There is an urgent need to develop therapeutics for inflammatory bowel disease (IBD) because up to 40% of patients with moderate-to-severe IBD are not adequately controlled with existing drugs. Glutamate carboxypeptidase II (GCPII) has emerged as a promising therapeutic target. This enzyme is minimally expressed in normal ileum and colon, but it is markedly up-regulated in biopsies from patients with IBD and preclinical colitis models. Here, we generated a class of GCPII inhibitors designed to be gut-restricted for oral administration, and we interrogated efficacy and mechanism using in vitro and in vivo models. The lead inhibitor, (S)-IBD3540, was potent (half maximal inhibitory concentration = 4 nanomolar), selective, gut-restricted (AUCcolon/plasma > 50 in mice with colitis), and efficacious in acute and chronic rodent colitis models. In dextran sulfate sodium-induced colitis, oral (S)-IBD3540 inhibited >75% of colon GCPII activity, dose-dependently improved gross and histologic disease, and markedly attenuated monocytic inflammation. In spontaneous colitis in interleukin-10 (IL-10) knockout mice, once-daily oral (S)-IBD3540 initiated after disease onset improved disease, normalized colon histology, and attenuated inflammation as evidenced by reduced fecal lipocalin 2 and colon pro-inflammatory cytokines/chemokines, including tumor necrosis factor-α and IL-17. Using primary human colon epithelial air-liquid interface monolayers to interrogate the mechanism, we further found that (S)-IBD3540 protected against submersion-induced oxidative stress injury by decreasing barrier permeability, normalizing tight junction protein expression, and reducing procaspase-3 activation. Together, this work demonstrated that local inhibition of dysregulated gastrointestinal GCPII using the gut-restricted, orally active, small-molecule (S)-IBD3540 is a promising approach for IBD treatment.

Department of Inflammation and Immunity Lerner Research Institute Cleveland Clinic Cleveland OH 44195 USA

Department of Medicine Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Department of Molecular and Comparative Pathobiology Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Department of Neurology Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Department of Neuroscience Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Department of Oncology Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Department of Pharmacology and Molecular Sciences Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Department of Psychiatry and Behavioral Sciences Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Institute of Organic Chemistry and Biochemistry Academy of Sciences of the Czech Republic 160 00 Prague Czechia

Johns Hopkins Drug Discovery Johns Hopkins University School of Medicine Baltimore MD 21205 USA

Zobrazit více v PubMed

GBD 2017 Inflammatory Bowel Disease Collaborators, The global, regional, and national burden of inflammatory bowel disease in 195 countries and territories, 1990-2017: A systematic analysis for the Global Burden of Disease Study 2017. Lancet Gastroenterol. Hepatol 5, 17–30 (2020). PubMed PMC

Dahlhamer JM, Zammitti EP, Ward BW, Wheaton AG, Croft JB, Prevalence of inflammatory bowel disease among Adults aged ≥18 years—United States, 2015. MMWR Morb. Mortal. Wkly Rep 65, 1166–1169 (2016). PubMed

Danese S, Allez M, van Bodegraven AA, Dotan I, Gisbert JP, Hart A, Lakatos PL, Magro F, Peyrin-Biroulet L, Schreiber S, Tarabar D, Vavricka S, Halfvarson J, Vermeire S, Unmet medical needs in ulcerative colitis: An expert group consensus. Dig. Dis 37, 266–283 (2019). PubMed

Denson LA, Curran M, McGovern DPB, Koltun WA, Duerr RH, Kim SC, Sartor RB, Sylvester FA, Abraham C, de Zoeten EF, Siegel CA, Burns RM, Dobes AM, Shtraizent N, Honig G, Heller CA, Hurtado-Lorenzo A, Cho JH, Challenges in IBD research: Precision medicine. Inflamm. Bowel Dis 25, S31–S39 (2019). PubMed

Daperno M, Armuzzi A, Danese S, Fries W, Liguori G, Orlando A, Papi C, Principi M, Rizzello F, Viscido A, Gionchetti P, Unmet medical needs in the management of ulcerative colitis: Results of an italian delphi consensus. Gastroenterol. Res. Pract 2019, 3108025 (2019). PubMed PMC

Gordon JP, McEwan PC, Maguire A, Sugrue DM, Puelles J, Characterizing unmet medical need and the potential role of new biologic treatment options in patients with ulcerative colitis and Crohn’s disease: A systematic review and clinician surveys. Eur. J. Gastroenterol. Hepatol 27, 804–812 (2015). PubMed PMC

Chandler CJ, Wang TT, Halsted CH, Pteroylpolyglutamate hydrolase from human jejunal brush borders. Purification and characterization. J. Biol. Chem 261, 928–933 (1986). PubMed

Wang TT, Chandler CJ, Halsted CH, Intracellular pteroylpolyglutamate hydrolase from human jejunal mucosa. Isolation and characterization. J Biol Chem 261, 13551–13555 (1986). PubMed

Troyer JK, Beckett ML, Wright GL Jr., Detection and characterization of the prostate-specific membrane antigen (PSMA) in tissue extracts and body fluids. Int. J. Cancer 62, 552–558 (1995). PubMed

Mhawech-Fauceglia P, Zhang S, Terracciano L, Sauter G, Chadhuri A, Herrmann FR, Penetrante R, Prostate-specific membrane antigen (PSMA) protein expression in normal and neoplastic tissues and its sensitivity and specificity in prostate adenocarcinoma: An immunohistochemical study using mutiple tumour tissue microarray technique. Histopathology 50, 472–483 (2007). PubMed

Haffner MC, Kronberger IE, Ross JS, Sheehan CE, Zitt M, Muhlmann G, Ofner D, Zelger B, Ensinger C, Yang XJ, Geley S, Margreiter R, Bander NH, Prostate-specific membrane antigen expression in the neovasculature of gastric and colorectal cancers. Hum. Pathol 40, 1754–1761 (2009). PubMed

Zhang T, Song B, Zhu W, Xu X, Gong QQ, Morando C, Dassopoulos T, Newberry RD, Hunt SR, Li E, An ileal Crohn’s disease gene signature based on whole human genome expression profiles of disease unaffected ileal mucosal biopsies. PLOS ONE 7, e37139 (2012). PubMed PMC

Noble CL, Abbas AR, Lees CW, Cornelius J, Toy K, Modrusan Z, Clark HF, Arnott ID, Penman ID, Satsangi J, Diehl L, Characterization of intestinal gene expression profiles in Crohn’s disease by genome-wide microarray analysis. Inflamm. Bowel Dis 16, 1717–1728 (2010). PubMed

Ben-Shachar S, Yanai H, Baram L, Elad H, Meirovithz E, Ofer A, Brazowski E, Tulchinsky H, Pasmanik-Chor M, Dotan I, Gene expression profiles of ileal inflammatory bowel disease correlate with disease phenotype and advance understanding of its immunopathogenesis. Inflamm. Bowel Dis 19, 2509–2521 (2013). PubMed

Haberman Y, Tickle TL, Dexheimer PJ, Kim MO, Tang D, Karns R, Baldassano RN, Noe JD, Rosh J, Markowitz J, Heyman MB, Griffiths AM, Crandall WV, Mack DR, Baker SS, Huttenhower C, Keljo DJ, Hyams JS, Kugathasan S, Walters TD, Aronow B, Xavier RJ, Gevers D, Denson LA, Pediatric Crohn disease patients exhibit specific ileal transcriptome and microbiome signature. J. Clin. Invest 124, 3617–3633 (2014). PubMed PMC

Yanai H, Ben-Shachar S, Baram L, Elad H, Gitstein G, Brazowski E, Tulchinsky H, Pasmanik-Chor M, Dotan I, Gene expression alterations in ulcerative colitis patients after restorative proctocolectomy extend to the small bowel proximal to the pouch. Gut 64, 756–764 (2015). PubMed

Perez K, Ngollo M, Rabinowitz K, Hammoudi N, Seksik P, Xavier RJ, Daly MJ, Dotan I, Le Bourhis L, Allez M, Meta-analysis of IBD gut samples gene expression identifies specific markers of ileal and colonic diseases. Inflamm. Bowel Dis 28, 775–782 (2022). PubMed

Rais R, Jiang W, Zhai H, Wozniak KM, Stathis M, Hollinger KR, Thomas AG, Rojas C, Vornov JJ, Marohn M, Li X, Slusher BS, FOLH1/GCPII is elevated in IBD patients, and its inhibition ameliorates murine IBD abnormalities. JCI Insight 1, e88634 (2016). PubMed PMC

Date AA, Rais R, Babu T, Ortiz J, Kanvinde P, Thomas AG, Zimmermann SC, Gadiano AJ, Halpert G, Slusher BS, Ensign LM, Local enema treatment to inhibit FOLH1/GCPII as a novel therapy for inflammatory bowel disease. J. Control. Release 263, 132–138 (2017). PubMed PMC

Peters DE, Norris LD, Slusher BS, Spontaneous loss-of-function Dock2 mutation alters murine colitis sensitivity and is a confounding variable in inflammatory bowel disease research. Crohn’s Colitis 360, otz030 (2019). PubMed PMC

Vornov JJ, Hollinger KR, Jackson PF, Wozniak KM, Farah MH, Majer P, Rais R, Slusher BS, Still NAAG’ing after all these years: The continuing pursuit of GCPII inhibitors. Adv. Pharmacol 76, 215–255 (2016). PubMed

Vornov JJ, Peters D, Nedelcovych M, Hollinger K, Rais R, Slusher BS, Looking for drugs in all the wrong places: Use of GCPII inhibitors outside the brain. Neurochem. Res 45, 1256–1267 (2020). PubMed PMC

Ward JBJ, Lajczak NK, Kelly OB, O’Dwyer AM, Giddam AK, Ni Gabhann J, Franco P, Tambuwala MM, Jefferies CA, Keely S, Roda A, Keely SJ, Ursodeoxycholic acid and lithocholic acid exert anti-inflammatory actions in the colon. Am. J. Physiol. Gastrointest. Liver Physiol 312, G550–G558 (2017). PubMed

Martinez-Moya P, Romero-Calvo I, Requena P, Hernandez-Chirlaque C, Aranda CJ, Gonzalez R, Zarzuelo A, Suarez MD, Martinez-Augustin O, Marin JJ, de Medina FS, Dose-dependent antiinflammatory effect of ursodeoxycholic acid in experimental colitis. Int. Immunopharmacol 15, 372–380 (2013). PubMed

Van den Bossche L, Hindryckx P, Devisscher L, Devriese S, Van Welden S, Holvoet T, Vilchez-Vargas R, Vital M, Pieper DH, Vanden Bussche J, Vanhaecke L, Van de Wiele T, De Vos M, Laukens D, Ursodeoxycholic acid and its taurine- or glycine-conjugated species reduce colitogenic dysbiosis and equally suppress experimental colitis in mice. Appl. Environ. Microbiol 83, e02766–16 (2017). PubMed PMC

Sinha SR, Haileselassie Y, Nguyen LP, Tropini C, Wang M, Becker LS, Sim D, Jarr K, Spear ET, Singh G, Namkoong H, Bittinger K, Fischbach MA, Sonnenburg JL, Habtezion A, Dysbiosis-induced secondary bile acid deficiency promotes intestinal inflammation. Cell Host Microbe 27, 659–670.e5 (2020). PubMed PMC

Hamilton JP, Xie G, Raufman JP, Hogan S, Griffin TL, Packard CA, Chatfield DA, Hagey LR, Steinbach JH, Hofmann AF, Human cecal bile acids: Concentration and spectrum. Am. J. Physiol. Gastrointest. Liver Physiol 293, G256–G263 (2007). PubMed

Devlin AS, Fischbach MA, A biosynthetic pathway for a prominent class of microbiota-derived bile acids. Nat. Chem. Biol 11, 685–690 (2015). PubMed PMC

Pavlidis P, Powell N, Vincent RP, Ehrlich D, Bjarnason I, Hayee B, Systematic review: Bile acids and intestinal inflammation-luminal aggressors or regulators of mucosal defence? Aliment. Pharmacol. Ther 42, 802–817 (2015). PubMed

Baars A, Oosting A, Knol J, Garssen J, van Bergenhenegouwen J, The gut microbiota as a therapeutic target in IBD and metabolic disease: A role for the bile acid receptors FXR and TGR5. Microorganisms 3, 641–666 (2015). PubMed PMC

Heinken A, Ravcheev DA, Baldini F, Heirendt L, Fleming RMT, Thiele I, Systematic assessment of secondary bile acid metabolism in gut microbes reveals distinct metabolic capabilities in inflammatory bowel disease. Microbiome 7, 75 (2019). PubMed PMC

Kruis W, Kalek HD, Stellaard F, Paumgartner G, Altered fecal bile acid pattern in patients with inflammatory bowel disease. Digestion 35, 189–198 (2004). PubMed

Duboc H, Rajca S, Rainteau D, Benarous D, Maubert MA, Quervain E, Thomas G, Barbu V, Humbert L, Despras G, Bridonneau C, Dumetz F, Grill JP, Masliah J, Beaugerie L, Cosnes J, Chazouilleres O, Poupon R, Wolf C, Mallet JM, Langella P, Trugnan G, Sokol H, Seksik P, Connecting dysbiosis, bile-acid dysmetabolism and gut inflammation in inflammatory bowel diseases. Gut 62, 531–539 (2013). PubMed

Lajczak-McGinley NK, Porru E, Fallon CM, Smyth J, Curley C, McCarron PA, Tambuwala MM, Roda A, Keely SJ, The secondary bile acids, ursodeoxycholic acid and lithocholic acid, protect against intestinal inflammation by inhibition of epithelial apoptosis. Physiol. Rep 8, e14456 (2020). PubMed PMC

Wirtz S, Popp V, Kindermann M, Gerlach K, Weigmann B, Fichtner-Feigl S, Neurath MF, Chemically induced mouse models of acute and chronic intestinal inflammation. Nat. Protoc 12, 1295–1309 (2017). PubMed

Vitharana D, France JE, Scarpetti D, Bonneville GW, Majer P, Tsukamoto T, Synthesis and biological evaluation of (R)-and (S)-2-(phosphonomethyl) pentanedioic acids as inhibitors of glutamate carboxypeptidase II. Tetrahedron Asym. 13, 1609–1614 (2002).

Tsukamoto T, Majer P, Vitharana D, Ni C, Hin B, Lu XC, Thomas AG, Wozniak KM, Calvin DC, Wu Y, Slusher BS, Scarpetti D, Bonneville GW, Enantiospecificity of glutamate carboxypeptidase II inhibition. J. Med. Chem 48, 2319–2324 (2005). PubMed

Rais R, Rojas C, Wozniak K, Wu Y, Zhao M, Tsukamoto T, Rudek MA, Slusher BS, Bioanalytical method for evaluating the pharmacokinetics of the GCP-II inhibitor 2-phosphonomethyl pentanedioic acid (2-PMPA). J. Pharm. Biomed. Anal 88, 162–169 (2014). PubMed PMC

Bowes J, Brown AJ, Hamon J, Jarolimek W, Sridhar A, Waldron G, Whitebread S, Reducing safety-related drug attrition: The use of in vitro pharmacological profiling. Nat. Rev. Drug Discov 11, 909–922 (2012). PubMed

Axelsson LG, Landstrom E, Bylund-Fellenius AC, Experimental colitis induced by dextran sulphate sodium in mice: Beneficial effects of sulphasalazine and olsalazine. Aliment. Pharmacol. Ther 12, 925–934 (1998). PubMed

Shin MR, Park HJ, Seo BI, Roh SS, Newapproach of medicinal herbs and sulfasalazine mixture on ulcerative colitis induced by dextran sodium sulfate. World J. Gastroenterol 26, 5272–5270 (2020). PubMed PMC

De Vries LCS, Duarte JM, De Krijger M, Welting O, Van Hamersveld PHP, Van Leeuwen-Hilbers FWM, Moerland PD, Jongejan A, D’Haens GR, De Jonge WJ, Wildenberg ME, A JAK1 selective kinase inhibitor and tofacitinib affect macrophage activation and function. Inflamm. Bowel Dis 25, 647–660 (2019). PubMed

Texler B, Zollner A, Reinstadler V, Reider SJ, Macheiner S, Jelusic B, Pfister A, Watschinger C, Przysiecki N, Tilg H, Oberacher H, Moschen AR, Tofacitinib-induced modulation of intestinal adaptive and innate immunity and factors driving cellular and systemic pharmacokinetics. Cell. Mol. Gastroenterol. Hepatol 13, 383–404 (2022). PubMed PMC

Jones GR, Bain CC, Fenton TM, Kelly A, Brown SL, Ivens AC, Travis MA, Cook PC, MacDonald AS, Dynamics of colon monocyte and macrophage activation during colitis. Front. Immunol 9, 2764 (2018). PubMed PMC

Alex P, Zachos NC, Nguyen T, Gonzales L, Chen TE, Conklin LS, Centola M, Li X, Distinct cytokine patterns identified from multiplex profiles of murine DSS and TNBS-induced colitis. Inflamm. Bowel Dis 15, 341–352 (2009). PubMed PMC

Keubler LM, Buettner M, Hager C, Bleich A, A multihit model: Colitis lessons from the interleukin-10-deficient mouse. Inflamm. Bowel Dis 21, 1967–1975 (2015). PubMed PMC

Iyer SS, Cheng G, Role of interleukin 10 transcriptional regulation in inflammation and autoimmune disease. Crit. Rev. Immunol 32, 23–63 (2012). PubMed PMC

Rennick DM, Fort MM, Lessons from genetically engineered animal models. XII. IL-10-deficient IL-10(−/−) mice and intestinal inflammation. Am. J. Physiol. Gastrointest. Liver Physiol 278, G829–G833 (2000). PubMed

Pearson T, Caporaso JG, Yellowhair M, Bokulich NA, Padi M, Roe DJ, Wertheim BC, Linhart M, Martinez JA, Bilagody C, Hornstra H, Alberts DS, Lance P, Thompson PA, Effects of ursodeoxycholic acid on the gut microbiome and colorectal adenoma development. Cancer Med. 8, 617–628 (2019). PubMed PMC

Winston JA, Rivera A, Cai J, Patterson AD, Theriot CM, Secondary bile acid ursodeoxycholic acid alters weight, the gut microbiota, and the bile acid pool in conventional mice. PLOS ONE 16, e0246161 (2021). PubMed PMC

Yan Q, Wi YM, Thoendel MJ, Raval YS, Greenwood-Quaintance KE, Abdel MP, Jeraldo PR, Chia N, Patel R, Evaluation of the CosmosID bioinformatics platform for prosthetic joint-associated sonicate fluid shotgun metagenomic data analysis. J. Clin. Microbiol 57, e01182–18 (2019). PubMed PMC

Lax S, Smith DP, Hampton-Marcell J, Owens SM, Handley KM, Scott NM, Gibbons SM, Larsen P, Shogan BD, Weiss S, Metcalf JL, Ursell LK, Vázquez-Baeza Y, Van Treuren W, Hasan NA, Gibson MK, Colwell R, Dantas G, Knight R, Gilbert JA, Longitudinal analysis of microbial interaction between humans and the indoor environment. Science 345, 1048–1052 (2014). PubMed PMC

Hasan NA, Young BA, Minard-Smith AT, Saeed K, Li H, Heizer EM, McMillan NJ, Isom R, Abdullah AS, Bornman DM, Faith SA, Choi SY, Dickens ML, Cebula TA, Colwell RR, Microbial community profiling of human saliva using shotgun metagenomic sequencing. PLOS ONE 9, e97699 (2014). PubMed PMC

Mostafa HH, Fissel JA, Fanelli B, Bergman Y, Gniazdowski V, Dadlani M, Carroll KC, Colwell RR, Simner PJ, Metagenomic next-generation sequencing of nasopharyngeal specimens collected from confirmed and suspect COVID-19 patients. MBio 11, e01969–20 (2020). PubMed PMC

Thoendel M, Jeraldo P, Greenwood-Quaintance KE, Yao J, Chia N, Hanssen AD, Abdel MP, Patel R, Comparison of three commercial tools for metagenomic shotgun sequencing analysis. J. Clin. Microbiol 58, e00981–19 (2020). PubMed PMC

Wang Y, Chiang IL, Ohara TE, Fujii S, Cheng J, Muegge BD, Ver Heul A, Han ND, Lu Q, Xiong S, Chen F, Lai CW, Janova H, Wu R, Whitehurst CE, VanDussen KL, Liu TC, Gordon JI, Sibley LD, Stappenbeck TS, Long-term culture captures injury-repair cycles of colonic stem cells. Cell 179, 1144–1159.e15 (2019). PubMed PMC

Sun X, Winglee K, Gharaibeh RZ, Gauthier J, He Z, Tripathi P, Avram D, Bruner S, Fodor A, Jobin C, Microbiota-derived metabolic factors reduce campylobacteriosis in mice. Gastroenterology 154, 1751–1763.e2 (2018). PubMed PMC

Nielsen OH, Gionchetti P, Ainsworth M, Vainer B, Campieri M, Borregaard N, Kjeldsen L, Rectal dialysate and fecal concentrations of neutrophil gelatinase-associated lipocalin, interleukin-8, and tumor necrosis factor-alpha in ulcerative colitis. Am. J. Gastroenterol 94, 2923–2928 (1999). PubMed

Oikonomou KA, Kapsoritakis AN, Theodoridou C, Karangelis D, Germenis A, Stefanidis I, Potamianos SP, Neutrophil gelatinase-associated lipocalin (NGAL) in inflammatory bowel disease: Association with pathophysiology of inflammation, established markers, and disease activity. J. Gastroenterol. 47, 519–530 (2012). PubMed

Chassaing B, Srinivasan G, Delgado MA, Young AN, Gewirtz AT, Vijay-Kumar M, Fecal lipocalin 2, a sensitive and broadly dynamic non-invasive biomarker for intestinal inflammation. PLOS ONE 7, e44328 (2012). PubMed PMC

Hsieh H, Morin J, Filliettaz C, Varada R, LaBarre S, Radi Z, Fecal lipocalin-2 as a sensitive and noninvasive biomarker in the TNBS Crohn’s inflammatory bowel disease model. Toxicol. Pathol 44, 1084–1094 (2016). PubMed

Zdravkovic ND, Jovanovic IP, Radosavljevic GD, Arsenijevic AN, Zdravkovic ND, Mitrovic S, Arsenijevic NN, Potential dual immunomodulatory role of VEGF in ulcerative colitis and colorectal carcinoma. Int. J. Med. Sci 11, 936–947 (2014). PubMed PMC

Scaldaferri F, Vetrano S, Sans M, Arena V, Straface G, Stigliano E, Repici A, Sturm A, Malesci A, Panes J, Yla-Herttuala S, Fiocchi C, Danese S, VEGF-A links angiogenesis and inflammation in inflammatory bowel disease pathogenesis. Gastroenterology 136, 585–595.e5 (2009). PubMed

D’Haens GR, van Deventer S, 25 years of anti-TNF treatment for inflammatory bowel disease: Lessons from the past and a look to the future. Gut 70, 1396–1405 (2021). PubMed

Neurath MF, IL-12 family members in experimental colitis. Mucosal Immunol. 1, S28–S30 (2008). PubMed

Kashani A, Schwartz DA, The expanding role of anti-IL-12 and/or anti-IL-23 antibodies in the treatment of inflammatory bowel disease. Gastroenterol. Hepatol (N Y) 15, 255–265 (2019). PubMed PMC

Fujino S, Andoh A, Bamba S, Ogawa A, Hata K, Araki Y, Bamba T, Fujiyama Y, Increased expression of interleukin 17 in inflammatory bowel disease. Gut 52, 65–70 (2003). PubMed PMC

Guimbaud R, Abitbol V, Bertrand V, Quartier G, Chauvelot-Moachon L, Giroud J, Couturier D, Chaussade DC, Leukemia inhibitory factor involvement in human ulcerative colitis and its potential role in malignant course. Eur. Cytokine Netw 9, 607–612 (1998). PubMed

Zhang YS, Xin DE, Wang Z, Song X, Sun Y, Zou QC, Yue J, Zhang C, Zhang JM, Liu Z, Zhang X, Zhao TC, Su B, Chin YE, STAT4 activation by leukemia inhibitory factor confers a therapeutic effect on intestinal inflammation. EMBO J. 38, 1–20 (2019). PubMed PMC

Kwon JH, Keates AC, Anton PM, Botero M, Goldsmith JD, Kelly CP, Topical antisense oligonucleotide therapy against LIX, an enterocyte-expressed CXC chemokine, reduces murine colitis. Am. J. Physiol. Gastrointest. Liver Physiol 289, G1075–G1083 (2005). PubMed

Uguccioni M, Gionchetti P, Robbiani DF, Rizzello F, Peruzzo S, Campieri M, Baggiolini M, Increased expression of IP-10, IL-8, MCP-1, and MCP-3 in ulcerative colitis. Am. J. Pathol 155, 331–336 (1999). PubMed PMC

Mayer L, Sandborn WJ, Stepanov Y, Geboes K, Hardi R, Yellin M, Tao X, Xu LA, Salter-Cid L, Gujrathi S, Aranda R, Luo AY, Anti-IP-10 antibody (BMS-936557) for ulcerative colitis: A phase II randomised study. Gut 63, 442–450 (2014). PubMed PMC

Ohtsuka Y, Lee J, Stamm DS, Sanderson IR, MIP-2 secreted by epithelial cells increases neutrophil and lymphocyte recruitment in the mouse intestine. Gut 49, 526–533 (2001). PubMed PMC

Shibahara T, Wilcox JN, Couse T, Madara JL, Characterization of epithelial chemoattractants for human intestinal intraepithelial lymphocytes. Gastroenterology 120, 60–70 (2001). PubMed

Kulkarni N, Pathak M, Lal G, Role of chemokine receptors and intestinal epithelial cells in the mucosal inflammation and tolerance. J. Leukoc. Biol 101, 377–394 (2017). PubMed

Marshman E, Ottewell PD, Potten CS, Watson AJ, Caspase activation during spontaneous and radiation-induced apoptosis in the murine intestine. J. Pathol 195, 285–292 (2001). PubMed

Hagiwara C, Tanaka M, Kudo H, Increase in colorectal epithelial apoptotic cells in patients with ulcerative colitis ultimately requiring surgery. J. Gastroenterol. Hepatol 17, 758–764 (2002). PubMed

Iwamoto M, Koji T, Makiyama K, Kobayashi N, Nakane PK, Apoptosis of crypt epithelial cells in ulcerative colitis. J. Pathol 180, 152–159 (1996). PubMed

Di Sabatino A, Ciccocioppo R, Luinetti O, Ricevuti L, Morera R, Cifone MG, Solcia E, Corazza GR, Increased enterocyte apoptosis in inflamed areas of Crohn’s disease. Dis. Colon Rectum 46, 1498–1507 (2003). PubMed

Chen L, Park SM, Turner JR, Peter ME, Cell death in the colonic epithelium during inflammatory bowel diseases: CD95/Fas and beyond. Inflamm. Bowel Dis 16, 1071–1076 (2010). PubMed

Blander JM, Death in the intestinal epithelium-basic biology and implications for inflammatory bowel disease. FEBS J. 283, 2720–2730 (2016). PubMed PMC

Araki Y, Mukaisyo K, Sugihara H, Fujiyama Y, Hattori T, Increased apoptosis and decreased proliferation of colonic epithelium in dextran sulfate sodium-induced colitis in mice. Oncol. Rep 24, 869–874 (2010). PubMed

Kuo WT, Shen L, Zuo L, Shashikanth N, Ong M, Wu L, Zha J, Edelblum KL, Wang Y, Wang Y, Nilsen SP, Turner JR, Inflammation-induced occludin downregulation limits epithelial apoptosis by suppressing caspase-3 expression. Gastroenterology 157, 1323–1337 (2019). PubMed PMC

Zuo G, Zhang T, Huang L, Araujo C, Peng J, Travis Z, Okada T, Ocak U, Zhang G, Tang J, Lu X, Zhang JH, Activation of TGR5 with INT-777 attenuates oxidative stress and neuronal apoptosis via cAMP/PKCε/ALDH2 pathway after subarachnoid hemorrhage in rats. Free Radic. Biol. Med 143, 441–453 (2019). PubMed PMC

Wang XX, Edelstein MH, Gafter U, Qiu L, Luo Y, Dobrinskikh E, Lucia S, Adorini L, D’Agati VD, Levi J, Rosenberg A, Kopp JB, Gius DR, Saleem MA, Levi M, G protein-coupled bile acid receptor TGR5 activation inhibits kidney disease in obesity and diabetes. J. Am. Soc. Nephrol 27, 1362–1378 (2016). PubMed PMC

Zhuang L, Ding W, Zhang Q, Ding W, Xu X, Yu X, Xi D, TGR5 attenuated liver ischemia-reperfusion injury by activating the Keap1-Nrf2 signaling pathway in mice. Inflammation 44, 859–872 (2021). PubMed

Sorrentino G, Perino A, Yildiz E, El Alam G, Bou Sleiman M, Gioiello A, Pellicciari R, Schoonjans K, Bile acids signal via TGR5 to activate intestinal stem cells and epithelial regeneration. Gastroenterology 159, 956–968.e8 (2020). PubMed

Cao Y, Gao Y, Xu S, Bao J, Lin Y, Luo X, Wang Y, Luo Q, Jiang J, Neale JH, Zhong C, Glutamate carboxypeptidase II gene knockout attenuates oxidative stress and cortical apoptosis after traumatic brain injury. BMC Neurosci. 17, 15 (2016). PubMed PMC

Zhang W, Zhang Z, Wu L, Qiu Y, Lin Y, Suppression of glutamate carboxypeptidase II ameliorates neuronal apoptosis from ischemic brain injury. J. Stroke Cerebrovasc. Dis 25, 1599–1605 (2016). PubMed

Queisser A, Hagedorn SA, Braun M, Vogel W, Duensing S, Perner S, Comparison of different prostatic markers in lymph node and distant metastases of prostate cancer. Mod. Pathol 28, 138–145 (2015). PubMed

Huang H, Guma SR, Melamed J, Zhou M, Lee P, Deng FM, NKX3.1 and PSMA are sensitive diagnostic markers for prostatic carcinoma in bone metastasis after decalcification of specimens. Am. J. Clin. Exp. Urol 6, 182–188 (2018). PubMed PMC

Holleran G, Lopetuso L, Petito V, Graziani C, Ianiro G, McNamara D, Gasbarrini A, Scaldaferri F, The innate and adaptive immune system as targets for biologic therapies in inflammatory bowel disease. Int. J. Mol. Sci 18, 10.3390/ijms18102020, (2017). PubMed DOI PMC

Corridoni D, Chapman T, Ambrose T, Simmons A, Emerging mechanisms of innate immunity and their translational potential in inflammatory bowel disease. Front. Med. (Lausanne) 5, 32 (2018). PubMed PMC

Rais R, Wozniak K, Wu Y, Niwa M, Stathis M, Alt J, Giroux M, Sawa A, Rojas C, Slusher BS, Selective CNS uptake of the GCP-II inhibitor 2-PMPA following intranasal administration. PLOS ONE 10, e0131861 (2015). PubMed PMC

Rojas C, Frazier ST, Flanary J, Slusher BS, Kinetics and inhibition of glutamate carboxypeptidase II using a microplate assay. Anal. Biochem 310, 50–54 (2002). PubMed

Robinson MB, Blakely RD, Couto R, Coyle JT, Hydrolysis of the brain dipeptide N-acetyl-l-aspartyl-l-glutamate. Identification and characterization of a novel N-acetylated alpha-linked acidic dipeptidase activity from rat brain. J. Biol. Chem 262, 14498–14506 (1987). PubMed

Erben U, Loddenkemper C, Doerfel K, Spieckermann S, Haller D, Heimesaat MM, Zeitz M, Siegmund B, Kuhl AA, A guide to histomorphological evaluation of intestinal inflammation in mouse models. Int. J. Clin. Exp. Pathol 7, 4557–4576 (2014). PubMed PMC

Nedelcovych MT, Kim BH, Zhu X, Lovell LE, Manning AA, Kelschenbach J, Hadas E, Chao W, Prchalova E, Dash RP, Wu Y, Alt J, Thomas AG, Rais R, Kamiya A, Volsky DJ, Slusher BS, Glutamine antagonist JHU083 normalizes aberrant glutamate production and cognitive deficits in the EcoHIV murine model of HIV-associated neurocognitive disorders. J. Neuroimmune Pharmacology 14, 391–400 (2019). PubMed PMC

Baldwin KT, Carbajal KS, Segal BM, Giger RJ, Neuroinflammation triggered by β-glucan/dectin-1 signaling enables CNS axon regeneration. Proc. Natl. Acad. Sci. U.S.A 112, 2581–2586 (2015). PubMed PMC

VanDussen KL, Marinshaw JM, Shaikh N, Miyoshi H, Moon C, Tarr PI, Ciorba MA, Stappenbeck TS, Development of an enhanced human gastrointestinal epithelial culture system to facilitate patient-based assays. Gut 64, 911–920 (2015). PubMed PMC

Boccellato F, Woelffling S, Imai-Matsushima A, Sanchez G, Goosmann C, Schmid M, Berger H, Morey P, Denecke C, Ordemann J, Meyer TF, Polarised epithelial monolayers of the gastric mucosa reveal insights into mucosal homeostasis and defence against infection. Gut 68, 400–413 (2019). PubMed PMC

Novakova Z, Foss CA, Copeland BT, Morath V, Baranova P, Havlinova B, Skerra A, Pomper MG, Barinka C, Novel monoclonal antibodies recognizing human prostate-specific membrane antigen (PSMA) as research and theranostic tools. Prostate 77, 749–764 (2017). PubMed PMC