High frequency of tumor-infiltrating FOXP3+ regulatory T cells predicts improved survival in mismatch repair-proficient colorectal cancer patients†
Corresponding Author
Daniel M. Frey
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Daniel M. Frey and Raoul A. Droeser contributed equally to the study.
Tel: +41-61-265-25-25, Fax: +41-61-265-72-50
Department of Surgery, University Hospital of Basel, Basel, SwitzerlandSearch for more papers by this authorRaoul A. Droeser
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Daniel M. Frey and Raoul A. Droeser contributed equally to the study.
Search for more papers by this authorCarsten T. Viehl
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorInti Zlobec
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorAlessandro Lugli
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorUrs Zingg
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorDaniel Oertli
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorChristoph Kettelhack
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorLuigi Terracciano
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorLuigi Tornillo
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorCorresponding Author
Daniel M. Frey
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Daniel M. Frey and Raoul A. Droeser contributed equally to the study.
Tel: +41-61-265-25-25, Fax: +41-61-265-72-50
Department of Surgery, University Hospital of Basel, Basel, SwitzerlandSearch for more papers by this authorRaoul A. Droeser
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Daniel M. Frey and Raoul A. Droeser contributed equally to the study.
Search for more papers by this authorCarsten T. Viehl
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorInti Zlobec
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorAlessandro Lugli
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorUrs Zingg
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorDaniel Oertli
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorChristoph Kettelhack
Department of Surgery, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorLuigi Terracciano
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorLuigi Tornillo
Institute of Pathology, University Hospital of Basel, Basel, Switzerland
Search for more papers by this authorThe authors declare that they have no financial competing interests.
Tel: +41-61-265-25-25, Fax: +41-61-265-72-50
Abstract
Regulatory T cells (Treg) inhibit the generation of host-versus-tumor immunity via suppression of tumor-specific effector T-cell responses and development of immune tolerance to neoplastic cells. The transcription factor forkhead box P3 (FOXP3) is an intracellular key molecule for Treg development and function and is considered to represent the most specific Treg cell marker. The aim of this study was to analyze the frequency and prognostic impact of tumor-infiltrating FOXP3+ Treg in colorectal cancer (CRC) stratified by mismatch-repair (MMR) status. Using the tissue microarray technique, 1,420 tumor samples were immunohistochemically stained for FOXP3 and stratified into 1,197 MMR-proficient and 223 MMR-deficient CRCs. Additionally, the 1,197 MMR-proficient CRCs were randomized into 2 subgroups (Test Groups 1 and 2; n = 613 and 584, respectively). In both MMR-proficient CRC subgroups high frequency tumor-infiltrating FOXP3+ Treg was associated with early T stage (p = 0.001 and <0.001), tumor location (p = 0.01 and 0.045) and increased 5-year survival rate (p = 0.004 and <0.001), whereas in MMR-deficient CRCs an association between FOXP3+ Treg and absence of lymph node involvement (p = 0.023), absence of vascular invasion (p = 0.023) and improved 5-year survival rate (p = 0.029) could be detected. In a multivariable analysis including age, gender, T stage, N stage, tumor grade, vascular invasion, and tumor border configuration, a high FOXP3+ Treg frequency was an independent prognostic factor in both MMR-proficient CRC subsets (p = 0.019 and p = 0.007), but not in the MMR-deficient CRCs (p = 0.13). Therefore, high frequency of tumor-infiltrating FOXP3+ Treg is associated with early T stage and independently predicts improved disease-specific survival in MMR-proficient CRC patients.
References
- 1 Pages F, Berger A, Camus M, Sanchez-Cabo F, Costes A, Molidor R, Mlecnik B, Kirilovsky A, Nilsson M, Damotte D, Meatchi T, Bruneval P, et al. Effector memory T cells, early metastasis, and survival in colorectal cancer. N Engl J Med 2005; 353: 2654–66.
- 2 Galon J, Costes A, Sanchez-Cabo F, Kirilovsky A, Mlecnik B, Lagorce-Pages C, Tosolini M, Camus M, Berger A, Wind P, Zinzindohoue F, Bruneval P, et al. Type, density, and location of immune cells within human colorectal tumors predict clinical outcome. Science 2006; 313: 1960–4.
- 3 Popat S, Hubner R, Houlston RS. Systematic review of microsatellite instability and colorectal cancer prognosis. J Clin Oncol 2005; 23: 609–18.
- 4 Sakaguchi S. Naturally arising Foxp3-expressing CD25+CD4+ regulatory T cells in immunological tolerance to self and non-self. Nat Immunol 2005; 6: 345–52.
- 5 Wang HY, Wang RF. Regulatory T cells and cancer. Curr Opin Immunol 2007; 19: 217–23.
- 6 Hori S, Nomura T, Sakaguchi S. Control of regulatory T cell development by the transcription factor Foxp3. Science 2003; 299: 1057–61.
- 7 Wolf D, Wolf AM, Rumpold H, Fiegl H, Zeimet AG, Muller-Holzner E, Deibl M, Gastl G, Gunsilius E, Marth C. The expression of the regulatory T cell-specific forkhead box transcription factor FoxP3 is associated with poor prognosis in ovarian cancer. Clin Cancer Res 2005; 11: 8326–31.
- 8 Bates GJ, Fox SB, Han C, Leek RD, Garcia JF, Harris AL, Banham AH. Quantification of regulatory T cells enables the identification of high-risk breast cancer patients and those at risk of late relapse. J Clin Oncol 2006; 24: 5373–80.
- 9 Curiel TJ, Coukos G, Zou L, Alvarez X, Cheng P, Mottram P, Evdemon-Hogan M, Conejo-Garcia JR, Zhang L, Burow M, Zhu Y, Wei S, et al. Specific recruitment of regulatory T cells in ovarian carcinoma fosters immune privilege and predicts reduced survival. Nat Med 2004; 10: 942–9.
- 10 Petersen RP, Campa MJ, Sperlazza J, Conlon D, Joshi MB, Harpole DH, Jr, Patz EF, Jr. Tumor infiltrating Foxp3+ regulatory T-cells are associated with recurrence in pathologic stage I NSCLC patients. Cancer 2006; 107: 2866–72.
- 11 Liyanage UK, Moore TT, Joo HG, Tanaka Y, Herrmann V, Doherty G, Drebin JA, Strasberg SM, Eberlein TJ, Goedegebuure PS, Linehan DC. Prevalence of regulatory T cells is increased in peripheral blood and tumor microenvironment of patients with pancreas or breast adenocarcinoma. J Immunol 2002; 169: 2756–61.
- 12 Hiraoka N, Onozato K, Kosuge T, Hirohashi S. Prevalence of FOXP3+ regulatory T cells increases during the progression of pancreatic ductal adenocarcinoma and its premalignant lesions. Clin Cancer Res 2006; 12: 5423–34.
- 13 Badoual C, Hans S, Rodriguez J, Peyrard S, Klein C, Agueznay Nel H, Mosseri V, Laccourreye O, Bruneval P, Fridman WH, Brasnu DF, Tartour E. Prognostic value of tumor-infiltrating CD4+ T-cell subpopulations in head and neck cancers. Clin Cancer Res 2006; 12: 465–72.
- 14 Ling KL, Pratap SE, Bates GJ, Singh B, Mortensen NJ, George BD, Warren BF, Piris J, Roncador G, Fox SB, Banham AH, Cerundolo V. Increased frequency of regulatory T cells in peripheral blood and tumour infiltrating lymphocytes in colorectal cancer patients. Cancer Immunol 2007; 7: 7.
- 15 Miller AM, Lundberg K, Ozenci V, Banham AH, Hellstrom M, Egevad L, Pisa P. CD4+CD25high T cells are enriched in the tumor and peripheral blood of prostate cancer patients. J Immunol 2006; 177: 7398–405.
- 16 Grabenbauer GG, Lahmer G, Distel L, Niedobitek G. Tumor-infiltrating cytotoxic T cells but not regulatory T cells predict outcome in anal squamous cell carcinoma. Clin Cancer Res 2006; 12: 3355–60.
- 17 Carreras J, Lopez-Guillermo A, Fox BC, Colomo L, Martinez A, Roncador G, Montserrat E, Campo E, Banham AH. High numbers of tumor-infiltrating FOXP3-positive regulatory T cells are associated with improved overall survival in follicular lymphoma. Blood 2006; 108: 2957–64.
- 18 Salama P, Phillips M, Grieu F, Morris M, Zeps N, Joseph D, Platell C, Iacopetta B. Tumor-infiltrating FOXP3+ T regulatory cells show strong prognostic significance in colorectal cancer. J Clin Oncol 2009; 27: 186–92.
- 19 Sauter G, Simon R, Hillan K. Tissue microarrays in drug discovery. Nat Rev Drug Discov 2003; 2: 962–72.
- 20 Hampel H, Stephens JA, Pukkala E, Sankila R, Aaltonen LA, Mecklin JP, de la Chapelle A. Cancer risk in hereditary nonpolyposis colorectal cancer syndrome: later age of onset. Gastroenterology 2005; 129: 415–21.
- 21 Jass JR, Atkin WS, Cuzick J, Bussey HJ, Morson BC, Northover JM, Todd IP. The grading of rectal cancer: historical perspectives and a multivariate analysis of 447 cases. Histopathology 1986; 10: 437–59.
- 22 Zlobec I, Steele R, Terracciano L, Jass JR, Lugli A. Selecting immunohistochemical cut-off scores for novel biomarkers of progression and survival in colorectal cancer. J Clin Pathol 2007; 60: 1112–6.
- 23 Ohtani H. Focus on TILs: prognostic significance of tumor infiltrating lymphocytes in human colorectal cancer. Cancer Immunol 2007; 7: 4.
- 24 Galon J, Fridman WH, Pages F. The adaptive immunologic microenvironment in colorectal cancer: a novel perspective. Cancer Res 2007; 67: 1883–6.
- 25 Gao Q, Qiu SJ, Fan J, Zhou J, Wang XY, Xiao YS, Xu Y, Li YW, Tang ZY. Intratumoral balance of regulatory and cytotoxic T cells is associated with prognosis of hepatocellular carcinoma after resection. J Clin Oncol 2007; 25: 2586–93.
- 26 Fu J, Xu D, Liu Z, Shi M, Zhao P, Fu B, Zhang Z, Yang H, Zhang H, Zhou C, Yao J, Jin L, et al. Increased regulatory T cells correlate with CD8 T-cell impairment and poor survival in hepatocellular carcinoma patients. Gastroenterology 2007; 132: 2328–39.
- 27 Yang XH, Yamagiwa S, Ichida T, Matsuda Y, Sugahara S, Watanabe H, Sato Y, Abo T, Horwitz DA, Aoyagi Y. Increase of CD4+ CD25+ regulatory T-cells in the liver of patients with hepatocellular carcinoma. J Hepatol 2006; 45: 254–62.
- 28 Loddenkemper C, Schernus M, Noutsias M, Stein H, Thiel E, Nagorsen D. In situ analysis of FOXP3+ regulatory T cells in human colorectal cancer. J Transl Med 2006; 4: 52.
- 29 Nishikawa H, Jager E, Ritter G, Old LJ, Gnjatic S. CD4+ CD25+ regulatory T cells control the induction of antigen-specific CD4+ helper T cell responses in cancer patients. Blood 2005; 106: 1008–11.
- 30 Clarke SL, Betts GJ, Plant A, Wright KL, El-Shanawany TM, Harrop R, Torkington J, Rees BI, Williams GT, Gallimore AM, Godkin AJ. CD4+CD25+FOXP3+ regulatory T cells suppress anti-tumor immune responses in patients with colorectal cancer. PLoS ONE 2006; 1: e129.
- 31 Le Gouvello S, Bastuji-Garin S, Aloulou N, Mansour H, Chaumette MT, Berrehar F, Seikour A, Charachon A, Karoui M, Leroy K, Farcet JP, Sobhani I. High prevalence of Foxp3 and IL17 in MMR-proficient colorectal carcinomas. Gut 2008; 57: 772–9.
- 32 Alvaro T, Lejeune M, Salvado MT, Bosch R, Garcia JF, Jaen J, Banham AH, Roncador G, Montalban C, Piris MA. Outcome in Hodgkin's lymphoma can be predicted from the presence of accompanying cytotoxic and regulatory T cells. Clin Cancer Res 2005; 11: 1467–73.
- 33 Roncador G, Brown PJ, Maestre L, Hue S, Martinez-Torrecuadrada JL, Ling KL, Pratap S, Toms C, Fox BC, Cerundolo V, Powrie F, Banham AH. Analysis of FOXP3 protein expression in human CD4+CD25+ regulatory T cells at the single-cell level. Eur J Immunol 2005; 35: 1681–91.
- 34 Walker MR, Carson BD, Nepom GT, Ziegler SF, Buckner JH. De novo generation of antigen-specific CD4+CD25+ regulatory T cells from human CD4+CD25- cells. Proc Natl Acad Sci USA 2005; 102: 4103–8.
- 35 Wang J, Ioan-Facsinay A, van der Voort EI, Huizinga TW, Toes RE. Transient expression of FOXP3 in human activated nonregulatory CD4+ T cells. Eur J Immunol 2007; 37: 129–38.
- 36 Baker K, Zlobec I, Tornillo L, Terracciano L, Jass JR, Lugli A. Differential significance of tumour infiltrating lymphocytes in sporadic mismatch repair deficient versus proficient colorectal cancers: a potential role for dysregulation of the transforming growth factor-beta pathway. Eur J Cancer 2007; 43: 624–31.
- 37 Michel S, Benner A, Tariverdian M, Wentzensen N, Hoefler P, Pommerencke T, Grabe N, von Knebel Doeberitz M, Kloor M. High density of FOXP3-positive T cells infiltrating colorectal cancers with microsatellite instability. Br J Cancer 2008; 99: 1867–73.
- 38 Gryfe R, Kim H, Hsieh ET, Aronson MD, Holowaty EJ, Bull SB, Redston M, Gallinger S. Tumor microsatellite instability and clinical outcome in young patients with colorectal cancer. N Engl J Med 2000; 342: 69–77.
- 39 Buckowitz A, Knaebel HP, Benner A, Blaker H, Gebert J, Kienle P, von Knebel Doeberitz M, Kloor M. Microsatellite instability in colorectal cancer is associated with local lymphocyte infiltration and low frequency of distant metastases. Br J Cancer 2005; 92: 1746–53.
- 40 Dolcetti R, Viel A, Doglioni C, Russo A, Guidoboni M, Capozzi E, Vecchiato N, Macri E, Fornasarig M, Boiocchi M. High prevalence of activated intraepithelial cytotoxic T lymphocytes and increased neoplastic cell apoptosis in colorectal carcinomas with microsatellite instability. Am J Pathol 1999; 154: 1805–13.
- 41 Lynch HT, Smyrk TC, Watson P, Lanspa SJ, Lynch JF, Lynch PM, Cavalieri RJ, Boland CR. Genetics, natural history, tumor spectrum, and pathology of hereditary nonpolyposis colorectal cancer: an updated review. Gastroenterology 1993; 104: 1535–49.
- 42 Ishikawa T, Fujita T, Suzuki Y, Okabe S, Yuasa Y, Iwai T, Kawakami Y. Tumor-specific immunological recognition of frameshift-mutated peptides in colon cancer with microsatellite instability. Cancer Res 2003; 63: 5564–72.
- 43 Viehl CT, Moore TT, Liyanage UK, Frey DM, Ehlers JP, Eberlein TJ, Goedegebuure PS, Linehan DC. Depletion of CD4+CD25+ regulatory T cells promotes a tumor-specific immune response in pancreas cancer-bearing mice. Ann Surg Oncol 2006; 13: 1252–8.
- 44 Liyanage UK, Goedegebuure PS, Moore TT, Viehl CT, Moo-Young TA, Larson JW, Frey DM, Ehlers JP, Eberlein TJ, Linehan DC. Increased prevalence of regulatory T cells (Treg) is induced by pancreas adenocarcinoma. J Immunother 2006; 29: 416–24.
- 45 Yu P, Lee Y, Liu W, Krausz T, Chong A, Schreiber H, Fu YX. Intratumor depletion of CD4+ cells unmasks tumor immunogenicity leading to the rejection of late-stage tumors. J Exp Med 2005; 201: 779–91.
- 46 Koenen HJ, Smeets RL, Vink PM, van Rijssen E, Boots AM, Joosten I. Human CD25highFoxp3pos regulatory T cells differentiate into IL-17-producing cells. Blood 2008; 112: 2340–52.
- 47 Radhakrishnan S, Cabrera R, Schenk EL, Nava-Parada P, Bell MP, Van Keulen VP, Marler RJ, Felts SJ, Pease LR. Reprogrammed FoxP3+ T regulatory cells become IL-17+ antigen-specific autoimmune effectors in vitro and in vivo. J Immunol 2008; 181: 3137–47.
- 48 Nam JS, Terabe M, Kang MJ, Chae H, Voong N, Yang YA, Laurence A, Michalowska A, Mamura M, Lonning S, Berzofsky JA, Wakefield LM. Transforming growth factor beta subverts the immune system into directly promoting tumor growth through interleukin-17. Cancer Res 2008; 68: 3915–23.
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