Profiling TCRβ Repertoires in NY-ESO-1 T-cell Therapy

Genomic DNA was isolated from FFPE tumor biopsy shavings on an Anaprep 12 nucleic acid extraction platform (BioChain), and from FACS-sorted NY-ESO-1 dextramer-positive T cells from final manufacture products of all patients, and from patient-matched infusion products and post-infusion peripheral PBMCs recovered at day +70 (+/− 10 days) where available, with an AllPrep DNA/RNA isolation kit according to the manufacturer’s instructions (Qiagen). TCRβ alleles were sequenced at 100,000 reads by Adaptive Biotechnologies (Seattle, WA). Productive TCRβ sequences, i.e. those that could be translated into open reading frames and did not contain a stop codon, were reported. Productive clonality and Shannon entropy calculations for each sample were generated as previously described (22 (link)). The transgenic NY-ESO-1 TCR was identified based on comparison of reads with the known TCRβ sequence for the transgenic product.

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Nowicki T.S., Berent-Maoz B., Cheung-Lau G., Huang R.R., Wang X., Tsoi J., Kaplan-Lefko P., Paula C.a.b.r.e.r.a., Tran J., Pang J., Macabali M., Garcilazo I.P., Carretero I.B., Kalbasi A., Cochran A.J., Grasso C.S., Hu-Lieskovan S., Chmielowski B., Comin-Anduix B., Singh A, & Ribas A. (2018). A Pilot Trial of the Combination of Transgenic NY-ESO-1-reactive Adoptive Cellular Therapy with Dendritic Cell Vaccination With or Without Ipilimumab. Clinical cancer research : an official journal of the American Association for Cancer Research, 25(7), 2096-2108.

Publication 2018

AllPrep DNA/RNA isolation kit

Adaptive Alleles Biopsy Clonality Entropy Genomic Isolation Nucleic acid Open reading frames Patient Stop codon T cells Tcrβ Transgenic Tumor

Corresponding Organization : Broad Center

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Variable analysis

independent variables

Isolation of genomic DNA from FFPE tumor biopsy shavings
Isolation of genomic DNA from FACS-sorted NY-ESO-1 dextramer-positive T cells from final manufacture products
Isolation of genomic DNA from patient-matched infusion products
Isolation of genomic DNA from post-infusion peripheral PBMCs recovered at day +70 (+/− 10 days)

dependent variables

Productive TCRβ sequences
Productive clonality
Shannon entropy calculations

control variables

Isolation of genomic DNA using an AllPrep DNA/RNA isolation kit according to the manufacturer's instructions (Qiagen)
TCRβ alleles sequenced at 100,000 reads by Adaptive Biotechnologies (Seattle, WA)

controls

Positive control: Comparison of reads with the known TCRβ sequence for the transgenic NY-ESO-1 TCR
Negative control: Not explicitly mentioned

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