Gene therapy in pancreatic cancer

Table 1 Comparison of characters of common viral vectors

Virus	Viral genome	Insertion capacity	Host range of infection	State of integration into host genome	Efficiency of gene transfection and expression	Immunogenicity	Titers of preparation in vitro(PFU/mL)	Bio-safety
AdV	Double-stranded DNA	38 kb	Broad spectrum (both dividing and non-dividing cells)	No integration	High	High	1011-1012	Safe
AAV	Single-stranded DNA	4.9 kb	Broad spectrum	Site-specific integration on chromosome 19q13.3	High	Low	1012, dependent on helper virus	Safe
RV	Single-stranded RNA	8 kb	Dividing cells only	Integrate randomly	Low	Low	106-107	Risk of insertional mutagenesis
Lentivirus	Single-stranded RNA	8 kb	Broad spectrum	Integrate randomly	High	Low	109-1010	Risk of viral infection and insertional mutagenesis
Pox virus	Souble-stranded DNA	25 kb	Broad spectrum	No integration	High	High, function as immunologic adjuvant	106-107	Safe
HSV	Double-stranded DNA	15-30 kb	Nerve cells and epithelial cells, especially neuro- tropic speciality	No integration	High	Moderate	1011-1012	Risk of viral infection

Table 2 Target genes of gene therapy in pancreatic cancer

Strategy	Categories	Examples
Gene augumentation	Tumor suppressor genes	p16INK4a, p21CIP1/WAF1, p14ARF, Retinoblastoma Protein (pRb), p53, p84 /Thoc1, p73, Smad4/DPC4
	Drug sensitivity genes/ suicide genes	Herpes Simplex Virus Thymidine Kinase (HSV-TK), Cytosine Deaminase (CD), Nitroreductase (NTR), Cytochrome P450 (CYP)
	Anti-angiogenesis genes	Soluble VEGFR, Soluble FGFR, Endostatin, Thrombospondin-1, Angiostatin, Vasostatin, NK4, Matrix metalloproteinases inhibitors (MMIPs/TIMPs), Somatostatin receptors (SSTR)
	Immune related genes	MHC molecules, Co-stimulatory molecules (B7 family, ICAM-1, LFA-3), Inflammatory cytokines (IL-2, IL-12, GM-CSF, IFN-α, IFN-β, IFN-γ, TNF-α), Tumor antigen (CEA, MUC-1, etc.)
	Apoptosis related genes	TRAIL
Gene blockade/antisense therapy	Oncogenes	K-ras, LSM1/CaSm, HER-2/EerB-2
	MDR	MDR1, MRP family, BCRP
	Proliferation related genes	VEGF, hTERT, COX-2

MDR: Multidrug resistance.

Table 3 Active and negative regulatory factors of angiogenesis

Stimulators	Inhibitors
Hypoxia	Angiostatin
Oncogenic proteins such as Ras	TSP-1
Inflammatory cytokines such as IL-8 and IL-6	Endostatin
FGF	Arrestin
TGF-β	Canstatin
HGF	MMPIs
PDGF	Somatostatin
G-CSF	Tumstatin
Angiogenin	VEGI
Leptin	Decoy receptors such as soluble VEGFR
Proliferin	Inflammatory cytokines such as IL-12

G-CSF: Granulocyte colony-stimulating factor; PDGF: Platelet-derived growth factor; HGF: Hepatocyte growth factor; TGF-β: Transforming growth factor-β; FGF: Fibroblast growth factor; TSP-1: Thrombospondin-1; MMPIs: Matrix metalloproteinases inhibitors; VEGI: Vascular endothelial growth inhibitor.

Table 4 Antigenic sources used in pancreatic cancer vaccine trials

Peptide or protein vaccines	Mutant K-ras peptide
	WT1 peptide
	CAP1-6D
	G17DT
	HLA-A*0201 restricted VEGF receptor-1 and -2 peptides
	HLA-A24 restricted survivin-2B80-88 peptide (AYACNTSTL)
	Autologous heat shock protein HSPPC-96
	TELOVAC
Whole cell vaccines or dendritic cell vaccines	α-Gal transferase transfected allogeneic tumor cells (including Algenpantucel-L)
	GVAX
	MUC-1 pulsed autologous dendritic cells
Viral or bacterial vaccines	PANVAC-VF-MUC-1, CEA, and TRICOM transfected virus
	Mesothelin (CRS-207) transfected live-attenuated Listeria
DNA vaccines	VXMO1

VXMO1: VEGF receptor-2 DNA vaccine; GVAX: GM-CSF transfected allogeneic tumor cells; TELOVAC: Telomerase peptide GV1001; CAP1-6D: CEA peptide; WT1: Wilms tumor 1; G17DT: Gastrin peptide.

Table 5 Closed clinical trials of gene therapy in pancreatic cancer

Trial ID and investigator	Phase	Clinical indication	Patients(n)	Transgenes	Vectors and target cells	Administration route	Combined therapy	Results
DE-0009 Löhr et al[67]	I/II	Inoperable PC	14	Cytochrome p450	Naked/plasmid DNA transfected allogeneic human 293 embryonic kidney cells	Inject into the tumor vasculature via supraselective angiography	In combination with low-dose ifosfamide	4 patients showed tumor regression, the other ten individuals remained stable. Median survival was doubled compared with historic controls. 1-yr survival rate was three times better
DE-0024 Pecher et al[207]	I/II	Advanced breast cancer, PC and gallbladder carcinoma	10 (2 PC)	MUC-1	Naked/plasmid DNA transfected autologus dendritic cells	Subcutaneous injection	None	9 patients showed signs of progression. Only one remained stable for 3 mo until she was transferred to another therapy. 3 of 10 patients developed vaccine-specific delayed-type hypersensitivity reaction (DTH). 4 of 10 patients showed increased mucinspecific INF-gamma- secreting CD8+ T cells
DE-0063 Kubuschok et al[208]	I	PC	3 healthy donors and 1 PC patient	Mutated ras oncoprotein	EB virus transformed autologous lymphoblastoid cells	Subcutaneous injection	None	All the subjects showed strong vaccine-induced muRas-specific cytotoxic T lymphocytes
DE-0083 Niethammer et al[209]	I	Advanced PC	45	VEGFR-2	Naked/plasmid DNA (oral DNA vaccine)	Oral administration	In combination with gemcitabine	Not reported
ES-0004 Mazzolini et al[210]	I	Liver cancer, PC, colorectal cancer	17 (3 PC)	Interleukin-12 (IL-12)	Adenovirus transfected autologous dendritic cells	Intratumoral injection	None	Treatment was well tolerated. 11 of 17 were assessable for response. A partial response was observed in 1 case with PC. Stable disease was observed in 2 patients and progression in 8 patients
FR-0018 Gilly et al[211]	I/II	Unresectable digestive cancer	6 (3 PC)	Interleukin-2 (IL-2)	Adenovirus	Intratumoral injection	None	Good safety. But final results of tumor responses were not reported
US-0853 Le et al[212]	I	Ovarian cancer, PC, lung cancer, mesothelioma	28	Mesothelin	Listeria monocytogenes	Intravenous injection	1: Live attenuated Listeria vaccine (n = 9); 2: live attenuated mesothein expressing Listeria vaccine ( n = 17)	In arm 2, Listeriolysin O and mesothelin-specific T-cell responses were seen and 37% of subjects lived ≥ 15 mo
US-0700 Galanis et al[213]	I	Gemcitabine-refractory, metastatic PC	12	Cyclin G1	Retrovirus	Intravenous injection	None	Good safety. But there was no evidence of anti-tumor activity
Kaufman et al[214]	I	Advanced PC	10	CEA, MUC-1, and TRICOM (including B7.1, ICAM-1, LFA-3)	Poxvirus	Subcutaneous injection	In combination with GM-CSF	Antigen-specific T-cell responses in 5 of 8 evaluated patients. 15.1 mo of median survival in responders vs 3.9 mo in non-responders. Overall median survival is 6.3 mo
Jaffee et al[141]	I	Resected PC	14	GM-CSF	Naked/plasmid DNA transfected allogeneic tumor cell vaccine	Intradermal injection	In combination with standard chemoradiation	3 of 14 patients developed DTH, and 3 patients had increased disease-free survival time (at least 25 mo after diagnosis)
US-0475 Lutz et al[215]	II	Resected PC	60	GM-CSF	Naked/plasmid DNA transfected allogeneic tumor cell vaccine	Intradermal injection	In combination with standard chemoradiation	The median disease-free survival is 17.3 mo with median survival of 24.8 mo. Induction of CD8+ mesothelin-specific T cells correlated with disease-free survival
Laheru et al[142]	A pilot study	Advanced PC	50	GM-CSF	Naked/plasmid DNA transfected allogeneic tumor cell vaccine	Intradermal injection	A: vaccine alone (n = 20) B: vaccine plus cyclophosphamide (n = 30)	Cohort B showed enhanced mesothelin-specific T-cell responses. Median survival values in cohort A and cohort B were 2.3 and 4.3 mo
Le et al[143]	Ib	Advanced PC	30	CM-CSF	Naked/plasmid DNA transfected allogeneic tumor cell vaccine	Intradermal injection	1: Ipilimumab alone (n = 15); 2: Ipilimumab plus vaccine (n = 15)	Median overall survival was 3.6 mo for arm 1 and 5.7 mo for arm 2. Mesothelin-specific T cells responses were associated with increased disease-free survival in arm 2

Table 6 Ongoing clinical trials of gene therapy in pancreatic cancer

Phase	Transgenes
I	Oncolytic adenovirus and oncolytic herpesvirus1
	Somatostatin receptor 2 (sst2), Deoxycitidine kinase :: uridylmonophosphate kinase (dck::umk)
	Somatostatin receptor 2 (sst2)
	Mesothelin-scFv with signaling domains comprised of TCR , CD28, and 4-1BB (CD137) cDNA
	GM-CSF
	PANVAC (CEA, MUC-1, and TRICOM)
	Cytosine deaminase, Herpes simplex virus thymidine kinase (HSV-TK)
	Cyclin G1
	p53
	BikDD (DOTAP-cholesterol mediated gene transfection)
	Mutated Ras
I/II	Cytochrome p450
	AEG 35156: antisense oligonucleotide to X-linked inhibitor of apoptosis protein (XIAP)
	Herpes simplex virus thymidine kinase (HSV-TK)
	Diphtheria Toxin A Chain (DTA) gene with H19 promoter
	Alpha-(1,3) galactosyltransferase
II	CEA, MUC-1, and PANVAC
	Mutated Ras
	Alpha-(1,3) galactosyltransferase
III	Alpha-(1,3) galactosyltransferase
	PANVAC (CEA, MUC-1, and TRICOM)2

¹Clinical trials of oncolytic viruses were completed or are ongoing in Spain and Japan, and their details were not clear;

²This clinical trial has closed, but details and results have not been published yet.

Table 7 Clinical trials of unspecific tumor gene therapy that may involve pancreatic cancer

Phase	Clinical indication	Transgenes
II	Adenocarcinoma	CEA, TCRzeta and CD28
I	Cachexia	GHRH
I	CEA- or HER-2-expressing malignancies	HER-2, CEA
I	CEA-expressing malignancies	CEA
I/II	CEA-expressing malignancies	CAP-1 peptide from CEA
I/II	CEA-expressing malignancies	T cell receptor alpha and beta chains cDNA
I	CEA-expressing malignancies	CEA, B7.1, ICAM-1, LFA-3, GM-CSF
I	CEA-expressing malignancies	Anti-CEA-SFv-Zeta T cell receptor
I	CEA-expressing malignancies	B7.1 (CD80)
II	Advanced cancer with overexpression of p53	Anti-p53 T cell receptor
I	MUC-1- expressing tumors	MUC-1, IL-2
I	Advanced cancer	Cytochrome P450
I	Advanced cancer	Endostatin
I	Advanced cancer	Heat shock protein 70
I	Advanced cancer	T cell receptor alpha and beta chain
I	Advanced cancer	Bifunctional shRNA specific for stathmin 1 oncoprotein
I/II	Advanced cancer	GM-CSF
I	Advanced cancer	IL-12
I/II	Advanced cancer	CYP1B1
I	Advanced cancer	GM-CSF, CD154 (CD40-ligand)
I/II	Advanced cancer	IL-2
I	Advanced cancer	p53
I	Advanced cancer	AMEP
I	Advanced cancer	B7.1 (CD80) ,ICAM-1, LFA-3
I/II	Advanced cancer	Cytosine deaminase
I/II	Advanced cancer	CEA
I	Solid tumors	Tumor antigen
I	Solid tumors	Interferon-gamma
I	Solid tumors	TNF
I	Solid tumors	Brachyury oncoprotein
I	Solid tumors	Human telomerase reverse transcriptase
I/II	Solid tumors	Oncolytic virus (no transgene)
I	Solid tumors	Retinoblastoma 94
I	Solid tumors	p53
I	Solid tumors	O6-methylguanine DNA methyltransferase (MGMT)
I	Solid tumors	GM-CSF
I	Solid tumors	GM-CSF, bi-shRNA-furin
I	Solid tumors	GM-CSF, TGF-beta 2 antisense
I	Solid tumors	GM-CSF, humanized Escherichia coli beta-galactosidase

CEA: Carcinoembryonic antigen; IL-2: Interleukin-2; AMEP: Antiangiogenic metargidin peptide; CYP1B1: Cytochrome P450 isoenzyme 1B1; TNF: Tumor necrosis factor; hTERT: Human telomerase reverse transcriptase; GHRH: Human growth hormone releasing hormone.