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Multi-Omic Analysis Finds SCID Gene Therapy Rebuilds Patients' Immune Cells, Microbiomes

NEW YORK (GenomeWeb) – A University of Pennsylvania-led team has used a multi-omic approach to examine how gene therapy influences both the immune cell repertoires and microbiomes of patients with X-linked severe combined immunodeficiency.

SCID-X1 affects about 1 in 50,000 to 100,000 newborns, according to the US National Institute of Health. Patients harbor mutations in the IL2RG gene, which encodes a key protein needed for the growth and development of the immune T and NK cells. Because they lack these immune cells, patients experience persistent infections.

Gene therapy with vectors carrying a corrected IL2RG gene has been used to treat some SCID-X1 patients. Researchers led by UPenn's Frederic Bushman analyzed blood, oral, fecal, and nasal samples from 14 patients who took part in one of the two SCID-X1 gene therapy trials, and as they reported yesterday in Genome Medicine, they were able to follow how patients' T cell repertoires and microbiomes changed following treatment.

"In this study, we used several high throughput sequence-based methods to analyze samples from the SCID-X1 trials, with the goal of probing immune mechanisms and the resulting effects on microbial communities," Bushman and his colleagues wrote.

The researchers collected samples from five patients from the first gene therapy trial to treat SCID-X1 (SCIDn1) and nine patients from the second trial, SCIDn2. The second trial, they noted, relied on an improved self-inactivated vector. The researchers also collected samples from six healthy children and three adults for comparison.

With these samples, the researchers sought to determine how gene therapy rebuilt patients' T cell populations. By performing targeted sequencing of viral vector integration sites, they estimated the number of gene-corrected progenitor cells. The number of integration sites varied from 263 to 682 in SCIDn1 patients, while SCIDn2 patients had about 1,000 such sites. Based on this, the researchers estimated the minimal number of progenitor cells to between 144 and 6,018 cells.

Through sequencing the recombined T cell receptor (TCR) beta locus from CD3+ cells, the researchers could begin to tease out the T cell population structure. Healthy adults, they noted, harbored between 18,000 and 27,000 productive rearrangements per sample, while healthy children had between 18,000 and 22,000 per sample. SCIDn2 patients had a similar range of 12,000 to 33,000 per sample, though SCIDn1 patients had a much lower range. They estimated a population size of 26,000 to 2.6 million CDR3 variants and noted that the patients' T cell repertoires became richer over time.

They also estimated that the median number of cell divisions needed to go from a lymphoid progenitor cell to a circulating T cell was about 9.1.

At the same time, the researchers extracted DNA from the swab and stool samples they collected for shotgun metagenomic sequencing and analysis. Bacteroides predominantly colonized the gut microbiomes of healthy subjects, while SCID-X1 patients harbored a range of colonists. One patient was dominated by Bifidobacteria, which is typically found in the guts of breast-fed infants, while two others had Veillonella, which is typically an oral bacterium. As expected, the oral microbiomes of healthy controls contained the expected oral bacteria, though the oral microbiomes of patients included Streptococcus and Rothia.

However, when the researchers analyzed the samples over time, they noted that the patients' microbiomes began to more closely resemble those of the healthy children. This suggested to Bushman and his colleagues that gene therapy helped rebuild patients' immune systems, which then lead to the normalization of their microbiomes.

"This study illustrates some of the uses of multi-omic data in assessing outcome in human gene therapy, including specifying aspects of T cell development and normalization of the microbiota with restoration of T cell function," the authors concluded.

However, they also cautioned that their sample size was small, limiting their ability to make broad conclusions.