Epigenetics and pancreatic cancer: Pathophysiology and novel treatment aspects

Table 1 Histomorphologic subtypes of pancreatic cancer related to their genetic/epigenetics and possible prognostic characteristics

Variant (source)	Morphology	Genetics/epigenetics	Prognosis
MC	Well defined pushing border, syncytial growth pattern, and poorly differentiated cancer cells	Germline or somatic mutations as well as epigenetic silencing by promoter methylation of mismatch repair genes MLH1 and MSH2	BetterMC: Overall 2- and 5-yr survival rate of 29% and 13%[23]CC: 2-yr and 5-yr survival rate of 70% and 57%[153]
CC	Suspension of well-differentiated cancer cells in extracellular mucin pools (at least 80%)	Unknown
AC	Combination of glandular and squamous (at least 30%) components	K-RAS2 mutations, inactivation of CDKN2A/p16, SMAD4/DPC4 and TP53	WorseAC/UC: Median survival of 5 mo after resection[154,155]
UC	Noncohesive cancer, lacking histological features of differentiation	K-RAS2 gene mutations, loss of E-cadherin protein expression (promoter hypermethylation)
		Expression of L1CAM, COX2, and EGFR
		Subtype with osteoclast-like giant cells shows mutations like noninvasive precursor lesions

MC: Medullary carcinoma; CC: Colloid carcinoma; AC: Adenosquamous carcinoma; UC: Undifferentiated carcinoma; COX: Cyclooxygenase; L1CAM: L1 cell adhesion molecule; EGFR: Epidermal growth factor receptor; MSH2: MutS homolog 2.

Table 2 Activated and deactivated key genes in pancreatic cancer according to hallmark of cancer, frequency as well as to kind of possible genetic and epigenetic alteration

Gene symbol (source)	Associated function according the hallmarks of cancer1[35,36]	Frequency	Type of genetic alteration	Evidence for epigenetic regulation (reference)
Activation
K-RAS2[131]	a, f, g	> 90	Point mutation
AKT2[162,163]	a, j, g	10-20	Amplification
BRAF[164]	a, b, g	5	Point mutation
Deactivation
CDKN2A/p16[165]	b, d, i	95	Homozygous deletion, intragenic mutation	Yes[166]
TP53[167-169]	b, d, i, h	50-70	Intragenic mutation an one allele and loss in the other allele
SMAD4/DPC4[170,171]	b, c, f	55	Homozygous deletion, intragenic mutation
MLH1[23,172]	h	3-15	Heterozygote mutations	Yes[34]
BRCA2[173]	a	7	Heterozygote mutations
STK11/LKB1[174]	i	5	Homozygous deletion, intragenic mutation
TGFBR2[175]	a, f	4	Homozygous deletion, homozygous frameshift mutation
MAP2K4[176,177]	a	2	Homozygous deletion, missense mutation

¹Hallmarks of cancer: Sustaining proliferative signaling (a), evading growth suppressors (b), avoiding immune destruction (c), enabling replicative immortality (d), tumour promoting inflammation (e), activating invasion and metastasis (f), inducing angiogenesis (g), genome instability and mutations (h), resisting cell death (i), deregulating cellular energetics (j)[35,36].

Table 3 Overview of DNA hyper-/hypomethylation involved in pancreatic cancer

DNA modification	Material			Gene affected	Ref.
	Cancer samples	Cell lines	Other
DNA hyper-methylation	√			p16	[166]
	√			RASSF1A	[178]
	√			MDFI, hsa-miR-9-1, ZNF415, CNTNAP2, ELOVL4	[179]
	√			SOX15	[180]
	√			HOP hoemobox (HOPX)	[181]
	√			KLF10	[182]
	√			hMLH1	[183]
	√			miR-34a/b/c	[184]
	√			SPARC	[185]
	√			FoxE1, NPTX2, CLDN5, P16, TFPI-2, SPARC, ppENK	[186]
	√			SFRP	[187]
	√	AsPC1, Hs766T, MiaPaCa2, Panc1		UCHL1, NPTX2, SARP2, CLDN5, reprimo, LHX1, WNT7A, FOXE1, TJP2, CDH3, ST14	[30,159]
	√	AsPC1, BxPC3, CFPAC1, Panc1		NPTX2	[188]
	√	Panc1, SW1990		miR-132	[121]
	√	BxPC3, Capan2, CFPAC1, HPACI, HPAFII, MiaPaCa2, Panc1, PL45		FOXA1/2	[81]
		MiaPaCa2		ARID1B	[189]1
		Panc1		NPTX2	[190]1
		AsPC1, BxPC3, Panc1, MIA PaCa-2		Dkk3	[191]
		BXPC3, HPAFII, HPAC, hTERT-HPDE, Panc1		Cldn18	[192]
		BxPC3, CFPAC1, Panc1, SW1990		TNFRSF10C	[193]
			Pancreatic juice	Neuronal pentraxin II (NPTX2)	[194]
			Pancreatobiliary fluid	UCHL1, RUNX3	[195]
			PanIN	p16	[196]
			IPMNs	BNIP3, PTCHD2, SOX17, NXPH1, EBF3, SPARC, SARP2, TSLC1, RELN, TFPI2, CLDN5, UCHL1	[157,197]
			Blood, brush cytology	NPTX2	[198,199]
DNA hypo-methylation	√			VAV1	[79]
	√			Claudin4, lipocalin2, 14-3-3 sigma, trefoil factor 2, S100A4, mesothelin, prostate stem cell antigen	[78]
	√			MUC4	[77]
		SW1990		ABCB1/MDR1, ABCC1/MRP1, ABCG2/BCRP	[200]

¹Only abstract available.

Table 4 Overview of miRNAs associated with specific targets/functions in pancreatic cancer

miRNA/function		Cell lines	Target gene(s)	Cellular effects	Ref.
Function as oncogene	-10a	AsPC1, Capan1, Capan2, MiaPaCa2, Panc1, Patu8988T, Patu8988S, Patu8902	HOXB1, 3	Metastasis ↑	[201]
	-21	AsPC1, BxPC3, Capan1, Capan2, CFPAC1, Hs776T, H48N, KP-1N, KP-2, KP-3, MiaPaCa2, NOR-P1, Panc1, SUIT-2, SW1990	HOXA1	Invasion ↑	[202]
		AsPC1, Capan1, Capan2, CFPAC1, H48N, HS766T, KP-1N, KP-2, KP-3, MiaPaCa2, NOR-P1, Panc1, SUIT-2, SW1990		Proliferation ↑, invasion ↑, chemoresistance ↑	[203]
		BxPC3		Proliferation ↑	[204]
		Capan1, HS766T, MiaPaCa2, MPanc96, Panc1, PL45, SW1990	PTEN, RECK	After miRNA inhibition: cell cycle arrest ↑, apoptosis ↑	[205]
	-132, -212	Panc1	Rb1	Proliferation ↑	[206]
	-155	Capan2, MiaPaCa2, MCF7, MEFs, 293T	TP53INP1	Apoptosis ↓	[207]
	-194, -200b, -200c, -429	AsPC1, A818, BxPC3, Capan1, Capan2, HPAFII, MiaPaCa2, MPanc96, Panc1, Patu8902, Patu8988T, Patu8988S, PT45, Suit 007, Su.86.86, Sut00281	EP300	Metastasis ↑	[208]
	-197	AsPC1, Panc1	p120 catenin	EMT ↑	[209]
	-210	Panc1, MiaPaCa2, SUIT-2		Migration ↓, vimentin ↓, snai-1 ↓, membraneous β-catenin ↑	[210]
	-221	Capan1, HS766T, MiaPaCa2, MPanc96, Panc1, PL45, SW1990	p27	Chemosensitivity ↑	[205]
	-224, -486	AsPC1, A818, BxPC3, Capan1, Capan2, HPAFII, Su 86.86, MPanc96, MiaPaCa2, Panc1, Patu8902, Patu8988T, PT45, Patu8988S, Suit 007, Suit 00281	CD40	Invasion ↑, metastasis ↑	[211]
Function as tumor suppressor	-301a	BxPC3, Hs766T	Bim	Proliferation ↑,	[212]
	-320c	AsPC1, Panc12	SMARCC1	Chemoresistance ↑	[213]
	-421	SW1990, Panc1	DPC4/Smad4	Proliferation ↑, colony formation ↑	[214]
	-491-5p	AsPC1, Capan1, MiaPaCa2, SW1990	Bcl-XL, TP53	Proliferation ↓, apoptosis ↑, STAT3 ↓, PI-3K/Akt ↓	[215]
	let-7let-7a	BxPC3, Capan1, Capan2, human HPNE (human pancreatic nestin-positive) cells MiaPaCa2, Panc1		Proliferation ↓, K-RAS ↓, MAPK ↓	[216]
		AsPC1	RAS	K-RAS ↓, radiosensitivity ↑	[217]
	-22	BxPC3	SP1, ESR1	Tumourigenesis ↓	[218]
	-26a	SW1990, Panc1	HMGA1	Proliferation ↓, invasion ↓, migration ↓, apoptosis ↑	[219]
	-34	BxPC3, MiaPaCa2	Bcl-2, Notch-1/2	Clonogenicity ↓, invasion ↓, apoptosis ↑, cell cycle arrest ↑, chemosensitivity ↑, radiosensitivity ↑, CSC ↓	[220]
	-34a	Panc1		Cell cycle arrest ↑, apoptosis ↑, migration ↓, E2F3 ↓, Bcl-2 ↓, c-myc ↓, cyclin D1 ↓	[221]
	-34b	AsPC1, MiaPaCa2	Notch-1	Proliferation, apoptosis	[222]
			Smad3	Progression in vivo↑	[223]3
	-107	MiaPaCa2, Panc1	CDK6	Proliferation ↓	[224]
	-126	AsPC1, BxPC3, KLM-1, MiaPaCa2, Panc1	ADAM9	Migration ↓, invasion ↓, E-cadherin ↑	[225]
	-132	BxPC3, HPAFII, HPAC, Panc1		Proliferation ↓, colony formation ↓, Akt ↓	[121]
	-143	AsPC1, BxPC3, Capan2, HPAFII, MiaPaCa2, Panc1	COX-2	Proliferation ↓, MEK/MAPK ↓	[226]
		Panc1	ARHGEF1 (GEF1), ARHGEF2 (GEF2), K-RAS	Migration ↓, invasion ↓, metastasis ↓, E-cadherin ↑	[227]
	-148a	IMIM-PC2	CDC25B	Proliferation ↓, colony formation ↓	[228]
	-148b	AsPC1, BxPC3, MiaPaCa2, Panc1, SW1990	AMPKα1	Proliferation ↓, apoptosis ↑, cell cycle arrest ↑, invasion ↓, chemosensitivity ↑, tumourigenicity ↓	[229]
	-150	Colo357, HPAF, Panc10.05	MUC4	Proliferation ↓, clonogenicity ↓, migration ↓, invasion ↓, cellular adhesion ↑	[230]
	-200	AsPC1, BxPC3, Colo357, HPAC, MiaPaCa2, L3.6pl, Panc12		EMT ↓ (ZEB1 ↓, slug ↓, vimentin ↓)	[231]
	-375			Proliferation ↓, cell cycle arrest ↑, apoptosis ↑	[232]3
	-548d	Panc1		Proliferation ↓, apoptosis ↑, cell cycle arrest ↑	[233]

¹Including an orthotopic murine model;

²Gemcitabine-resistant cell line;

³Only abstract available. Based on and updated from Park et al[120] 2011.

Table 5 Overview of Epigenetic mechanisms - see text for details and references

Mechanism	Enzyme	Subclasses/components	Effect on target gene expression
DNA (de-) methylation	DNMT	DNMT1 (methylation maintenance)	↓
		DNMT3A and -3B (de novo methylation)
	DNA de-methylase	Not known	↑
Histone (de-) acetylation	HAT	e.g., CBP, p300	↑
Histone methylation	HDAC	Class I (HDACs-1-3, -8), class IIa (HDACs-4, -5, -7, -9), class IIb (HDACs-6, -10), class III (SIRT1-7), class IV (HDAC-11)	↑
	PcG→ H3-K27-me3	PRC1: CBX-2, 4, or 9, PHC-1, 2, or 3, BMI1, RING1A/B or RNF2	↓
		→ H3-K27-me3 maintenance
		PRC2: EZH2, SUZ12, EED
		→de novo H3-K27-me3 maintenance
	TrxG → H3-K4-me3	Several members	↑
Post-transcriptional	miRNAs	2578 mature miRNA (miRBase v20)	↓

HAT: Histone acetylase; HDAC: Histone deactylase; DNMT: DNA methyltransferase.

Table 6 Trials using epigenetic agents in pancreatic cancer

Compound	Combination	Phase	Endpoint	ClincialTrials.gov	Treatment
DNMT inhibitors
Azacitidine	+ Gemcitabine	I	MTD	NCT01167816
Azacitidine		II	PFS	NCT01845805	2
Decitabine		Various stages of development for solid tumors			3
Zebularine		Preclinical
HDAC inhibitors
Vorinostat	+ Marizomib	I	MTD	NCT00667082[234]
Vorinostat	+ Radiation	I/II	MTD, PFS	NCT00948688
	+ 5-FU
Vorinostat	+ Radiation	I	MTD	NCT00983268
	+ Capecitabine
Vorinostat	+ Radiation	I/II	MTD	NCT00831493
Belinostat		Various stages of development for solid tumors
Entinostat		I	MTD	NCT00020579[235]
Entinostat	13-cis retinoic acid	I	MTD
Panobinostat	+ Bortezomib	II	PFS	NCT01056601[236]	1
HAT inhibitors
Curcumin		II	Survival	NCT00094445[147]	1
Curcumin	+ Gemcitabine	II	TTP	NCT00192842	1
Curcumin	+ Gemcitabine	III		NCT00486460	1
	+ Celecoxib
Curcumin		I	MTD	-[237]
Curcumin	+ Gemcitabine	I	MTD	-[238]
Curcumin	+ Gemcitabine	I/II	MTD	-[239]

1: Palliative; 2: Postoperative adjuvant; 3: Both. MTD: Maximum tolerated dose; PFS: Progression-free survival; HDAC: Histone deacetylase; HAT: Histone acetyltransferase.