THP-1 Monocytic Cells Are Polarized to More Antitumorigenic Macrophages by Serial Treatment with Phorbol-12-Myristate-13-Acetate and PD98059.

Background and Objectives: As modulators of the tumor microenvironment, macrophages have been extensively studied for their potential in developing anticancer strategies, particularly in regulating macrophage polarization towards an antitumorigenic (M1) phenotype rather than a protumorigenic (M2) one in various experimental models. Here, we evaluated the effect of PD98059, a mitogen-activated protein kinase kinase MAPKK MEK1-linked pathway inhibitor, on the differentiation and polarization of THP-1 monocytes in response to phorbol-12-myristate-13-acetate (PMA) under various culture conditions for tumor microenvironmental application. Materials and Methods: Differentiation and polarization of THP-1 were analyzed by flow cytometry and RT-PCR. Polarized THP-1 subsets with different treatment were compared by motility, phagocytosis, and so on. Results: Clearly, PMA induced THP-1 differentiation occurs in adherent culture conditions more than nonadherent culture conditions by increasing CD11b expression up to 90%, which was not affected by PD98059 when cells were exposed to PMA first (post-PD) but inhibited when PD98059 was treated prior to PMA treatment (pre-PD). CD11bhigh THP-1 cells treated with PMA and PMA-post-PD were categorized into M0 (HLA-DRlow and CD206low), M1 (HLA-DRhigh and CD206low), and M2 (HLA-DRlow and CD206high), resulting in an increased population of M1 macrophages. The transcription levels of markers of macrophage differentiation and polarization confirmed the increased M1 polarization of THP-1 cells with post-PD treatment rather than with PMA-only treatment. The motility and cytotoxicity of THP-1 cells with post-PD treatment were higher than THP-1 cells with PMA, suggesting that post-PD treatment enhanced the anti-tumorigenicity of THP-1 cells. Confocal microscopy and flow cytometry showed the effect of post-PD treatment on phagocytosis by THP-1 cells. Conclusions: We have developed an experimental model of macrophage polarization with THP-1 cells which will be useful for further studies related to the tumor microenvironment.

Loss of zinc-finger protein 143 contributes to tumour progression by interleukin-8-CXCR axis in colon cancer.

Several studies have shown that expression of zinc-finger protein 143 (ZNF143) is closely related to tumour progression including colon cancer. However, it remains unclear how ZNF143 expression is related to tumour progression within the tumour microenvironment. Here, we investigated whether ZNF143 expression affects the tumour microenvironment and tumour progression by screening molecules secreted by colon cancer cells stably expressing short-hairpin RNAs against ZNF143 or control RNAs. We observed that secretion of interleukin (IL)-8 was increased when ZNF143 expression was reduced in two colon cancer cell lines. The mRNA and protein levels of IL-8 were increased in cells following ZNF143 knockdown, and this effect was reversed when ZNF143 expression was restored. The Janus tyrosine kinase/signal transducer and activator of transcription (JAK/STAT) and extracellular signal-regulated kinase pathways were also shown to contribute to IL-8 expression in ZNF143-knockdown cells. The expression levels of ZNF143 and IL-8 were inversely correlated with three-dimensionally grown spheroids and colon cancer tissues. THP-1 cells were differentiated when cells were incubated with condition media from colon cancer cell with less ZNF143, drastically. Loss of ZNF143 may contribute to the development of colon cancer by regulating intracellular and intercellular signalling for cell plasticity and the tumour microenvironment respectively.

Integrated molecular characterization of adult soft tissue sarcoma for therapeutic targets.

BACKGROUND: Several studies have investigated the molecular drivers and therapeutic targets in adult soft tissue sarcomas. However, such studies are limited by the genomic heterogeneity and rarity of sarcomas, particularly in those with complex and unbalanced karyotypes. Additional biomarkers are needed across sarcoma types to improve therapeutic strategies. To investigate the molecular characteristics of complex karyotype sarcomas (CKSs) for therapeutic targets, we performed genomic profiling. RESULTS: The mutational landscape showed that TP53, ATRX, and PTEN genes were highly mutated. CKS samples were categorized into three groups based on copy number variations that were associated with CDK4 and RB1 signatures. Integrated analysis of genomic and transcriptomic data revealed several pathways related to PDGFR, which could be a strategic target for anti-sarcoma therapy. CONCLUSIONS: This study provides a detailed molecular classification of CKSs and proposes several therapeutic targets. Targeted or combinational therapies for treating CKS should be considered before chemotherapy.

A role of zinc-finger protein 143 for cancer cell migration and invasion through ZEB1 and E-cadherin in colon cancer cells.

To investigate the role of zinc-finger protein 143 in cancer cells, we stably introduced ZNF143 expression knockdown by infecting colon cancer cells with short hairpin (sh) RNA-lentiviral particles against ZNF143 (HCT116 sh-ZNF143). Compared to sh-control cells, HCT116 sh-ZNF143 cells showed faster wound healing, increased migration through Transwell chambers, and increased invasion through Matrigel in Transwell chambers. ZNF143 knockdown increased transcriptional expression of ZEB1. Additionally, ZNF143 regulated E-cadherin transcriptional expression. Small interfering-RNA-mediated silencing of ZEB1 expression affected motility in HCT116 sh-ZNF143 cells. These data suggest that ZNF143 is involved in cellular motility through a ZEB1-E-cadherin-linked pathway in colon cancer cells.

Sarcoma Immunotherapy: Confronting Present Hurdles and Unveiling Upcoming Opportunities.

Sarcomas are rare and heterogeneous mesenchymal neoplasms originating from the bone or soft tissues, which pose significant treatment challenges. The current standard treatment for sarcomas consists of surgical resection, often combined with chemo- and radiotherapy; however, local recurrence and metastasis remain significant concerns. Although immunotherapy has demonstrated promise in improving long-term survival rates for certain cancers, sarcomas are generally considered to be relatively less immunogenic than other tumors, presenting substantial challenges for effective immunotherapy. In this review, we examine the possible opportunities for sarcoma immunotherapy, noting cancer testis antigens expressed in sarcomas. We then cover the current status of immunotherapies in sarcomas, including progress in cancer vaccines, immune checkpoint inhibitors, and adoptive cellular therapy and their potential in combating these tumors. Furthermore, we discuss the limitations of immunotherapies in sarcomas, including a low tumor mutation burden and immunosuppressive tumor microenvironment, and explore potential strategies to tackle the immunosuppressive barriers in therapeutic interventions, shedding light on the development of effective and personalized treatments for sarcomas. Overall, this review provides a comprehensive overview of the current status and potential of immunotherapies in sarcoma treatment, highlighting the challenges and opportunities for developing effective therapies to improve the outcomes of patients with these rare malignancies.

New established cell lines from undifferentiated pleomorphic sarcoma for in vivo study.

As a high-grade soft-tissue sarcoma (STS), undifferentiated pleomorphic sarcoma (UPS) is highly recurrent and malignant. UPS is categorized as a tumor of uncertain differentiation and has few options for treatment due to its lack of targetable genetic alterations. There are also few cell lines that provide a representative model for UPS, leading to a dearth of experimental research. Here, we established and characterized new cell lines derived from two recurrent UPS tissues. Cells were obtained from UPS tissues by mincing, followed by extraction or dissociation using enzymes and culture in a standard culture environment. Cells were maintained for several months without artificial treatment, and some cell clones were found to be tumorigenic in an immunodeficient mouse model. Interestingly, some cells formed tumors in vivo when injected after aggregation in a non-adherent culture system for 24 h. The tissues from in vivo study and tissues from patients shared common histological characteristics. Pathways related to the cell cycle, such as DNA replication, were enriched in both cell clones. Pathways related to cell-cell adhesion and cell-cell signaling were also enriched, suggesting a role of the mesenchymal-to-epithelial transition for tumorigenicity in vivo. These new UPS cell lines may facilitate research to identify therapeutic strategies for UPS. [BMB Reports 2023; 56(4): 258-264].

USP15 and USP4 facilitate lung cancer cell proliferation by regulating the alternative splicing of SRSF1.

The deubiquitinating enzyme USP15 is implicated in several human cancers by regulating different cellular processes, including splicing regulation. However, the underlying molecular mechanisms of its functional relevance and the successive roles in enhanced tumorigenesis remain ambiguous. Here, we found that USP15 and its close paralog USP4 are overexpressed and facilitate lung cancer cell proliferation by regulating the alternative splicing of SRSF1. Depletion of USP15 and USP4 impair SRSF1 splicing characterized by the replacement of exon 4 with non-coding intron sequences retained at its C-terminus, resulting in an alternative isoform SRSF1-3. We observed an increased endogenous expression of SRSF1 in lung cancer cells as well, and its overexpression significantly enhanced cancer cell phenotype and rescued the depletion effect of USP15 and USP4. However, the alternatively spliced isoform SRSF1-3 was deficient in such aspects for its premature degradation through nonsense-mediated mRNA decay. The increased USP15 expression contributes to the lung adenocarcinoma (LUAD) development and shows significantly lower disease-specific survival of patients with USP15 alteration. In short, we identified USP15 and USP4 as key regulators of SRSF1 alternative splicing with altered functions, which may represent the novel prognostic biomarker as well as a potential target for LUAD.

Molecular Mechanisms and Current Treatment Options for Cancer Cachexia.

Cancer cachexia is a condition marked by functional, metabolic, and immunological dysfunctions associated with skeletal muscle (SM) atrophy, adipose tissue loss, fat reduction, systemic inflammation, and anorexia. Generally, the condition is caused by a variety of mediators produced by cancer cells and cells in tumor microenvironments. Myostatin and activin signaling, IGF-1/PI3K/AKT signaling, and JAK-STAT signaling are known to play roles in cachexia, and thus, these pathways are considered potential therapeutic targets. This review discusses the current state of knowledge of the molecular mechanisms underlying cachexia and the available therapeutic options and was undertaken to increase understanding of the various factors/pathways/mediators involved and to identify potential treatment options.

TJP1 Contributes to Tumor Progression through Supporting Cell-Cell Aggregation and Communicating with Tumor Microenvironment in Leiomyosarcoma.

Leiomyosarcoma (LMS) is a mesenchymal malignancy with a complex karyotype. Despite accumulated evidence, the factors contributing to the development of LMS are unclear. Here, we investigated the role of tight-junction protein 1 (TJP1), a membrane-associated intercellular barrier protein during the development of LMS and the tumor microenvironment. We orthotopically transplanted SK-LMS-1 cells and their derivatives in terms of TJP1 expression by intramuscular injection, such as SK-LMS-1 Sh-Control cells and SK-LMS-1 Sh-TJP1. We observed robust tumor growth in mice transplanted with LMS cell lines expressing TJP1 while no tumor mass was found in mice transplanted with SK-LMS-1 Sh-TJP1 cells with silenced TJP1 expression. Tissues from mice were stained and further analyzed to clarify the effects of TJP1 expression on tumor development and the tumor microenvironment. To identify the TJP1-dependent factors important in the development of LMS, genes with altered expression were selected in SK-LMS-1 cells such as cyclinD1, CSF1 and so on. The top 10% of highly expressed genes in LMS tissues were obtained from public databases. Further analysis revealed two clusters related to cell proliferation and the tumor microenvironment. Furthermore, integrated analyses of the gene expression networks revealed correlations among TJP1, CSF1 and CTLA4 at the mRNA level, suggesting a possible role for TJP1 in the immune environment. Taken together, these results imply that TJP1 contributes to the development of sarcoma by proliferation through modulating cell-cell aggregation and communication through cytokines in the tumor microenvironment and might be a beneficial therapeutic target.

Leukotriene B4 receptor-2 contributes to KRAS-driven lung tumor formation by promoting interleukin-6-mediated inflammation.

Although lung cancer is the leading cause of cancer-related deaths worldwide and KRAS is the most frequently mutated oncogene in lung cancer cases, the mechanism by which KRAS mutation drives lung cancer has not been fully elucidated. Here, we report that the expression levels of leukotriene B4 receptor-2 (BLT2) and its ligand-producing enzymes (5-LOX, 12-LOX) were highly increased by mutant KRAS and that BLT2 or 5-/12-LOX blockade attenuated KRAS-driven lung cell proliferation and production of interleukin-6 (IL-6), a principal proinflammatory mediator of lung cancer development. Next, we explored the roles of BLT2 and 5-/12-LOX in transgenic mice with lung-specific expression of mutant KRAS (KrasG12D) and observed that BLT2 or 5-/12-LOX inhibition decreased IL-6 production and tumor formation. To further determine whether BLT2 is involved in KRAS-driven lung tumor formation, we established a KrasG12D/BLT2-KO double-mutant mouse model. In the double-mutant mice, we observed significantly suppressed IL-6 production and lung tumor formation. Additionally, we observed high BLT2 expression in tissue samples from patients with KrasG12D-expressing lung adenocarcinoma, supporting the contributory role of BLT2 in KRAS-driven human lung cancer. Collectively, our results suggest that BLT2 is a potential contributor to KRAS-driven lung cancer and identify an attractive therapeutic target for KRAS-driven lung cancer.

Characterization of Proteins Regulated by Androgen and Protein Kinase a Signaling in VCaP Prostate Cancer Cells.

Androgen signaling via the androgen receptor (AR) is involved in normal prostate development and prostate cancer progression. In addition to androgen binding, a variety of protein kinases, including cyclic AMP-dependent protein kinase A (PKA), can activate the AR. Although hormone deprivation, especially that of androgen, continues to be an important strategy for treating prostate cancer patients, the disease ultimately progresses to castration-resistant prostate cancer (CRPC), despite a continuous hormone-deprived environment. To date, it remains unclear which pathways in this progression are active and targetable. Here, we performed a proteomic analysis of VCaP cells stimulated with androgen or forskolin to identify proteins specific for androgen-induced and androgen-bypassing signaling, respectively. Patterns of differentially expressed proteins were quantified, and eight proteins showing significant changes in expression were identified. Functional information, including a Gene Ontology analysis, revealed that most of these proteins are involved in metabolic processes and are associated with cancer. The mRNA and protein expression of selected proteins was validated, and functional correlations of identified proteins with signaling in VCaP cells were assessed by measuring metabolites related to each enzyme. These analyses offered new clues regarding effector molecules involved in prostate cancer development, insights that are supported by the demonstration of increased expression levels of the eight identified proteins in prostate cancer patients and assessments of the progression-free interval. Taken together, our findings show that aberrant levels of eight proteins reflect molecular changes that are significantly regulated by androgen and/or PKA signaling pathways, suggesting possible molecular mechanisms of CRPC.

Rapid Androgen-Responsive Proteome Is Involved in Prostate Cancer Progression.

Androgen exerts its functions by binding with an androgen receptor (AR). It can activate many signaling pathways that are important to the progression of castration-resistant prostate cancer (CRPC). Here, we characterized the rapid proteomic changes seen at 5, 15, 30, and 60 min after the androgen treatment of VCaP cells via the tandem mass tag (TMT) labeling strategy. A total of 5529 proteins were successfully identified and quantified. Dynamic time profiling of protein expression patterns allowed us to identify five protein clusters involved in various stages of androgen-initiated signal transmission and processing. More details of protein functions and localization patterns, and our elucidation of an AR-interacting protein network, were obtained. Finally, we validated the expression level of AR-regulated proteins known to be significantly regulated in CRPC patients using the mouse xenograft model and patient samples. Our work offers a systematic analysis of the rapid proteomic changes induced by androgen and provides a global view of the molecular mechanisms underlying CRPC progression.

O-GlcNAcylation of Sox2 at threonine 258 regulates the self-renewal and early cell fate of embryonic stem cells.

Sox2 is a core transcription factor in embryonic stem cells (ESCs), and O-GlcNAcylation is a type of post-translational modification of nuclear-cytoplasmic proteins. Although both factors play important roles in the maintenance and differentiation of ESCs and the serine 248 (S248) and threonine 258 (T258) residues of Sox2 are modified by O-GlcNAcylation, the function of Sox2 O-GlcNAcylation is unclear. Here, we show that O-GlcNAcylation of Sox2 at T258 regulates mouse ESC self-renewal and early cell fate. ESCs in which wild-type Sox2 was replaced with the Sox2 T258A mutant exhibited reduced self-renewal, whereas ESCs with the Sox2 S248A point mutation did not. ESCs with the Sox2 T258A mutation heterologously introduced using the CRISPR/Cas9 system, designated E14-Sox2TA/WT, also exhibited reduced self-renewal. RNA sequencing analysis under self-renewal conditions showed that upregulated expression of early differentiation genes, rather than a downregulated expression of self-renewal genes, was responsible for the reduced self-renewal of E14-Sox2TA/WT cells. There was a significant decrease in ectodermal tissue and a marked increase in cartilage tissue in E14-Sox2TA/WT-derived teratomas compared with normal E14 ESC-derived teratomas. RNA sequencing of teratomas revealed that genes related to brain development had generally downregulated expression in the E14-Sox2TA/WT-derived teratomas. Our findings using the Sox2 T258A mutant suggest that Sox2 T258 O-GlcNAc has a positive effect on ESC self-renewal and plays an important role in the proper development of ectodermal lineage cells. Overall, our study directly links O-GlcNAcylation and early cell fate decisions.

Chronic activation of 4-1BB signaling induces granuloma development in tumor-draining lymph nodes that is detrimental to subsequent CD8+ T cell responses.

The antitumor capabilities of agonistic anti-4-1BB mAbs have made them an attractive target for tumor immunotherapy. However, the adverse side effects associated with agonist antibodies have hindered their clinical development. Here, we aimed to study the immune-related adverse events of repeated doses and long-term use of agonistic anti-4-1BB mAbs. We show that chronic activation of 4-1BB signals induced the accumulation of IFN-γ-producing PD-1+CD8+ T cells in the secondary lymphoid organs of tumor-bearing mice by increasing the number of dividing CD8+ T cells, which was beneficial for suppressing tumor growth in the early phase of anti-4-1BB induction. However, repeated exposure to anti-4-1BB mAbs led to granuloma development in tumor-draining lymph nodes (TDLNs) of mice due to recruitment and accumulation of macrophages via the CD8+ T cell-IFN-γ axis. This was accompanied by excessive lymph node swelling, which impaired the sequential activation of CD8+ T cells. Our data provide insights into the immune-related adverse events of long-term agonist 4-1BB antibody dosing, which should be considered during the clinical development of immunomodulating therapy.

The Role of Nuclear Receptor Subfamily 1 Group H Member 4 (NR1H4) in Colon Cancer Cell Survival through the Regulation of c-Myc Stability.

Nuclear receptor subfamily group H member 4 (NR1H4), also known as farnesoid X receptor, has been implicated in several cellular processes in the liver and intestine. Preclinical and clinical studies have suggested a role of NR1H4 in colon cancer development; however, how NR1H4 regulates colon cancer cell growth and survival remains unclear. We generated NR1H4 knockout (KO) colon cancer cells using clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein-9 nuclease (CAS9) technology and explored the effects of NR1H4 KO in colon cancer cell proliferation, survival, and apoptosis. Interestingly, NR1H4 KO cells showed impaired cell proliferation, reduced colony formation, and increased apoptotic cell death compared to control colon cancer cells. We identified MYC as an important mediator of the signaling pathway alterations induced by NR1H4 KO. NR1H4 silencing in colon cancer cells resulted in reduced MYC protein levels, while NR1H4 activation using an NR1H4 ligand, chenodeoxycholic acid, resulted in time- and dose-dependent MYC induction. Moreover, NR1H4 KO enhanced the anti-cancer effects of doxorubicin and cisplatin, supporting the role of MYC in the enhanced apoptosis observed in NR1H4 KO cells. Taken together, our findings suggest that modulating NR1H4 activity in colon cancer cells might be a promising alternative approach to treat cancer using MYC-targeting agents.

Targeting TJP1 attenuates cell-cell aggregation and modulates chemosensitivity against doxorubicin in leiomyosarcoma.

Tight junction protein 1 (TJP1) is a membrane-associated cytosolic protein important for cell-cell communication in intercellular barriers in epithelial and non-epithelial cells. Here, we explored the functional involvement of TJP1 in non-epithelial tumors such as soft tissue sarcoma, especially in leiomyosarcoma (LMS). TJP1 expression in soft tissue sarcoma was analyzed in normal and tumor tissues as well as from public datasets such as the TCGA provisional dataset, in which TJP1 expression was compared with other subtypes such as undifferentiated sarcomas, and myxofibrosarcomas. SK-LMS-1 cell lines with reduced TJP1 expression showed attenuated anchorage-independent colony formation as well as reduced intercellular aggregation on non-coated culture plates compared with control as well as parental SK-LMS-1 cells. Transcriptome profiling following TJP1 knockdown in SK-LMS-1 cells suggested the involvement of several signaling pathways, including NF-κB pathway and growth factor receptor signaling. In addition, TJP1 downregulation induced enhanced response against anti-cancer agents, doxorubicin and gefitinib. Taken together, these results suggest that TJP1 contributes to sarcoma genesis and might be useful therapeutic target. KEY MESSAGES: • TJP1 expression at RNA level higher in tumor than in normal tissues of sarcoma. • Targeting TJP1 attenuates cell-cell aggregation and anchorage-independent growth. • Targeting TJP1 is beneficial in anti-cancer therapy in LMS.

The type III transforming growth factor-beta receptor negatively regulates nuclear factor kappa B signaling through its interaction with beta-arrestin2.

Transforming growth factor-beta (TGF-beta) increases or decreases nuclear factor kappa B (NFkappaB) signaling in a context-dependent manner through mechanisms that remain to be defined. The type III transforming growth factor-beta receptor (TbetaRIII) is a TGF-beta superfamily co-receptor with emerging roles in both mediating and regulating TGF-beta superfamily signaling. We have previously reported a novel interaction of TbetaRIII with the scaffolding protein, beta-arrestin2, which results in TbetaRIII internalization and downregulation of TGF-beta signaling. beta-arrestin2 also scaffolds interacting receptors with the mitogen-activated protein kinase and NFkappaB-signaling pathways. Here, we demonstrate that TbetaRIII, through its interaction with beta-arrestin2, negatively regulates NFkappaB signaling in MCF10A breast epithelial and MDA-MB-231 breast cancer cells. Increasing TbetaRIII expression reduced NFkappaB-mediated transcriptional activation and IkappaBalpha degradation, whereas a TbetaRIII mutant unable to interact with beta-arrestin2, TbetaRIII-T841A, had no effect. In a reciprocal manner, short hairpin RNA-mediated silencing of either TbetaRIII expression or beta-arrestin2 expression increased NFkappaB-mediated transcriptional activation and IkappaBalpha degradation. Functionally, TbetaRIII-mediated repression of NFkappaB signaling is important for TbetaRIII-mediated inhibition of breast cancer cell migration. These studies define a mechanism through which TbetaRIII regulates NFkappaB signaling and expand the roles of this TGF-beta superfamily co-receptor in regulating epithelial cell homeostasis.

Prediction of IDH1 Mutation Status in Glioblastoma Using Machine Learning Technique Based on Quantitative Radiomic Data.

OBJECTIVE: Isocitrate dehydrogenase 1 (IDH1) mutation status is an independent favorable prognostic factor for glioblastoma (GBM) and is usually determined by sequencing or immunohistochemistry. An accurate prediction of IDH1 mutation status via noninvasive methods helps establish the appropriate treatment strategy. We aimed to predict IDH1 mutation status using quantitative radiomic data in patients with GBM. METHODS: Between May 2010 and June 2015, we retrospectively identified 88 patients with newly diagnosed GBM. After semiautomatic segmentation of the lesions, we extracted 31 features from preoperative multiparametric magnetic resonance images. We also determined IDH1 mutation status using targeted sequencing and immunohistochemistry. A training cohort (n = 88) was used to train machine learning-based classifiers, with internal validation. The machine-learning technique was then validated in an external dataset of 35 patients with GBM. RESULTS: We detected the IDH1 mutation in 12 out of 88 GBMs. Multiparametric radiomic profiles revealed that the IDH1 mutation was associated with a smaller enhancing area volume and a larger necrotic area volume. Using the machine learning-based classification algorithms, we identified 70.3%-87.3% of prediction rate of IDH1 mutation status and found 66.3%-83.4% accuracy in the external validation set. CONCLUSIONS: We demonstrate that machine learning algorithms can predict IDH1 mutation status in GBM using preoperative multiparametric magnetic resonance images.

The Role of ZNF143 in Breast Cancer Cell Survival Through the NAD(P)H Quinone Dehydrogenase 1⁻p53⁻Beclin1 Axis Under Metabolic Stress.

Autophagy is a cellular process that disrupts and uses unnecessary or malfunctioning components for cellular homeostasis. Evidence has shown a role for autophagy in tumor cell survival, but the molecular determinants that define sensitivity against autophagic regulation in cancers are not clear. Importantly, we found that breast cancer cells with low expression levels of a zinc-finger protein, ZNF143 (MCF7 sh-ZNF143), showed better survival than control cells (MCF7 sh-Control) under starvation, which was compromised with chloroquine, an autophagy inhibitor. In addition, there were more autophagic vesicles in MCF7 sh-ZNF143 cells than in MCF7 sh-Control cells, and proteins related with the autophagic process, such as Beclin1, p62, and ATGs, were altered in cells with less ZNF143. ZNF143 knockdown affected the stability of p53, which showed a dependence on MG132, a proteasome inhibitor. Data from proteome profiling in breast cancer cells with less ZNF143 suggest a role of NAD(P)H quinone dehydrogenase 1(NQO1) for p53 stability. Taken together, we showed that a subset of breast cancer cells with low expression of ZNF143 might exhibit better survival via an autophagic process by regulating the p53⁻Beclin1 axis, corroborating the necessity of blocking autophagy for the best therapy.

A Mutation in ZNF143 as a Novel Candidate Gene for Endothelial Corneal Dystrophy.

Corneal dystrophies (CDs) are a diverse group of inherited disorders with a heterogeneous genetic background. Here, we report the identification of a novel ZNF143 heterozygous missense mutation in three individuals of the same family with clinical and pathological features that are consistent with endothelial CD. Ophthalmologic examination revealed diffuse corneal clouding and edema with decreased endothelial cell density. Pathological findings showed increased corneal thickness due to edema of basal epithelial cells and stroma, and abnormal metaplastic endothelium with stratified epithelium-like changes. Patients' metaplastic corneal endothelial cells expressed predominantly cytokerain 7, cytokeratin 19, and E-cadherin. Although Sanger sequencing did not detect any mutation associated with endothelial CDs, whole exome sequencing identified the ZNF143 c.937G>C p.(Asp313His) mutation as a candidate gene for our patients' endothelial CD. In-vitro functional studies demonstrated that mutant ZNF143 promoted the mesenchymal-to-epithelial transition; it upregulated the expression of genes associated with epithelialization in human corneal endothelial cells. Additionally, proinflammatory cytokine responsive genes were significantly enriched after mutant ZNF143 transfection, which may contribute to the severe phenotype of the three patients. These findings link a mutation in ZNF143 with endothelial CD for the first time.