PKCδ promotes the invasion and migration of colorectal cancer through c-myc/NDRG1 pathway.

Objective: Colorectal cancer (CRC) is the third cause of expected cancer deaths both in men and women in the U.S. and the third most commonly diagnosed cancer in China Targeted therapy has been proven to improve overall survival for unresectable metastatic CRC. But the location of the primary tumor or the presence of various core driver gene mutations that confer resistance may limit the utility of targeted therapy. Therefore, it is of great significance to further elucidate novel mechanisms of invasion and metastasis of CRC and find potential novel therapeutic targets. Protein Kinase C Delta (PKCδ) plays an important role in various diseases, including tumors. In CRC, the function of PKCδ on proliferation and differentiation is mostly studied but various research results were reported. Therefore, the role of PKCδ in CRC needs to be further studied, especially in tumor invasion and metastasis in CRC which few studies have looked into. Methods: The expression of PRKCD was analyzed by the Genotype-Tissue Expression (GTEx) and The Cancer Genome Atlas (TCGA) databases and Immunohistochemical (IHC). Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Gene Set Enrichment Analysis (GSEA) enrichment analysis were used to explore the biological functions and pathways related to PRKCD. Lentivirus transfection was used to construct CRC cell lines with overexpression and knock-down of PKCδ or N-myc Downstream Regulated Gene 1 (NDRG1). Cell invasion and migration assay, wound healing assay were used to detect the function of PKCδ and NDRG1 in the invasion and migration of cells. Flow cytometry analysis was used to detect the influence of PKCδ on the CRC cell cycles .Immunofluorescence histochemistry ,Immunoprecipitation Assay and qPCR were used to detect the relationship of PKCδ and NDRG1. Xenograft model was used to verify the role of PKCδ in vivo. Results: PKCδ is overexpressed in CRC and could promote Epithelial-Mesenchymal Transition (EMT) and the invasion and migration of CRC in vitro. We confirmed that PKCδ and the tumor suppressor factor NDRG1 had a co-localization relationship in CRC. PKCδ inhibited NDRG1 transcription and protein expression. Overexpressing NDRG1 could inhibit the function of PKCδ in promoting tumor invasion and migration. PKCδ could regulate c-Myc, one transcription factor of NDRG1, to down-regulate NDRG1. In vivo, overexpressing PKCδ could promote xenograft growth and volume. Thus, our results showed that PKCδ reduced the expression of NDRG1 through c-Myc, promoting the invasion and migration of CRC through promoting EMT. Conclusion: The increased expression of PKCδ in CRC tumor tissue could promote the invasion and migration of tumor cells, and one of the mechanisms may be regulating c-Myc to inhibit the expression of NDRG1 and promote EMT.

Detection and Clinical Value of Circulating Tumor Cells as an Assisted Prognostic Marker in Colorectal Cancer Patients.

BACKGROUND: Circulating tumor cells (CTCs) are cells that have been shed into the vasculature from a primary tumor and circulate in the bloodstream. It has been suggested that detecting CTCs could help the clinician to detect early metastasis or recurrence more effectively. This trial sets out to assess the detection and clinical value of CTCs as an assisted prognostic marker in patients with colon cancer and rectal cancer. METHODS: A prospective cohort of patients with colorectal cancer (CRC) was enrolled from July 2015 to February 2018 in Shanghai Minimally Invasive Surgery Center, Shanghai, China. In this study, 149 patients with CRC were enrolled and underwent surgical treatment. There were 79 cases of colon cancer and 70 cases of rectal cancer, including 93 males and 56 females. To investigate the correlativity and clinical value of CTCs, the patients were statistically analyzed in different subgroups: colon cancer group vs rectal cancer group, and left hemicolon cancer group vs right hemicolon cancer group. RESULTS: The results of analysis comparing CTC counts and clinical pathological features in colon and rectal cancer indicated that with increased tumor stage, the number of CTCs also increased, with significant statistical differences. CTC counts in patients with colon and rectal cancer showed positive correlations with TNM staging (P=0.001, 0.013, respectively), T staging (P=0.021, 0.001), N staging (P=0.014, 0.035) and M staging (P=0.018, 0.203). Detection of serum biomarkers in CTC-positive and CTC-negative groups indicated a significantly increasing expression in the CTC-positive group. To confirm the correlations between CTCs and histoembryological differences, analysis was conducted with the patients in two subgroups: left hemicolon cancer group and right hemicolon cancer group. The results showed that the positive rate of CTCs increased in both groups with the increase in tumor stage. The survival analysis indicated that there was a steep gradient in survival in the follow-up period, particularly in the CTC-positive group (P=0.000). Risk assessment curves showed that the change escalated more rapidly in the CTC-positive group. Furthermore, with the increase in T stage, changes in the survival curve and risk curve escalated more rapidly in the CTC-positive group. CONCLUSION: It was confirmed that in the left hemicolon cancer group, a much higher coincidence rate could be found on CTC-positive rate and clinicopathological features, than in the right hemicolon cancer group. The sensitivity of CTCs may be related to the histoembryological location of the tumor, lymphatic metastasis and the depth of infiltration. Monitoring CTCs may have value in evaluating clinical staging and estimating clinical prognosis.

Altered structure and anion transport properties of band 3 (AE1, SLC4A1) in human red cells lacking glycophorin A.

We have studied the properties of band 3 in different glycophorin A (GPA)-deficient red cells. These red cells lack either both GPA and glycophorin B (GPB) (M(k)M(k) cells) or GPA (En(a-) cells) or contain a hybrid of GPA and GPB (MiV cells). Sulfate transport was reduced in all three red cell types to approximately 60% of that in normal control red cells as a result of an increased apparent K(m) for sulfate. Transport of the monovalent anions iodide and chloride was also reduced. The reduced iodide transport resulted from a reduction in the V(max) for iodide transport. The anion transport site was investigated by measuring iodide fluorescence quenching of eosin-5-maleimide (EMA)-labeled band 3. The GPA-deficient cells had a normal K(d) for iodide binding, in agreement with the unchanged K(m) found in transport studies. However, the apparent diffusion quenching constant (K(q)) was increased, and the fluorescence polarization of band 3-bound EMA decreased in the variant cells, suggesting increased flexibility of the protein in the region of the EMA-binding site. This increased flexibility is probably associated with the decrease in V(max) observed for iodide transport. Our results suggest that band 3 in the red cell can take up two different structures: one with high anion transport activity when GPA is present and one with lower anion transport activity when GPA is absent.