Immune-regulated IDO1-dependent tryptophan metabolism is source of one-carbon units for pancreatic cancer and stellate cells

•IFNγ and attachment-independent growth promote IDO1 expression in PDAC cells•Metabolism of tryptophan by contributes one-carbons to the THF cycle•Tryptophan can substitute for serine as a one-carbon source•Serine glycine restriction enhances anti-tumor activity an inhibitor Cancer cells adapt their metabolism support elevated energetic anabolic demands proliferation. Folate-dependent is critical metabolic process underpinning cellular proliferation supplying carbons synthesis nucleotides incorporated into DNA RNA. Recent research has focused on nutrients that supply folate cycle, particularly serine. Tryptophan theoretical source units through IDO1, enzyme intensively investigated context tumor immune evasion. Using vitro vivo pancreatic cancer models, we show highly dependent, influenced canonical activator IFNγ. In IDO1-expressing cells, bona fide donor purine nucleotide vivo. 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This surprising, given outputs known importance metabolism: reactive oxygen species (superoxide), metabolism, NAD(P)+, alanine, entry (via α-ketoadipate) tricarboxylic (TCA) present study, sought investigate consequences ductal adenocarcinoma (PDAC). tumors extremely aggressive, clinical outcomes. Characteristically, exhibit hypovascularization deranged contain large proportion complex stroma. Non-cancerous stromal provision alanine (Sousa 2016Sousa Biancur D.E. Halbrook C.J. Sherman M.H. Kremer Hwang R.F. Witkiewicz Ying al.Pancreatic autophagic secretion.Nature. 536: 479-483Crossref (494) Unlike other PDAC-bearing (KPC) mice unresponsive (Maddocks There evidence may enhanced additional contributing factor ability alternative sources tryptophan. Intriguingly, recently one most depleted interstitial fluid KPC (Sullivan 2019aSullivan Danai L.V. Gui D.Y. Vander Heiden Quantification metabolites murine determinants nutrient availability.eLife. 8: e44235Crossref (134) Analysis public data shows types, cancer, high-IDO1-expressing subsets, expressed genetically engineered mouse models PDAC. We find well represented standard culture conditions, induced IFNγ attachment conditions. when promotes generation Under act combination increase epacadostat. formate released (a component stroma) capture exogenously and, similarity channel To perform analyses expressing physiologically relevant evaluate IDO1/Ido1 across contexts (Figure 1A). Direct analysis Pdx1-Cre;LSL-KrasG12D/+;Trp53fl/+ Pdx1-Cre;LSL-KrasG12D/+;LSL-Trp53R172H/+ showed Ido1 versus normal pancreas tissue, certain high (Figures 1B 1C). Compared model (GEMM) tumor-derived primary cultured under conditions displayed undetectable 1D). Addition IFNγ, Ido1, vitro. expected, human did affect assess could restore expression, injected mixed background pure C57Bl6/J subcutaneous allografts three separate experiments. first experiment, very immunocompromised CD1-nude contrast, second 3 lines immunocompetent syngeneic instances strong 2 S1A), albeit intra-animal variability, seen autochthonous Figure 1B. For third implanted (C57Bl6/J) (CD-1 nude) simultaneously compared (using line no. western blot control expression) 1E). side-by-side experiment clearly lack mice, robust comparable +IFNγ. suggest (immunocompetent), growth, there sufficient immune-dependent signaling drive recapitulated similar supplementing 1 ng/mL 1E S1A). infiltration microenvironment observed phenomenon, vary considerably tumor. Genomic demonstrates macrophages (both secretion; Gao 2003Gao W. Pan Scully Girardi Augenlicht L.H. 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Opitz Sahm Trump Winter Ochs Lutz Liu al.Constitutive IDO sustained autocrine loop involving IL-6, STAT3 AHR.Oncotarget. 1038-1051Crossref (166) correlates worse 1F). Next, assessed panel Similar predicted MERAV data, addition (human form) consistently 1G). upregulated during formation setting. Given interactions 2A), immune-independent stimuli expression. potentially linked two ways: (1) mitochondrial production superoxide (Murphy, 2009Murphy M.P. How mitochondria produce species.Biochem. 2009; 417: 1-13Crossref (4507) (2) TCA α-ketoadipate. Exposure rotenone—both oxidative phosphorylation (OXPHOS) modulate levels—had little impact 2B 2C). Similarly, substitution glucose galactose (to OXPHOS) 2D). Unexpectedly, found transferring two-dimensional (2D) monolayer AI 3D (without any adjustments conditions) caused BxPC-3, CFPAC-1, HPAF-II, SU.86.86 2E, 2F, S2A, S2B). accompanied dramatic measured efflux, ablated 2G). While less than establish mechanisms were independent. Treatment proteasome MG132 S2C) lysosomal bafilomycin S2D) no effect levels. degradation proteasomal systems. JAK/STAT cascade (Mojic 2017Mojic Takeda Hayakawa Dark Side IFN-γ: Its Promoting Immunoevasion.Int. Mol. Sci. 19: 89Crossref (114) questioned activate leading upon 1H S2E), indicating activation. appeared specific STAT3, detected STAT1 S2F S2G). Upregulation AI-grown blocked JAK (JAKi) 2H S2E). stimulates both activation, activates only smaller bind promoter transcription (Sun 2009Sun Chin Y.E. Weisiger Malter Tawara Toubai Gatza Mascagni Dinarello Reddy Cutting edge: negative acetylation nonhistone STAT-3.J. 182: 5899-5903Crossref (111) 2014Yu Yan Li Zhao Ren Noncanonical NF-κB STAT3-stimulated upregulation myeloid-derived suppressor cancer.J. 193: 2574-2586Crossref (117) confirm level. qRT-PCR confirmed S2H). During number formed important roles 2A). presence 13C11-tryptophan performed liquid chromatography mass spectrometry (LC-MS) analysis. readily took up 13C11-tryptophan, over 24 h, pool fully labeled. fraction labeling 3A S3B). Downstream small amount identified alanine; however, fraction. metabolized acetyl-coenzyme (CoA) routes: α-ketoadipate acetyl group), (which ring) S3C). substantial acetyl-CoA, S3A S3B), illustrating acetyl-CoA labeled ring coenzyme A, rather group low/negligible kynurenine-dependent (m+6) NAD/H NADP/H. tryptophan, destination From there, carbon pathways, clear entering biosynthesis legitimate Another THF-dependent fat