Abstract Number: 2951 ($ISIS)
Potent antisense pharmacology of highly optimized antisense oligonucleotides in multiple transgenic, spontaneous and patient derived xenograft models of cancer reveals antitumor activity for the non-coding RNA MALAT-1.
Abstract Body: Antisense technology holds great promise as a novel drug discovery platform that can rapidly translate discoveries from cancer genomics into highly selective therapeutics. Antisense oligonucleotices (ASOs) are particularly attractive, as they can be applied to difficult to drug target classes currently not tractable by other therapeutic modalities. Recent clinical demonstration of activity for ASOs in cancer patients supports the potential of this drug class, however, challenges in demonstrating robust antisense pharmacology in preclinical cancer models has slowed the progress of this technology as cancer therapeutics. Here we have employed a high resolution in-situ hybridization-based methodology (QuantigeneTM) among other detection methods, to demonstrate visually and quantitatively the activity of systemically administered, high potency next generation antisense oligonucleotides, in multiple preclinical cancer models. Cancer models evaluated include, transgenic models, chemically induced tumor, genetically predisposed mouse strains, cell line derived xenograft and patient derived xenograft models. As a test antisense target RNA sequence we chose the non-coding RNA MALAT1 (also called NEAT2) because it is ubiquitously expressed at high levels in most cell types and thus RNA levels could be readily visualize at the cellular level by Quantigene method. In addition, as MALAT-1 is overexpressed in many human tumors it also had the potential to be a therapeutically relevant target. We screened >1000 ASO sequences in vitro and identified highly potent mouse and mouse/human cross reactive MALAT-1 ASOs that reduced target RNA in cells in culture with IC50 values in the low nanomalor range (10-50 nM), without any lipid mediated delivery vehicles (ASO free uptake). Systemic delivery (s.c. administration of ASOs formulated in saline) of MALAT1 ASOs in vivo were well tolerated in all animals tested, and reduced target RNA by 70->90% in the tumor cells of APC/min- mice, prostate tumor cells of the TRAMP model, DEN-induced HCC tumors as well as in the tumor cells of several human tumor xenograft models and in a patient derived NSCLC primary tumor explants model. Interestingly, MALAT-1 inhibition by ASOs was also associated with significant antitumor effects including inhibition of tumor formation and decreased BrdU positive cells in the polyps of APC/min- mice, decrease tumor growth in TRAMP prostate tumors and DEN HCC tumors and significant tumor growth delays in several xenograft and human tumor explant models. These data demonstrate unequivocal, potent, ASO mediated antisense activity of highly optimized next generation ASOs targeting MALAT-1 by systemic administration and highlight a previously uncharacterized role of the ncRNA MALAT1 as regulator of tumor growth in vivo.
Abstract Number: LB-363 ($ALNY)
Intravesical delivery of lipid nanoparticle formulated p21(Waf1/Cip1) activating dsRNA induces tumor regression and enhances animal survival in a orthotopic bladder cancer model
Abstract Body: Small RNA molecules are a promising new class of drugs by offering expanded targets not druggable by conventional therapies with high target specificity and low toxicity. However, their clinical use is significantly hindered by the lack of vehicles that deliver the molecules efficiently to target tissues and cells. The bladder is an easily accessible hollow organ and is ideal for local delivery of drugs or molecules such as small RNA. RNA activation (RNAa) is a newly discovered mechanism of gene induction triggered by promoter targeted double-stranded RNA, also known as small activating RNA (saRNA). In this study, we investigate, in an orthotopic bladder cancer model, antitumor effects of a saRNA (dsP21) targeting the promoter of the p21WAF1/CIP1 (p21) gene, a key negative regulator of the cell cycle rarely mutated or deleted in bladder cancer. Introducing dsP21 into bladder cancer cells activated p21 expression with subsequent inhibition of cell proliferation, arrest of the cell cycle and induction of apoptosis accompanied by the activation of caspase 3 and PARP. Chemical modification (2’-Fluoro) and subsequent formulation of dsP21 in lipid nanoparticles (LNPs) retained its RNAa activity with minimal immunostimulatory effect and extended its stability in urine, and when delivered intravesically it could well diffuse into the bladder wall. Delivery of LNP-formulated dsP21 (LNP-dsP21) into mouse bladder with established human bladder cancer significantly inhibited tumor growth and extended animal survival with demonstrated p21 activation in vivo. Of particular significance, LNP-dsP21 treatment caused the regression or disappearance of established tumors in 30% of the treated mice. Our results provide proof-of-principle that targeted activation of p21 can be applied to the treatment of bladder cancer and LNP-formulated small RNA can be successfully delivered to the bladder by intravesical instillation. Further clinical development of RNAa-based intravesical therapy is warranted for the treatment of residual and recurring bladder cancer in humans. Acknowledgement: financial support from the AACR Henry Shepard Bladder Cancer Research Grants (09-60-30-LI).