Personalised Medicine in Colorectal Cancer? Mr Arfon G M T Powell MB ChB MSc MRCSEd Clinical Research Fellow in Surgery

Colorectal cancer is the third most common cancer in the UK 39,991 new cases in 2008 Cancer Reseach UK. Bowel cancer statistics – UK,

CRC is the 2 nd most common cause of cancer-death Accounting for 16,259 deaths in 2009 Cancer Reseach UK. Cancer Mortality – UK Statistics

Treatment Treatment regimens are currently based on disease stage Surgery Chemotherapy – Curative – Palliative Biological therapy

Prognosis Prognosis still remains stage dependent – Dukes’ A  93% – Dukes’ B  77% – Dukes’ C  48% Cancer Reseach UK. Bowel cancer statistics – UK, bowel-cancer#outlook

Surgical approach to colorectal cancer

Variation in biomarker prognostic value

Colorectal cancer development Accumulation of genetic alterations – Vogelstein

Microsatellite Instability Phenotype Distinct genomic instability pathway Microsatellite repeats Associated with loss of mismatch repair protein (MMR) function Improved outcome

Söreide K, Janssen EA, Söiland H, Körner H, Baak JP. Microsatellite instability in colorectal cancer. Br J Surg 2006; 93:

CpG Island Methylator Phenotype Hypermethylation of cytosine- and guanine-rich stretches of DNA, called CpG islands, in the promoter region of genes causes transcriptional silencing and has been implicated in carcinogenesis

MSI/CIMP+ MSI/CIMP- Microsatellite stability status CIMP status CIMP +ve CIMP -ve MSI MSS MSS/CIMP+ MSS/CIMP- CIMP +ve CIMP -ve

MSI/CIMP+ MSI/CIMP- Microsatellite stability status CIMP status CIMP +ve CIMP -ve MSI MSS MSS/CIMP+ MSS/CIMP- CIMP +ve CIMP -ve Good survival Poor survival

MSI/CIMP+ MSI/CIMP- Microsatellite stability status CIMP status CIMP +ve CIMP -ve MSI MSS MSS/CIMP+ MSS/CIMP- CIMP +ve CIMP -ve Prognostic information remains unclear

Serrated Adenocarcinoma Proximal location MSI positive Outcome variable which depends on tumour site Serrated Adenocarcinoma Non Serrated Adenocarcinoma

Our experience with performing MSI and CIMP status analysis on colorectal tumours

Study design Retrospective study of 750 FFPE tumours IHC for MMR proteins (MLH1, MSH2, MSH6 and PMS2) 40% tumour required within the section for PCR MSI PCR analysis of: – BAT 25 – BAT 26 – MONO 27 – NR-21 – NR-24

Technical issues 55% of patients required macroscopic dissection to the equivalent of 2 10micron sections

Technical issues 55% of patients required macroscopic dissection to the equivalent of 2 10micron sections

MSI +ve MSI -ve

Preliminary results on 233 patients Not significantly associated with: – Increasing age (P=0.168) – Dukes stage (P=0.054) – Poor differentiation (P=0.362) – Vascular invasion (P=0.176) – Anaemia (P=0.192) – Raised CRP (P=0.374) – Hypoalbuminaemia (P=0.541) – Emergency presentation (P=0.943) Significantly associated with: – Right colon location (P<0.001) – Polypoid morphology (P=0.031) – Lower lymph node ratio (P=0.040) – Mucin production (P=0.009) – Serrated adenocarcinoma (P<0.001)

P=0.042 The relationship between MSI status and cancer-specific survival

CIMP study design Extracted DNA requires bisulfite conversion Followed by a methylight PCR assay for – CACNA1G – IGF2 – NEUROG1 – RUNX3 – SOCS1

DNA recovery following bisulfite treatment DNA recovery following bisulfite treatment is variable and does not reach the projected > 75% PatientInput (ng)nano drop (ng/ul)DNA recovered (ng)Percentage recovered (%)

MSI and CIMP status as predictors of response to treatment

Treatment A Curative resection surgery B CResection surgery + adjuvant therapy (eg. Chemothearpy) DDependent on tumour characteristics

Treatment of colorectal cancer Surgery remains the primary modality for cure Chemotherapy for high risk patients – Lymph node involvement – Locally advanced tumours MDT decision Difficulty identifying patients that benefit from chemotherapy

Adjuvant Chemotherapy 2 major regimens for CRC treatment: – FOLFIRI (5-FU, folinic acid [Leucovorin], and irinotecan [Campostar]) – FOLFOX (5-FU, folinic acid [Leucovorin], and oxaliplatin [Eloxatin])

Adjuvant Chemotherapy 2 major regimens for CRC treatment: – FOLFIRI (5-FU, folinic acid [Leucovorin], and irinotecan [Campostar]) – FOLFOX (5-FU, folinic acid [Leucovorin], and oxaliplatin [Eloxatin]) Results in context of MSI is conflicting

Conclusions Colorectal cancer tumour heterogeneity exists Techniques validated MSI+/CIMP+ confers improved survival Response to treatment remains unclear

Acknowledgments I would like to thank Dr David Baty and Christine Black (Molecular Genetics, Dundee) for their expertise with the MSI analysis I would like to thank Rachael Ellis (Molecular Genetics, Glasgow) for her help with the bisulfite treatments I would like to thank Clare Orange for her continued help over the last 3 years!

Thank you!