The field of oligonucleotide therapeutics holds great promise for the advancement of medicine. To date, there are at least ten oligonucleotide therapeutics in the clinic, and many more under pre-clinical development. However, widespread and sustained RNA interference in the brain, using chemically modified small interfering RNAs (siRNAs) has just begun to be realized.

The most successful example of approved oligonucleotides for the central nervous system (CNS) is SPINRAZA (Nusinersen) (Finkel et al. 2017). Nusinersen is a fully modified antisense oligonucleotide (ASO) that functions via splicing modulation and has been a transformative treatment for Spinal Muscular Atrophy (Finkel et al. 2017). Another ASO, Tominersen works via a different mechanism (RNAse H) and is under development for the treatment of Huntington’s disease (Tabrizi et al. 2019).  Through the development of these ASOs, IONIS, in collaboration with partners, is responsible for many of the technological and chemical advances in the field. Despite the success and promise of ASOs, their clinical utility is validated for diseases where modulation of disease-causing genes is crucial in the spinal cord. Thus, the limited distribution of ASOs to other regions of the CNS, particularly in larger animals, might hinder their utility in treating diseases with wider brain involvement. 

Last year our team published a paper where we developed a novel, di-valent small interfering RNA (siRNA) chemistry platform for efficacious target silencing throughout the CNS in mice and non-human primates (Alterman et al. 2019). Like previously developed oligonucleotides, the di-siRNA relies on phosphorothioate-mediated uptake, but we also believe the large size (~27 kDa vs ~ 7kDa for ASOs) and chemical configuration of the di-siRNA is  responsible for slower CSF clearance, observed CNS retention, widespread distribution, and long-term silencing of the di-siRNA.

The beauty of RNA-based medicines, and chemically modified oligonucleotides in particular, is their “informational” nature. Unlike small molecules, informational drugs are made up of separate dianophore, which largely determines the pharmacokinetics and dynamics, and pharmocophore, the targeting sequence which causes the gene specific silencing (Khvorova and Watts 2017). Thus, once an effective and safe chemical configuration is identified, it can be applied to theoretically any target and with minimal impact on pharmacokinetic or dynamic properties.

Our publication explored the bio-distribution and efficacy of the di-siRNA targeting Huntingtin, the gene whose CAG repeat expansion is causative of Huntington’s disease, and Apolipoprotein E, a risk factor for Alzheimer disease, in wild-type mice, but left opportunities for further investigation: 

1. Impact of silencing target genes on pathology: This publication provided us with key tools to studying the role of risk genes in CNS diseases and pathology. With the tools in hand, we are currently exploring the impact of silencing CNS ApoE in animal models of Alzheimer disease. Consistent with ApoE knockout models, we observed a reduction in amyloid pathology in Alzheimer disease mouse models, that is greatest in female animals. Interestingly, transcriptomic analysis identified genes involved in the immune response as potential mechanistic mediators, and resulted in no detectable changes in App, Psen1, Mapt, or Bace1 RNA expression. We are currently drafting this manuscript for publication.
2. Informational nature of the di-siRNA chemistry: Excitingly, we are further validating the informational nature of the di-siRNA and are able to achieve similar silencing efficacy in mice for additional targets including alpha-synuclein and tau.
3. Investigation of novel chemical configurations: Our initial results were exceptionally exciting, the fully modified di-siRNA resulted in distribution to previously inaccessible brain regions such as the caudate and putamen and showed an unprecedented duration of effect after a single, bolus injection. These results serve as a solid foundation to build upon as we investigate novel chemical modifications and configurations that may increase the siRNA stability, therapeutic index, and duration of effect in vivo.

We are fortunate to perform translational biomedical research with dual goals: scientific discovery and the development of therapeutics for individuals with debilitating conditions. In providing a platform for gene modulation in the CNS, this publication had a large impact on our team and hopefully many others. Two of the first authors, Bruno Godinho and Matthew Hassler, are playing integral roles in building a new biotech company, Atalanta Therapeutics (https://www.atalantatx.com), focusing on the preclinical development of the di-siRNA. Julia Alterman is currently overseeing the development of a new direction at UMASS focusing on oligonucleotide therapeutics for multiple indications. I am working towards achieving my MD/PhD and returning to medical school in the spring. I hope to practice as a physician-scientist, aiding in the development and application of novel therapeutics for some of the most challenging diseases, specifically neurological conditions. The work in this publication truly laid the ground for us to achieve both our personal goals and greater scientific goals. 

Our lab continues to work on siRNA therapeutic development in a highly collaborative manner both within the lab and with our academic and industry partners. With the power of many minds, diverse experiences, and complementary skill sets, we hope to bring disease modifying therapies to those in the greatest of need and revolutionize the field of medicine.

Banner photo: Khvorova Lab, Fall 2019. 

Link to the paper: https://www.nature.com/articles/s41587-019-0205-0 

Link to the Khvorova Lab, University of Massachusetts Medical School, Worcester MA. https://www.umassmed.edu/khvorovalab/ 

References: 

Alterman, J. F., Bmdc Godinho, M. R. Hassler, C. M. Ferguson, D. Echeverria, E. Sapp, R. A. Haraszti, A. H. Coles, F. Conroy, R. Miller, L. Roux, P. Yan, E. G. Knox, A. A. Turanov, R. M. King, G. Gernoux, C. Mueller, H. L. Gray-Edwards, R. P. Moser, N. C. Bishop, S. M. Jaber, M. J. Gounis, M. Sena-Esteves, A. A. Pai, M. DiFiglia, N. Aronin, and A. Khvorova. 2019. 'A divalent siRNA chemical scaffold for potent and sustained modulation of gene expression throughout the central nervous system', Nat Biotechnol, 37: 884-94.

Finkel, R. S., E. Mercuri, B. T. Darras, A. M. Connolly, N. L. Kuntz, J. Kirschner, C. A. Chiriboga, K. Saito, L. Servais, E. Tizzano, H. Topaloglu, M. Tulinius, J. Montes, A. M. Glanzman, K. Bishop, Z. J. Zhong, S. Gheuens, C. F. Bennett, E. Schneider, W. Farwell, D. C. De Vivo, and Endear Study Group. 2017. 'Nusinersen versus Sham Control in Infantile-Onset Spinal Muscular Atrophy', N Engl J Med, 377: 1723-32.

Khvorova, A., and J. K. Watts. 2017. 'The chemical evolution of oligonucleotide therapies of clinical utility', Nat Biotechnol, 35: 238-48.

Tabrizi, S. J., B. R. Leavitt, G. B. Landwehrmeyer, E. J. Wild, C. Saft, R. A. Barker, N. F. Blair, D. Craufurd, J. Priller, H. Rickards, A. Rosser, H. B. Kordasiewicz, C. Czech, E. E. Swayze, D. A. Norris, T. Baumann, I. Gerlach, S. A. Schobel, E. Paz, A. V. Smith, C. F. Bennett, R. M. Lane, and Ionis-HTTRx Study Site Teams Phase 1-2a. 2019. 'Targeting Huntingtin Expression in Patients with Huntington's Disease', N Engl J Med, 380: 2307-16.