Manufacturing hurdles, including changes in production sites and manufacturing processes, pose challenges to developers regarding reproducibility and comparability of results for gene therapy. Introduction of an international standard for gene therapy is especially important, given the usually orphan nature of the diseases to be treated, hampering the comparison of cross-trial and cross-manufacturing results. This presentation will discuss challenges and regulatory perspectives in quality control and standardization of gene therapy and an international effort in developing the 1st WHO International Standard for gene therapy products.

We have investigated the performance of membrane chromatography as a polishing step for purification of AAV5. We compare the performance for contaminant removal compared to commonly used resin based approaches to show the potential of membrane chromatography as a platform-able polishing step for AAV purification. Membrane chromatography can be loaded 40 times more quickly than conventional chromatography resins, which can lead to much higher productivities and greatly speed the purification step

To-date, gene therapy development has focused on orphan conditions. This talk will look at both the challenges and opportunities associated with moving gene therapies to large, non-orphan diseases.

This presentation will discuss several strategies to commercialize cell and gene therapy products. Particular attention will be given to the manufacturing of these products and how early planning can increase the speed to market and enable lower COGS. Both allogeneic and autologous therapies as well as gene therapy strategies will be discussed.

RNA plays important and diverse roles in biology, but molecular tools to manipulate and measure RNA are limited. We demonstrate that RNA-targeting CRISPR effector, Cas13, can be engineered for mammalian cell RNA knockdown, binding, and RNA editing. Cas13 can be heterologously expressed in mammalian and plant cells for targeted knockdown of either reporter or endogenous transcripts, targeted RNA binding for transcript imaging, and programmable RNA editing. We also show that both Cas13 and Cas12 can be used for sensitive nucleic acid diagnostics and show that CRISPR diagnostics (CRISPR-dx) can be used for point-of-care COVID-19 detection. Our results establish CRISPR-Cas13 as a flexible platform for RNA targeting with wide applicability for studying RNA, diagnostics, and therapeutics.

Pre-existing immunity is a major challenge for AAV gene therapy, preventing many patients from being treated by therapies under development today. Artificial intelligence (AI) presents new opportunities for overcoming such challenges, especially when applied to the design of synthetic AAV capsids. This approach combines three advanced technologies: i) next-gen library synthesis; ii) measurement via next-gen sequencing; and iii) library analysis powered by machine learning. Such a workflow can dramatically accelerate the search for synthetic capsids with superior targeting and immunological profiles relative to capsids derived from natural AAV isolates. I will review the technological advances that are pushing the field of AAV capsid engineering toward AI-powered methods, describe and explore the promise of this approach, and discuss anticipated challenges. In the near future, AI will revolutionize our ability to design safe, targeted, and robust delivery vectors for the treatment of many new genetic conditions, and importantly, enable all patients to benefit from such therapies.

Small Nanobodies® with their modular design show a different pharmacokinetic profile compared to conventional antibodies. This difference in exposure can be exploited by alternative delivery methods. DNA-based gene transfer of Nanobodies and biopharmaceuticals in general is an appealing alternative to conventional protein therapy. We demonstrate that multivalent Nanobodies encoded as DNA results in a stronger, longer-term and more localized exposure compared to conventional protein therapy.

The high-resolution size capabilities of the Zetasizer Ultra have enabled a new assay to rapidly characterize (AAV) viral concentration.  Utilizing (MADLS), Zetasizer Ultra can determine both AAV viral concentration and size in a single measurement that is cuvette-based, rapid, label-free, low volume, and non-destructive.  

A highly sensitive iLite^® reporter assay based on the establishment of a cell line stably transfected with a luciferase reporter-gene placed under the control of an AAV-responsive chimeric promoter, provides a highly sensitive, rapid and precise method for quantifying the immune response to a wide range of recombinant AAV vectors.

We developed an anticancer drug and implanted it directly into solid tumors, suppressing the oncogenes and genes responsible for activation of telomerase. The drug is continuously available in the DNA promoter regions for long periods of time to prevent overexpression and reactivation. In 4 separate experiments, we implanted the drug, using, an intratumoral sustained polymeric system, directly into 100 mouse tumors, reducing the tumor volume 5-fold. Kaplan-Meier p-value was 0.0001.

Aggregation of misfolded proteins into toxic species is implicated in the pathogenesis of many neurodegenerative proteinopathies, including Alzheimer’s disease, Parkinson’s disease, and prion disease. Lowering pathogenic protein expression at the DNA level using zinc finger protein transcription factors (ZFP-TFs) has the potential to slow or stop disease progression. We present data from iPSC-derived human neurons, rodents, and nonhuman primates demonstrating durable, highly specific targeted gene regulation following a single ZFP-TF administration.

Neurodegenerative disorders are the leading cause of disability in the aging population and predicted to soon surpass cancer and become the second leading cause of death worldwide. Our group aims to harness the highly versatile genome-modifying technologies to develop corrective gene therapies for such neurodegenerative conditions. This talk will highlight our most recent efforts on using recently emerged precision gene-editing tools to treat amyotrophic lateral sclerosis and Huntington’s disease, two devastating and relentlessly progressing neurodegenerative disorders.