BC-Monitor: Towards a Routinely Accessible Circulating Tumor DNA-Based Tool for Real-Time Monitoring Breast Cancer Progression and Treatment Effectiveness

, László Tordai 4 , Rozália Tóth 4 , Zsuzsanna Kahán 4 and Lajos Haracska 5,

Loss of CDX2 gene expression is associated with DNA repair proteins and is a crucial member of the Wnt signaling pathway in liver metastasis of colorectal cancer

Caudal type homeobox 2 (CDX2) has been well-established as a diagnostic marker for colorectal cancer (CRC); however, less is known about its regulation, particularly its potential interactions with the DNA repair proteins, adenomatous polyposis coli (APC) and beta-catenin, in a non-transcriptional manner. In the present study, the protein expression of CDX2 was analyzed, depending on the expression of the DNA repair proteins, mismatch repair (MMR), O6-methylguanine DNA methyltransferase (MGMT) and excision repair cross-complementing 1 (ERCC1), and its importance in Wnt signaling was also determined. A total of 101 liver metastases were punched into tissue microarray (TMA) blocks and serial sections were cut for immunohistochemistry. For each protein, an immunoreactive score was generated according to literature data and the scores were fitted to TMA. Subsequently, statistical analysis was performed to compare the levels of expression with each other and with clinical data. CDX2 loss of expression was observed in 38.5% of the CRC liver metastasis cases. A statistically significant association between CDX2 and each of the investigated MMRs was observed: MutL Homolog 1 (P<0.01), MutS protein Homolog (MSH) 2 (P<0.01), MSH6 (P<0.01), and postmeiotic segregation increased 2 (P=0.040). Furthermore, loss of MGMT and ERCC1 was also associated with CDX2 loss (P=0.039 and P<0.01, respectively). In addition, CDX2 and ERCC1 were inversely associated with metastatic tumor size (P=0.038 and P=0.027, respectively). Sustained CDX2 expression was associated with a higher expression of cytoplasmic/membranous beta-catenin and with nuclear APC expression (P=0.042 and P<0.01, respectively). In conclusion, CDX2 loss of expression was not a rare event in liver metastasis of CRC and the results suggested that CDX2 may be involved in mechanisms resulting in the loss of DNA repair protein expression, and in turn methylation; however, its exact function in this context remains to be elucidated

Expression of ERCC1, RRM1, TUBB3 in correlation with apoptosis repressor ARC, DNA mismatch repair proteins and p53 in liver metastasis of colorectal cancer

Liver metastasis in colorectal cancer is common and the primary treatment is chemotherapy. To date, there is no routinely used test in clinical practice to predict the effectiveness of conventional chemotherapy. Therefore, biomarkers with predictive value for conventional chemotherapy would be of considerable benefit in treatment planning. We analysed three proteins [excision repair cross-complementing 1 (ERCC1), ribonucleoside-diphosphate reductase 1 (RRM1) and class III beta-tubulin (TUBB3)] in colorectal cancer liver metastasis. We used tissue microarray slides with 101 liver metastasis samples, stained for ERCC1, RRM1 and TUBB3 and established scoring systems (fitted for tissue microarray) for each protein. In statistical analysis, we compared the expression of ERCC1, RRM1 and TUBB3 to mismatch proteins (MLH1, MSH2, MSH6 and PMS2), p53 and to apoptosis repressor protein (ARC). Statistically significant correlations were found between ERCC1, TUBB3 and MLH1, MSH2 and RRM1 and MSH2, MSH6. Noteworthy, our analysis revealed a strong significant correlation between cytoplasmic ARC expression and RRM1, TUBB3 (p=0.000 and p=0.001, respectively), implying an additional role of TUBB3 and RRM1 not only in therapy resistance, but also in the apoptotic machinery. Our data strengthens the importance of ERCC1, TUBB3 and RRM1 in the prediction of chemotherapy effectiveness and suggest new functional connections in DNA repair, microtubule network and apoptotic signaling (i.e. ARC protein). In conclusion, we showed the importance and need of predictive biomarkers in metastasized colorectal cancer and pointed out the relevance not only of single predictive markers but also of their interactions with other known and newly explored relations between different signaling pathways