Prognostic significance and relationship of SMAD3 phosphoisoforms and VEGFR-1 in gastric cancer:A clinicopathological study

BACKGROUND The TGF-β/SMAD3 and VEGFR-1 signaling pathways play important roles in gastric cancer metastasis.SMAD3 phosphorylation is a crucial prognostic marker in gastric cancer.AIM To determine the prognostic value and relationship of SMAD3 phospho-isoforms and VEGFR-1 in gastric cancer.METHODS This was a single-center observational study which enrolled 98 gastric cancer patients and 82 adjacent normal gastric tissues from patients aged 32-84 years(median age 65)between July 2006 and April 2007.Patients were followed up until death or the study ended(median follow-up duration of 28.5 mo).The samples were used to generate tissue microarrays(TMAs)for immunohistochemical(IHC)staining.The expressions of TGF-β1,pSMAD3C(S423/425),pSMAD3L(S204),and VEGFR-1 in gastric cancer(GC)tumor tissue and normal tissue were measured by IHC staining using TMAs obtained from 98 GC patients.Prognosis and survival information of the patients was recorded by Outdo Biotech from May 2007 to July 2015.The relationship between TGF-β1,pSMAD3C(S423/425),pSMAD3L(S204),and VEGFR-1 protein expression levels was analyzed using Pearson's correlation coefficient.The relationship between protein expression levels and clinicopathological parameters was analyzed using the Chi-squared test.A survival curve was generated using the Kaplan-Meier survival analysis.RESULTS TGFβ-1 and VEGFR-1 expression was significantly upregulated in gastric cancer tissue compared to adjacent noncancerous tissue.The positive expression of phosphorylated isoforms of Smad3 varied depending on the phosphorylation site[pSMAD3C(S423/425):51.0%and pSMAD3L(S204):31.6%].High expression of pSMAD-3L(S204)was significantly correlated with larger tumors(P=0.038)and later N stages(P=0.035).Additionally,high expression of VEGFR-1 was closely correlated with tumor size(P=0.015)and pathological grading(P=0.013).High expression of both pSMAD3L(S204)and VEGFR-1 was associated with unfavorable outcomes in terms of overall survival(OS).Multivariate analysis indicated that high expression of pSMAD3L(S204)and VEGFR-1 were independent risk factors for prognosis in GC patients.VEGFR-1 protein expression was correlated with TGF-β1(r=0.220,P=0.029),pSMAD3C(S423/425)(r=0.302,P=0.002),and pSMAD3L(S204)(r=0.201,P=0.047),respectively.Simultaneous overexpression of pSMAD3L(S204)and VEGFR-1 was associated with poor OS in gastric cancer patients.CONCLUSION Co-upregulation of pSMAD3L(S204)and VEGFR-1 can serve as a predictive marker for poor gastric cancer prognosis,and pSMAD3L(204)may be involved in enhanced gastric cancer metastasis in a VEGFR-1-dependent manner.

Smad3 and its phosphoisoforms are prognostic predictors of hepatocellular carcinoma after curative hepatectomy

BACKGROUND:Progression of hepatocellular carcinoma (HCC) often leads to vascular invasion and intrahepatic metastasis,which correlate with recurrence after surgical treatment and poor prognosis.HCC may be an unusual cancer affected by continuous inflammation that can lead to consistent upregulation of transforming growth factor-β (TGF-β).Chronic inflammation shifts hepatocytic TGF-β signaling from the tumor-suppressive pSmad3C pathway to the oncogenic pSmad3L pathway.In this study,we investigated the functional roles of Smad3 and its phosphoisoforms in the progression of HCC.METHODS:Tumor tissue microarrays of samples from 272 HCC patients who underwent curative surgical resection were used to detect the expression of Smad3,Smad4,pSmad3C (S423/425),pSmad3L (T179),pSmad3L (S204),and pSmad3L (S213).Disease-specific death was defined as 1) tumor occupying more than 80% of the liver,2) portal venous tumor thrombus (PVTT) proximal to the second bifurcation,3) obstructive jaundice due to tumor,4) distant metastases,or 5) variceal hemorrhage with PVTT proximal to the first bifurcation.At the time of analysis,tumor recurrence was detected in 184 (67.6%) patients,and 96 (35.3%) had died of HCC.RESULTS:Nuclear and cytoplasmic localization of Smad3,and nuclear localization of Smad4 were observed in 18.0%,9.9%,and 9.2% of HCCs,respectively.The rates of Smad3 phosphoisoform-immunoreactive HCC varied according to the location of phosphorylation:pSmad3C (S423/425) 8.1%,pSmad3L (T179) 2.6%,pSmad3L (S204) 2.2%,and pSmad3L (S213) 10.3%.Multivariate analyses revealed that pSmad3C (S423/425) (P=0.022) was an independent predictor of longer recurrence-free survival.pSmad3L (S213) (P=0.006),intrahepatic metastasis,multicentric occurrence,and liver cirrhosis were independent predictors of shorter recurrence- free survival.Cytoplasmic Smad3 (P=0.006),larger tumor size,and intrahepatic metastasis were independent predictors of shorter disease-specific survival.Only pSmad3L (S213) did not show an unfavorable influence on recurrence-free survival (P=0.331) on univariate analysis.CONCLUSIONS:pSmad3C (S423/425),pSmad3L (S213),and Smad3 may be predictors of prognosis in HCC patients after curative hepatectomy.pSmad3C (S423/425) and pSmad3L (S213) may be used as immunohistochemical biomarkers to identify patients with a high risk of recurrence.