Carol Burke, MD, FACG, FASGE, AGAF, FACPSanford R Weiss Center for Hereditary Colorectal NeoplasiaDepartment of Gastroenterology and HepatologyCleveland Clinic, Cleveland, OhioSeptember 19, 2015

New Era of Genetic Testing in Colon Cancer

Overview

• Recognize clues suggestive of a genetic colorectal cancer syndrome

• Understand the genetic testing process for hereditary colorectal cancer syndromes

Pathways to CRC

AdenomaAdenoma SessileSessile Serrated Polyp Serrated Polyp

Sporadic CRC

CIN-MSS CIMPCIN-MSS CIMP

MSI MSIMSI MSI

MLH1 promotor methylation BRAF mutation

Lynch Syndrome

FAP MAP

Chromosomal Instability

Pino MS, et al. NEJM 2010;339;1277

CpG Island Methylation (CIMP)

Gene Expression

Gene Silencing

Turns off MLH1

Multi Target Stool DNA Testing vs FIT • Assay: Methylation of BMP3 and NDRG4, KRAS mutations ,

B-actin and a fecal immunochemical test

Imperiale T, 2014;370:1287

P < 0.001%

P < 0.001P = 0.002

Microsatellite Instability

• Repeated nucleotide sequences called “microsatellites”

• DNA fidelity maintained by Mismatch Repair Proteins (MMR)

Boland CR, Gastroenterology 2010;138:2073

MLH1 PMS2

MSH2 MSH6

Mismatch Repair Protein Function

Nucleotide

mismatch

Normal MMR

Defective MMR Etiology: 1.MLH1 promoter methylation2.Germline MMR mutation- Lynch Syndrome

TT CCTT AA CC

A G C T GA G C T G

T C G A T C G A CCA G C T GA G C T G

TT CCTT AA CC

A G C T GA G C T G A G A G AA T GT G

T C T C TT A C A C

Microsatellite Instability (MSI)

Microsatellite Stable (MSS)

8

Tumor MSI TestingNR21 BAT25 Mono27

Normal Tissue

Tumor Tissue

MSI-H: > 2/5 (30%) consensus MSI sequences

MLH1 MSH2

Immunohistochemistry

Multi Society Task Force Universal Testing of CRC for dMMR

11

Giardiello FM , Am J Gastro 2014;109:1159

Universal Tumor Testing for LS• 1066 unselected tumors assessed for MSI/MMR

• 19.5% had MSI– 11% (21 patients) diagnosed with LS

• Phenotype:– 43% diagnosed > 50 years

– 22% did not Amsterdam II or revised Bethesda guidelines

Hampel H et al. NEJM 2005;352;18

Germline Testing Results In 21 Proband’s Relatives

Relationship Tested Positive Negative

First degree 54 25 29

Second degree 22 10 12

> Third degree 41 17 24

Total 117 52 65

Typical Genetic Counseling Appointment

• Collect personal and family history

• Perform risk assessment (including breast cancer risk models)

• Educate about genetic syndromes, management options, genetic testing process

• Informed consent and coordination of genetic testing

• Psychosocial support and counseling

Who Should Have Genetic Counseling for Lynch Syndrome?

• Abnormal MSI/IHC testing–(Unless MLH1 methylation is proven)

• Colon or endometrial cancer < 50

• > 2 Lynch Syndrome cancers in individual

• > 2 relatives with LS cancer, 1 < 50

• > 3 relatives with LS cancer at any age

• > 10 colon adenomas

• Peutz-Jeghers Polyp

• Juvenile/Inflammatory Polyps

• Colon cancer < age 50

• Close relative diagnosed < age 50 or >2 close relatives with colon cancer

Who Should Have Genetic Counseling for other Colon Cancer Syndromes?

Hereditary Colon Cancer SyndromesSyndrome Gene(s) Features

Lynch MLH1, MSH2, MSH6, PMS2, EPCAM

Colon, endometrial, ovarian, gastric, urinary tract, small bowel cancers, brain tumors, sebaceous neoplasms

Li Fraumeni TP53 Childhood cancers, sarcoma, leukemia, brain tumors, breast cancer, colon cancer

Familial Adenomatous Polyposis (FAP)

APC Adenomas, colon cancer, thyroid cancer, osteomas and soft tissue tumors, desmoid tumors

MYH-Associated Polyposis (MAP)

MUTYH* Adenomas, colon cancer, thyroid cancer

Peutz-Jeghers STK11 Mucocutaneous melanin spots, hamartomas, breast, GI, pancreatic, and rare gyn cancers

Cowden PTEN Hamartomas, derm lesions, macrocephaly, breast, thyroid, and endometrial cancers

Juvenile Polyposis Syndrome

BMPR1A, SMAD4

Hamartomas, colon cancer, some with SMAD4 have HHT

HNPCC/Lynch syndrome!

colon ca 50

d. uterine ca 61

d. stroke 80

colon ca 47

57 6059 62

32 302427 35

d. colon ca 47

Traditional Cancer Risk Assessment and Genetic Testing

Traditional Genetic Testing

• Utilizes Sanger sequencing and large rearrangement analysis

• Testing often limited to 1-2 syndromes based on assessment of personal/family history

• Only testing for high-risk, well known syndrome

• Variant of uncertain significance rate is low

• Results take 2-3 weeks

Next Generation Sequencing

• Whole genome, or several genes can be analyzed at once

• Allows for testing many genes relatively inexpensively

• Used for panel genetic testing

– Cancer specific vs

– Pan cancer

Multi-Gene Panels

• Benefits–Increased mutation

positive rate

–Identification of conditions of low clinical suspicion

–Cost effective

–Lower turn-around-time then reflex testing

• Limitations–Not all tests are

equal

–Various levels of gene coverage

–Increased VUS rate

–Moderate-risk genes

–Longer turn-around-time than single gene

Insurance Coverage & Cost

• Most labs offer insurance pre-authorization

• Many labs billing with same CPT codes as BRCA, Lynch testing

• Costs range from $1500-$4400

What Are Testing Options?

• 10 panels (5-28 genes)

• 10 panels (7-29 genes)–Build your own

• 7 panels (7-29 genes)–Build your own

• 1 panel (25 genes)

• 2 panels (20-52 genes)

Example from Invitae

Myriad Genetics Lab myRisk Gene Panel

High Risk Genes

Breast Cancer

• BRCA1/BRCA2 (HBOC)

• CDH1 (Hereditary Diffuse Gastric Cancer)

• PTEN (Cowden)

• STK11 (Peutz-Jeghers)

• TP53 (Li-Fraumeni)

Colon Cancer

• APC (Familial Adenomatous Polyposis)

• BMPR1A/SMAD4 (Juvenile Polyposis)

• MLH1/MSH2/MSH6/PMS2/EPCAM (Lynch Syndrome)

• MYH (MYH-Associated Polyposis)

High Risk Genes

• Significant risk of developing certain types of cancer

• Considerable research and professional society guidelines for screening and surgery

• Family members can be tested for the same mutation

Moderate Risk Genes

• ATM (Ataxia Telangiectasia)

–Breast, pancreatic, colon

• CDKN2A (Familial Atypical Mole Malignant Melanoma)

–Pancreatic, melanoma

• CHEK2 (Li-Fraumeni Like Syndrome)

–Breast, prostate, colon

• PALB2 (Fanconi Anemia)

–Breast, pancreatic

Moderate Risk Genes

• 2- to 4-fold risk over the general population risk.

• Cancer risk is not as elevated as a mutation in a high risk gene.

• Limited research and no professional society screening or surgical recommendations

• Family history is often better for risk stratification

• Unclear if it is beneficial to test other family members for these mutations

Potential Results from myRisk

• Positive

–High risk gene

–Moderate risk gene

–New moderate risk gene

• Negative

• Variant of uncertain significance

Incidental Finding Case 2

42dx br ca

42

45 34

64 6555 d. youngaccident

61 test ca

85dx female ca 65

90 d. 83dx br ca 60s

no info

Caucasian/N. American IrishAJ

n

Lynch Syndrome PMS2

NCCN 2.2014

2926

55 d.55 b/l br ca 44 BRCA -

47 50

8189CRC 89

88br ca 45

d. 64

Hungary Sicily

61 51

p

2

d. 40 Panc ca

d. 54 Panc ca

Incidental Finding Case 3

Hereditary Diffuse Gastric Cancer

• CDH1 mutations

• 39% breast cancer risk by age 80

• Diffuse gastric cancer

– 67% for men and 83% for women by age 80

– Average onset 38 years (range of 14-69 years)

– Options for intense screening or prophylactic gastrectomy/mastectomy

Variants of Uncertain Significance: Our results

• 235 VUS in 171 patients (41.4%)

– majority in moderate risk genes (62.2%)

• VUS rate lower in those of European ancestry than African, Asian, and Middle Eastern (p=0.001)

Important Insurance Updates

• Companies requiring pre-test genetic counseling:

–CareSource

–CCF Employee Health Plan

–Cigna

–Medical Mutual of Ohio

Conclusions

• Variety of “Genetic” tests to determine cause of hereditary colon cancer syndromes

• Currently, test the tumor first in patients with CRC

– Germline testing for polyposis or when tumor not available

• Genetic testing in transformation:

• NGS lowering costs and driving panel based testing

– Not all panel tests are created equal

• Caution: panel testing “easy” to order but complicated to interpret

• Get a lot of information we might not understand

• Find unanticipated mutations that highly impact patient care