Background
Pneumonitis is an immune-related adverse event (irAE) associated with immune checkpoint inhibitor (ICI) therapy that is increasing in incidence and carries a risk of hospitalisation, treatment complexity and mortality. Biomarker monitoring, early identification and novel, mechanistically driven treatment options would improve outcomes.
Methods
Patients enrolled on the translational HYST study (IRAS 105915) who developed pneumonitis and other irAEs were identified. Samples taken at the onset of post-ICI irAEs were evaluated along with patient-matched baseline samples and tolerant controls. Following PBMC gDNA and RNA extraction, high throughput sequencing of the TCR and BCR adaptome was conducted using iR-RepSeq+.1 RNA also underwent immunophenotyping. Libraries were multiplexed and sequenced on the Illumina NextSeq 1000 P1 600-cycle. iRepertoire IR-Map software and Kruskal-Wallis testing were used for analysis.
Results
Samples were grouped: baseline ‘no-tox’ (n = 5), baseline ‘tox’ (n = 10), post-treatment ‘no-tox’ (n = 5), and post-treatment ‘tox’ (n = 10). Within the two ‘tox’ groups there were pneumonitis (n=5) and hepatitis (‘other-tox’) (n=5). Several significant differences between those with and without toxicity were observed including the largest clone percentage in both TRD and IGK and several class-switch recombination indices on IgA, IgD, IgG and IgM (p < 0.05). Compared to ‘no-tox’, both ‘tox’ groups showed a significant subset of expanding and contracting CDR3 peptide sequences for receptor chains TRA, TRB, IGK, and IGL (edgeR, FDR < 0.05) (figures 1,2). The largest clone percentage of TRG showed significant baseline differences. Pneumonitis generally showed a trend towards lower immune repertoire diversity at baseline compared to controls (figure 3). Unsupervised learning of 146 immunophenotyping genes revealed strong two-cluster sample separation with 49 genes showing inter-cluster significance (FDR < 0.05). 5/5 pneumonitis, 4/5 hepatitis and 3/5 baseline pneumonitis samples fell into a single cluster. The most impactful genes included S100A8, ITGAL, CD247, and RUNX3 (FDR = 3.64E-04).
Conclusions
Comparing samples across different timepoints/toxicity status’ regarding pneumonitis/irAEs illustrated four main findings in this exploratory cohort. There was a baseline difference in the largest clone percentage of TRG suggesting the potential for prediction; there were differentiating TCR and BCR repertoire cohorts between ‘tox’ and ‘non-tox’ groups; only the ‘tox’ group illustrated repertoire time-dynamic changes and delineating signals in immunophenotyping were suggestive of mechanistic features including S100A8, which is infection-associated and hypothetically, would link pre-infection and pneumonitis. Whilst this is a small sample size there are several significant findings that may be factors in determining predisposition and mechanistic propagation of pneumonitis/irAEs which warrant further investigation.
Acknowledgements
This analysis was funded by F. Hoffmann-La Roche Ltd and Biotech Support provided by iRepertoire Inc.
Reference
Han J, Lotze MT. The adaptome as a biomarker for assessing cancer immunity and immunotherapy. In Methods in Molecular Biology 2020;2055:369–397. Springer. https://doi.org/10.1007/978-1-4939-9773-2_2.
Ethics Approval
Ethic approval within A Mechanistic Investigation into Drug and Chemical Induced Hypersensitivity Reactions (IRAS 105915) with UK Ethics COmmittee. All samples were obtained from patients following informed, written consent.
Abstract 1174 Figure 1
Clonal dynamics looking at the number of significantly expanding or contracting CDR3 peptides
Abstract 1174 Figure 2
TCR/BCR methodology
Abstract 1174 Figure 3
Illustrating diversity by group. For diversity and richness calculations a bootstrapping approach was used utilizing unique molecular indexes (UMIs). For each subsample the index is calculated, with the final score being the median of 1000 iterations