Disabled-2 Research Articles

The scaffold protein disabled 2 (DAB2) and its role in tumor development and progression.

Disabled 2 (DAB2) is a multifunctional protein that has emerged as a critical component in the regulation of tumor growth. Its dysregulation is implicated in various types of cancer, underscoring its importance in understanding the molecular mechanisms underlying tumor development and progression. This review aims to unravel the intricate molecular mechanisms by which DAB2 exerts its tumor-suppressive functions within cancer signaling pathways. We conducted a comprehensive review of the literature focusing on the structure, expression, physiological functions, and tumor-suppressive roles of DAB2. We provide an overview of the structure, expression, and physiological functions of DAB2. Evidence supporting DAB2's role as a tumor suppressor is explored, highlighting its ability to inhibit cell proliferation, induce apoptosis, and modulate key signaling pathways involved in tumor suppression. The interaction between DAB2 and key oncogenes is examined, elucidating the interplay between DAB2 and oncogenic signaling pathways. We discuss the molecular mechanisms underlying DAB2-mediated tumor suppression, including its involvement in DNA damage response and repair, regulation of cell cycle progression and senescence, and modulation of epithelial-mesenchymal transition (EMT). The review explores the regulatory networks involving DAB2, covering post-translational modifications, interactions with other tumor suppressors, and integration within complex signaling networks. We also highlight the prognostic significance of DAB2 and its role in pre-clinical studies of tumor suppression. This review provides a comprehensive understanding of the molecular mechanisms by which DAB2 exerts its tumor-suppressive functions. It emphasizes the significance of DAB2 in cancer signaling pathways and its potential as a target for future therapeutic interventions.

Disable 2, A Versatile Tissue Matrix Multifunctional Scaffold Protein with Multifaceted Signaling: Unveiling Role in Breast Cancer for Therapeutic Revolution.

The Disabled-2 (DAB2) protein, found in 80-90% of various tumors, including breast cancer, has been identified as a potential tumor suppressor protein. On the contrary, some hypothesis suggests that DAB2 is associated with the modulation of the Ras/MAPK pathway by endocytosing the Grb/Sos1 signaling complex, which produces oncogenes and chemoresistance to anticancer drugs, leading to increased tumor growth and metastasis. DAB2 has multiple functions in several disorders and is typically under-regulated in several cancers, making it a potential target for treatment of cancer therapy. The primary function of DAB2 is the modulation of transforming growth factor- β (TGF-β) mediated endocytosis, which is involved in several mechanisms of cancer development, including tumor suppression through promoting apoptosis and suppressing cell proliferation. In this review, we will discuss in detail the mechanisms through which DAB2 leads to breast cancer and various advancements in employing DAB2 in the treatment of breast cancer. Additionally, we outlined its role in other diseases. We propose that upregulating DAB2 could be a novel approach to the therapeutics of breast cancer.

STAT3–mediated up-regulation of DAB2 via SRC-YAP1 signaling axis promotes Helicobacter pylori-driven gastric tumorigenesis

BackgroundHelicobacter pylori (H pylori) infection is the primary cause of gastric cancer (GC). The role of Disabled-2 (DAB2) in GC remains largely unclear. This study aimed to investigate the role of DAB2 in H pylori-mediated gastric tumorigenesis.MethodsWe screened various datasets of GC to analyze DAB2 expression and cell signaling pathways. DAB2 expression was assessed in human GC tissue microarrays. H pylori infection in vivo and in vitro models were further explored. Immunostaining, immunofluorescence, chromatin immunoprecipitation, co-immunoprecipitation, Western blot, quantitative polymerase chain reaction, and luciferase reporter assays were performed in the current study.ResultsThe bioinformatic analysis verified that DAB2 was 1 of the 8 genes contributed to tumorigenesis and associated with poor prognosis in GC. The median overall survival and disease-free survival rates in DAB2high group were significantly less than those in DAB2low group. These findings demonstrated that H pylori transcriptionally activated DAB2 expression via signal transducer and activator of transcription 3 (STAT3)-dependent pathway. By bioinformatics analysis and knockdown or overexpression of DAB2, we found that DAB2 upregulated Yes-associated protein 1 (YAP1) transcriptional activity. Mechanistically, DAB2 served as a scaffold protein for integrin beta 3 (ITGB3) and SRC proto-oncogene non-receptor tyrosine kinase (SRC), facilitated the phosphorylation of SRC, promoted the small GTPase ras homolog family member A (RHOA) activation and phosphorylation of YAP1, and ultimately enhanced the YAP1 transcriptional activity.ConclusionsAltogether, these findings indicated that DAB2 is a key mediator in STAT3–regulated translation of YAP1 and plays crucial roles in H pylori-mediated GC development. DAB2 might serve as a novel therapeutic target for GC.

Disabled-2: a protein up-regulated by high molecular weight hyaluronan has both tumor promoting and tumor suppressor roles in ovarian cancer

Although the pro-tumorigenic functions of hyaluronan (HA) are well documented there is limited information on the effects and targets of different molecular weight HA. Here, we investigated the effects of 27 kDa, 183 kDa and 1000 kDa HA on ES-2 ovarian cancer cells overexpressing the stem cell associated protein, Notch3. 1000 kDA HA promoted spheroid formation in ES-2 cells mixed with ES-2 overexpressing Notch3 (1:3). We report disabled-2 (DAB2) as a novel protein regulated by 1000 kDa HA and further investigated its role in ovarian cancer. DAB2 was downregulated in ovarian cancer compared to normal tissues but increased in metastatic ovarian tumors compared to primary tumors. High DAB2 expression was associated with poor patient outcome and positively correlated with HA synthesis enzyme HAS2, HA receptor CD44 and EMT and macrophage markers. Stromal DAB2 immunostaining was significantly increased in matched ovarian cancer tissues at relapse compared to diagnosis and associated with reduced survival. The proportion of DAB2 positive macrophages was significantly increased in metastatic ovarian cancer tissues compared to primary cancers. However, DAB2 overexpression significantly reduced invasion by both A2780 and OVCAR3 cells in vivo. Our research identifies a novel relationship between HA signalling, Notch3 and DAB2. We highlight a complex relationship of both pro-tumorigenic and tumor suppressive functions of DAB2 in ovarian cancer. Our findings highlight that DAB2 has a direct tumor suppressive role on ovarian cancer cells. The pro-tumorigenic role of DAB2 may be mediated by tumour associated macrophages and requires further investigation.

LncRNA MEG3 Inhibits Tumor Progression by Modulating Macrophage Phenotypic Polarization via miR-145-5p/DAB2 Axis in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the predominant histological type of primary liver cancer, which ranks sixth among the most common human tumors. Tumor-associated macrophages (TAMs) are an important component of tumor microenvironment (TME) and the M2 macrophage polarization substantially contributes to tumor growth and metastasis. Long non-coding RNA (lncRNA) MEG3 was reported to restrain HCC development. However, whether MEG3 regulates macrophage phenotypic polarization in HCC remains unclear. Bone marrow derived macrophages (BMDMs) were treated with LPS/IFNγ and IL4/IL13 to induce the M1 and M2 macrophage polarization, respectively. M2-polarized BMDMs were simultaneously transfected with adenovirus vector overexpressing MEG3 (Adv-MEG3). Subsequently, M2-polarized BMDMs were cultured for 24 h with serum-free medium, the supernatants of which were harvested as conditioned medium (CM). HCC cell line Huh7 was cultured with CM for 24 h. F4/80+CD68+ and F4/80+CD206+ cell percentages in M1-and M2-polarized BMDMs were calculated using flow cytometry. Huh7 cell migration, invasion and angiogenesis were determined via Transwell assay and tube formation experiment. Nude mice were implanted with Huh7 cells and Adv-MEG3-transfected M2-polarizd BMDMs, and tumor growth and M2 macrophage polarization markers were assessed. The binding between miR-145-5p and MEG3 or disabled-2 (DAB2) was verified by luciferase reporter assay. MEG3 presented lower expression in HCC tissues than in normal controls, and low expression of MEG3 was correlated to poorer prognosis of HCC patients. MEG3 expression was enhanced during LPS/IFNγ-induced M1 polarization, but was reduced during IL4/IL13-induced M2 polarization. MEG3 overexpression inhibited the expression of M2 polarization markers in both M2-polarized BMDMs and mice. Mechanically, MEG3 bound with miR-145-5p to regulate DAB2 expression. Overexpressing MEG3 suppressed M2 polarization-induced HCC cell metastasis and angiogenesis by upregulating DAB2 and inhibited in vivo tumor growth. LncRNA MEG3 curbs HCC development by repressing M2 macrophage polarization via miR-145-5p/DAB2 axis.

DAB2 promotes pulmonary fibrosis and may act as an intermediate between IGF‑1R and PI3K/AKT signaling pathways.

Idiopathic pulmonary fibrosis (IPF) is a heterogeneous lung disease associated with high mortality. Disabled-2 (DAB2), an adapter protein, regulates cell-fibrinogen adhesion and fibrinogen uptake. DAB2 is differentially expressed in mouse fibrotic lungs induced by bleomycin according to a genome microarray analysis based on Gene Expression Omnibus database. However, the role of DAB2 in IPF has not been revealed. A bleomycin-induced mouse model of pulmonary fibrosis was constructed in the present study. It found that the expression of DAB2 was upregulated in bleomycin-induced fibrotic lung tissue with collagen fiber deposition and pulmonary interstitium thickening. Colocalization of DAB2 with α-smooth muscle actin (SMA) was observed in lung tissue sections. In vitro, human lung fibroblast MRC-5 cells were treated with TGF-β1 and the expression of DAB2 was increased. Knockdown of DAB2 suppressed cell proliferation and the expression of α-SMA, collagen I, collagen IV and fibronectin in TGF-β1-treated MRC-5 cells. The phosphorylation levels of PI3K and AKT were suppressed in DAB2-knockdown cells. IGF-1/IGF-1R has been reported to promote pulmonary fibrosis and activate the PI3K/Akt signaling. In the present study, the activation of IGF-1/IGF-1R signaling pathways in bleomycin-induced fibrotic lung tissues were positively associated with DAB2 expression. The phosphorylation level of IGF-1R was increased in MRC-5 cells with TGF-β1 treatment, and DAB2 expression was decreased by silencing of IGF-1R. This suggested that DAB2 might be a downstream target of the IGF-1R pathway and thus induced PI3K/AKT signaling activation and fibrogenesis. The current study demonstrated the importance of DAB2 in pulmonary fibrosis and suggested the potential of IGF-1R/DAB2/PI3K in the pathogenesis of IPF.

Disabled-2 (DAB2): A Key Regulator of Anti- and Pro-Tumorigenic Pathways.

Disabled-2 (DAB2), a key adaptor protein in clathrin mediated endocytosis, is implicated in the regulation of key signalling pathways involved in homeostasis, cell positioning and epithelial to mesenchymal transition (EMT). It was initially identified as a tumour suppressor implicated in the initiation of ovarian cancer, but was subsequently linked to many other cancer types. DAB2 contains key functional domains which allow it to negatively regulate key signalling pathways including the mitogen activated protein kinase (MAPK), wingless/integrated (Wnt) and transforming growth factor beta (TGFβ) pathways. Loss of DAB2 is primarily associated with activation of these pathways and tumour progression, however this review also explores studies which demonstrate the complex nature of DAB2 function with pro-tumorigenic effects. A recent strong interest in microRNAs (miRNA) in cancer has identified DAB2 as a common target. This has reignited an interest in DAB2 research in cancer. Transcriptomics of tumour associated macrophages (TAMs) has also identified a pro-metastatic role of DAB2 in the tumour microenvironment. This review will cover the broad depth literature on the tumour suppressor role of DAB2, highlighting its complex relationships with different pathways. Furthermore, it will explore recent findings which suggest DAB2 has a more complex role in cancer than initially thought.

Chemotherapy induces feedback up-regulation of CD44v6 in colorectal cancer initiating cells through β-catenin/MDR1 signaling to sustain chemoresistance.

Chemoresistance in colorectal cancer initiating cells (CICs) involves the sustained activation of multiple drug resistance (MDR) and WNT/β-catenin signaling pathways, as well as of alternatively spliced-isoforms of CD44 containing variable exon-6 (CD44v6). In spite of its importance, mechanisms underlying the sustained activity of WNT/β-catenin signaling have remained elusive. The presence of binding elements of the β-catenin-interacting transcription factor TCF4 in the MDR1 and CD44 promoters suggests that crosstalk between WNT/β-catenin/TCF4-activation and the expression of the CD44v6 isoform mediated by FOLFOX, a first-line chemotherapeutic agent for colorectal cancer, could be a fundamental mechanism of FOLFOX resistance. Our results identify that FOLFOX treatment induced WNT3A secretion, which stimulated a positive feedback loop coupling β-catenin signaling and CD44v6 splicing. In conjunction with FOLFOX induced WNT3A signal, specific CD44v6 variants produced by alternative splicing subsequently enhance the late wave of WNT/β-catenin activation to facilitate cell cycle progression. Moreover, we revealed that FOLFOX-mediated sustained WNT signal requires the formation of a CD44v6-LRP6-signalosome in caveolin microdomains, which leads to increased FOLFOX efflux. FOLFOX-resistance in colorectal CICs occurs in the absence of tumor-suppressor disabled-2 (DAB2), an inhibitor of WNT/β-catenin signaling. Conversely, in sensitive cells, DAB2 inhibition of WNT-signaling requires interaction with a clathrin containing CD44v6-LRP6-signalosome. Furthermore, full-length CD44v6, once internalized through the caveolin-signalosome, is translocated to the nucleus where in complex with TCF4, it binds to β-catenin/TCF4-regulated MDR1, or to CD44 promoters, which leads to FOLFOX-resistance and CD44v6 transcription through transcriptional-reprogramming. These findings provide evidence that targeting CD44v6-mediated LRP6/β-catenin-signaling and drug efflux may represent a novel approach to overcome FOLFOX resistance and inhibit tumor progression in colorectal CICs. Thus, sustained drug resistance in colorectal CICs is mediated by overexpression of CD44v6, which is both a functional biomarker and a therapeutic target in colorectal cancer.

Disabled‐2 overexpression mediates immune suppression in systemic lupus erythematosus by modulating Treg/Th17 cell differentiation

Systemic lupus erythematosus (SLE) is an autoimmune disorder. T helper 17 (Th17) and regulatory T (Treg) cells play key roles in SLE progression. Disabled-2 (DAB2) exhibits immunomodulatory effects in inflammatory diseases. However, the role of DAB2 in SLE and the precise mechanisms remain unknown. Here, a decreased DAB2 expression and an increased miR-448-3p level were observed in peripheral blood mononuclear cells (PBMCs) from SLE patients. DAB2level was negatively correlated with SLE Disease Activity Index (SLEDAI), suggesting a functional correlation between DAB2 and SLE. To test this, we used 8-week-old MRL/lpr mice and treated them with lentivirus-mediated DAB2 (LV-DAB2) or its negative control (LV-NC). LV-DAB2 treatment increased DAB2 expression and reduced serum immunoglobulin G (IgG) and anti-dsDNA IgG levels. DAB2 upregulation alleviated splenomegaly and lymphadenopathy and SLE-related organ damage. Moreover, DAB2 enhanced the percentage of CD25+ Foxp3+ Treg cells, but reduced Th17 cell frequency in lupus, along with the reduction in tumour necrosis factor α (TNF-α), interleukin (IL)-6 (IL-6) and IL-17A levels, and the elevation in IL-10. In vitro, naive CD4+ T cells isolated from MRL/lpr mice were polarized into Th17 or Treg phenotypes and treated with lentivirus. LV-DAB2 treatment downregulated IL-17A expression and inhibited the generation of CD4+ IL-17A+ Th17 cells. DAB2 triggered the production of IL-10 and the activation of Treg cells. Furthermore, DAB2 was verified as a direct target for miR-448-3p. MiR-448-3p overexpression cancelled the promoting effect of DAB2 on Treg cell differentiation. Taken together, DAB2 exerts an immunosuppressive effect on SLE through promoting Treg cell activation and inhibiting Th17 cell differentiation, which may be modulated by miR-448-3p.

MicroRNA‑93 knockdown inhibits acute myeloid leukemia cell growth via inactivating thePI3K/AKT pathway by upregulating DAB2.

Acute myeloid leukemia (AML) is associated with a poor prognosis in elderly adults and currently lacks optimal treatment strategies. MicroRNAs (miRNAs or miRs) have increasingly been reported to be associated with AML progression; however, the mechanisms of action of miR-93 in AML with the involvement of disabled 2 (DAB2) are currently unknown. In the present study, miR-93 expression was assessed in patients with AML and in AML cell lines. The association between miR-93 expression and the pathological characteristics of patients with AML was analyzed. AML cells were then transfected to knockdown or overexpress miR-93 in order to elucidate its function in AML progression. The target gene of miR-93 was assessed using a dual-luciferase reporter gene assay. The expression levels of miR-93, DAB2 and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway-related proteins were measured and in vivo experiments were conducted to confirm the results. It was observed that miR-93 was highly expressed in patients with AML and in AML cells. The knockdown of miR-93 in HL-60 cells inhibited AML cell proliferation and resistance to apoptosis, while the overexpression of miR-93 in THP-1 cells led to contrasting results. Moreover, miR-93 targeted DAB2 to inactivate the PI3K/AKT pathway, and the overexpression of DAB2 reversed the effects of miR-93 on THP-1 cell growth. Tumor volume, tumor weight, and the positive expression of Ki67, survivin and p53 were increased in THP-1 cells overexpressing miR-93. On the whole, the present study demonstrates that miR-93 is highly expressed in AML cells, and that the suppression of miR-93 inhibits AML cell growth by targeting DAB2 and inhibiting the PI3K/AKT pathway.

Involvement of Disabled-2 on skin fibrosis in systemic sclerosis

BackgroundSystemic sclerosis (SSc) is a connective tissue disease characterized by inflammation and fibrosis. Our previous research found Disabled-2 (DAB2) expression was significantly downregulated by salvianolic acid B, a small molecular medicine which attenuated experimental skin fibrosis of SSc. These suggest that DAB2 plays an important role in SSc skin fibrosis, but the role of DAB2 in SSc remains unclear. ObjectivesTo investigate the role of DAB2 in SSc. MethodsDAB2 expression level was detected in the skin and peripheral blood mononuclear cells of SSc patients. Bleomycin (BLM)-induced SSc mice and primary SSc skin fibroblasts were used to investigate the effect of DAB2 downregulation on fibrosis. RNA-seq transcriptome analysis was performed to underlie the mechanism of DAB2 in fibroblasts. ResultsDAB2 expression was enhanced in SSc lesion skin and was positively correlated with fibrotic genes, such as α-SMA and PAI-1. The in vivo study revealed that DAB2 downregulation alleviated skin fibrosis, alleviating skin thickness and reducing collagen deposition, and DAB2 knockdown ameliorated the inflammatory cell infiltration. The in vitro study showed that DAB2 knockdown reduced extracellular matrix genes and proteins expression. Moreover, Transcriptome analysis revealed TGF-β and focal adhesion signaling pathways were the main downregulated pathways involved in DAB2 siRNA treated fibroblasts. ConclusionsTaken together, our results revealed that DAB2 was increased in SSc skin, and DAB2 downregulation inhibited BLM-induced mouse skin fibrosis and SSc skin fibroblasts activation. DAB2 played an important role in the pathogenesis of SSc and DAB2 modulation may represent a potential therapeutic method for SSc.

DAB2 suppresses gastric cancer migration by regulating the Wnt/β-catenin and Hippo-YAP signaling pathways.

BackgroundDisabled-2 (DAB2), a potential tumor suppressor, plays an in important role in cancer development and cellular differentiation. Its lower expression levels have founded in many cancers. In addition, DAB2 is involved in multiple signaling pathways, including TGF-β and Wnt signal pathways. Gastric cancer (GC) is a common gastrointestinal malignant tumor. Nonetheless, the role of DAB2 in GC remains unclear.MethodsThirty-seven clinical specimens of GC tissues and adjacent non-tumor tissues were examined by immunohistochemistry. Proteins were extracted from two of them to perform Western blot analysis. Then, CMV-MCS-3FLAG-SV40-DAB2 and si-DAB2 were transfected into MGC and SGC cell line, respectively. The migration of GC cells was evaluated by transwell migration assay. And, the expression of migration related proteins was detected by Western blot and immunofluorescence (IF).ResultsEighty-six percent (32/37) of patients DAB2 staining was reduced in GC tissues compared to adjacent normal tissues. Further studies showed that in six human GC cell lines, the level of DAB2 expression was lower than normal gastric epithelial cells, and that DAB2 was closely related to cell migration in vitro. In DAB2 silenced cells, the Wnt/β-catenin signaling was increased and the Hippo-YAP pathway was affected. In addition, lower DAB2 level led to nuclear translocation of β-catenin and Yap.ConclusionsThe lower expression of DAB2 regulates cell migration in GC via interfering with the Wnt and Hippo signaling pathway. Our findings suggested that DAB2 played an important role in the migration of GC.

MiR-134-5p Promotes Stage I Lung Adenocarcinoma Metastasis and Chemoresistance by Targeting DAB2

Despite surgery and adjuvant therapy, early-stage lung adenocarcinoma (LUAD) treatment often fails due to local or metastatic recurrence. However, the mechanism is largely unknown. Here, we report that increased expression levels of miR-134-5p and decreased levels of disabled-2 (DAB2) were significantly correlated with recurrence in stage I LUAD patients. Our data show that miR-134-5p overexpression or DAB2 silencing strongly stimulated LUAD cell metastasis and chemoresistance. In contrast, inhibition of miR-134-5p or overexpression of DAB2 strongly suppressed LUAD cell metastasis and overcame the insensitivity of chemoresistant LUAD cells to chemotherapy. In addition, we demonstrated that DAB2 is a target of miR-134-5p and that miR-134-5p stimulates chemoresistance and metastasis through DAB2 in LUAD. Taken together, these findings suggest that miR-134-5p and its target gene DAB2 have potential as a biomarker for predicting recurrence in stage I LUAD patients. Additionally, miR-134-5p inhibition or DAB2 restoration may be a novel strategy for inhibiting LUAD metastasis and overcoming LUAD cell resistance to chemotherapy.

Disabled-2 (DAB2) Overexpression Inhibits Monocyte-Derived Dendritic Cells' Function in Vogt-Koyanagi-Harada Disease.

Recent studies reported that the tumor suppressor disabled-2 (DAB2) is a negative regulator of immune function. In this study, we investigated the role of DAB2 in monocyte-derived dendritic cells (DCs) from Vogt-Koyanagi-Harada disease (VKH) patients. The mRNA and protein levels of DAB2 were quantified by quantitative real-time PCR and Western blot. The Sequenom MassARRAY system was used to detect the promoter methylation level. An adenovirus carrying the DAB2 gene was transduced into immature DCs, isolated, and induced from active VKH patients. The surface markers of DCs, the frequency of T helper (Th) type 1 (Th1) and Th17 cells in CD4+T cells, which were cocultured with DCs, were tested by flow cytometry. ELISA was used to analyze the inflammatory cytokines produced by DC and CD4+T cell cocultures. The mRNA and protein expression levels of DAB2 in DCs obtained from active VKH patients were decreased, while the DAB2 promoter methylation level was marginally increased when compared with inactive VKH patients and normal controls. The expression of CD86 on DCs was significantly downregulated by DAB2 overexpression. The DC-related inflammatory factors IL-6 and TNF-α were also decreased. The frequency of Th1 and Th17 cells and their related cytokines were reduced significantly after coculture with DAB2 overexpressing DCs. DAB2 overexpression did not affect autophagy in DCs from VKH patients. These results suggest that the decreased expression of DAB2 in DCs plays a role in the pathogenesis of VKH disease. DAB2 overexpression inhibits DC function, but this is not mediated via autophagy.

MiR-143/145 differentially regulate hematopoietic stem and progenitor activity through suppression of canonical TGF\u03b2 signaling

Expression of miR-143 and miR-145 is reduced in hematopoietic stem/progenitor cells (HSPCs) of myelodysplastic syndrome patients with a deletion in the long arm of chromosome 5. Here we show that mice lacking miR-143/145 have impaired HSPC activity with depletion of functional hematopoietic stem cells (HSCs), but activation of progenitor cells (HPCs). We identify components of the transforming growth factor β (TGFβ) pathway as key targets of miR-143/145. Enforced expression of the TGFβ adaptor protein and miR-145 target, Disabled-2 (DAB2), recapitulates the HSC defect seen in miR-143/145−/− mice. Despite reduced HSC activity, older miR-143/145−/− and DAB2-expressing mice show elevated leukocyte counts associated with increased HPC activity. A subset of mice develop a serially transplantable myeloid malignancy, associated with expansion of HPC. Thus, miR-143/145 play a cell context-dependent role in HSPC function through regulation of TGFβ/DAB2 activation, and loss of these miRNAs creates a preleukemic state.

MiR-191/DAB2 axis regulates the tumorigenicity of estrogen receptor-positive breast cancer.

Disabled-2 (DAB2) has been shown to be downregulated in a variety of human cancer types including breast tumors. However, the role of DAB2 in estrogen receptor positive (ER+) breast cancer cells has not been reported. In this context, we demonstrated that DAB2 expression was significantly decreased in ER+ breast cancer cell lines and ER+ clinical specimens, compared with ER- breast cancer cell lines and ER- tissues, respectively. Depletion of estrogen significantly elevated DAB2 expression in ER+ MCF7 and T-47D cells. Treatment with estradiol (E2) reduced the expression of DAB2 and administration of tamoxifen upregulated DAB2 expression in a dose-dependent manner. Functionally, silencing of DAB2 in hormone-starved MCF7 and T-47D cells promoted cellular proliferation and enforced expression of DAB2 in normal-cultured or E2-treated cells suppressed cellular proliferation. Mechanistically, estrogen-induced miR-191 was identified as a direct upstream regulator of DAB2 in ER+ cells. Luciferase reporter assay indicated that miR-191 inhibited DAB2 expression by directly targeting the 3'-UTR of DAB2. Importantly, the expression and function of miR-191 showed the opposite tendency with DAB2 in ER+ cells. In vivo, inhibition of miR-191 significantly suppressed the xenograft growth induced by E2, and silencing of DAB2 could restored the growth arrest induced by miR-191 inhibition. Taken together, our data unveil that the miR-191-DAB2 axis seems to be an important pathway associated with estrogen signaling in breast cancer and may serve as a potential diagnostic biomarker and a powerful therapeutic target for ER+ breast cancer patients. © 2017 IUBMB Life, 70(1):71-80, 2018.

Expression and Histopathological Significance of Disabled-2 in Aldosterone-Producing Adenoma.

The current pathological diagnosis of aldosterone-producing adenoma (APA) is challenging because no histological markers of aldosterone production are available in routine practice. A previous study demonstrated that Disabled-2 (DAB2) is a specific marker of the zona glomerulosa (ZG) in rodents. The aim of the present study was to investigate the significance of immunohistochemical staining to detect DAB2 in the adrenal tissue of patients with APA. We investigated the expression of DAB2 in 36 adrenal glands with APA, 23 adrenal glands with cortisol-producing adenoma (CPA), and 33 adrenal glands with non-functioning adenoma (NFA). Immunohistochemical staining was performed using anti-DAB2 antibodies on paraffin-embedded sections. We analysed the expression of DAB2 semi-quantitatively by scoring staining intensity, and assessed the correlation of this information with the clinical findings. DAB2 mRNA expression in adenoma tissues was evaluated by RT-PCR. DAB2 was highly expressed in the ZG in normal human adrenal glands. DAB2 expression was heterogeneous in APA, with spotted, strong staining noted in most samples (25 of 36 APA). CPA and NFA also exhibited extensive low or moderate DAB2 expression. DAB2 mRNA was significantly increased and positively correlated with CYP11B2 in APA (p<0.05). In APA, the DAB2 score adjusted for tumour volume was positively correlated with plasma aldosterone (p<0.05). Patients with low or moderate DAB2 staining more frequently exhibited high blood pressure and were diagnosed at a younger age compared with patients with high DAB2 staining. The present study clearly demonstrates that DAB2 is a specific marker of the ZG in normal human adrenal glands but that DAB2 immunostaining is not sufficiently powerful for histopathological diagnosis of APA. DAB2 might be involved in excessive aldosterone biosynthesis and correlate with specific clinical characteristics of APA patients.

Low disabled-2 expression promotes tumor progression and determines poor survival and high recurrence of esophageal squamous cell carcinoma.

Patients with esophageal squamous cell carcinomas (ESCCs) have poor survival and high recurrence rate, but lack a prognostic biomarker. Disabled-2 (DAB2) is a crucial tumor suppressor, but its roles in ESCCs are uncertain. We investigated whether low DAB2 expression in ESCCs could lead into tumor progression and poor prognosis. Our results found patients with low-DAB2 expression ESCCs had significantly larger tumor size, deeper tumor invasion depth, lymph node metastasis, worse survival, and higher recurrence rate (P<0.05). The Cox-regression model revealed low-DAB2 expression was an independent factor of poor survival (P<0.05), and also of tumor recurrence with the predictive performance superior to clinical TNM stage (P<0.05). Low-DAB2 cancer cells, validated by DAB2 knockdown or over-expression, had higher phosphorylated ERK and migration abilities, which could be suppressed by ERK inhibitor treatment. TGF-β-induced epithelial-to-mesenchymal transition (EMT) only existed in the high-DAB2 cells, and related to worse prognosis of high-DAB2 ESCCs (P<0.05). In conclusion, DAB2 can suppress the ERK signaling, but correlate to have TGF-β-induced EMT in ESCCs. DAB2 expression could be a biomarker to identify patients with worse survival and high recurrence. Our data suggest DAB2 expression can stratify patients in need of aggressive surveillance and with possible benefit from anti-ERK or anti-TGF-β therapies.

Quantitative proteomics identify DAB2 as a cardiac developmental regulator that inhibits WNT/β-catenin signaling

To reveal the molecular mechanisms involved in cardiac lineage determination and differentiation, we quantified the proteome of human embryonic stem cells (hESCs), cardiac progenitor cells (CPCs), and cardiomyocytes during a time course of directed differentiation by label-free quantitative proteomics. This approach correctly identified known stage-specific markers of cardiomyocyte differentiation, including SRY-box2 (SOX2), GATA binding protein 4 (GATA4), and myosin heavy chain 6 (MYH6). This led us to determine whether our proteomic screen could reveal previously unidentified mediators of heart development. We identified Disabled 2 (DAB2) as one of the most dynamically expressed proteins in hESCs, CPCs, and cardiomyocytes. We used clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) mutagenesis in zebrafish to assess whether DAB2 plays a functional role during cardiomyocyte differentiation. We found that deletion of Dab2 in zebrafish embryos led to a significant reduction in cardiomyocyte number and increased endogenous WNT/β-catenin signaling. Furthermore, the Dab2-deficient defects in cardiomyocyte number could be suppressed by overexpression of dickkopf 1 (DKK1), an inhibitor of WNT/β-catenin signaling. Thus, inhibition of WNT/β-catenin signaling by DAB2 is essential for establishing the correct number of cardiomyocytes in the developing heart. Our work demonstrates that quantifying the proteome of human stem cells can identify previously unknown developmental regulators.

Abstract A1-07: Identification of base pair mutations and structural rearrangements acquired in breast cancer metastases including a novel hyperactive ESR1-DAB2 fusion gene in hormone-resistant progression

Abstract DNA structural variations (SVs) are a major source of genetic instability in cancer, but they remain understudied. Large-insert mate-pair sequencing (MPS) is a powerful method designed to detect SVs, even in highly repetitive regions. Using MPS and other methods, we performed a comprehensive analysis of genomic alterations in breast cancer progression. Matched primary/recurrent frozen tumor samples from 6 patients, including two patients from our rapid autopsy program with multiple metastatic tissues (20 total samples; average 5.5 years to recurrence) were examined by multiple large-insert library (3-5, 5-8, 8-12kb) MPS to identify metastatic acquired SVs. This was supplemented with RNAseq (n=15), whole exome sequencing (n=18;~75x), whole genome sequencing (n=3; 40-65x), and SNP arrays. A relatively small fraction (~10%) of somatic single nucleotide variants (SNVs) in the primary tumor were identified in matched metastatic samples, and the majority of metastatic SNVs were not found in the matched primary tumor. This indicates that a rare sub-clone colonizes the metastatic site and evolves extensively before becoming clinically evident. For example, in one patient with an ER+ tumor who initially declined anti-estrogen therapy, the recently described ESR1 Y537S mutation was not present in the primary tumor or in metastatic disease 5 years later. However, after extensive anti-estrogen treatment for metastatic disease, the mutation was identified at rapid autopsy, indicating that this mutation can be acquired even after initial metastatic spread. Chromatin immunoprecipitation assays in metastatic tissue from tumors with mutant ERα show strong enrichment for ERα at classical ERα target genes and we are currently assessing the genome-wide binding pattern of ERα to identify novel binding sites. We observed extensive patient-to-patient variability in the number and types of SVs. In general, the overall patterns of SVs were remarkably similar between matched primary and metastatic samples indicating that these events likely occurred early in tumorigenesis and are stable throughout disease progression. We identified a number of metastatic specific SVs that likely contribute to disease progression. Specifically, in one patient with an ER+ primary tumor treated with adjuvant Tamoxifen, we identified a novel fusion gene between ESR1 (estrogen receptor-α, ERα) and DAB2 (disabled-2) only in a lymph node recurrence. RT-PCR and western blot analysis confirmed that the fusion RNA/protein was expressed/translated only in the recurrent disease. The fusion retains the DNA-binding domain (DBD) and hinge region of ERα while the ligand-binding domain (LBD) is replaced with the majority of DAB2. We hypothesized that this is a functional genetic alteration conferring ligand-independent ERα-mediated signaling and growth. Confirming this, in vitro ERE-Tk-luc reporter assays showed that the ESR1-DAB2 fusion has ligand-independent activity that is 13-290x higher than wild-type ERα. We are currently assessing the genome-wide binding of ESR1-DAB2 and the functional contribution of DAB2 to the fusion protein. This study represents the most comprehensive analysis to date of genomic changes in breast cancer progression and indicates extensive changes occur during metastatic spread. A number of acquired changes likely represent therapeutically targetable metastatic dependencies. Citation Format: Ryan James Hartmaier, Amir Bahreini, Shannon L. Puhalla, Steffi Oesterreich, Aju Mathew, Nancy E. Davidson, Adam M. Brufsky, Adrian V. Lee. Identification of base pair mutations and structural rearrangements acquired in breast cancer metastases including a novel hyperactive ESR1-DAB2 fusion gene in hormone-resistant progression. [abstract]. In: Proceedings of the AACR Special Conference on Translation of the Cancer Genome; Feb 7-9, 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 1):Abstract nr A1-07.