Monitoring immunotherapy response in non-small cell lung cancer patients using 5-hydroxymethylcytosine signatures in circulating cell free DNA

e21505 Background: Liquid biopsies are gaining prominence for not only cancer diagnosis but also patient monitoring. Mutational signals derived from cell-free DNA (cfDNA) show promise to assess response to cancer treatment, including immunotherapy. However, reliance of these methods on mutational data from tissue biopsies limit their applicability when a tumor biopsy is unavailable, or when mutational landscape of tumor changes under the selective pressures of cancer drug treatment. Epigenomic approaches have the potential to address these shortcomings. Methods: Blood draws were obtained from a cohort of non-small cell lung cancer (NSCLC) patients (n = 19) who went on to anti-PD1 treatment prior to therapy start and while on therapy. cfDNA was isolated from plasma and was subsequently processed to generate 5hmC genome-wide profiles. Results: We analyzed cfDNA from NSCLC patients undergoing anti-PD1 therapy to investigate whether immunotherapy response can be detected from plasma. Using a predictive model trained on lung cancer and non-cancer samples, we were able to detect changes in prediction scores in patient treated with immunotherapy that were consistent with RECIST. Patients with progressive disease (n = 3), determined by RECIST, had prediction scores that increased while they received treatment. On the other hand, majority of the patients that exhibited partial response to treatment (n = 12) had predictive scores that decreased with treatment, again consistent with RECIST. Furthermore, score changes consistent with RECIST was observed one cycle prior to the RECIST timepoint in all except one patient, where an extra blood draw after baseline was available (n = 7). Annotation of the regions that account for differential scoring identified enhancer, 5’UTR and promoter regions. Comparison of partial responders to patients with progressive disease revealed genes involved in metastasis, oncogenes and tumor suppressors that change in opposing directions between these patient groups, consistent with the underlying biology. Conclusions: Our results suggest that 5hmC profiles from cfDNA can be used to determine immunotherapy response in non-small cell lung cancer patients. Compared with mutation based liquid biopsy methods to assess response, epigenomics-based methods have the advantage of being agnostic to starting tumor mutations, and not relying on a mutational analysis from tumor biopsy. Future work will help determine applicability of this method to other kinds of therapies and cancer types.