MiR-30c/MTDH Axis Regulates Polarity Reversal And Cell Clustered Invasive And Metastasis In Invasive Micropapillary Carcinoma Of Breast

ObjectiveInvasive micropapillary carcinoma of breast(IMPC)is an aggressive subtype of breast cancer,characterized by a special growth pattern of reversed polarity cell clusters and high propensity of lymphovascular invasion and axillary lymph node metastasis.Recent study showed that IMPC could serve as a model to investigate the underlying molecular mechanism of tumor cell polarity abnormalities and metastasis.microRNAs regulate multiple process during carcinogenesis such as cell proliferation,apoptosis,invasion,metastasis and angiogenesis.Here by bioinformatic prediction,we found miR-30c could target MTDH.Despite the results of these previous molecular mechanisms,the pathological relevance and biological alterations of miRNAs leading to aggressive characters of IMPC remain largely unknown.Therefore,it is important to explore the clinical and molecular mechanisms of miR-30c/MTDH axis in regulation of polarity reversal and metastasis of IMPC.MethodsWe analyzed miR-30c and MTDH expressions in 124 cases of IMPC tissues and the corresponding 149 case of IDC-NST tissues by ISH and IHC at the tissue level.We analyzed miR-30c and MTDH expressions were whether associated with the clinical pathological characteristics and prognosis of IMPC patients.At the molecular level,we measured miR-30c could whether regulate MTDH protein expression directly by the dual luciferase reporter gene experiment and western blots.By SRB,clone formation,and 3D matrigel test,miR-30c/MTDH signal axis could whether effects on cell proliferation ability.By transwell and wound healing assey,miR-30c/MTDH signal axis could whether impact on cell migration ability.Nude mice model to evaluate whether miR-30c could inhibit tumor formation and lung metastasis.By immunofluorescence to find whether miR-30c/MTDH could regulate the primary cell polarity of IMPC.ResultsHere by bioinformatic prediction,reporter assay and western blots,we found miR-30c could target MTDH and thereby regulate breast cancer cell proliferation,invasion and metastasis.In vivo experiment showed miR-30c could inhibit breast tumor growth and lung metastasis.In clinical pathological characteristics,IMPC compare to IDC-NST have larger tumor size,higher lymph node metastasis,higher tumor stage and positive ER receptor status.Moreover,we revealed miR-30c was down regulated in IMPC(n=124)comparing with IDC-NST(n=149)and highly negatively correlated with MTDH expression,which was upregulated in IMPC specimens by in situ hybridization and immunohistochemistry analysis(r_s=-0.221,P=0.013).miR-30c and MTDH expression levels were highly correlated with tumor size(P=0.042,P=0.038),lymph nodules(P<0.01,P<0.01)and tumor grade(P<0.01,P<0.01),overall(P=0.015,P<0.01)and disease-free survival(P<0.01,P<0.01)of IMPCs.Multivariate survival analyses showed tumor stage(P=0.043)and MTDH(P=0.028)expression were independent risk factors for OS,nodal stage(P=0.026)and MTDH(P=0.01)expression were independent risk factors for DFS in IMPC.The experiments show that the overexpression of MUC1 could improve the inside-out stain of MUC1,while overexpression of miR-30c could abolish the effect by multi-dimensional invasive spherical masses analysis(P<0.05).We found that miR-30c/MTDH axis could regulate IMPC cluster formation through downstream MUC1.ConclusionIn present study,we revealed the downregulation of miR-30c could upregulate the protein level of MTDH and thereby contribute to special polarity buildup and aggressive character of IMPC.Overall,we reveal a potential key regulatory role of polarity reversal and metastasis through miR-30c/MTDH axis which may facilitate the development of novel therapeutic strategies for IMPC of breast.