Session 6: RNAi/ASO Preclinical Studies
Tuesday, October 30, 2012
10:30 AM-11:00 AM
Targeting alpha1-antitrypsin for the Treatment of A1AT Liver Disease
Shuling Guo1, Sheri L. Booten1, Gene Hung1, Andrew Watt1, Keith Blomenkamp2, Jeffery H. Teckman2, Susan M. Freier1, Michael L. McCaleb1, Brett P. Monia1
2St. Louis University School of Medicine
Alpha-1 antitrypsin deficiency (AATD) is a rare genetic disease due to mutations of the alpha-1 antitrypsin (A1AT) gene. Emphysema associated with AATD is caused by the deficiency of systemic A1AT protein (loss-of-function phenotype) and can be treated with augmentation therapy. In contrast, AATD Liver Disease, which is caused by aggregation and retention of mutant A1AT protein in the liver, can only be treated by liver transplantation. The hepatic pathology is the result of a toxic gain-of-function of the mutant Z protein, which affects both children and adults. We have developed optimized 2’-O-methoxyethyl (“2nd Generation”) phosphorothioate antisense oligonucleotide (ASO) drugs targeted against A1AT to determine if reducing the expression of A1AT production in liver would improve AATD-related liver disease. ASOs were designed to utilize an RNaseH-dependent mechanism of action, which promotes cleavage of A1AT mRNA. Administration of A1AT ASOs to PIZ transgenic mice expressing the mutant human gene resulted in significant reductions in hepatic A1AT mRNA and circulating A1AT levels that were dose dependent. Importantly, ASO treatment of PIZ mice produced significant reductions in A1AT protein aggregates in liver, reduced markers of fibrosis (e.g., TIMP1), and slowed fibrosis progression. Futhermore, administration of A1AT ASOs in non-human primates (50 mg/kg x 12 wk) resulted in marked reductions in hepatic and circulating A1AT mRNA and protein levels. Our data demonstrate that antisense oligonucleotide treatment can potently reduce mutant A1AT protein production and represents a potential treatment for AATD liver disease.
Session 7: Alternate Mechanisms Preclinical Studies
Tuesday, October 30, 2012
3:00 PM-3:30 PM
Correction of myotonic dystrophy in mouse models by antisense targeting of nuclear-retained RNA
Charles Thornton1, Thurman M. Wheeler1, Andrew J. Leger3, Sanjay K. Pandey2, A. Robert MacLeod2, Masayuki Nakamori1, Seng H. Cheng3, Bruce M. Wentworth3, C. Frank Bennet2
1University of Rochester Medical Center, Rochester, NY
2Isis Pharmaceuticals, Carlsbad, CA
3Genzyme Corporation, Framingham, MA
Myotonic dystrophy type 1 (DM1) is one of several dominantly-inherited disorders in which genomic expansions of tandem repeats lead to expression of repetitive RNAs that have deleterious effects. This RNA gain-of-function mechanism is implicated in both types of DM, several forms of cerebellar degeneration, and the most common form of familial ALS. In DM1 the repetitive segment is an expanded CUG repeat (CUGexp) in the 3' untranslated region of DM protein kinase (DMPK) mRNA. The mutant RNA accumulates in nuclear foci, trapping proteins that have poly(CUG) binding affinity. This leads to misregulated alternative splicing and other perturbations of the muscle transcriptome. Antisense drugs would seem ideally suited for treating RNA toxicity, were it not for the problem of low biodistribution to muscle. This presentation will focus on evidence that nuclear-retained transcripts are unusually sensitve to RNase H-active antisense oligonucleotides (ASOs), enabling therapeutic knockdown in skeletal muscle. In transgenic mouse models of DM1, the systemic administration of 2′-0-methoxyethyl gapmer ASOs produced 80% knockdown of CUGexp RNA in skeletal muscle. Strong knockdown was achieved within 4 weeks (8 subcutaneous injections) and persisted for one year after cessation of treatment. No specific changes of ASO design or formulation were required, and the effect did not depend on enhanced entry of ASOs into muscle fibers. Many of the physiological, morphological, and biochemical features of the disease were corrected, without evidence for toxicity or off-target knockdown. These results suggest that the ASO-RNase H mechanism may provide an effective strategy for treating RNA dominant diseases.
Session 8: Advances in Oligonucleotide Clinical Development I
Wednesday, October 31, 2012
9:30 AM-10:00 AM
Development of Antisense Drugs for Cancer
Oncology drug discovery and development is rapidly moving towards a “Personalized Medicine” approach in which cancer therapy is tailored to the genetic or epigenetic cause of an individual’s cancer. The foundation for this approach is genomic sequencing of cancer cells to identify causal mutations, which facilitates both choice of treatment and the identification of novel targets for cancer drug discovery and development. Antisense technology is particularly well suited to exploit this new paradigm for cancer drug discovery due to its ability to rapidly identify drugs against all targets regardless of their function, and to impact target function with unprecedented specificity. However, antisense drug development for cancer has historically had mixed results, and the need for greater clinical success is high. Research into the discovery of new antisense chemistries that are more effective in penetrating and producing robust antisense effects in cancer cells is progressing rapidly, and resulting in more reproducible and robust effects in preclinical cancer models and in the clinic. We have been developing a bicyclic nucleic acid chemistry, referred to as constrained ethyl (cET), for a number of disease indications including cancer. The most advanced cET antisense program is the ISIS-STAT3rx program for cancer. STAT3 is a transcription factor that resides within the JAK-STAT signaling pathway, and mutations leading to the chronic activiation of STAT3 have been strongly linked to a variety of hematological and solid cancers. ISIS-STAT3rx potently and reproducibly lowers levels of STAT3 in cell culture, and in tumor cells and tumor-associated stromal cells in preclinical models, and produces robust antitumor activity in STAT3-dependent tumor models. ISIS-STAT3rx is currently under evaluation in a Phase I study in patients with cancer and early results are encouraging. Results will be presented demonstrating the utility of the cET antisense approach for cancer including a review of the ISIS-STAT3rx program.