IND-enabling Programs for Gene and Cell Therapies Webinar Questions

7 June 2021

Get insightful answers to some common questions on IND-enabling studies for cell and gene therapies (CGTs) from our recent presentation. For more information on our solutions, visit our Cell & Gene Therapy Education Center here.


An estimated 362 CGTs were in clinical development in the US in early 2020, targeting over 100 diseases. Translating a lab-based innovation into an effective therapy can be challenging; thus having a robust preclinical study program to advance a therapeutic candidate into clinical trials is vital. Read on to get some answers to common questions related to Investigational New Drug (IND)-enabling CGT studies.

1. Can canine models be used for in vivo IND-enabling studies?

In vivo model selection is a key consideration for CGT IND-enabling studies, and one we often receive questions about. When possible, we typically recommend a hybrid approach that uses both healthy and disease models with the choice of species being dictated by a number of factors; most notably the nature of the CGT and the availability of the appropriate in vivo models. It is possible to use canine models and they may be more appropriate for regenerative products rather than viral-based therapies. When considering viral-based products, the NHP immune system may respond in a similar way to the human immune system and may tolerate the presence of human protein better than canine models. It’s important to note that one of the US-marketed gene therapies used a canine model because there was a recognized relevant canine model in existence. The ultimate decision about in vivo model choice is driven by what is known about the therapeutic candidate.

2. What NHP immunosuppression protocols do you recommend for AAV vector-based products?

A challenge in using NHPs is that they may have been exposed to a variety of other pathogens (e.g. viruses, parasites, etc.) before entering a study. That’s why, even before getting into recommendations about the immunosuppression protocol, we take great care to source ’clean’ animals and ensure we have insight into their viral status. In addition, we reduce the risk of pathogen exposure through irradiation of their food and other procedures. For AAV vector-based products, we often treat with immunosuppressants such as prednisone/prednisolone prior to dosing with the aim of “knocking down” the immune system. We would not knock it out completely, as having some underlying inflammatory activity helps to control for any harbored viruses (e.g. CMV, etc.). In some cases, we’ve also make use of immunosuppressants such as rapamycin, mycophenolate mofetil, or cyclosporin A (alone, or in combination) to ensure appropriate suppression.

3. What safety assessments are needed for explanted primary progenitor cells to ensure that all in vitro transfection virus is removed before implantation?

The focus of the requirement here is around assessment of viral load. The simplest approach would be a PCR assay of the cell and/or supernatant over time, or even an infectivity assay.

4. What types of safety pharmacology assessments do you include for CNS-delivered therapies in NHPs?

There are a host of physiology, neurological and behavioral assessments we make on NHPs receiving CNS-delivered therapies. Jacket telemetry provides good basic physiological and CNS data, including autonomic tone information. From a neurological perspective, we examine induced reflexes (e.g. toe pinch, patellar reflex, pupillary response, menace reflex, etc.) and can assess nerve conduction velocity using evoked potentials. We also conduct in-cage behavioral assessments of general movement and activity and observations such as changes in limb use or favoring of one limb over another.

5. What approaches – for example biopsy or imaging biomarkers – can be used to demonstrate maintenance of therapy both preclinically and clinically?

Preclinically, the amount of tissue needed for cell therapy assays is usually greater than would be available via biopsy, thereby potentially limiting its use to assess maintenance of a cell therapy. However, for systemic gene therapies, in particular for those targeting the liver, routine biopsies are possible. As an alternative to biopsy, for some preclinical cell biodistribution and homing studies in small animals it is possible to label cells of interest with a bioluminescent marker and use in vivo imaging to assess cell distribution. Other useful imaging modalities include ultrasound, which can assess potential tumorigenicity, and DEXA scans, which provide information on muscle mass and bone mineral density. Both of these latter analytical techniques can be used in the preclinical phase as well as in clinical settings.

6. Do all cell therapies (CAR-Ts, eTCRs, NKs and TILs) require in vivo biodistribution studies?

Yes – biodistribution studies are one of the tenets of IND-enabling programs for all cell (and gene) therapies. Biodistribution studies provide a detailed understanding of where the administered cells distribute within the body, and potentially their status (e.g. differentiated cells, tumor formation, etc). These dynamics are important to confirm that the administered cellular medication is correctly maintained, if it is rejected, expanding as expected, the level of persistence, and also to assess what happens if the cells wane.

7. Is the frozen tissue microarray for CAR-T therapies run with the CAR-Ts themselves or with a purified scFv?

Frozen tissue microarrays are generally run with antibody fragments, although they can be run with T-cells themselves, but antibody fragments are the most appropriate approach. Using the purified fragment provides high throughput analysis of cross-reactivity to non-target tissues which is important for assessing off-target binding. The same antibody fragments, or the external antibody domain, can then be used subsequently for microscopic analysis of tissue sample slides.