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Adoptive T cell therapy targeting a widespread inducible product of aberrant mRNA translation

ABSTRACT: Deregulated protein production by oncogenic pathways fuels tumor development. Consequently, the induction of tryptophan shortage by Indoleamine 2,3-Dioxygenase 1 (IDO1) enzyme, following interferon-gamma (IFNg) secretion by activated T cells, induces tryptophan to phenylalanine (W>F) substitutants1,2. These defective protein products are processed and presented on Human Leukocyte Antigen (HLA) molecules as neoepitopes. Here, we addressed the potential of substitutant neoepitopes to improve cancer immunotherapy. Adoptive cancer immunotherapy is an attractive treatment mode but suffers from the lack of widespread immunogenic targets, as somatic genetic aberrations in cancer are mostly private and counter-selected for strong immunogenicity3,4. Using immunopeptidomics, we identified IFNg-inducible common W>F substitutant neoepitopes presented on HLA-A*24:02, an abundant receptor allele in the Asian population that favors binding to peptides with phenylalanine in their sequence. TMBIM6W>F neoepitope from this list showed the broadest and second-highest expressed host gene in cancer transcriptome datasets. Therefore, we screened T cells and identified one TCR (TCRTMBIM6W>F.1) possessing high affinity and specificity towards TMBIM6W>F/HLA-A*24:02, compared to its wild-type counterpart. We further demonstrate that TCRTMBIM6W>F.1 T cells are reactivated by IFNg-mediated tryptophan-depleted cancer cells, provided they express HLA-A*24:02, TMBIM6, and IDO1, and are proficient in peptide presentation. Finally, in vivo xenograft experiments confirmed the ability of TCRTMBIM6W>F.1 T cells to suppress tumor progression. Thus, substitutant neoepitopes are widespread inducible products of aberrant mRNA translation that provide novel means to overcome current limitations in T-cell transfer therapy.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): T Cell, Cell Culture

DISEASE(S): Cell Type Cancer

SUBMITTER: Onno Bleijerveld

LAB HEAD: Onno Bleijerveld

PROVIDER: PXD041893 | Pride | 2025-01-10

REPOSITORIES: pride

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			Action	DRS
	20220525_COL040-00007-HLAIp_R01.raw	Raw
	20220525_COL040-00007-HLAIp_R02.raw	Raw
	20220525_COL040-00008-HLAIp_R01.raw	Raw
	20220525_COL040-00008-HLAIp_R02.raw	Raw
	20220525_COL040-00009-HLAIp_R01.raw	Raw

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Project description:Many tumors escape by activating multiple cellular pathways that induce immunosuppression. One pivotal immune-suppressive mechanism is the production of tryptophan metabolites along the kynurenine pathway by IFNγ-induced IDO1 enzyme production 4-8. Phase III clinical trials using chemical inhibition of IDO1 in combination with PD1 pathway blockade, however, failed to improve melanoma treatment 9-12. This points at an incomplete understanding of the role of IDO1 and the consequent tryptophan degradation on mRNA translation and cancer progression. Here, we investigated the effects of prolonged IFNγ treatment on mRNA translation in melanoma cells by ribosome profiling. Surprisingly, we observed a massive accumulation of ribosomes ~20 amino acids downstream of tryptophan codons (termed here as W-Bumps) along with their expected stalling at the tryptophan codon itself. This indicated ribosomal bypass of the tryptophan codons in the absence of tryptophan. Detailed examination of W-Bumps position and its corresponding peptide sequences pinpointed towards ribosomal frameshifting events and their effects in the ribosome exit tunnel. In particular, W-Bumps strength was associated with the disorderedness level of potential out-of-frame peptides predicted downstream of tryptophan codons. Indeed, reporter assays demonstrated the induction of ribosomal frameshifting, and the generation of trans-frame proteins and their presentation at the cell surface after IFNγ treatment. Proteomics and immunopeptidomics analyses verified the production of IFNγ-induced trans-frame and out-of-frame aberrant peptides and their presentation on HLA class I molecules. Priming of naïve T cells from healthy donors with aberrant peptides resulted in identification of reactive, peptide-specific T cells. Altogether, our results suggest that IFNγ-induced IDO1-mediated tryptophan depletion plays a role in the immune recognition of melanoma cells by contributing to the diversity of the peptidome landscape, and by inducing the presentation of aberrant peptides.

Project description:Intracellular parasites reprogram the host functions for their survival and reproduction. Conversely, the infected host attempts to defend the microbial insult. The extent and relevance of parasite-mediated host response in vivo remains poorly studied. We utilized Eimeria falciformis, an obligate intracellular parasite completing its entire life cycle in the mouse intestinal epithelium, to identify and validate the host determinants of the parasite infection. The most prominent mouse genes induced during the onset of asexual (24 hrs) and sexual (144 hrs) parasite cycle include IFNg-regulated factors, e.g., immunity-related GTPases IRGA6/B6/D/M2/M3, guanylate-binding proteins GBP2/3/5/8, chemokines CxCL9-11 and several enzymes of the kynurenine pathway including indoleamine 2,3-dioxygenase 1 (IDO1). These results indicated a multifarious innate defense (tryptophan catabolism, IRG, GBP, chemokines signaling) mounted by epithelial cells, and a consequential adaptive immune response (chemokines-cytokines signaling, lymphocyte recruitment). A notable increase in the inflammation- and immunity-associated transcripts correlated with the severity of infection and influx of B-cells, T-cells and macrophages to the parasitized tissue. Indeed, parasite growth was enhanced in the animals inhibited for CxCr3, a major chemokine receptor on immune cells. Interestingly, despite a prominent induction, the mouse IRGB6 failed to recognize and disrupt the parasitophorous vacuole in the parasite cultures, implying an immune evasion by E. falciformis. Likewise, the oocyst output was impaired in IFNg-R-/- and IDO1-/- mice, which signifies a subversion of IFNg-signaling by the parasite to promote its growth. In brief, the Eimeria-rodent model shows contrasting roles of IFNg-signaling for parasite development, identifies a retinue of potential host determinants, and epitomizes its efficacy for in vivo parasite-host interaction studies. Microarray experiments were performed as dual-color hybridizations on Agilent mouse whole genome catalog 44K arrays. To compensate for dye-specific effects, a dye-reversal color-swap was applied.

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Adoptive T cell therapy targeting a widespread inducible product of aberrant mRNA translation

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