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Title: The significance of molecular biomarkers in the recurrence of rectal tumors after Transanal Endoscopic Microsurgery
Author: Sideris, Michail
ISNI:       0000 0004 7656 4435
Awarding Body: King's College London
Current Institution: King's College London (University of London)
Date of Award: 2018
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Local excision (LE) of rectal cancer has been practiced as a treatment for 30 years on a highly selected group of patients and tumours. A method of local excision which has recently gained wider acceptance in the treatment of low-grade CRC (T1) is transanal endoscopic microsurgery (TEMS). TEMS offers advantages for operative access and oncological clearance compared to those of transanal resection (TAR). A number of studies have shown that TEMS can have almost equal results to radical surgery for early rectal cancer. Radical surgery has the disadvantage of an approximate mortality rate of 5% and a complication rate of around 20%, and it impacts on quality of life due to stomas and of sexual dysfunction. However, TEMS has a higher local recurrence rate, and efforts have been made to classify risk with morphological and histological criteria. Molecular biomarkers in the evaluation of CRC prognosis and treatment stratification have been extensively discussed in the literature. Until now, 3 pathways have been identified to explain the background of CRC molecular pathogenesis. Microsatellite instability (MSI) refers to point mutations in the DNA mismatch repair genes. MSI is currently responsible for 15−20% of CRC cases, and there is enough evidence to support its association with prognosis. Chromosomal instability (CIN) is another pathway of carcinogenesis which affects 85% of CRC cases and has been flagged as a poor prognostic marker. CIN encompasses any structural chromosomal abnormality that results in aneuploidy or polyploidy. The third pathway is related to aberrant hypermethylation of suppressor promoter CpG islands, commonly known as CIMP. v-raf murine sarcoma viral oncogene homolog B (BRAF) encodes a serine-threonine protein kinase that acts as a downstream effector of Kirsten rat sarcoma viral oncogene homolog (KRAS) pathways. Various studies have revealed that v-raf murine sarcoma viral oncogene homolog B V600E (BRAF V600E) mutations appear to be valid prognostic indicators. KRAS is a proto-oncogene that encodes a GTPase, which is involved in facilitating cellular response to extracellular stimuli. KRAS point mutations appear in 40% of CRC cases, and their presence is associated with poor response to anti-epidermal growth factor receptor (anti-EGFR) chemotherapy. However, to date there has been no literature linking biomarkers with stratification of risk for the treatment of rectal cancer following TEMS. Aim: The aim of this dissertation is to assess the significance of these molecular biomarkers in the prediction of local recurrence and prognosis of early rectal cancer following TEMS. Materials and Methods: Initially, we performed a narrative review of the literature to consolidate the evidence available for molecular biomarkers in the evaluation of CRC. We then designed a retrospective pilot study to identify the molecular biomarker status of 41 confirmed CRC cases among 1,446 consecutive referrals for suspected cancer. As part of this study, we retrospectively analysed clinical, biochemical, and histopathological data. Gene profile analysis (KRAS, BRAF) of the specimens was also performed. Following this, we proceeded to analyse data from a series of patients who had undergone TEMS for Stage I rectal cancer at King’s College Hospital (KCH). Demographic, biochemical, histopathological, and follow-up data were prospectively collected. Molecular analysis was prospectively performed in the Advanced Diagnostics Laboratory of KCH to identify the status of BRAF, KRAS, p16 O6-methylguanine-DNA methyltransferase (MGMT), and β-catenin. Finally, we retrospectively collected equivalent data on a 4-year series of 135 confirmed Stage I−IV rectal cancer cases who underwent radical surgery +/- neoadjuvant chemoradiotherapy. Data on the status of the same molecular biomarkers were retrospectively collected. In both cohorts of rectal cancer cases (TEMS/radical surgery +/- additional treatment), the biomarker status was compared with the histopathology and follow-up outcomes, including recurrence and overall cancer-related survival. Results: In our pilot study (41 cases), there was no significant correlation of presenting haemoglobin (Hb) levels with eventual disease staging (p>0.05 for all associations). Patients with right-sided tumours were found to have a lower Hb level than patients with either left-sided or rectal tumours. Hb levels were also significantly lower in patients with the BRAF V600E mutation, although this may be because all 3 patients with the mutation had right-sided tumours. Neither KRAS status nor lymphovascular invasion (LVI) status had a specific correlation with Hb levels. Of 29 specimens of cases who underwent TEMS, there was a statistically significant association between KRAS mutant status and local recurrence (n=6, p=0.037). P16 expression > 5% (mean=10.8%, min=0, max=95) was associated with earlier recurrence within 11.70 months (n=7, p=0.004). Membranous β-catenin expression (n=12, 48%) was also related to KRAS mutant (mt) status (p=0.006) but not to survival (p>0.05). BRAF gene was found to be wild type in all cases tested (n=23). With regard to the specimens of rectal cancer cases who underwent radical excision, 28 cases were Stage I (20.9%), n=30; Stage II (22.4%), n=45; Stage III (33.6%) and n=31 Stage IV (23.1. KRAS mt status was associated with female gender (n=20, p=0.021) and older age (69.62 vs. 62.27, p=0.005). Stage I early cancer subgroup analysis showed that KRAS mt status was associated with distant recurrence of disease (n=4, p=0.045). Conclusions: BRAF V600E mutation seems to be associated with right-sided CRC and iron-deficiency anaemia. This could be used as an adjunct to diagnostic molecular tests for early diagnosis. KRAS, p16, and β-catenin could be used as biomarkers for prediction of local recurrence and stratification of the risk for further surgery in Stage I rectal cancer. Further to this, KRAS may be a predictor of distant recurrence in cases of early stage rectal cancer.
Supervisor: Bjarnason, Ingvar Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available