Significance Of Cytochrome C Oxidase Subunit 1 In The Early Diagnosis And Treatment Of Colorectal Cancer

Colorectal cancer (CRC) is one of the most common gastrointestinal cancers, incidence of CRC rand the third of totoal malignant tumors, and CRC is the fourth leading cause of cancer death. Although morbidity of colorectal cancer tends to be stable or even decreases in western world, the morbidity of ulcerative colitis-associated colorectal cancer (UCACC) is expected to increase along with increase of morbidity of ulcerative colitis (UC). Therefore, the early diagnosis of colorectal cancer and identification of high-risk population is of great clinical importance. Currently tumor biomarker is one of the focused research areas of life sciences fields. However, the current tumor biomarkers of colorectal cancer have great limitation and are lack of clinical utility. Therefore, it is high time to develop high-sensitivity and high-specificity biomarkers for detecting colorectal cancer and high-risk population among ulcerative colitis patients. So that high-risk population can be guided to increase colonoscopy screen test frequency, change life styles and eating habits, make better chemical prevention strategy, and treatment therapy etc. to achieve early diagnosis、treatment and improvement of prognosis of colorectal cancer.Investigating the mechanism of UCACC tumorigenosis helps to better understand initiation and progression of cancer, so that to provide useful information for making early prevention scheme and selecting treatment target. Currently the tumorigenosis research have focused on alteration of genes. Studies have proved that alteration of genes happened in UC before historical changes of dysplasia appeared. Recently, it has been proved that mitochondrial genes is closed involved in tumorigenosis, studies suggested alteration of cytochrome c oxidase function is a good reflection of alteration of mitochondrial genes and mitochondrial function。In first chapter, immunohistochemistry with Nuance multispectral imaging quantification was performed to detect expression changes of mitochondrial protein— cytochrome c oxidase subunit 1 (CCO1) along course of sporadic colorectal cancer (SCC) tumorigenosis. CCO1 expression was high in normal colonic crypts、tumor and organ/lymph node metastasis, decreased in adenoma and lowest in para-carcinoma tissue, which suggests that mitochondrial genes is related with colorectal tumorigenosis. Next we choose UCACC as study model to investigate the role of mitochondria in "Inflammation (non-dysplasia)- dysplasia - cancer" pathway. Different Pathological degrees of biopsies from same UCACC colectomy were selected, immunohistochemistry of CCO1 was performed. CCO1 expression in non-dysplasia decreases with increasing proximity to tutmor; along with the disease progression, CCO1 expression increased in low-grade dysplasia、high-grade dysplasia and tumor.Therefore, we assumed CCO1 loss precedes the development of dysplasia in UC and it could be used to detect and potentially predict cancer.In chapter two, CCO1 immunostaining was performed in three non-dysplasia biopsies from six UCACCs (UC progressor) and six UC patients (UC non-progressor). At the same time, NanoString nCounter was performed in these samples to assess 70 genes mRNA expression including 13 mitochondrial genes and 57 nuclear encoded genes related to mitochondrial function. CCO1 protein and mitochondrial genes expression were significantly lower in UC progressor than in UC non-progressor. Then multiple statistical methods were applied and diagnostic models based on CCO1 protein and several mitochondrial mRNA biomarker panels for UC progressor were trained.In chapter three, NanoString nCounter was performed in a larger independent cohort consists of six non-dysplasia biopsies from 28 UC progressors and 28 UC non-progressors. Receiver operating characteristic (ROC) curve analysis was used to examine the validity of diagnostic models based on different mitochondrial mRNA biomarker panels. Based on area under curve (AUC) of 0.92, highest sensitivity of 91.67% and specificity of 82.14%, the mean MT_ CO1 mRNA expression of six non-dysplasia biopsies was selected as the most valuable diagnostic marker, which could distinguish UC progressors from UC non-progressors. In addition, reproducibility test was done to evaluate performance of NanoString nCounter system. NanoString nCounter was proved to provide high specificity、accuracy and reproducibiliby results.In chapter four, based on previous finding that mitochondrial function is upregulated in tumor, hydrogen peroxide-induced apoptosis of HCT116 cells in vitro was established to investigate the role of mitochondrial in tumor apoptosis.0.8mM H2O2 at 6 hours significantly suppressed cell proliferation and enhanced cell apoptosis percentage; mitochondrial protein and mRNA expression were significantly decreased and activitiy of cytochrome c oxidase were significantly inhibited. It is assumed that mitochondrial depletion might have an important role in tumor apoptosis. Therefore, mitochondrial could be potentially used as a treatment target for colorectal cancer in the near future.In conclusion, mitochondrial loss preceds UC tumorigenosis; then mitochondria restored in UC associated dysplasia/cancer. MT_ CO1 mRNA level in non-dysplasia tissues is an effective diagnostic biomarker for UC progressor; targeted inhibition of mitochondrial genes might be a potential treatment for colorectal cancer.