SMAD4 as a Prognostic Marker in Colorectal Cancer

Abstract

More than 50% of patients with Dukes C colorectal cancer have disease recurrence and die within 5 years after surgical removal of their primary tumor. It is currently not possible to distinguish patients with good and bad prognosis. SMAD4 is an important tumor suppressor gene that mediates transforming growth factor-h superfamily signaling and is located in chromosome 18q21, a region with frequent genetic losses in these tumors. Allelic imbalance in 18q has been linked to poor prognosis in a subset of colorectal cancer patients. Therefore, we generated a tissue microarray containing triplicate tumor samples from 86 Dukes C patients and used immunohistochemistry to assess the relative expression level of SMAD4 and its value as a prognostic marker. In addition, SMAD4 was screened for mutations and two polymorphic microsatellite markers were used to assess the presence of allelic imbalance in these tumors. Patients with tumors expressing high SMAD4 levels had significantly better overall (P < 0.025) and disease-free (P < 0.013) survival than patients with low levels. This identifies SMAD4 as a prognostic marker for Dukes C colorectal cancer. Although all tumors with absent SMAD4 staining showed allelic imbalance in 18q21, tumors with 18q21 allelic imbalance as a group showed no difference in SMAD4 levels compared with tumors without allelic imbalance, suggesting that additional mechanisms of SMAD4 down-regulation exist. In addition, although SMAD4 mutations were found in five tumors, they were not associated with shorter survival. In conclusion, the level of expression of SMAD4 was found to be a more sensitive marker than 18q21 allelic imbalance and SMAD4 mutations, which were of no prognostic significance for these patients.One of the most common cytogenetic abnormalities in colorectal tumors is the loss of genetic material in chromosome 18q. This is believed to be indicative of an important tumor suppressor gene in this genomic location (1, 2). The identity of the gene targeted by these deletions in 18q has remained elusive and several genes, including SMAD2 and DCC (deleted in colorectal carcinoma), have been proposed (3 -5). Recently, SMAD4, a key transducer of transforming growth factor-h (TGF-h) superfamily signaling that is located in chromosome 18q21 and regulates cell proliferation, differentiation, and apoptosis (6 -8), has attracted considerable attention due to several findings. SMAD4 mutations have recently been shown to be associated with the occurrence of juvenile polyposis, an autosomal dominant syndrome predisposing to hamartomatous polyps and colorectal cancer (9, 10). In addition, frequent somatic mutations have been found in human colorectal tumors in several studies, further suggesting an important role for this gene in colorectal carcinogenesis (11,12). Moreover, animal studies have shown that SMAD4 inactivation is involved in the malignant transformation of gastrointestinal adenomas (13) and a reduction in SMAD4 mRNA levels has been observed during tumor progression (14).A significan...

An integrated computational and experimental study uncovers FUT 9 as a metabolic driver of colorectal cancer

“…(iii) Finally, a Kaplan–Meier survival analysis further narrowed down this list to 177 candidate tumor suppressors whose downregulation is negatively correlated with patient survival (and thus, likely to enhance tumor progression; see Materials and Methods, Fig 1A and Table EV3). Reassuringly, the resulting list includes several known colon tumor suppressors such as APC (Fearnhead et al , 2001; Aoki & Taketo, 2007), TCF7L2 (Hazra et al , 2008; Slattery et al , 2008), MCC (Kinzler et al , 1991), PTEN (Nassif et al , 2004; Song et al , 2012), and SMAD4 (Miyaki et al , 1999; Alazzouzi et al , 2005). It also includes 34 metabolic genes that are present in the human metabolic model (Table EV4), and which we further studied in the next modeling step.…”

Chromosome 18q deletion and Smad4 protein inactivation correlate with liver metastasis: a study matched for T- and N- classification

“…They also classified patients into two groups according to the status of liver metastasis, and indicated a borderline significant correlation between Smad4 staining and liver metastasis (P ¼ 0.05) (Maitra et al, 2000). Alazzouzi et al (2005) studied 86 Dukes' C colorectal cancer patients and showed that patients with Dukes' C tumours expressing high Smad4 protein levels had significantly better overall (Po0.025) and disease-free (Po0.013) survival than patients with low levels by immunohistochemical staining. We, too, recently discussed the association between Smad4 level and prognosis in colorectal cancers.…”

“…The samples were evaluated as reported previously (Alazzouzi et al, 2005). To evaluate the intensity of Smad4 immunohistochemical staining, we used a semiquantitative scale, where 0 ¼ no Smad4 staining and 4 was the highest staining.…”

“…Smad4 gene, which mediates the intracellular signalling pathway of transforming growth factor (TGF)-beta receptor, has been detected as one of the target genes at 18q21 (Wang et al, 1995;Eppert et al, 1996;Hahn et al, 1996;Thiagalingam et al, 1996;Carethers et al, 1998;Markowitz, 2000;Fink et al, 2003;Alazzouzi et al, 2005;Li et al, 2005). Recently, an immunohistochemical method for evaluating Smad4 protein has been developed and several studies found higher frequency of Smad4 protein inactivation in the cases with liver metastasis and in the cases presenting unfavourable survival (Maitra et al, 2000;Alazzouzi et al, 2005).Currently, recurrences of colorectal cancers appear mainly as lymph node metastasis or liver metastasis. This means that both lymph node metastasis and liver metastasis have an influence on survival after surgery.…”

Quantitative expression analysis and prognostic significance of L-DOPA decarboxylase in colorectal adenocarcinoma

“…In an attempt to refine prognostication and predict the benefit derived from systemic treatment, several protein and genetic markers have been evaluated in patients with CRC, including allelic loss of chromosome 18q (Jen et al, 1994;Martinez-Lopez et al, 1998;Ogunbiyi et al, 1998;, absence of the deleted in colorectal carcinoma (DCC) protein (Shibata et al, 1996;Reymond et al, 1998;, decreased SMAD4 mRNA expression (Boulay et al, 2002;Alazzouzi et al, 2005), expression and/or abnormalities of cytoplasmic oncoprotein p53 (TP53) (Sun et al, 1992;Munro et al, 2005;Russo et al, 2005), protein levels and/or gene haplotype of thymidylate synthetase (TYMS) (Popat et al, 2004;Suh et al, 2005;Tsourouflis et al, 2008), microsatellite instability (MSI) (Gryfe et al, 2000;, and chromosomal instability (CIN) (Walther et al, 2008). Nevertheless, none of these biomarkers has been prospectively validated and established so far in clinical practice.…”