carcinoma (HCC) is limited by treatment resistance. However, the mechanisms underlying immunotherapy resistance remain elusive. We aimed to identify the role of CT10 regulator of kinase-like (CRKL) in resistance to anti-PD-1 therapy in HCC. METHODS: Gene expression in HCC specimens from 10 patients receiving anti-PD-1 therapy was identified by RNA-sequencing. A total of 404 HCC samples from tissue microarrays were analyzed by immunohistochemistry. Transgenic mice (Alb-Cre/Trp53fl/fl) received hydrodynamic tail vein injections of a CRKL-overexpressing vector. Mass cytometry by time of flight was used to profile the proportion and status of different immune cell lineages in the mouse tumor tissues. RESULTS: CRKL was identified as a candidate anti-PD-1-resistance gene using a pooled genetic screen. CRKL overexpression nullifies anti-PD-1 treatment efficacy by mobilizing tumor-associated neutrophils (TANs), which block the infiltration and function of CD8+ T cells. PD-L1+ TANs were found to be an essential subset of TANs that were regulated by CRKL expression and display an immunosuppressive phenotype. Mechanistically, CRKL inhibits APC (adenomatous polyposis coli)-mediated proteasomal degradation of ß-catenin by competitively decreasing Axin1 binding, and thus promotes VEGFα and CXCL1 expression. Using human HCC samples, we verified the positive correlations of CRKL/ß-catenin/VEGFα and CXCL1. Targeting CRKL using CRISPR-Cas9 gene editing (CRKL knockout) or its downstream regulators effectively restored the efficacy of anti-PD-1 therapy in an orthotopic mouse model and a patient-derived organotypic tumor spheroid model. CONCLUSIONS: Activation of the CRKL/ß-catenin/VEGFα and CXCL1 axis is a critical obstacle to successful anti-PD-1 therapy. Therefore, CRKL inhibitors combined with anti-PD-1 could be useful for the treatment of HCC. IMPACT AND IMPLICATIONS: Here, we found that CRKL was overexpressed in anti-PD-1-resistant hepatocellular carcinoma (HCC) and that CRKL upregulation promotes anti-PD-1 resistance in HCC. We identified that upregulation of the CRKL/ß-catenin/VEGFα and CXCL1 axis contributes to anti-PD-1 tolerance by promoting infiltration of tumor-associated neutrophils. These findings support the strategy of bevacizumab-based immune checkpoint inhibitor combination therapy, and CRKL inhibitors combined with anti-PD-1 therapy may be developed for the treatment of HCC.

Inhibition of ADAM9 promotes the selective degradation of KRAS and sensitizes pancreatic cancers to chemotherapy.

Kirsten rat sarcoma virus (KRAS) signaling drives pancreatic ductal adenocarcinoma (PDAC) malignancy, which is an unmet clinical need. Here, we identify a disintegrin and metalloproteinase domain (ADAM)9 as a modulator of PDAC progression via stabilization of wild-type and mutant KRAS proteins. Mechanistically, ADAM9 loss increases the interaction of KRAS with plasminogen activator inhibitor 1 (PAI-1), which functions as a selective autophagy receptor in conjunction with light chain 3 (LC3), triggering lysosomal degradation of KRAS. Suppression of ADAM9 by a small-molecule inhibitor restricts disease progression in spontaneous models, and combination with gemcitabine elicits dramatic regression of patient-derived tumors. Our findings provide a promising strategy to target the KRAS signaling cascade and demonstrate a potential modality to enhance sensitivity to chemotherapy in PDAC.

MRCK as a Potential Target for Claudin-Low Subtype of Breast Cancer.

To find new molecular targets for triple negative breast cancer (TNBC), we analyzed a large-scale drug screening dataset based on breast cancer subtypes. We discovered that BDP-9066, a specific MRCK inhibitor (MRCKi), may be an effective drug against TNBC. After confirming the efficacy and specificity of BDP-9066 against TNBC in vitro and in vivo, we further analyzed the underlying mechanism of specific activity of BDP-9066 against TNBC. Comparing the transcriptome of BDP-9066-sensitive and -resistant cells, the activation of the focal adhesion and YAP/TAZ pathway were found to play an important role in the sensitive cells. Furthermore, YAP/TAZ is indeed repressed by BDP-9066 in the sensitive cells, and active form of YAP suppresses the effects of BDP-9066. YAP/TAZ expression and activity are high in TNBC, especially the Claudin-low subtype, consistent with the expression of focal adhesion-related genes. Interestingly, NF-κB functions downstream of YAP/TAZ in TNBC cells and is suppressed by BDP-9066. Furthermore, the PI3 kinase pathway adversely affected the effects of BDP-9066 and that alpelisib, a PI3 kinase inhibitor, synergistically increased the effects of BDP-9066, in PIK3CA mutant TNBC cells. Taken together, we have shown for the first time that MRCKi can be new drugs against TNBC, particularly the Claudin-low subtype.

Genetic insights into carbohydrate sulfotransferase 8 and its impact on the immunotherapy efficacy of cancer.

Immune checkpoint blockade (ICB) is a promising therapy for solid tumors, but its effectiveness depends on biomarkers that are not precise. Here, we utilized genome-wide association study to investigate the association between genetic variants and tumor mutation burden to interpret ICB response. We identified 16 variants (p < 5 × 10-8) probed to 17 genes on 9 chromosomes. Subsequent analysis of one of the most significant loci in 19q13.11 suggested that the rs111308825 locus at the enhancer is causal, as its A allele impairs KLF2 binding, leading to lower carbohydrate sulfotransferase 8 (CHST8) expression. Breast cancer cells expressing CHST8 suppress T cell activation, and Chst8 loss attenuates tumor growth in a syngeneic mouse model. Further investigation revealed that programmed death-ligand 1 (PD-L1) and its homologs could be sulfated by CHST8, resulting in M2-like macrophage enrichment in the tumor microenvironment. Finally, we confirmed that low-CHST8 tumors have better ICB response, supporting the genetic effect and clinical value of rs111308825 for ICB efficacy prediction.

DriverDBv4: a multi-omics integration database for cancer driver gene research.

Advancements in high-throughput technology offer researchers an extensive range of multi-omics data that provide deep insights into the complex landscape of cancer biology. However, traditional statistical models and databases are inadequate to interpret these high-dimensional data within a multi-omics framework. To address this limitation, we introduce DriverDBv4, an updated iteration of the DriverDB cancer driver gene database (http://driverdb.bioinfomics.org/). This updated version offers several significant enhancements: (i) an increase in the number of cohorts from 33 to 70, encompassing approximately 24 000 samples; (ii) inclusion of proteomics data, augmenting the existing types of omics data and thus expanding the analytical scope; (iii) implementation of multiple multi-omics algorithms for identification of cancer drivers; (iv) new visualization features designed to succinctly summarize high-context data and redesigned existing sections to accommodate the increased volume of datasets and (v) two new functions in Customized Analysis, specifically designed for multi-omics driver identification and subgroup expression analysis. DriverDBv4 facilitates comprehensive interpretation of multi-omics data across diverse cancer types, thereby enriching the understanding of cancer heterogeneity and aiding in the development of personalized clinical approaches. The database is designed to foster a more nuanced understanding of the multi-faceted nature of cancer.

Gasdermin C sensitizes tumor cells to PARP inhibitor therapy in cancer models.

Several poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) are approved by FDA to treat cancer with BRCA mutations. BRCA mutations are considered to fuel a PARPi killing effect by inducing apoptosis. However, resistance to PARPi is frequently observed in the clinic due to an incomplete understanding on the molecular basis of PARPi function and a lack of good markers, beyond BRCA mutations, to predict response. Here, we show that gasdermin C (GSDMC) sensitized tumor cells to PARPi in vitro and in immunocompetent mice and caused durable tumor regression in an immune-dependent manner. A high expression level of GSDMC predicted better response to PARPi treatment in patients with triple-negative breast cancer (TNBC). PARPi treatment triggered GSDMC/caspase-8-mediated cancer cell pyroptosis (CCP) that enhanced PARPi killing of tumor cells. GSDMC-mediated CCP increased memory CD8+ T cell population in lymph node (LN), spleen, and tumor and, thus, promoted cytotoxic CD8+ T cell infiltration in the tumor microenvironment. T cell-derived granzyme B (GZMB) activated caspase-6, which subsequently cleaved GSDMC to induce pyroptosis. Interestingly, IFN-γ induced GSDMC expression, which, in turn, enhanced the cytotoxicity of PARPi and T cells. Importantly, GSDMC promoted tumor clearance independent of BRCA deficiency in multiple cancer types with PARPi treatment. This study identifies a general marker and target for PARPi therapy and offers insights into the mechanism of PARPi function.

Epigenetic regulation of asymmetric cell division by the LIBR-BRD4 axis.

Asymmetric cell division (ACD) is a mechanism used by stem cells to maintain the number of progeny. However, the epigenetic mechanisms regulating ACD remain elusive. Here we show that BRD4, a BET domain protein that binds to acetylated histone, is segregated in daughter cells together with H3K56Ac and regulates ACD. ITGB1 is regulated by BRD4 to regulate ACD. A long noncoding RNA (lncRNA), LIBR (LncRNA Inhibiting BRD4), decreases the percentage of stem cells going through ACD through interacting with the BRD4 mRNAs. LIBR inhibits the translation of BRD4 through recruiting a translation repressor, RCK, and inhibiting the binding of BRD4 mRNAs to polysomes. These results identify the epigenetic regulatory modules (BRD4, lncRNA LIBR) that regulate ACD. The regulation of ACD by BRD4 suggests the therapeutic limitation of using BRD4 inhibitors to treat cancer due to the ability of these inhibitors to promote symmetric cell division that may lead to tumor progression and treatment resistance.

Development of triazole-based PKC-inhibitors to overcome resistance to EGFR inhibitors in EGFR-mutant lung cancers.

Protein kinase C delta (PKCδ) is prominently expressed in the nuclei of EGFR-mutant lung cancer cells, and its presence correlates with poor survival of the patients undergoing EGFR inhibitor treatment. The inhibition of PKCδ has emerged as a viable approach to overcoming resistance to EGFR inhibitors. However, clinical-grade PKCδ inhibitors are not available, highlighting the urgent needs for the development of effective drugs that target PKCδ. In this study, we designed and synthesized a series of inhibitors based on the chemical structure of a pan PKC inhibitor sotrastaurin. This was achieved by incorporating a triazole ring group into the original sotrastaurin configuration. Our findings revealed that the sotrastaurin derivative CMU-0101 exhibited an elevated affinity for binding to the ATP-binding site of PKCδ and effectively suppressed nuclear PKCδ in resistant cells in comparison to sotrastaurin. Furthermore, we demonstrated that CMU-0101 synergistically enhanced EGFR TKI gefitinib sensitivity in resistant cells. Altogether, our study provides a promising strategy for designing and synthesizing PKCδ inhibitors with improved efficacy, and suggests CMU-0101 as a potential lead compound to inhibit PKCδ and overcome TKI resistance in lung cancers.

Inhibition of Galectin-9 sensitizes tumors to anthracycline treatment via inducing antitumor immunity.

Anthracyclines are a class of conventionally and routinely used first-line chemotherapy drugs for cancer treatment. In addition to the direct cytotoxic effects, increasing evidence indicates that the efficacy of the drugs also depends on immunomodulatory effects with unknown mechanisms. Galectin-9 (Gal-9), a member of the ß-galactoside-binding protein family, has been demonstrated to induce T-cell death and promote immunosuppression in the tumor microenvironment. Here, we asked whether anthracycline-mediated immunomodulatory activity might be related to Gal-9. We found that combining doxorubicin with anti-Gal-9 therapy significantly inhibited tumor growth and prolonged overall survival in immune-competent syngeneic mouse models. Moreover, Gal-9 expression was increased in response to doxorubicin in various human and murine cancer cell lines. Mechanistically, doxorubicin induced tumoral Gal-9 by activating the STING/interferon ß pathway. Clinically, Gal-9 and p-STING levels were elevated in the tumor tissues of breast cancer patients treated with anthracyclines. Our study demonstrates Gal-9 upregulation in response to anthracyclines as a novel mechanism mediating immune escape and suggests targeting Gal-9 in combination with anthracyclines as a promising therapeutic strategy for cancer treatment.

PD-1 blockade in combination with dasatinib potentiates induction of anti-acute lymphocytic leukemia immunity.

Immunotherapy, in the form of hematopoietic stem cell transplantation (HSCT), has been part of the standard of care in the treatment of acute leukemia for over 40 years. Trials evaluating novel immunotherapeutic approaches, such as targeting the programmed death-1 (PD-1) pathway, have unfortunately not yielded comparable results to those seen in solid tumors. Major histocompatibility complex (MHC) proteins are cell surface proteins essential for the adaptive immune system to recognize self versus non-self. MHC typing is used to determine donor compatibility when evaluating patients for HSCT. Recently, loss of MHC class II (MHC II) was shown to be a mechanism of immune escape in patients with acute myeloid leukemia after HSCT. Here we report that treatment with the tyrosine kinase inhibitor, dasatinib, and an anti-PD-1 antibody in preclinical models of Philadelphia chromosome positive B-cell acute lymphoblastic leukemia is highly active. The dasatinib and anti-PD-1 combination reduces tumor burden, is efficacious, and extends survival. Mechanistically, we found that treatment with dasatinib significantly increased MHC II expression on the surface of antigen-presenting cells (APC) in a tumor microenvironment-independent fashion and caused influx of APC cells into the leukemic bone marrow. Finally, the induction of MHC II may potentiate immune memory by impairing leukemic engraftment in mice previously cured with dasatinib, after re-inoculation of leukemia cells. In summary, our data suggests that anti-PD-1 therapy may enhance the killing ability of dasatinib via dasatinib driven APC growth and expansion and upregulation of MHC II expression, leading to antileukemic immune rewiring.

PD-L1 and CD58 co-regulated by CMTM6 play yin and yang to shape anti-tumor immunity.

The CD58-CD2 axis regulates T cell-mediated cancer immunity, but little is known about the regulation of CD58. In two recent papers pubished in Cancer Cell, Miao et al. and Ho et al. define a mechanism of CD58 regulation by CMTM6 and show an unexpected yin-yang link between PD-L1 and CD58.

Gasdermins and cancers.

The identification of gasdermin as the executor of pyroptosis has opened new avenues for the study of this process. Although pyroptosis research has mainly focused on immune cells since it was discovered three decades ago, accumulating evidence suggests that pyroptosis plays crucial roles in many biological processes. One example is the discovery of gasdermin-mediated cancer cell pyroptosis (CCP) which has become an important and frontier field in oncology. Recent studies have shown that CCP induction can heat tumor microenvironment (TME) and thereby elicit the robust anti-tumor immunity to suppress tumor growth. As a newly discovered form of tumor cell death, CCP offers promising opportunities for improving tumor treatment and developing new drugs. Nevertheless, the research on CCP is still in its infancy, and the molecular mechanisms underlying the expression, regulation and activation of gasdermins are not yet fully understood. In this review, we summarize the recent progress of gasdermin research in cancer area, and propose that the anti-tumor effect of immune cell pyroptosis (ICP) and CCP depends on their duration, intensity, and the type of cells undergoing pyroptosis within TME.

Ribonuclease 1 Enhances Antitumor Immunity against Breast Cancer by Boosting T cell Activation.

The secretory enzyme human ribonuclease 1 (RNase1) is involved in innate immunity and anti-inflammation, achieving host defense and anti-cancer effects; however, whether RNase1 contributes to adaptive immune response in the tumor microenvironment (TME) remains unclear. Here, we established a syngeneic immunocompetent mouse model in breast cancer and demonstrated that ectopic RNase1 expression significantly inhibited tumor progression. Overall changes in immunological profiles in the mouse tumors were analyzed by mass cytometry and showed that the RNase1-expressing tumor cells significantly induced CD4+ Th1 and Th17 cells and natural killer cells and reduced granulocytic myeloid-derived suppressor cells, supporting that RNase1 favors an antitumor TME. Specifically, RNase1 increased expression of T cell activation marker CD69 in a CD4+ T cell subset. Notably, analysis of cancer-killing potential revealed that T cell-mediated antitumor immunity was enhanced by RNase1, which further collaborated with an EGFR-CD3 bispecific antibody to protect against breast cancer cells across molecular subtypes. Our results uncover a tumor-suppressive role of RNase1 through adaptive immune response in breast cancer in vivo and in vitro, providing a potential treatment strategy of combining RNase1 with cancer immunotherapies for immunocompetent patients.