Cell and gene therapy has phenomenal potential to change lives and has already shown promise in reversing previously incurable and torturous diseases such as b-thalassemia and sickle cell anemia. Unsurprisingly, there has been an explosion of clinical trials to validate new products over the last 5 years. Cell and gene therapies, while miraculous, are incredibly complex and require much more rigorous manufacturing and analytical testing than traditional biologics/large molecule drugs. Safety concerns have plagued the field from its inception and developing today’s therapies contains a multitude of challenges, which could ultimately lead to failed drug development and fewer treatments available to patients who desperately require them. In this blog post, we explore some of the major challenges in the cell and gene therapy world and how single-cell multi-omics analysis can overcome many of these issues.
1. Safety Concerns
The first gene therapy trial was conducted over 20 years ago to treat a four-year-old girl, named Ashanti DeSilva, who had a rare metabolic disorder (called severe combined immune deficiency; SCID). Ashanti is living an active life today, thanks to the experimental treatment she received at the NIH in 1990. While cell and gene therapies present life-changing possibilities, their complexity and lack of deep characterization tools raise serious concerns around safety. This has caused the FDA to issue several clinical holds to trials underway. The FDA has recently discussed safety issues, such as identifying if and where viral genomes have integrated into the host cell and how many times they have done so (vector copy number). However, the methods of analytical characterization of these drug products often fall short and do not reveal rare but critical events that can cause safety concerns. In addition, as discussed in the next section, the complexity of these new therapeutic approaches makes it challenging, if not impossible to standardize assays and individual drug companies are often forced to identify the most fit-for-purpose methodology and present it to regulators for approval.
Ashanti DeSilva, the first recipient of a gene therapy.
Source: 25 Hour News
2. Inherent Complexity
The field of cell and gene therapies encompasses a wide range of approaches to treating disease, including introducing genes using adeno-associated viruses (AAV) to replace a faulty gene, modifying patient cells to introduce new biology (such as chimeric antigen receptor-T cell therapy or CAR-T) or creating new cell populations from induced pluripotent stem cells (iPSCs) to replace non-functional host cells. Each of these approaches brings unique challenges and can be compounded by the manipulation of cells ex-vivo. This includes genomic modification using techniques such as CRISPR to introduce a function, enhance efficacy, or improve safety. Due to limited sample sizes, it is often difficult to interrogate the entire population of cells or materials to be introduced into patients. This can have deleterious effects, especially when leveraging a powerful genomic editing technique like CRISPR that can cause rearrangements and create unintended genetic changes that could contribute to tumor growth. Regardless of the therapy type, all of these modalities are not synthetic entities and do not function like standard biologics, rather they have cellular-wide effects that could have many biological influences.
3. Short Timelines
Unlike biologics and oral solid dose drug products, current cell and gene therapies are often patient-specific and/or time-sensitive. This reduces the timeframe to comprehensively test the drug product to ensure it is safe and stable. Identification of transduction efficiency and viral copy number often involves creating clonal populations, which requires cells to be sparsely grown for several weeks to form single-cell derived populations, which can take several weeks. During this time, the often very sick patient could be deteriorating quickly. For some assays, such as karyotyping, it can take 2-4 weeks for results to be released and these assays are exceedingly manual and low-throughput.
4. Many Assays & a Lack of Standardization
The FDA requires multiple, different yet complementary assays to show the drug product is pure, safe and potent. However, given the complexity and composition of these new therapeutics, it can be difficult to interpret how the results from one assay correspond to another. Many assays require normalization using a standard curve, which is an indirect measure and could make data integration more difficult. Given the precious nature of the sample, especially for autologous therapies, researchers often do not have many cells to spare for analytical assays, leading to small numbers of cells being analyzed as a surrogate for the entire sample. For example, for karyotyping, which assesses chromosomal structure, often only 20 cells are analyzed as a representation of a whole sample – that could contain millions of cells. As mentioned above, compounding this issue is the lack of standardization within the field, limiting the number of standard procedures that can be performed for similar cell and gene therapies.
Single-cell Analysis Can Help
With new innovative single-cell technologies, it is now possible to measure multiple cell characteristics, such as DNA sequence and protein expression, simultaneously within the same cells. Such advancements have the potential to greatly improve the analytical characterization of cell and gene therapies, in terms of time, simplicity, and sensitivity.
For example, as discussed above, many conventional assays can take weeks to provide results. However, because single-cell analysis does not require clonal isolation, a typical 4-6 week timeframe is decreased to just two days. In addition, as multiple readouts are available from the same cell, the ability to reconcile the data from these two different assays is much simpler, due to a reduction in the variability from trying to analyze two separate cell populations with two different assays.
Lastly, many assays provide in-direct and low sensitivity analysis of aspects such as transduction efficiency, on/off-target effects, and viral copy number. With single-cell technology, even rare events can be identified and genomic changes directly visualized. This is incredibly important, as the vast majority of safety issues plaguing the current therapies are rare events, which require sensitive yet efficient analytical assays to identify and triage.