Association of microbial metabolism of tryptophan with resistance to immune checkpoint (ICB) therapy in renal cell cancer (RCC).

Background: Metastatic RCC has a poor prognosis. Despite improvement in treatment outcomes with ICB and targeted therapy, many patients fail to respond to first line therapy and durable responses remain elusive. The tryptophan metabolite kynurenine, an agonist of the immunosuppressive aryl hydrocarbon receptor (AhR), is associated with resistance to ICB. However, IDO (Indole 2,3 dioxygenase) blockers which inhibit tryptophan metabolism to kynurenine failed to show benefit in clinical trials, suggesting the presence of alternate AhR activation. We hypothesize that microbial metabolism of tryptophan to indole metabolites may be associated with AhR activation and ICB resistance. Methods: We prospectively collected paired stool and blood samples of treatment naïve metastatic RCC patients, treated with ICB +/- Tyrosine kinase inhibitors (TKI) at treatment initiation and at time of first response assessment (12+/-3 weeks). We evaluated stool metagenomics and untargeted stool and plasma metabolomics among responders (R) and non-responders (NR). We focused on kynurenine/tryptophan and indoles/tryptophan ratio to evaluate differential host and microbial metabolism of tryptophan. A responder was classified as progression free survival (PFS) greater than 6 months. We also performed global metabolomics on tumor and plasma of germ-free (GF) & specific pathogen free (SPF) mice to identify microbial & host metabolites. Results: Among 120 patients accrued, baseline paired samples were analyzed from 99 patients, 39 were treated with combination ICB, while 60 patients were treated with ICB + TKI. Median follow up was 15 months. 65% of patients had PFS > 6 months. Using unpaired t test comparing baseline relative abundance of metabolites between R vs NR - plasma Kynurenine/tryptophan ratio was significantly higher in NR vs R, at baseline and at 3 months (baseline-0.042 vs 0.01, p=0.01, 3 months-0.058 vs 0.02, p=0.02). Microbial metabolites of Tryptophan -Indole pyruvic acid (IPA), Indole carboxylic acid (IcA), and Indole acetaldehyde (IAAld) were differentially associated with ICB resistance and was significantly higher in NR (IcA: 8.0 vs 5.2, p=0.03; IPA: 82.4 vs 64.9, p=0.01; IAAld: 78.7 vs 64.6, p=0.03). We also noted differential enrichment of microbial KEGG enzymes associated with IPA, IAAld and IcA production in the stool of NR patients compared to R. Microbial origin of these metablites was confirmed by its absence in tumor of GF mice. Conclusions: This is first study in metastatic RCC which shows the association of indoles (microbial metabolites of tryptophan) with resistance to ICB. These indoles are known to be potent AhR agonists and are associated with immunosuppression suggesting that microbial metabolism of tryptophan may represent a novel pathway for ICB resistance in RCC patients.

Duke Scholars

Author Andrew John Armstrong Medicine, Medical Oncology

Author Jose Ramon Conejo-Garcia Integrative Immunobiology

Author Daniel James George Medicine, Medical Oncology