There are two types of cyclic AMP (cAMP)-dependent protein kinase (PKA), type I (PKA-I) and type II (PKA-II), which contain an identical catalytic (C) subunit but have distinct regulatory (R) subunits, RI and RII, respectively. Evidence suggests that increased expression of PKA-I and its regulatory subunit (RIα) underlies tumorigenesis and tumor growth. Our studies include: (1) Antisense strategy. Antisense oligonucleotide targeted against the RIα subunit of PKA is used to demonstrate that the sequence-specific inhibition of RIα gene expression inhibits tumor growth. The results are confirmed by the use of siRNA, and the clinical utility of antisense RIα is confirmed by the findings that antisense RIα acts synergistically with cytotoxic drugs currently in use and CpG immune-modulatory oligonucleotides. Our laboratory was the first to show the genome-wide effects of antisense inhibition of gene expression in microarrays. Using in vivo tumor models of PC3M human prostate carcinoma grown in nude mice, the specificity of RIα antisense effects on gene expression signatures was critically assessed using three oligonucleotides that differed in sequence or chemical modification. Antisense treatment was found to downregulate one cluster, or signature, of genes involved in cell proliferation and upregulate another involved in cell differentiation. Importantly, these expression signatures were quiescent and unaltered in the livers of antisense-treated animals. Thus, cDNA microarray analysis defined the molecular portrait of a reverted tumor phenotype in antisense-treated regressing tumors. (2) Retroviral vectormediated overexpression of wild-type and point-mutated R and C subunits of PKA is used to study cell growth, differentiation, and reverse transformation of tumor cells. The results obtained from the study of PC3M cells via classical biochemical analysis, DNA microarrays, and confocal microscopy demonstrated that cancer cells exhibit abnormal expression of PKA isozymes that constitute malignant phenotype, and correction of this abnormal PKA profile achieved by differential expression of the PKA R subunit genes provides a novel tumor targetbased gene therapy of cancer. In OVAR-8 ovarian carcinoma cells, the molecular portraits of the distinct phenotype induced by differential R subunit gene overexpression identified the distinct disease-dependent expression profiles of clinical tumors. Many of these genes had been previously identified in clinical studies as upregulated in progressively transformed ovarian tissues as compared to normal counterparts. Analysis of pathway involvement showed active involvement of the differential signaling genes contained within the ERK/MAPK signaling pathway versus genes in the Wnt/β signaling pathway. This result suggests that perturbation of the PKA-I to PKA-II ratio in the cell is an abnormal/dangerous signal that may underlie tumorigenesis and tumor progression. This study thus suggests that protein kinase A isozyme switching in the cell might provide a novel tumor targetbased gene therapy of human cancer. (3) Effect of CRE-transcription factory decoy oligonucleotides: Our previous studies showed that the CRE-decoy inhibits the CRE- and Ap1-directed transcription and inhibits cancer cell growth selectively without harming normal cell growth (Park, et al., J Biol Chem, 1999). It was found that in cancer cells, CREB (CRE-binding protein) phosphorylation by ERK/MAPK signaling overrides that by cAMP/PKA signaling, whereas, growth factormediated CREB phosphorylation is minimal in normal cells. Blocking of CRE/CREB effect by the decoy oligonucleotide resulted in profound growth inhibition exceeding that by Tomoxifen, on the MCF7 breast tumor in nude mice without producing systemic cytotoxicity (appeared in NewsClipsBiocentury, Part II, Belmont, CA weekly Part II Edition Nov. 18, 2002). (4) Extracellular PKA (ECPKA) as a molecular marker for cancer diagnosis and prognosis. (5) Development of an autoantibody-detection method (in contrast to the conventional antigen-detection method) for cancer diagnostics that gives greater sensitivity and specificity than existing cancer diagnostics (U.S. patent application no. 60/550,348 March 8, 2004). (6) Development of nanocapsule delivery of oligonucleotides and other drugs (in collaboration with Claude Malvy, Ph.D., University of Paris, France)

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Intramural Research (Z01)
Project #
1Z01BC005216-36
Application #
7592514
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
36
Fiscal Year
2007
Total Cost
$113,753
Indirect Cost
Name
National Cancer Institute Division of Basic Sciences
Department
Type
DUNS #
City
State
Country
United States
Zip Code
Nesterova, M; Johnson, N; Cheadle, C et al. (2006) Autoantibody biomarker opens a new gateway for cancer diagnosis. Biochim Biophys Acta 1762:398-403
Cho-Chung, Yoon S (2006) Autoantibody biomarkers in the detection of cancer. Biochim Biophys Acta 1762:587-91
Nesterova, Maria V; Cho-Chung, Yoon S (2004) Antisense protein kinase A RIalpha inhibits 7,12-dimethylbenz(a)anthracene-induction of mammary cancer: blockade at the initial phase of carcinogenesis. Clin Cancer Res 10:4568-77
Cho-Chung, Yoon S (2004) Antisense protein kinase A RI alpha-induced tumor reversion: portrait of a microarray. Biochim Biophys Acta 1697:71-9
Cho-Chung, Yoon S (2003) cAMP signaling in cancer genesis and treatment. Cancer Treat Res 115:123-43
Cho, Yee Sook; Cho-Chung, Yoon S (2003) Antisense protein kinase A RIalpha acts synergistically with hydroxycamptothecin to inhibit growth and induce apoptosis in human cancer cells: molecular basis for combinatorial therapy. Clin Cancer Res 9:1171-8
Kim, Young Hoon; Lim, Do Sun; Lee, Ji Hye et al. (2003) Gene expression profiling of oxidative stress on atrial fibrillation in humans. Exp Mol Med 35:336-49
Cheadle, Chris; Cho-Chung, Yoon S; Becker, Kevin G et al. (2003) Application of z-score transformation to Affymetrix data. Appl Bioinformatics 2:209-17
Cho-Chung, Yoon S (2003) CRE-enhancer DNA decoy: a tumor target-based genetic tool. Ann N Y Acad Sci 1002:124-33
Mani, S; Goel, S; Nesterova, M et al. (2003) Clinical studies in patients with solid tumors using a second-generation antisense oligonucleotide (GEM 231) targeted against protein kinase A type I. Ann N Y Acad Sci 1002:252-62

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