Genetic analysis of subsequent second primary malignant neoplasms in long-term pancreatic cancer survivors suggests new potential hereditary genetic alterations

Figures & data

Table 1 Baseline patient characteristics (entire cohort)

Parameters	Number of patients*	%
Sex
Male	75	64
Female	43	36
TNM stage
I	20	17
IIA	34	29
IIB	54	46
III	2	2
IV	8	7
Histologic grade
G1 + G2 (well to moderate)	62	52
G3 (poor)	51	44
Not available	5	4
Lymphovascular invasion
pL0	74	63
pL1	38	32
Not available	6	5
Perineural invasion
pP0	35	30
pP1	77	65
Not available	6	5
Angioinvasion
pA0	91	77
pA1	21	18
Not assessed	6	5
Adjuvant therapy
Yes	79	68
No	37	31
Unknown	2	2

Note:

* 118 patients in total.

Table 2 Clinical data of patients with SMN

Sex	Age	pT	pN	Grade	Perineural invasion	Angioinvasion	Lymphovascular invasion	Adjuvant treatment	Family history of PDAC	Family history of other cancers	DFS	SMN	TTS	Treatment of SMN	TTT	OS	Status
Male	68	3	0	3	Yes	No	No	GEM	No	No	64	Rectal cancer	60	Surgery	60	64	Died
Male	69	2	1	3	No	No	No	GEM	No	No	105	Urinary bladder cancer	17	Surgery	63	105	Alive
Male	67	3	1	3	No	No	No	GEM	Yes	No	14	Malignant melanoma	45	Surgery	45	104	Alive
Male	51	3	0	2	Yes	No	Yes	GEM	No	No	92	Prostate cancer	87	Hormonal therapy	87	92	Alive
Male	75	2	0	1	No	No	No	R/5FU	No	No	62	Rectal cancer	61	None	NA	62	Died
Female	70	3	0	2	No	No	Yes	GEM	No	No	73	Breast cancer	9	Surgery	9	73	Alive

Abbreviations: pT, pathologic tumor size; pN, pathologic lymph node metastasis; DFS, disease-free survival (months); NA, not applicable; SMN, subsequent secondary malignant neoplasm; TTS, time to diagnosis of SMN (months); TTT, time to therapy of SMN (months); OS, overall survival (months); GEM, gemcitabine (six cycles); R/5 FU, concomitant chemoradiotherapy with 5-fluorouracil; PDAC, pancreatic ductal adenocarcinoma.

Table 3 List of genes analyzed by targeted next-generation sequencing

Abbreviation	Gene name (alternative denominations)
AIP	Aryl hydrocarbon receptor interacting protein
ALK	Anaplastic lymphoma kinase
APC	Adenomatous polyposis coli
APEX1	APEX nuclease (multifunctional DNA repair enzyme) 1
ATM	Ataxia telangiectasia mutated
ATMIN	ATM interactor
ATR	Ataxia telangiectasia and Rad3 related
ATRIP	ATR interacting protein
AURKA	Aurora kinase A
AXIN1	Axin 1
BABAM1	BRISC and BRCA1 A complex member 1
BAP1	BRCA1-associated protein-1 (ubiquitin carboxy-terminal hydrolase)
BARD1	BRCA1-associated RING domain 1
BLM	Bloom syndrome, RecQ helicase-like
BMPR1A	Bone morphogenetic protein receptor, type IA
BRAP	BRCA1-associated protein
BRCA1	Breast cancer 1, early onset
BRCA2	Breast cancer 2, early onset
BRCC3	BRCA1/BRCA2-containing complex, subunit 3
BRE	Brain and reproductive organ-expressed (TNFRSF1A modulator)
BRIP1	BRCA1 interacting protein C-terminal helicase 1
BUB1B	Budding uninhibited by benzimidazoles 1 homolog beta (yeast)
C11orf30	Chromosome 11 open reading frame 30 (EMSY)
C19orf40	Chromosome 19 open reading frame 40 (FAAP24)
CASP8	Caspase 8, apoptosis-related cysteine peptidase
CCND1	Cyclin D1
CDC73	Cell division cycle 73, Paf1/RNA polymerase II complex component, homolog (Saccharomyces cerevisiae)
CDH1	Cadherin 1, type 1, E-cadherin (epithelial)
CDK4	Cyclin-dependent kinase 4
CDKN1B	Cyclin-dependent kinase inhibitor 1B (p27, Kip1)
CDKN1C	Cyclin-dependent kinase inhibitor 1C (p57, Kip2)
CDKN2A	Cyclin-dependent kinase inhibitor 2A
CEBPA	CCAAT/enhancer binding protein (C/EBP), alpha
CEP57	Centrosomal protein 57 kDa
CLSPN	Claspin
CSNK1D	Casein kinase 1, delta
CSNK1E	Casein kinase 1, epsilon
CWF19L2	CWF19-like 2, cell cycle control (Schizosaccharomyces pombe)
CYLD	Cylindromatosis (turban tumor syndrome)
DCLRE1C	DNA cross-link repair 1C
DDB2	Damage-specific DNA binding protein 2, 48 kDa
DHFR	Dihydrofolate reductase
DICER1	Dicer 1, ribonuclease type III
DMC1	DMC1 dosage suppressor of mck1 homolog, meiosis-specific homologous recombination (yeast)
DNAJC21	DnaJ (Hsp40) homolog, subfamily C, member 21
DPYD	Dihydropyrimidine dehydrogenase
EGFR	Epidermal growth factor receptor
EPCAM	Epithelial cell adhesion molecule
EPHX1	Epoxide hydrolase 1, microsomal (xenobiotic)
ERCC1	Excision repair cross-complementing rodent repair deficiency, complementation group 1
ERCC2	Excision repair cross-complementing rodent repair deficiency, complementation group 2
ERCC3	Excision repair cross-complementing rodent repair deficiency, complementation group 3
ERCC4	Excision repair cross-complementing rodent repair deficiency, complementation group 4
ERCC5	Excision repair cross-complementing rodent repair deficiency, complementation group 5
ERCC6	Excision repair cross-complementing rodent repair deficiency, complementation group 6
ESR1	Estrogen receptor 1
ESR2	Estrogen receptor 2 (ER beta)
EXO1	Exonuclease 1
EXT1	Exostosin 1
EXT2	Exostosin 2
EYA2	Eyes absent homolog 2 (Drosophila)
EZH2	Enhancer of zeste homolog 2 (Drosophila)
FAM175A	Family with sequence similarity 175, member A
FAM175B	Family with sequence similarity 175, member B
FAN1	FANCD2/FANCI-associated nuclease 1
FANCA	Fanconi anemia, complementation group A
FANCB	Fanconi anemia, complementation group B
FANCC	Fanconi anemia, complementation group C
FANCD2	Fanconi anemia, complementation group D2
FANCE	Fanconi anemia, complementation group E
FANCF	Fanconi anemia, complementation group F
FANCG	Fanconi anemia, complementation group G
FANCI	Fanconi anemia, complementation group I
FANCL	Fanconi anemia, complementation group L
FANCM	Fanconi anemia, complementation group M
FBXW7	F-box and WD repeat domain containing 7, E3 ubiquitin protein ligase
FH	Fumarate hydratase
FLCN	Folliculin
GADD45A	Growth arrest and DNA-damage-inducible, alpha
GATA2	GATA binding protein 2
GPC3	Glypican 3
GRB7	Growth factor receptor-bound protein 7
HELQ	Helicase, POLQ-like
HNF1A	HNF1 homeobox A
HOXB13	Homeobox B13
HRAS	v-Ha-ras Harvey rat sarcoma viral oncogene homolog
HUS1	HUS1 checkpoint homolog (S. pombe)
CHEK1	Checkpoint kinase 1
CHEK2	Checkpoint kinase 2
KAT5	K(lysine) acetyltransferase 5
KCNJ5	Potassium inwardly rectifying channel, subfamily J, member 5
KIT	V-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog
LIG1	Ligase I, DNA, ATP-dependent
LIG3	Ligase III, DNA, ATP-dependent
LIG4	Ligase IV, DNA, ATP-dependent
LMO1	LIM domain only 1 (rhombotin 1)
LRIG1	Leucine-rich repeats and immunoglobulin-like domains 1
MAX	MYC-associated factor X
MCPH1	Microcephalin 1
MDC1	Mediator of DNA-damage checkpoint 1
MDM2	Mdm2, p53 E3 ubiquitin protein ligase homolog (mouse)
MDM4	Mdm4 p53 binding protein homolog (mouse)
MEN1	Multiple endocrine neoplasia I
MET	Met proto-oncogene (hepatocyte growth factor receptor)
MGMT	O-6-methylguanine-DNA methyltransferase
MLH1	mutL homolog 1, colon cancer, nonpolyposis type 2 (Escherichia coli)
MLH3	mutL homolog 3 (E. coli)
MMP8	Matrix metallopeptidase 8 (neutrophil collagenase)
MPL	Myeloproliferative leukemia virus oncogene
MRE11A	MRE11 meiotic recombination 11 homolog A (S. cerevisiae)
MSH2	mutS homolog 2, colon cancer, nonpolyposis type 1 (E. coli)
MSH3	mutS homolog 3 (E. coli)
MSH5	mutS homolog 5 (E. coli)
MSH6	mutS homolog 6 (E. coli)
MSR1	Macrophage scavenger receptor 1
MUS81	MUS81 endonuclease homolog (S. cerevisiae)
MUTYH	mutY homolog (E. coli)
NAT1	N-acetyltransferase 1 (arylamine N-acetyltransferase)
NBN	Nibrin
NCAM1	Neural cell adhesion molecule 1
NELFB	Cofactor of BRCA1
NF1	Neurofibromin 1
NF2	Neurofibromin 2 (merlin)
NFKBIZ	Nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, zeta
NHEJ1	Nonhomologous end-joining factor 1
NSD1	Nuclear receptor binding SET domain protein 1
OGG1	8-oxoguanine DNA glycosylase
PALB2	Partner and localizer of BRCA2
PARP1	Poly (ADP-ribose) polymerase 1
PCNA	Proliferating cell nuclear antigen
PHB	Prohibitin
PHOX2B	Paired-like homeobox 2b
PIK3CG	Phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit gamma
PLA2G2A	Phospholipase A2, group IIA (platelets, synovial fluid)
PMS1	PMS1 postmeiotic segregation increased 1 (S. cerevisiae)
POLB	Polymerase (DNA directed), beta
POLD1	Polymerase (DNA directed), delta 1, catalytic subunit
POLE	Polymerase (DNA directed), epsilon, catalytic subunit
PPM1D	Protein phosphatase, Mg2+/Mn2+ dependent, 1D
PREX2	Phosphatidylinositol-3,4,5-trisphosphate-dependent Rac exchange factor 2
PRF1	Perforin 1 (pore forming protein)
PRKAR1A	Protein kinase, cAMP-dependent, regulatory, type I, alpha
PRKDC	Protein kinase, DNA-activated, catalytic polypeptide
PTEN	Phosphatase and tensin homolog
PTCH1	Patched 1
PTTG2	Pituitary tumor-transforming 2
RAD1	RAD1 homolog (S. pombe)
RAD17	RAD17 homolog (S. pombe)
RAD18	RAD18 homolog (S. cerevisiae)
RAD23B	RAD23 homolog B (S. cerevisiae)
RAD50	RAD50 homolog (S. cerevisiae)
RAD51	RAD51 homolog (S. cerevisiae)
RAD51AP1	RAD51 associated protein 1
RAD51B	RAD51 homolog B (S. cerevisiae)
RAD51C	RAD51 homolog C (S. cerevisiae)
RAD51D	RAD51 homolog D (S. cerevisiae)
RAD52	RAD52 homolog (S. cerevisiae)
RAD54B	RAD54 homolog B (S. cerevisiae)
RAD54L	RAD54-like (S. cerevisiae)
RAD9A	RAD9 homolog A (S. pombe)
RB1	Retinoblastoma 1
RBBP8	Retinoblastoma binding protein 8
RECQL	RecQ protein-like (DNA helicase Q1-like)
RECQL4	RecQ protein-like 4
RECQL5	RecQ protein-like 5
RET	Ret proto-oncogene
RFC1	Replication factor C (activator 1) 1, 145 kDa
RFC2	Replication factor C (activator 1) 2, 40 kDa
RFC4	Replication factor C (activator 1) 4, 37 kDa
RHBDF2	Rhomboid 5 homolog 2 (Drosophila)
RNF146	Ring finger protein 146
RNF168	Ring finger protein 168, E3 ubiquitin protein ligase
RNF8	Ring finger protein 8, E3 ubiquitin protein ligase
RPA1	Replication protein A1, 70 kDa
RUNX1	Runt-related transcription factor 1
SDHAF2	Succinate dehydrogenase complex assembly factor 2
SDHB	Succinate dehydrogenase complex, subunit B, iron sulfur (Ip)
SETBP1	SET binding protein 1
SETX	Senataxin
SHPRH	SNF2 histone linker PHD RING helicase, E3 ubiquitin protein ligase
SLX4	SLX4 structure-specific endonuclease subunit homolog (S. cerevisiae)
SMAD4	SMAD family member 4
SMARCA4	SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a, member 4
SMARCB1	SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily b, member 1
SMARCE1	SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily e, member 1
STK11	Serine/threonine kinase 11
SUFU	Suppressor of fused homolog (Drosophila)
TCL1A	T-cell leukemia/lymphoma 1A
TELO2	TEL2, telomere maintenance 2, homolog (S. cerevisiae)
TERF2	Telomeric repeat binding factor 2
TERT	Telomerase reverse transcriptase
TLR2	Toll-like receptor 2
TLR4	Toll-like receptor 4
TMEM127	Transmembrane protein 127
TOPBP1	Topoisomerase (DNA) II binding protein 1
TP53	Tumor protein p53
TP53BP1	Tumor protein p53 binding protein 1
TSC1	Tuberous sclerosis 1
TSC2	Tuberous sclerosis 2
TSHR	Thyroid stimulating hormone receptor
UBE2A	Ubiquitin-conjugating enzyme E2A
UBE2B	Ubiquitin-conjugating enzyme E2B
UBE2I	Ubiquitin-conjugating enzyme E2I
UBE2V2	Ubiquitin-conjugating enzyme E2 variant 2
UBE4B	Ubiquitination factor E4B
UIMC1	Ubiquitin interaction motif containing 1
VHL	Von Hippel–Lindau tumor suppressor, E3 ubiquitin protein ligase
WRN	Werner syndrome, RecQ helicase-like
WT1	Wilms tumor 1
XPA	Xeroderma pigmentosum, complementation group A
XPC	Xeroderma pigmentosum, complementation group C
XRCC1	X-ray repair complementing defective repair in Chinese hamster cells 1
XRCC2	X-ray repair complementing defective repair in Chinese hamster cells 2
XRCC3	X-ray repair complementing defective repair in Chinese hamster cells 3
XRCC4	X-ray repair complementing defective repair in Chinese hamster cells 4
XRCC5	X-ray repair complementing defective repair in Chinese hamster cells 5
XRCC6	X-ray repair complementing defective repair in Chinese hamster cells 6
ZNF350	Zinc finger protein 350
ZNF365	Zinc finger protein 365

Table 4 Table of identified variants classified as likely pathogenic/pathogenic according to the ClinVar database

Patient	Gene	Nucleotide	Protein	ClinVar classification	Sex/age primary	Personal history (age at diagnosis)	Family history
With SMN
OL0138	CHEK2	c.349A>G	p.Arg117Gly	Class 4–5	Female/70	Breast (71)	0
Without SMN
OL0130	RAD51D	c.345+2T>G	–	Class 4	Male/62	0	Mother – gastric
OL0132	MLH1	c.390C>G	p.Tyr130Ter	Class 5	Female/52	0	Father – colon, father’s mother – brain
	ATM	c.3849delA	p.Leu1283fs	Class 5
PCI77	CHEK2	c.1100delC	p.Thr367fs	Class 5	Male/55	0	0

Note: All variants are heterozygous.

Abbreviation: SMN, subsequent malignant neoplasm after pancreatic ductal adenocarcinoma (PDAC).

Table 5 List of identified variants of unknown significance

Patient	Gene	Nucleotide	Protein	rs number	EXaC MAF	ClinVar/VarSome classification	SIFT	PP2	MA	Damag. acc. to ≥2 software
With SMN
OL0134	BLM	c.11T>C	p.Val4Ala	rs144706057	0.0017	1–3/3	0	0.132	2.14	Y
OL0135	PTCH1	c.2597G>A	p.Gly866Glu	NA	NA	3/3	0.08	0.999	2.31	Y
	ATM	c.3208G>A	p.Val1070Ile	NA	NA	3/3	0.35	0.026	2.135	N
OL0136	PLA2G2A	c.185G>A	p.Arg62His	NA	8.34E-05	NA/3	0.02	0.888	3.005	Y
	LRIG1	c.2195C>T	p.Pro732Leu	rs61746346	0.0022	NA/3	0	0.991	1.975	Y
	RECQL5	c.1801G>A	p.Val601Met	NA	NA	NA/3	0.3	0.04	1.905	N
OL0138	PREX2	c.C1672G	p.Pro558Ala	rs199541834	0.0001	NA/3	0.15	0.145	0.46	N
	PARP1	c.C659T	p.Ala220Val	rs139232092	0.0006	NA/3	0.15	0.003	1.155	N
Without SMN
OL0041	BUB1B	c.1042G>A	p.Ala348Thr	NA	8.24E-06	NA/3	0.33	0.85	2.175	N
	MRE11A	c.C1475A	p.Ala492Asp	rs61749249	0.0034	1–3/3	0.43	0.754	1.735	N
OL0130	XRCC1	c.632A>G	p.Tyr211Cys	NA	1.74E-05	NA/3	0.15	0.998	2.175	Y
OL0131	0
OL0132	GRB7	c.1439T>C	p.Val480Ala	rs143372931	0.0004	NA/3	0	0.848	3.07	Y
	RAD9A	c.215G>A	p.Arg72His	rs377299831	1.65E-05	NA/3	0.58	0.019	1.2	N
OL0133	EXT2	c.1859C>T	p.Thr620Met	rs138495222	0.0006	2–3/3	0.02	0.999	2.24	Y
	MLH3a	c.3281-1G>C	–	NA	NA	NA/3	–	–	–	–
OL0137	PREX2	c.2167A>G	p.Asn723Asp	NA	1.65E-05	NA/3	0.03	0.614	1.63	N
	HELQ	c.1418G>A	p.Arg473His	NA	2.48E-05	NA/3	0	1	4.545	Y
	RFC4	c.908C>T	p.Ala303Val	rs144238574	9.07E-05	NA/3	0.44	0.027	1.235	N
OL0139	RHBDF2	c.940G>A	p.Ala314Thr	rs140433374	0.0008	NA/3	0.33	0.952	1.78	N
	MDM4	c.1162C>G	p.Pro388Ala	rs61754765	0.0006	NA/3	0.92	0.997	1.1	N
OL0140	FANCM	c.3407T>C	p.Leu1136Ser	NA	1.65E-05	NA/3	0.01	0.963	1.905	Y
	POLE	c.1601T>C	p.Leu534Pro	NA	NA	NA/3	0	0.991	3.565	Y
OL0141	0
OL0142	RAD54L	c.1817G>A	p.Arg606Gln	rs374574941	2.47E-05	NA/3	0	1	4.735	Y
	POLD1	c.2116C>G	p.Pro706Ala	NA	NA	3/3	0.01	0.733	2.41	Y
OL0144	CWF19L2	c.2240A>C	p.Lys747Ther	NA	NA	NA/3	0.08	0.697	1.915	N
	SETX	c.967A>G	p.Ser323Gly	NA	1.65E-05	NA/3	0	0.994	0.975	Y
OL0157	TP53BP1	c.2226A>T	p.Glu742Asp	rs150423877	0.0004	NA/3	0.48	0.987	0.46	N
PCI77	0
PCI15	PTCH1	c.3376G>A	p.Val1126Ile	rs147025073	0.0005	3/3	0.26	0.927	1.77	N
	NCAM1	c.1481C>A	p.Thr494Asn	NA	NA	NA/3	0.01	0.347	NA	N
PCI39	0
PCO11	BRCA1	c.3929C>A	p.Thr1310Lys	rs80357257	8.24E-06	1–3/3	0.01	0.787	1.895	N
	AURKA	c.1028G>A	p.Arg343Gln	rs200181472	0.0002	NA/3	0.04	0.027	0.71	N
	EXO1	c.820G>A	p.Gly274Arg	rs149397534	0.0021	NA/3	0.16	0.999	1.295	N

Notes: The variants predicted to be damaging by at least two out of three prediction tools employed are represented in bold.

a The splice-site variant was analyzed by splicing prediction software spidex with a score −25.6359, suggesting that it is the damaging variant.

Abbreviation: NA, not applicable.