BRCA1 BRCA2 CDKN2A Etc

Registro español de cáncer de páncreas familiar (Pan-Gen-FAM): Análisis genómico de individuos afectos y cribado de personas de alto riesgo

C GUILLÉN-PONCE , J EARL , MT SALAZAR LOPEZ, C CALCEDO, R FERREIRO, E MOCCI, C GUERRERO, M MARQUEZ, C GONZÁLEZ GORDALIZA, MJ MUÑOZ BELTRÁN, E VAZQUEZ SEQUEIROS, A SANJUANBENITO, J MONTANS, SOLERA GARCÍA J, FX REAL, N MALATS AND A CARRATO.

Pancreatic Cancer Etiology

 10th most common cancer

 Incidence almost equals mortality: Worldwide 217,000 new cases and around 213,000 deaths.

Pancreatic Cancer Etiology

 10th most common cancer

 Incidence almost equals mortality: Worldwide 217,000 new cases and around 213,000 deaths.

 Overall 1 year survival rate 20%  Overall 5 year survival rate 5.4%

 80% of patients present with locally advanced or metastatic disease

Pancreatic cancer risk factors

• Smoking, type II diabetes, BMI, heavy alcohol consumption, family history.

Pancreatic cancer risk factors

• Smoking, type II diabetes, BMI, heavy alcohol consumption, family history.

• Sporadic PC 1:10,000, increased risk according to the number of affected family members: 1 : 2.3-4.51 fold, 2 : 6.4 fold and 3: 32 fold

• 5-10% of cases have a family history:

Pancreatic cancer risk factors

• Smoking, type II diabetes, BMI, heavy alcohol consumption, family history.

• Sporadic PC 1:10,000, increased risk according to the number of affected family members: 1 : 2.3-4.51 fold, 2 : 6.4 fold and 3: 32 fold

• 5-10% of cases have a family history:

• Familial Pancreatic Cancer (FPC): rare syndrome, autosomal dominant, defined as families with two or more first-degree affected relatives that do not fulfil the criteria of other inherited tumor syndromes with increased risks of pancreatic cancer

• Breast/ovarian, Peutz Jeghers Syndrome (PJS), Familial Atypical Multiple Mole Melanoma (FAMMM), HNPCC, hereditary pancreatitis (PRSS1)

• 15–20% of families carry germline mutations in BRCA2, PALB2 and ATM

Objectives: PANGEN-FAM

• Identification and creation of a national registry of families with FPC.

Objectives: PANGEN-FAM

• Identification and creation of a national registry of families with FPC. • Characterization of the phenotype of families and offer appropriate genetic testing.

Objectives: PANGEN-FAM

• Identification and creation of a national registry of families with FPC. • Characterization of the phenotype of families and offer appropriate genetic testing. • Whole exome sequencing of individuals in order to identify novel genes and/or variants associated with FPC. • Establish a screening program for at risk to detect a Pancreatic Cancer (PC) at a potentially curative stage. • Endoscopic Ultrasound (EUS) vs Magnetic Resonance Imaging (RMI) both conventional and with diffusion. • Evaluation of minimally invasive tumor marker detection techniques: Circulating Tumor Cells (CTCs), cfDNA y miRNAs in peripheral blood. • Compare the results with other international screening programs and establish collaborations.

Pancreatic cancer families inclusion criteria

 ≥ 2 cases of 1st degree relatives or ≥ 3 cases independent of relationship.

 Families that present with other criteria:

• Peutz-Jeghers Syndrome (SPJ), Familial Atypical Multiple Mole melanoma (FAMMM)* and Hereditary Breast and Ovarian Cancer (HBOC)*. • Families with Hereditary Pancreatitis (HP). • Families that present with a young case of PC (<=50 years old)**.

*For these syndromes only families that present with at least one case of PC will be included. **These families will only undergo genetic analysis and not screening.

Study Protocol

 Obtain informed consent.  Generate a family tree with a minimum of 3 generations.  Completion of an epidemiological questionnaire: demographic data, smoking, alcohol consumption and clinical data.  Collection of biological samples: • PC cases: blood and tumor tissue (fresh or paraffin embedded, depending upon the availability). • At risk family members: blood.  Screening program for at risk individuals.

Families registered in PAN-GEN-FAM

Number of families Family Phenotype registered FPC 21 FPC >= 3 PC 4 PC <= 50 Years of age 6 HBOC + PC 10 HBOC + PC <=50 Years of age 6 FAMMM-PC 1 FPC + Gastric 2 FPC (paternal)/HBOC + PC 1 (maternal) FPC+LYNCH+HBOC 1 Total 52

Sample Collection

20-50ml Sangre serum, plasma, lymphocytes, leucocytes and RBC

Sample Collection

20-50ml Sangre serum, plasma, lymphocytes, leucocytes and RBC

Routine genetic tests: BRCA1 BRCA2 CDKN2A etc.

Sample Collection

20-50ml Sangre serum, plasma, lymphocytes, leucocytes and RBC

Routine genetic Investigation project: tests: BRCA1 Whole exome sequencing BRCA2 in 1-2 cases per family in CDKN2A etc. germ line and tumor DNA.

Sample Collection

20-50ml Sangre serum, plasma, lymphocytes, leucocytes and RBC

Routine genetic Investigation project: Minimally invasive tumor tests: BRCA1 Whole exome sequencing markers: CTC, miRNAs, BRCA2 in 1-2 cases per family in cfDNA. CDKN2A etc. germ line and tumor DNA.

BRCA germline mutation screening

BRCA1 BRCA1 variant BRCA2 BRCA2 variant of Family Phenotype mutation of unknown mutation unknown positive significance positive significance FPC 0/6 5/6 0/6 2/6 FPC >= 3 PC 0/1 1/1 0/1 0/1 PC <= 50 Years of age 0/1 0/1 0/1 0/1 HBOC + PC 1/6 3/6 2/6 2/6 HBOC + PC <=50 Years 0/6 3/6 3/6 2/6 of age unclassified 0/1 1/1 0/1 1/1 Total 1/21 13/21 5/21 7/21

BRCA germline mutation screening

Screening Protocol

EUS & MRI

EUS + MRI EUS or RMI Normal Abnormal

Evaluation by a multidisciplinary Repeat imaging annually team (oncologists, surgeons, gastroenterologists, radiologists, pathologists)

Screening Results 57 at risk individuals from 24 families (1-13 participants per family) participate in the annual imaging screening program (EUS/MRI). The majority of anomalies identified are consisted with changes associated with pancreatitis. All Rounds Number of Number of Extra Abnormalities Technique Pancreatic Abnormalities detected pancreatic detected abnormalities lesions pancreatic heterogeneity (12), cysts (7), lobulation (5), gallstones, fatty EUS 28/78 (36%) 4/78 (5%) hyperechoic areas (3), liver, hiatal hernia pancreatitis (2) solid lesion (3) renal, biliary, MRI 17/51 (33%) Cysts (10) 29/51 (57%) hepatic cysts

Cystic Lesions One solid lesion and 13 cystic lesions. One cystic lesion was subsequently identified as a well differentiated neuroendocrine tumor and another was shown to have benign cytology and whilst a carcinoma was found in a third patient. Identification Other Observations CT CT/MRI EUS/MRI EUS/MRI FNA: benign EUS Following EUS: NET. Resected March 2013 Mucinous Tumor <1cm, PAAF 2014 carcinoma, resected June CT/EUS/MRI 2014 MRI Detected in round 3 MRI Detected in round 3 MRI CT/EUS/MRI RM MRI/EUS Two lesions (IPMN) MRI/EUS CT/EUS Solid lesion, 5x4mm in body-tail, resected 08/2015

57F093021001 (carcinoma)

 30/01/2012 EUS: Normal;  06/09/2012 MRI: cystic lesion 1 cm;  21/03/2013 MRI: cystic lesion 1cm unchanged from previous MRI;  05/04/2014 MRI: cystic lesion 1cm; unchanged from previous MRI;  07/04/2014 EUS+PAAF: 8,7 x 8,9 mm contact with the pancreatic 2o branch. Pathology : carcinoma.

57F093021001

Pathology of the resected lesion

A: (PAP, 400x) Images of cells from FNA B: (H-E, 200x) Histology images of the 0.4 of a cystic lesion of 0.6 cm. The cells cm lesion surgically removed showed have a large cytoplasm and nuclear characteristics of serous cystadenoma with atypia suspicious of carcinoma. atypical changes and PanIN-3

57F093049001 (malignant lesion)

 08/06/2015 EUS: solid lesion 5x4mm in body-tail; PAAF: atypical epithelial lesion, potentially malignant  18/06/2015 MRI: Normal  28/08/2015: Esplenopancreatectomía corporocaudal laparoscópica

Pathology: multiple PanIN-I and PanIN-II lesions

57F093049001

GRUPOS INTERNACIONALES Creación de nuevo grupo internacional; EU-Collaborative Group of Prevention of Pancreatic Cancer

EU- Collaborative Group of Prevention of Familial Pancreatic Cancer

GRUPOS INTERNACIONALES Creación de nuevo grupo internacional; EU-Collaborative Group of Prevention of Pancreatic Cancer

Screening protocols

 The surveillance program in Madrid included annual EUS and MRI.

 The surveillance programme at the Philipps University in Marburg included annual screening with MRI with MRCP and EUS between 2002 and 2010. Since January 2011 follow-up imaging consisted of annual MRI with MRCP and EUS every third year or when suspicious alterations were detected by MRI.

 IAR of FPC families at the Leiden University Medical Centre were invited for an annual MRI with MRCP. Beginning in 2011, EUS was also offered as an option in addition to annual MRI.

Characteristics of pancreatic lesions in IAR (n=133) Parameter All IAR with IAR with lesions IAR with lesions lesions first screening during follow-up (n=133) (n=88) (n=45) Type of lesion 126 / 2 / 5 83 / 0 / 5 43 /2 / 0 (cystic/solid/other) Detected by EUS 58 (44 %) 47 (53%) 11 (24%) Detected by MRI+MRCP 122 (92%) 79 (90%) 43 (96%) Detected EUS+MRI 45 (34%) 36 (41%) 9 (20%) Surgery 20 (15%) 9 (10%) 11 (24%) potentially relevant 14/20 (70%) 6/9 (67%) 8/11 (73%) histology* significant histology# 5/20 (25%) 2/9 (22%) 3/11 (27%)

* includes 1 PC, PanIN3, multifocal PanIN2 + BD-IPMN+AFL, IPMN with moderate/high dysplasia, pNEN; # includes only PC, PanIN3 and IPMN with high grade dysplasia

Prevalence of pancreatic lesions according to age (133 IAR) Age at first IAR with any IAR with lesions IAR with lesions screening pancreatic lesions at 1st screening during follow-up (%) (%) (%)

< 40 years 16/48 (33%) 13 (34%) 3 (6%) > 40 years 117/204 (57%) 75 (37%) 42 (20%) < 45 years 36/98 (37%) 26 (26%) 10 (10%) > 45 years 97/154 (63%)* 62 (40%) 35 (23%) < 50 years 67/152 (44%) 46 (30%) 21 (14%) > 50 years 66/100 (66%) 42 (42%) 24 (24%) < 55 years 86/183 (47%) 56 (31%) 30 (16%) > 55 years 47/69 (68%) 32 (46%) 15 (22%) < 60 years 107/217 (49%) 70 (32%) 37 (17%) > 60 years 26/35 (74%) 18 (51%) 8 (23%)

Total with lesions 133/252 (52.8%) 88 (35%) 45 (18%)

*-p=0.0001

Proportion of potentially relevant lesions according to age Age at first screening Relevant lesions# IAR with relevant IAR with relevant detected by lesions# at 1st lesions# during screening screening follow-up

< 40 years 0/48 (0%) 0/48 (0%) 0/48 (0%) > 40 years 14/204 (6.5%) 5/204 (2.4%) 9/204 (4.5%) < 45 years 0/98 (0%) 0/98 (0%) 0/98 (0%) > 45 years 14/154 (9.1%) 5/154 (3.2%) 9/154 (5.9%) < 50 years 2/152 (1.3%) 0/152 (0%) 2/152 (1.3%) > 50 years 12/100 (12%)* 5/100 (5%) 7/100 (7%) < 55 years 7/183 (3.8%) 1/183 (0.5%) 6/183 (3.3%) > 55 years 7/69 (10.1%) 4/69 (5.8%) 3/69 (4.3%) < 60 years 9/217 (4.1%) 1/217 (0.4%) 8/217 (3.7%) > 60 years 5/35 (14.3%) 4/35 (11.4%) 1/35 (2.9%) Total IAR with 14*/252 (5.6%) 5/252 (2%) 9/252 (3.6%) lesions

# potentially relevant lesions: PC, multifocal PanIN2/3 lesions with/without BD-IPMN or AFL, pNET; * p=0.0004

conclusions

 The age at which screening should be initiated is uncertain. Almost all programs start screening at the age of 40 to 50 years or 10 years below the youngest age of onset in the family.  Endoscopic ultrasound and magnetic resonance imaging are used complementarily for pancreatic imaging in the majority of screening programs, but it is unclear whether both are necessary at every screening visit.  The optimal intervals for follow-up imaging need to be determined, although most experts recommend annual imaging, if the pancreas is normal at baseline screening.

What are the new findings?  Significant and potentially relevant pancreatic lesions detected by screening before the age of 50 years are extremely rare.  MRI based screening supplemented by EUS at baseline and every third year or when changes in MRI occur appeared to be efficient.  Screening intervals of 24 months may be justified in IAR with an unremarkable pancreas at baseline imaging.

Acknowledgements

Coordinators: • Alfredo Carrato • Carmen Guillén  Pancreas Comittee (HRC) • Nuria Malats Participants:  • Francisco Real European Familial Cancer Group • Julie Earl • Mirari Marquez Contacts: • Carme Guerrero • Reyes Ferreiro [email protected] • Vanessa Pachón • Mª Teresa Salazar [email protected] • Celia Calcedo [email protected] Collaborators: [email protected] • Isabel Chirivella González (H. Clínico de Valencia) • Iván Márquez (H. Gregorio Marañón) [email protected] • Eva Martínez de Castro (H. Marqués de Valdecilla) • José María Mesa (H. Príncipe de Asturias) • Carmelo Loinaz y Luis Robles Díaz (H. Doce de Octubre) Phone: +34 913 369 085 • Nuria Rodríguez Salas (H. La Paz) • José Santiago Crespo (H. Virgen de la Luz)