DACH1 Expression Research Articles

DACH1 Attenuates Airway Inflammation in COPD by Activating NRF2 Signaling.

Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disease of the airways, it is characterized by impaired lung function induced by cigarette smoke (CS). Reduced DACH1 expression has a detrimental role in numerous disorders. However, its role in COPD remains understudied. This study aims to elucidate the role and underlying mechanism of DACH1 in airway inflammation of COPD. DACH1 expression was measured in lung tissues of patients with COPD. Airway epithelium-specific DACH1 knockdown mice and AAV-transfected DACH1 overexpressed mice were used to investigate its role and potential for therapeutic targeting in experimental COPD caused by CS. Furthermore, we discovered a potential mechanism of DACH1 in inflammation induced by cigarette smoke extract simulation (CSE) in vitro. Compared to non-smokers and smokers without COPD, COPD patients had reduced DACH1 expression, especially in the airway epithelium. Airway epithelium-specific DACH1 knockdown aggravated mice airway inflammation and lung function decline caused by CS, whereas DACH1 overexpression protected mice from airway inflammation and lung function decline. DACH1 knockdown and overexpression promoted and inhibited IL-6 and IL-8 secretion in 16 HBE cells after CSE simulation, respectively. Nuclear factor erythroid 2-related factor 2 (NRF2) was discovered to be a novel downstream target of DACH1, which binds directly to its promoter. By activating NRF2 signaling, DACH1 induction reduced inflammation. DACH1 levels are lower in smokers and nonsmoking COPD patients when compared to nonsmokers. DACH1 has protective effects against inflammation induced by CS by activating NRF2 signaling pathway. Targeting DACH1 is a potentially viable therapeutic approach for COPD treatment.

Abstract 5581: The DACH1 gene, frequently co-deleted with BRCA2 in prostate cancer, governs PARP inhibitor resistance

Abstract Background: Prostate cancer (PCa), the second leading cause of death in American men, includes distinct genetic subtypes with distinct therapeutic vulnerabilities (1,2). The DACH1 gene, in the 13q21.31-q21.33 region, encodes a winged helix/Forkhead DNA-binding protein that governs cell-fate determination that is deleted in ~ 18% of human PCa, where it is associated with poor prognosis (3). Prostate-specific deletion of the Dach1 gene enhanced prostatic intraepithelial neoplasia (PIN). Methods: Analysis of DACH1 gene deletion and expression was conducted from public data bases and human prostate cancer samples. Cells derived from Dach1 gene deletion mice and human prostate cancer cell lines with DACH1 overexpression were analyzed for PARP inhibitor responses and mechanisms of PARPi resistance. Transgenic prostate Oncomice were generated in which the Dach1 gene was deleted in the prostate. Results: Herein, DACH1 was shown to be co-deleted with BRCA2 in each of 8 separate cohorts (N=2,186 patients), in ~3-12% of patients. Dach1 gene deletion or knockdown conferred PARP inhibitor resistance (talazoparib > niraparib > olaparib > rucaparib > veliparib). DACH1 enhanced PARP sensitivity through several mechanisms affecting replication stress, PARP1 binding, inhibiting G9a (reducing H3K9me2), inhibiting CHK1p and CDK1p and inducing expression of snoRNA, specifically the snoRNA known to bind PARP. PARP1 has a BRCT (BRCA1 C-terminus 1) homology domain governing PARP1 dimerization and binding to intact DNA in a complex within a nucleosome. DACH1 binding to PARP1 requires the BRCT domain. Conclusions: There is a correlation between DACH1a-/- and PARPi resistance, correlating with PARPi trapping ability. As reduced Dach1a expression may define a subclass of PCa that warrants specific therapies, testing for DACH1a may be warranted.

S100A8/A9 as a risk factor for breast cancer negatively regulated by DACH1

BackgroundS100A8 and S100A9 are members of Ca2+-binding EF-hand superfamily, mainly expressed by macrophages and neutrophils. Limited by the poor stability of homodimers, they commonly exist as heterodimers. Beyond acting as antibacterial cytokines, S100A8/A9 is also associated with metabolic and autoimmune diseases such as obesity, diabetes, and rheumatoid arthritis. While the involvement of S100A8/A9 in breast cancer development has been documented, its prognostic significance and the precise regulatory mechanisms remain unclear.MethodsS100A8/A9 protein in breast cancer samples was evaluated by immunohistochemistry staining with tumor tissue microarrays. The serum S100A8 concentration in patients was measured by enzyme-linked immunosorbent assay (ELISA). The S100A8 secreted by breast cancer cells was detected by ELISA as well. Pooled analyses were conducted to explore the relationships between S100A8/A9 mRNA level and clinicopathological features of breast cancer patients. Besides, the effects of S100A8/A9 and DACH1 on patient outcomes were analyzed by tissue assays. Finally, xenograft tumor assays were adopted to validate the effects of DACH1 on tumor growth and S100A8/A9 expression.ResultsThe level of S100A8/A9 was higher in breast cancer, relative to normal tissue. Increased S100A8/A9 was related to poor differentiation grade, loss of hormone receptors, and Her2 positive. Moreover, elevated S100A8/A9 predicted a worse prognosis for breast cancer patients. Meanwhile, serum S100A8 concentration was upregulated in Grade 3, basal-like, and Her2-overexpressed subtypes. Additionally, the results of public databases showed S100A8/A9 mRNA level was negatively correlated to DACH1. Stable overexpressing DACH1 in breast cancer cells significantly decreased the generation of S100A8. The survival analysis demonstrated that patients with high S100A8/A9 and low DACH1 achieved the shortest overall survival. The xenograft models indicated that DACH1 expression significantly retarded tumor growth and downregulated S100A8/A9 protein abundance.ConclusionS100A8/A9 is remarkedly increased in basal-like and Her2-overexpressed subtypes, predicting poor prognosis of breast cancer patients. Tumor suppressor DACH1 inhibits S100A8/A9 expression. The combination of S100A8/A9 and DACH1 predicted the overall survival of breast cancer patients more preciously.

The DACH1 gene is frequently deleted in prostate cancer, restrains prostatic intraepithelial neoplasia, decreases DNA damage repair, and predicts therapy responses

Prostate cancer (PCa), the second leading cause of death in American men, includes distinct genetic subtypes with distinct therapeutic vulnerabilities. The DACH1 gene encodes a winged helix/Forkhead DNA-binding protein that competes for binding to FOXM1 sites. Herein, DACH1 gene deletion within the 13q21.31-q21.33 region occurs in up to 18% of human PCa and was associated with increased AR activity and poor prognosis. In prostate OncoMice, prostate-specific deletion of the Dach1 gene enhanced prostatic intraepithelial neoplasia (PIN), and was associated with increased TGFβ activity and DNA damage. Reduced Dach1 increased DNA damage in response to genotoxic stresses. DACH1 was recruited to sites of DNA damage, augmenting recruitment of Ku70/Ku80. Reduced Dach1 expression was associated with increased homology directed repair and resistance to PARP inhibitors and TGFβ kinase inhibitors. Reduced Dach1 expression may define a subclass of PCa that warrants specific therapies.

Abstract 2598: The DACH1 gene is frequently deleted in prostate cancer, restrains prostatic intraepithelial neoplasia, augments DNA damage repair and predicts therapy responses

Abstract Background: Prostate cancer (PCa), the second leading cause of death in American men, includes distinct genetic subtypes with distinct therapeutic vulnerabilities. The DACH1 gene encodes a winged helix/Forkhead DNA-binding protein that competes for binding to FOXM1 sites. Methods: Analysis of DACH1 gene deletion and gene expression was conducted from public data bases and human prostate cancer samples. Transgenic mice were generated in which the Dach1 gene was deleted in the prostate of prostate Oncomice. Cells derived from Dach1 gene deletion mice and human prostate cancer cell lines with Dach1 knockdown were analyzed for DNA damage repair responses. Results: DACH1 gene deletion within the 13q21.31-q21.33 region, occurred in up to 18% of human PCa, and was associated with increased AR activity, and poor prognosis. DACH1 homozygous deletions more frequent in the metastatic site than in the primary tumors (Mich: 10% vs. 18%, N=59; FHCRC: 4% vs.11%, N= 54, SU2C: not profiled vs. 3.3, N=150). The prevalence of DACH1 heterozygous deletions was higher in the metastatic lesions than in primary tumors within a given cohort for three of six cohorts (Mich: 27.3% vs. 36%, N=59; FHCRC: 15% vs. 59%, N=54; SU2C: 0% vs. 26%, N=150, respectively). The patients with homozygous DACH1 deletions had reduced overall survival (medians of 84 vs. 120 months, N=667, log rank test P=9.3x10-3). Low DACH1 gene expression (expressed as a z-score with a z-score threshold of -1.25) was significantly correlated with earlier biochemical recurrence (BCR, log rank p value = 4.7x10-4, n=79). In prostate OncoMice, prostate-specific deletion of the Dach1 gene enhanced prostatic intraepithelial neoplasia (PIN) and was associated with increased DNA damage. Reduced Dach1 increased DNA damage in responses to genotoxic stresses. DACH1 was recruited to sites of DNA damage, augmenting recruitment of Ku70/Ku80. Reduced Dach1 expression was associated with resistance to TGFβ kinase and PARP inhibitors. Conclusions: Reduced Dach1 expression may define a subclass of PCa that warrants specific therapies. Citation Format: Zhiping Li, Xuanmao Jiao, Gordon Robertson, Gabriele Di Sante, Anthony W. Ashton, Agnese DiRocco, Min Wang, Jun Zhao, Sankar Addya, Chenguang Wang, Peter A. McCue, Andrew P. South, Carlos Cordon-Cardo, Runzhi Liu, Kishan Patel, Rasha Hamid, Jorim Parmar, James B. DuHadaway, Nikolaus Schultz, Andrew Kossenkov, Lai Yee Phoon, Hao Chen, Li Lan, Yunguang Sun, Kenneth A. Iczkowski, Hallgeir Rui, Richard G. Pestell. The DACH1 gene is frequently deleted in prostate cancer, restrains prostatic intraepithelial neoplasia, augments DNA damage repair and predicts therapy responses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2598.

Identification and verification of the temozolomide resistance feature gene DACH1 in gliomas.

The most important chemotherapy treatment for glioma patients is temozolomide. However, the development of drug resistance severely restricts the use of temozolomide. Therefore, elucidating the mechanism of temozolomide resistance, enhancing temozolomide sensitivity, and extending patient survival are urgent tasks for researchers. Temozolomide resistance hub differential genes were identified using differential analysis and protein interaction analysis from the GEO datasets (GSE100736 and GSE113510). These genes were further studied in glioma patients treated with temozolomide in the TCGA and CGGA databases. Patients from the mRNAseq_325 dataset (CGGA) were considered as the training set to construct a risk model for predicting glioma sensitivity to temozolomide, while patients from the mRNAseq_693 dataset (CGGA) and TCGA-GBM dataset were considered as the validation set to evaluate the performance of models. PCR and western blot were performed to determine the difference in expression of the feature gene DACH1 between glioma cells and temozolomide-resistant glioma cells. The alterations in the sensitivity of tumor cells to temozolomide were also observed after DACH1 was silenced. The patients were then divided into two groups based on the expression of DACH1, and the differences in patient survival rates, molecular pathway activation, and level of immune infiltration were compared. Based on four signature genes (AHR, DACH1, MGMT, and YAP1), a risk model for predicting glioma sensitivity to temozolomide was constructed, and the results of timeROC in both the training and validation sets showed that the model had good predictive performance. The expression of the signature gene DACH1 was significantly downregulated in temozolomide-resistant cells, according to the results of the PCR and western blot experiments. The sensitivity of tumor cells to temozolomide was significantly reduced after DACH1 was silenced. DACH1 probably regulates temozolomide resistance in glioblastoma through the transcriptional dysregulation in cancer and ECM. This study constructs a risk model that can predict glioma susceptibility to temozolomide and validates the function of the feature gene DACH1, which provides a promising target for the research of temozolomide resistance.

Study of DACH1 Expression and its Epigenetic Regulators as Possible Breast Cancer-Related Biomarkers

Breast carcinogenesis involves both genetic and epigenetic changes. DNA methylation, as well as micro-RNA regulations, are the significant epigenetic phenomena dysregulated in breast cancer. Herein, the expression of DACH1 as a tumor suppressor gene and its promoter methylation status was analyzed in breast cancer tumors. Also, the expression of three micro RNAs (miR-217, miR-6807-3p, and miR-552), which had been previously reported to target DACH1, was assessed. The SYBR green-based Real-Time reverse transcription-PCR was used to determine DACH1 and micro-RNAs (miR-217, miR-6807-3p, and miR-552) expression in 120 ductal breast cancer tumors compared with standard control. Also, the promoter methylation pattern of DACH1 was investigated using the Methylation-specific PCR technique. DACH1 expression was significantly down-regulated in breast tumors (p<0.05). About 33.5% of tumors showed DACH1 promoter hyper-methylation. The studied micro-RNAs, expression was negatively correlated with DACH1 expression. The highest expressions of miRNAs and higher DACH1 promoter methylation were observed in advanced cancer situations. The Kaplan-Meier survival curves indicated that the overall survival was significantly poor in higher miRNAs and lower DACH1 expression in breast cancer patients (p<0.002). DACH1 down-regulation may be associated with a poor breast cancer prognosis. The DACH1 down-regulation may be due to epigenetic regulations such as promoter methylation, especially in triple-negative cases. Other factors, such as micro-RNAs (miR-217, miR-6807-3p, and miR-552), may also have an impact. The elevated expression of miR-217, miR-6807-3p, and miR-552, maybe candidates as possible poor prognostic biomarkers in breast cancer management for further consideration.

DACH1 inhibits the proliferation and migration of papillary thyroid carcinoma.

DACH1 is an important component of the retinal determinate gene network (RDGN), which regulates the expression of target genes by directly binding or interacting with other factors. DACH1 shows inhibitory effects in most tumors, but its role in papillary thyroid carcinoma is unclear and warrants further investigation. We assessed the expression of DACH1 in different tissues and correlation with immune infiltration by The Cancer Genome Atlas (TCGA) and Tumor Immune Estimation Resource (TIMMER2.0 databases). The effects of DACH1 on the proliferation and migration of TPC-1 and Bcpap cells were assessed by cell viability assay, colony formation assay, wound healing assay, transwell migration assay, and flow cytometry. Finally, the effects of DACH1 on CXCL8, CXCL10, and CXCL12 expression in Nthy-ori-3-1, TPC-1 and Bcpap cells were assessed by enzyme-linked immunosorbent assay kit and real-time polymerase chain reaction, respectively. The results showed that DACH1 was differentially expressed in different tumors and tissues. Basal expression of DACH1 was lower in thyroid and papillary thyroid carcinoma than in other normal tissues and corresponding tumors, and positively correlated with CD8+ T cell infiltration. In Nthy-ori-3-1, TPC-1 and Bcpap cells, overexpression of DACH1 inhibitedcell migration and proliferation, and the opposite results was obtained by knocking down DACH1 using small interfering RNA. We also demonstrated that DACH1 regulated chemokines CXCL8, CXCL10, and CXCL12, thereby modulating tumor immunity.

Abstract 2500: The Forkhead-like protein Dach1 governs sensitivity to WEE1 kinase and PARP inhibitors

Abstract Molecular analysis of all cancer types in the cancer genome atlas (TCGA) demonstrated that multiple pathways may be disrupted within a given tumor including PCa. Prostate cancer (PCa), the second leading cause of death in American men, includes distinct genetic subtypes with distinct therapeutic vulnerabilities. Substratification of PCa into genetic subtypes, forms the basis of rational therapy for PCa. Known genetic drivers to PCa include deletions of target genes (PTEN and NKX3.1 deletions), rearrangements/fusions of multiple genes (including TMPRSS2 and the oncogenic ETS transcription factor, ERG) and genetic predisposing factors (including germline DNA-repair gene mutations). An increasingly common finding in PCa are defects in DNA repair. The Drosophila Dac gene was initially cloned as dominant inhibitor of Elipse. The human homologue, DACH1, encodes a winged helix/Forkhead DNA-binding protein that binds in chromatin to FOXM1 sites. Herein, reduced expression of DACH1 is shown in PCa, correlating with DACH1 promoter methylation. Homozygous deletion of DACH1 occurred in up to 18% of human PCa, associated with increased AR activity, increased somatic copy-number alteration and poor prognosis. DACH1 increased DNA repair, as shown in comet assays, enhancing nonhomologous end joining (NHEJ) while reducing homology directed repair (HDR) in DNA repair reporter assays. Laser wounding showed DACH1 was recruited to sites of DNA damage, augmenting the recruitment of Ku70/Ku80. In Dach1 deletion cells reduced expression of Dach1 was associated with resistance to inhibitors of Poly-(ADP)-ribose polymerase (PARP) and enhanced sensitivity to inhibitors of WEE1 kinase and DNA-PK. The abundance of DACH1 in prostate cancer may define a subclass of prostate cancer warranting selective therapies. Citation Format: Zhiping Li, Xuanmao Jiao, A. Gordon Robertson, Anthony W. Ashton, Andrew Kossenkov, Hao Chen, Lai Yee Phoon, Li Lan, Richard G. Pestell. The Forkhead-like protein Dach1 governs sensitivity to WEE1 kinase and PARP inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2500.

DACH1 protects podocytes from experimental diabetic injury and modulates PTIP-H3K4Me3 activity.

Dachshund homolog 1 (DACH1), a key cell-fate determinant, regulates transcription by DNA sequence-specific binding. We identified diminished Dach1 expression in a large-scale screen for mutations that convert injury-resistant podocytes into injury-susceptible podocytes. In diabetic kidney disease (DKD) patients, podocyte DACH1 expression levels are diminished, a condition that strongly correlates with poor clinical outcomes. Global Dach1 KO mice manifest renal hypoplasia and die perinatally. Podocyte-specific Dach1 KO mice, however, maintain normal glomerular architecture at baseline, but rapidly exhibit podocyte injury after diabetes onset. Furthermore, podocyte-specific augmentation of DACH1 expression in mice protects from DKD. Combined RNA sequencing and in silico promoter analysis reveal conversely overlapping glomerular transcriptomic signatures between podocyte-specific Dach1 and Pax transactivation-domain interacting protein (Ptip) KO mice, with upregulated genes possessing higher-than-expected numbers of promoter Dach1-binding sites. PTIP, an essential component of the activating histone H3 lysine 4 trimethylation (H3K4Me3) complex, interacts with DACH1 and is recruited by DACH1 to its promoter-binding sites. DACH1-PTIP recruitment represses transcription and reduces promoter H3K4Me3 levels. DACH1 knockdown in podocytes combined with hyperglycemia triggers target gene upregulation and increases promoter H3K4Me3. These findings reveal that in DKD, diminished DACH1 expression enhances podocyte injury vulnerability via epigenetic derepression of its target genes.

Abstract PO009: Increased DACH1 mutation frequency in a Kentucky hospital uterine cancer population

Abstract Purpose: DACH1 is a transcriptional repressor and tumor suppressor gene frequently mutated in breast, bladder, and prostate cancer. Most studied in breast carcinoma, DACH1 expression was reduced or lost in invasive breast cancer patients, and its expression was inversely related to tumor diameter, stage, and nodal metastasis, and directly associated with increased survival time. While less studied in uterine cancer, nearly all normal endometrial samples show nuclear expression of DACH1, with DACH1 expression lost in more than half of uterine cancers. Loss of DACH1 expression has been associated with higher surgical stage, positive peritoneal cytology, and lymph node positivity in uterine cancer. In this study, the Oncology Research Information Exchange Network (ORIEN) Avatar database was used to determine the frequency of DACH1 mutations in our Kentucky hospital population with uterine cancer. Experimental Design: Clinical and genomic data were collected for 65 patients with uterine cancer from Markey Cancer Center (MCC) through Total Cancer Care (TCC) ®. TCC is a prospective cohort study with whole-exome tumor sequencing, RNA sequencing, germline sequencing, and lifetime follow-up. The clinical attributes, whole-exome sequencing, RNA sequencing, microsatellite instability, and tumor mutational burden were compared by DACH1 status. EdgeR was used to perform normalization, expression modeling, and difference testing. The network analysis was performed using Qiagen’s Ingenuity Pathway Analysis (IPA) system for core analysis of the RNA sequencing data and overlaid with the Global Molecular Network Overlay in the IPA knowledge base. The Cancer Genome Atlas Project (TCGA) PanCancer Atlas dataset was used for comparison through cBioPortal.org, utilizing endometrial cancer and carcinosarcoma subgroups for analysis totaling 586 patients. Results: Uterine cancer patients at MCC had an increased frequency of DACH1 mutations (12/65 patients, 18.5%) compared to the TCGA uterine cancer population (25/586 patients, 3.8%) with p-value = 1.04E-05. DACH1 mutations were associated with increased tumor mutation count in both TCGA (median 65 vs. 8972, p-value =7.35e-09) and MCC population (490 vs. 2160, p-value = 6E-04). DACH1 mutated patients had a higher tumor mutation burden compared to DACH1 wild-type (24 vs. 6.02, p-value = 4.288E-05). DACH1 mutations showed significant gene co-occurrence patterns with POLE, MLH1, and PMS2. Conclusions: DACH1 mutations are prevalent in Kentucky patients with uterine cancer. These mutations are associated with high tumor mutational burden and co-occur with genome destabilizing gene mutations. These findings suggest DACH1 could be a candidate biomarker for future trials with immunotherapy, particularly in uterine cancers, and may help explain the differences seen in uterine cancer incidence and mortality in Kentucky. Citation Format: McKayla Riggs, Nan Lin, Chi Wang, Dava Piecoro, Rachel Miller, Oliver Hampton, Mahadev Rao, Frederick Ueland, Jill Kolesar. Increased DACH1 mutation frequency in a Kentucky hospital uterine cancer population [abstract]. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO009.

Role of DACH1 on Proliferation, Invasion, and Apoptosis in Human Lung Adenocarcinoma Cells.

To investigate DACH1 protein expression in lung cancer tissue and matched paracancerous tissue and explore its effect on proliferation, invasion, and apoptosis in human lung adenocarcinoma cells (HLACs). Tumor tissue and matched paracancerous tissue were collected from 46 patients with pathologically diagnosed lung cancer. RT-PCR was performed to detect DACH1 mRNA expression and immunohistochemistry to measured DACH1 protein expression. To determine the effect of DACH1 on lung cancer behavior, small interfering RNA (siRNA) was used to silence DACH1 expression in A549 cells. The impact on the proliferation of tumor cells was then observed by MTT assay, changes in the invasion of tumor cells were identified using transwell chamber assay, and the effects on apoptosis in the cell line were detected using flow cytometry. The expression of DACH1 mRNA and DACH1 protein was significantly decreased in lung cancer tissue versus matched paracancerous control tissue. The silencing of DACH1 expression in A549 cells significantly enhanced cell proliferation, significantly increased cell invasion, and significantly reduced spontaneous apoptosis. DACH1 is downregulated in lung adenocarcinoma tissue. In vitro assessment shows that DACH1 functions as a tumor suppressor, suggesting its potential use as a new target for lung cancer treatment.

A new lymphoid-primed progenitor marked by Dach1 downregulation identified with single cell multi-omics.

A classical view of blood cell development is that multipotent hematopoietic stem and progenitor cells (HSPCs) become lineage-restricted at defined stages. Lin-c-Kit+Sca-1+Flt3+ cells, termed lymphoid-primed multipotent progenitors (LMPPs), have lost megakaryocyte and erythroid potential but are heterogeneous in their fate. Here, through single-cell RNA sequencing, we identify the expression of Dach1 and associated genes in this fraction as being coexpressed with myeloid/stem genes but inversely correlated with lymphoid genes. Through generation of Dach1-GFP reporter mice, we identify a transcriptionally and functionally unique Dach1-GFP- subpopulation within LMPPs with lymphoid potential with low to negligible classic myeloid potential. We term these 'lymphoid-primed progenitors' (LPPs). These findings define an early definitive branch point of lymphoid development in hematopoiesis and a means for prospective isolation of LPPs.

RDGN-based predictive model for the prognosis of breast cancer

BackgroundBreast cancer is the most diagnosed malignancy in females in the United States. The members of retinal determination gene network (RDGN) including DACH, EYA, as well as SIX families participate in the proliferation, apoptosis, and metastasis of multiple tumors including breast cancer. A comprehensive predictive model of RDGN might be helpful to herald the prognosis of breast cancer patients.MethodsIn this study, the Gene Expression Ominibus (GEO) and Gene Set Expression Analysis (GSEA) algorithm were used to investigate the effect of RDGN members on downstream signaling pathways. Besides, based on The Cancer Genome Atlas (TCGA) database, we explored the expression patterns of RDGN members in tumors, normal tissues, and different breast cancer subtypes. Moreover, we estimated the relationship between RDGN members and the outcomes of breast cancer patients. Lastly, we constructed a RDGN-based predictive model by Cox proportional hazard regression and verified the model in two separate GEO datasets.ResultsThe results of GSEA showed that the expression of DACH1 was negatively correlated with cell cycle and DNA replication pathways. On the contrary, the levels of EYA2 and SIX1 were significantly positively correlated with DNA replication, mTOR, and Wnt pathways. Further investigation in TCGA database indicated that DACH1 expression was lower in breast cancers especially basal-like subtype. In the meanwhile, SIX1 was remarkably upregulated in breast cancers while EYA2 level was increased in Basal-like and Her-2 enriched subtypes. Survival analyses demonstrated that DACH1 was a favorable factor while EYA2 and SIX1 were risk factors for breast cancer patients. Given the results of Cox proportional hazard regression analysis, two members of RDGN were involved in the present predictive model and patients with high model index had poorer outcomes.ConclusionThis study showed that aberrant RDGN expression was an unfavorable factor for breast cancer. This RDGN-based comprehensively framework was meaningful for predicting the prognosis of breast cancer patients.

CXCL1 as an Unfavorable Prognosis Factor Negatively Regulated by DACH1 in Non-small Cell Lung Cancer.

Background: Interaction between cancer cells with microenvironment is essential for cancer progression, therapeutic resistance and prognosis. Chemokine CXCL1 shows variable roles in the development of cancers. DACH1 has been considered as a tumor suppressor and represses the expressions of several chemokines. The relationship between CXCL1 and DACH1 in non-small cell lung cancer (SCLC) deserves further investigation.Methods: Immunohistochemistry staining was performed on tumor tissue microarrays from lung cancer patients to detect CXCL1 protein. The CXCL1 concentration in the serum of adenocarcinoma patients was measured by ELISA. The CXCL1 protein secreted by cancer cell lines was detected by SearchLight proteome array and human cytokine antibody array. The meta-analysis of CXCL1 expression form public databases was performed and correlation between CXCL1 and DACH1 was analyzed. Moreover, the association between clinicopathological features and prognosis with CXCL1 and DACH1 was analyzed by tissue array and KM-plotter from public database.Results: The protein abundance of CXCL1 in lung cancer tissues was significantly higher than that in adjacent normal tissues. CXCL1 was closely related to TNM stage, tumor size, and lymph node metastasis and predicted worse overall survival in adenocarcinoma. The level of CXCL1 in the peripheral blood of adenocarcinoma patients also significantly elevated and positively related with clinical stage. The meta-analysis demonstrated that CXCL1 mRNA level was increased in lung cancer tissues and high level of CXCL1 indicated tumor progression in lung adenocarcinoma. In addition, public database analyses showed that CXCL1 negatively correlated with DACH1. Stable overexpressing DACH1 in cultured lung cancer cells remarkably decreased CXCL1 protein. Moreover, ectopic expression of DACH1 significantly inhibited the expression of CXCL1, Ki67, and cyclin D1 in tumor tissues compared with A549 cells with empty vector. Survival analysis showed that high CXCL1 and low DACH1 indicated poor overall survival and progression-free survival.Conclusion: CXCL1 is closely associated with tumor progression and poor survival. DACH1 significantly inhibits the expression of CXCL1 and indicates good prognosis. Therefore, combined detection of CXCL1 and DACH1 could more precisely predict prognosis of lung adenocarcinoma.

633P - DACH1 induced stemness of intestinal organoids by directly suppressing bone morphogenetic proteins (BMP) signaling and contributes to intestinal tumorigenesis

BackgroundCrypt base cells are candidates for intestinal epithelial stem cells (ISC). But limited progress has been made concerning the modulation of stem cells. DACH1 is upregulated in LGR5+ ISC cells with stemness maintenance. Hence, we postulated that DACH1 may serve as a putative modulator of intestine stem cells leading to carcinogenesis. MethodsWe established colon normal, adenomas and cancers organoids, which supported stem cell proliferation and undifferentiation in vitro. A stable and efficient culture system for colon epithelial crypts and tumor organoids was use to facilitate investigation of the dynamics and factors affecting colon stem cell niche and carcinogenesis. ResultsWe showed that in the mouse and human intestine, DACH1 was expressed in discrete crypt base. In colorectal cancer, DACH1 expression became widespread, with increased expression and nuclear localization at all stages. ShRNA-mediated inhibition of DACH1 in colon cancer cells inhibited colony formation in vitro and tumor growth in vivo through affecting cancer stem cell signatures. What is more, DACH1 stimulated both colonosphere formation and tumor organoid formation in vitro. Mechanistic characterization indicated that DACH1 recruited and interacts with SMAD4 to suppress BMP signaling pathway. Eventually, DACH1 induced stemness of organoids through BMP-mediated transcriptional increasing of a large number of stem cell signature genes, including Notch1, Msi-2 and Sox9. Here we provided evidence for an essential role of DACH1 in inducing stem cell self-renewal through direct upregulating stem cell signature genes. In detail, DACH1 was shown by microarray analyses to repress BMP induction of Smad signaling, then DACH1 binding to Smad4 was required for repression of Smad signaling ConclusionsIn conclusion, our findings illustrate an essential role of DACH1 signaling in inducing stem cell expansion during intestinal homeostasis and tumor development. This study also provided a mechanistic insight into the function of DACH1 in maintaining stem cell self-renewal via Smad4. Legal entity responsible for the studySan-Jun Cai. FundingHas not received any funding. DisclosureAll authors have declared no conflicts of interest.

DACH1 DOWNREGULATION DELINEATE A LYMPHOID PRIMED PROGENITOR (LPP) POPULATION WITHIN THE MPP4 CLUSTER

Classical models of haematopoiesis dictate that a single hematopoietic stem cell (HSC) with self-renewal capacity can give rise to all the hematopoietic lineage through subsequent division and loss of potential. This loss of potential is captured through surface markers that classify different stem and progenitor stages including long-term (LT-) and short-term haematopoietic stem cells (ST-HSC) that give rise to 4 categories of multipotent progenitor (MPP) subpopulations 1-4. The MPP4 cluster Lin–c-kit+Sca1+Flt3+ is also known as the lymphoid multipotent primed progenitor (LMPPs) and lack of megakaryocyte and erythroid potential. We demonstrate through cellular barcoding and scRNA-seq that the MPP4 cluster is heterogenous for fate. In particular, Dach1 expression is heterogenous and co-expressed with myeloid and stem-like genes but inversely correlates with lymphoid genes. Through the generation of a Dach1-GFP reporter mouse we demonstrate that Dach1– MPP4s represents a novel progenitor subpopulation that definitively lacks myeloid potential but retains lymphoid potential, thus termed lymphoid-primed progenitors or LPPs. This work describes the earliest definitive branch point of lymphoid development in within haematopoiesis and shows the power of scRNA-seq in identifying progenitor subpopulations.

2037 – DACH1 DOWNREGULATION MARKS A ‘LYMPHOID-PRIMED PROGENITOR’ IN EARLY HAEMATOPOIESIS

A classical view of blood cell development is that multipotent haematopoietic stem and progenitor cells (HSPCs) become lineage-restricted at defined stages. The Lin–c-kit+Sca1+Flt3+ stage, termed lymphoid-primed multipotent progenitors (LMPPs), have lost megakaryocyte and erythroid potential but are heterogeneous in their fate. Through single cell RNA-sequencing, we identify heterogeneous expression of Dach1 and associated genes in this fraction where it co-expressed with myeloid/stem genes but inversely correlated with lymphoid genes. Through generation of Dach1-GFP reporter mice, we identify a transcriptionally and functionally unique Dach1– subpopulation within LMPPs with lymphoid potential but devoid of myeloid potential - both in vivo and in vitro. Dach1 based separation is distinct from to separation by Rag1-GFP, Flt3, CD27, CD48 or VCAM. We term these ‘lymphoid-primed progenitors’, or LPPs. These findings define the earliest branch point of lymphoid development in haematopoiesis and a means for their prospective isolation.

Role of Dach1 revealed using a novel inner ear-specific Dach1-knockdown mouse model.

ABSTRACTThe Dach1 gene is expressed in the inner ear of normal mouse embryos in the area that differentiates into the cochlear stria vascularis (SV). We hypothesised that Dach1 downregulation in the inner ear would lead to SV dysplasia. However, because Dach1 knockout is embryonic lethal in mice, the role of Dach1 in the inner ear is unclear. Here, we established inner ear-specific Dach1-knockdown mice and showed that Dach1 downregulation resulted in hearing loss, reduced endocochlear potential and secondary outer hair cell loss. There were no abnormalities in marginal cells and basal cells in the SV or spiral ligament in inner ear-specific Dach1-knockdown mature mice. However, intermediate cell dysplasia and thinning of the SV were observed. Moreover, dynamic changes in the expression of key genes related to the epithelial-mesenchymal transition were observed in the lateral wall of the cochlear epithelium, which differentiated into the SV in inner ear-specific Dach1-knockdown mice at embryonic stages. In summary, suppression of Dach1 expression in the inner ear caused the epithelial-mesenchymal transition in the lateral wall of cochlear epithelium, resulting in loss of intermediate cells in the SV and SV dysplasia.This article has an associated First Person interview with the first author of the paper.

Effect of DACH1 on proliferation and invasion of laryngeal squamous cell carcinoma

BackgroundTo investigate the effect of DACH1 over-expression on proliferation and invasion of laryngeal squamous cell carcinoma (LSCC).MethodsThe 120 cases of LSCC tumors and 114 adjacent non-neoplastic tissues were collected to detect the expression of DACH1 by immunohistochemistry. The changes of DACH1 expression from each group were assessed and correlated to the clinical parameters of the patients. Plasmid-DACH1 was transfected into Hep-2 cells to up-regulate the expression of DACH1C. Real-time PCR, Western blot, CCK8 and transwell assay were used to verify the cell proliferation and invasion after plasmid-DACH1 transfection.ResultsThe results indicated that DACH1 was downregulated in LSCC tissues as compared to corresponding adjacent non-neoplastic tissues. Decreased expression of DACH1 was found in the tumors upraglottic tumor, lymph node metastases, T3–4 stage and advanced clinical stage. In Hep-2 cells, transfection with plasmid-DACH1 could suppress cell proliferation, invasion and induce G1 phase extension in cell cycle.ConclusionsDACH1 may act as a tumor suppressor gene and could be a potential target for therapeutic intervention of LSCC.