Dual cancer-specific targeting strategy cures primary and distant breast carcinomas in nude mice

Abstract

Limitations of current viral-based gene therapies for malignant tumors include lack of cancer-specific targeting and insufficient tumor delivery. To ameliorate these problems and develop a truly effective adenovirus gene-based therapy for cancer, we constructed a conditionally replication competent adenovirus (CRCA) manifesting the unique properties of tumor-specific virus replication in combination with production of a cancer-selective cytotoxic cytokine, melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24), which embodies potent bystander antitumor activity. Cancer cell selective tropism was ensured by engineering the expression of the adenoviral E1A protein, necessary for viral replication, under the control of a minimal promoter region of progression elevated gene-3 (PEG-3), which functions selectively in diverse cancer cells with minimal activity in normal cells. In the E3 region of this CRCA, we introduced the mda-7/IL-24 gene, thereby mediating robust production of this cytokine as a function of adenovirus replication. Infection of this CRCA (designated Ad.PEG-E1A-mda-7) in normal mammary epithelial cells and breast cancer cells confirmed cancer cell selective adenoviral replication, mda-7/ IL-24 expression, growth inhibition, and apoptosis induction. Injecting Ad.PEG-E1A-mda-7 into human breast cancer xenografts in athymic nude mice completely eradicated not only the primary tumor but also distant tumors (established on the opposite flank of the animal) thereby implementing a cure. This dual cancer-specific targeting strategy provides an effective approach for treating breast and other human neoplasms with the potential for eradicating both primary tumors and metastatic disease. Additionally, these studies support the potential use of mda-7/IL-24 in the therapy of malignant cancers.bystander antitumor activity ͉ conditionally replication competent adenovirus ͉ PEG-Prom ͉ mda-7͞IL-24 ͉ in vivo tumorigenesis

Enhanced delivery of mda-7/IL-24 using a serotype chimeric adenovirus (Ad.5/3) improves therapeutic efficacy in low CAR prostate cancer cells

“…27 The primary antibodies used were anti-BiP/GRP78 (Santa Cruz Biotechnology, Inc, Santa Cruz, CA), anti-Bcl-x L , anti-Bcl-2, PARP (poly(ADP-ribose) polymerase-1), p38 MAP (mitogenactivated protein) kinase, p-p38MAP kinase, ERK Human prostate cancer xenografts in athymic nude mice DU-145 and PC-3 cells (3 Â 10 6 ) were injected subcutaneously in 100 ml of PBS in the left flank of male athymic nude mice (NCRnu/nu, 4 weeks of age, 20 g body weight). 27,28 After establishment of visible tumors of B75 mm 3 , requiring approximately 8-10 days, intratumoral injections of different Ads were administered only to tumors on the left flank at a dose of 4.5 Â 10 8 p.f.u. (plaqueforming units) in 100 ml.…”

A cancer terminator virus eradicates both primary and distant human melanomas

“…[9][10][11][12][13][14][15][16][17][18] These findings were extended to human-tumor xenograft nude mouse models, indicating that Ad.mda-7 suppresses tumor growth and cancer progression. [18][19][20][21][22] Through studies in multiple laboratories, it is now eminently clear that mda-7/IL-24 is one of the broadest-acting, cancer-specific and apoptosis-inducing cytokine genes, which also displays antiangiogenic, radiosensitizing, immune-stimulating and potent 'bystander' antitumor activity. [17][18][19][20][21][22][23][24][25][26] A momentous step in the evolution of mda-7/IL-24 as a potential therapeutic gene was the evaluation of Ad.mda-7 (INGN 241) in a phase-I clinical trial by intratumoral injection in patients with advanced solid tumors, including melanomas.…”

“…[18][19][20][21][22] Through studies in multiple laboratories, it is now eminently clear that mda-7/IL-24 is one of the broadest-acting, cancer-specific and apoptosis-inducing cytokine genes, which also displays antiangiogenic, radiosensitizing, immune-stimulating and potent 'bystander' antitumor activity. [17][18][19][20][21][22][23][24][25][26] A momentous step in the evolution of mda-7/IL-24 as a potential therapeutic gene was the evaluation of Ad.mda-7 (INGN 241) in a phase-I clinical trial by intratumoral injection in patients with advanced solid tumors, including melanomas. 13,16,18,[27][28][29] These studies indicated that mda-7/IL-24 was safe and could induce as much as 70% apoptosis in tumors following a single injection of recombinant virus, and with multiple injections, it promoted objective clinical responses, especially in melanoma patients.…”

“…This approach employs a genetically modified adenovirus, CTV, in which replication is controlled by a minimal active region of the promoter of progression-elevated gene-3 (PEG-3), which expresses selectively in diverse cancer cells with limited activity in normal cells. 21,[39][40][41] The PEG-3 gene was cloned using subtraction hybridization as an upregulated transcript from a transformation progression rodent cancer model. 42,43 Of direct relevance for gene therapy applications, activity of the PEG-3 promoter (PEG-Prom) is significantly and often markedly elevated not only in rodent but also in human cancer cells of diverse origin when compared to normal cells.…”

“…42,43 Of direct relevance for gene therapy applications, activity of the PEG-3 promoter (PEG-Prom) is significantly and often markedly elevated not only in rodent but also in human cancer cells of diverse origin when compared to normal cells. 21,[39][40][41] The mechanism underlying the cancer-specific expression of the PEG-Prom implicates two transcription factors, AP-1 and PEA-3, which are expressed at elevated levels, either singly or in combination, in virtually all types of cancers. 21,[39][40][41][42][43][44][45] Using the PEG-Prom to transcriptionally regulate green fluorescence protein (GFP) or luciferase gene expression via a replication-incompetent Ad confirmed targeted cancer-cell-selective transgene expression in human prostate and breast cancer cells, as well as in malignant glioma cells.…”