Tumor Biology

, Volume 37, Issue 2, pp 2461–2471 | Cite as

The chemokines CCR1 and CCRL2 have a role in colorectal cancer liver metastasis

  • Israa G. Akram
  • Rania Georges
  • Thomas Hielscher
  • Hassan Adwan
  • Martin R. Berger
Original Article


C-C chemokine receptor type 1 (CCR1) and chemokine C-C motif receptor-like 2 (CCRL2) have not yet been sufficiently investigated for their role in colorectal cancer (CRC). Here, we investigated their expression in rat and human CRC samples, their modulation of expression in a rat liver metastasis model, as well as the effects on cellular properties resulting from their knockdown. One rat and five human colorectal cancer cell lines were used. CC531 rat colorectal cells were injected via the portal vein into rats and re-isolated from rat livers after defined periods. Following mRNA isolation, the gene expression was investigated by microarray. In addition, all cell lines were screened for mRNA expression of CCR1 and CCRL2 by reverse transcription polymerase chain reaction (RT-PCR). Cell lines with detectable expression were used for knockdown experiments; and the respective influence was determined on the cells’ proliferation, scratch closure, and colony formation. Finally, specimens from the primaries of 50 patients with CRC were monitored by quantitative RT-PCR for CCR1 and CCRL2 expression levels. The microarray studies showed peak increases of CCR1 and CCRL2 in the early phase of liver colonization. Knockdown was sufficient at mRNA but only moderate at protein levels and resulted in modest but significant inhibition of proliferation (p < 0.05), scratch closure, and colony formation (p < 0.05). All human CRC samples were positive for CCR1 and CCRL2 and showed a significant pairwise correlation (p < 0.0004), but there was no correlation with tumor stage or age of patients. In summary, the data point to an important role of CCR1 and CCRL2 under conditions of organ colonization and both chemokine receptors qualify as targets of treatment during early colorectal cancer liver metastasis.


Colorectal cancer progression Colorectal cancer cell lines Gene knockdown Liver colonization Chemokine receptors 


Conflicts of interest


Supplementary material

13277_2015_4089_MOESM1_ESM.docx (15 kb)
ESM 1 (DOCX 15 kb)
13277_2015_4089_MOESM2_ESM.docx (15 kb)
ESM 2 (DOCX 14 kb)


  1. 1.
    Brenner H, Kloor M, Pox CP. Colorectal cancer. Lancet. 2014;383:1490–502.CrossRefPubMedGoogle Scholar
  2. 2.
    Kattan MW, Wheeler TM, Scardino PT. Postoperative nomogram for disease recurrence after radical prostatectomy for prostate cancer. J Clin Oncol Off J Am Soc Clin Oncol. 1999;17:1499–507.CrossRefGoogle Scholar
  3. 3.
    Gallagher DJ, Kemeny N. Metastatic colorectal cancer: from improved survival to potential cure. Oncology. 2010;78:237–48.CrossRefPubMedGoogle Scholar
  4. 4.
    Chambers AF, Groom AC, MacDonald IC. Dissemination and growth of cancer cells in metastatic sites. Nat Rev Cancer. 2002;2:563–72.CrossRefPubMedGoogle Scholar
  5. 5.
    Duffy MJ, McGowan PM, Gallagher WM. Cancer invasion and metastasis: changing views. J Pathol. 2008;214:283–93.CrossRefPubMedGoogle Scholar
  6. 6.
    Deryugina EI, Quigley JP. Matrix metalloproteinases and tumor metastasis. Cancer Metastasis Rev. 2006;25:9–34.CrossRefPubMedGoogle Scholar
  7. 7.
    Wang CL, Sun BS, Tang Y, Zhuang HQ, Cao WZ. CCR1 knockdown suppresses human non-small cell lung cancer cell invasion. J Cancer Res Clin Oncol. 2009;135:695–701.CrossRefPubMedGoogle Scholar
  8. 8.
    Lazennec G, Richmond A. Chemokines and chemokine receptors: new insights into cancer-related inflammation. Trends Mol Med. 2010;16:133–44.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Kruizinga RC, Bestebroer J, Berghuis P, de Haas CJ, Links TP, de Vries EG, et al. Role of chemokines and their receptors in cancer. Curr Pharm Des. 2009;15:3396–416.CrossRefPubMedGoogle Scholar
  10. 10.
    Erreni M, Bianchi P, Laghi L, Mirolo M, Fabbri M, Locati M, et al. Expression of chemokines and chemokine receptors in human colon cancer. Methods Enzymol. 2009;460:105–21.CrossRefPubMedGoogle Scholar
  11. 11.
    Horuk R. Chemokine receptor antagonists: overcoming developmental hurdles. Nat Rev Drug Discov. 2009;8:23–33.CrossRefPubMedGoogle Scholar
  12. 12.
    Pease JE, Horuk R. Chemokine receptor antagonists: part 1. Exp Opin Ther Patents. 2009;19:39–58.CrossRefGoogle Scholar
  13. 13.
    Pease JE, Horuk R. Chemokine receptor antagonists: part 2. Exp Opin Ther Patents. 2009;19:199–221.CrossRefGoogle Scholar
  14. 14.
    Strieter RM. Chemokines: not just leukocyte chemoattractants in the promotion of cancer. Nat Immunol. 2001;2:285–6.CrossRefPubMedGoogle Scholar
  15. 15.
    Vandercappellen J, Van Damme J, Struyf S. The role of CXC chemokines and their receptors in cancer. Cancer Lett. 2008;267:226–44.CrossRefPubMedGoogle Scholar
  16. 16.
    Luster AD. Chemokines–chemotactic cytokines that mediate inflammation. N Engl J Med. 1998;338:436–45.CrossRefPubMedGoogle Scholar
  17. 17.
    Wang JM, Deng X, Gong W, Su S. Chemokines and their role in tumor growth and metastasis. J Immunol Methods. 1998;220:1–17.CrossRefPubMedGoogle Scholar
  18. 18.
    Zlotnik A, Yoshie O. Chemokines: a new classification system and their role in immunity. Immunity. 2000;12:121–7.CrossRefPubMedGoogle Scholar
  19. 19.
    D’Ambrosio D, Panina-Bordignon P, Sinigaglia F. Chemokine receptors in inflammation: an overview. J Immunol Methods. 2003;273:3–13.CrossRefPubMedGoogle Scholar
  20. 20.
    Weber C, Weber KS, Klier C, Gu S, Wank R, Horuk R, et al. Specialized roles of the chemokine receptors CCR1 and CCR5 in the recruitment of monocytes and T(H)1-like/CD45RO(+) T cells. Blood. 2001;97:1144–6.CrossRefPubMedGoogle Scholar
  21. 21.
    Fioretti F, Fradelizi D, Stoppacciaro A, Ramponi S, Ruco L, Minty A, et al. Reduced tumorigenicity and augmented leukocyte infiltration after monocyte chemotactic protein-3 (MCP-3) gene transfer: perivascular accumulation of dendritic cells in peritumoral tissue and neutrophil recruitment within the tumor. J Immunol. 1998;161:342–6.PubMedGoogle Scholar
  22. 22.
    Gao JL, Wynn TA, Chang Y, Lee EJ, Broxmeyer HE, Cooper S, et al. Impaired host defense, hematopoiesis, granulomatous inflammation and type 1-type 2 cytokine balance in mice lacking CC chemokine receptor 1. J Exp Med. 1997;185:1959–68.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Revesz L, Bollbuck B, Buhl T, Eder J, Esser R, Feifel R, et al. Novel CCR1 antagonists with oral activity in the mouse collagen induced arthritis. Bioorg Med Chem Lett. 2005;15:5160–4.CrossRefPubMedGoogle Scholar
  24. 24.
    Shahrara S, Proudfoot AE, Woods JM, Ruth JH, Amin MA, Park CC, et al. Amelioration of rat adjuvant-induced arthritis by Met-RANTES. Arthritis Rheum. 2005;52:1907–19.CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Anders HJ, Ninichuk V, Schlondorff D. Progression of kidney disease: blocking leukocyte recruitment with chemokine receptor CCR1 antagonists. Kidney Int. 2006;69:29–32.CrossRefPubMedGoogle Scholar
  26. 26.
    Ninichuk V, Anders HJ. Chemokine receptor CCR1: a new target for progressive kidney disease. Am J Nephrol. 2005;25:365–72.CrossRefPubMedGoogle Scholar
  27. 27.
    Vielhauer V, Berning E, Eis V, Kretzler M, Segerer S, Strutz F, et al. CCR1 blockade reduces interstitial inflammation and fibrosis in mice with glomerulosclerosis and nephrotic syndrome. Kidney Int. 2004;66:2264–78.CrossRefPubMedGoogle Scholar
  28. 28.
    Ajuebor MN, Hogaboam CM, Le T, Proudfoot AE, Swain MG. CCL3/MIP-1alpha is pro-inflammatory in murine T cell-mediated hepatitis by recruiting CCR1-expressing CD4(+) T cells to the liver. Eur J Immunol. 2004;34:2907–18.CrossRefPubMedGoogle Scholar
  29. 29.
    Yang X, Lu P, Fujii C, Nakamoto Y, Gao JL, Kaneko S, et al. Essential contribution of a chemokine, CCL3, and its receptor, CCR1, to hepatocellular carcinoma progression. Int J Cancer J Int Cancer. 2006;118:1869–76.CrossRefGoogle Scholar
  30. 30.
    Reichel CA, Khandoga A, Anders HJ, Schlondorff D, Luckow B, Krombach F. Chemokine receptors Ccr1, Ccr2, and Ccr5 mediate neutrophil migration to postischemic tissue. J Leukoc Biol. 2006;79:114–22.CrossRefPubMedGoogle Scholar
  31. 31.
    Bonville CA, Lau VK, DeLeon JM, Gao JL, Easton AJ, Rosenberg HF, et al. Functional antagonism of chemokine receptor CCR1 reduces mortality in acute pneumovirus infection in vivo. J Virol. 2004;78:7984–9.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Ness TL, Carpenter KJ, Ewing JL, Gerard CJ, Hogaboam CM, Kunkel SL. CCR1 and CC chemokine ligand 5 interactions exacerbate innate immune responses during sepsis. J Immunol. 2004;173:6938–48.CrossRefPubMedGoogle Scholar
  33. 33.
    Yun JJ, Whiting D, Fischbein MP, Banerji A, Irie Y, Stein D, et al. Combined blockade of the chemokine receptors CCR1 and CCR5 attenuates chronic rejection. Circulation. 2004;109:932–7.CrossRefPubMedGoogle Scholar
  34. 34.
    Gao W, Topham PS, King JA, Smiley ST, Csizmadia V, Lu B, et al. Targeting of the chemokine receptor CCR1 suppresses development of acute and chronic cardiac allograft rejection. J Clin Investig. 2000;105:35–44.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Yoshimura T, Oppenheim JJ. Chemokine-like receptor 1 (CMKLR1) and chemokine (C-C motif) receptor-like 2 (CCRL2); two multifunctional receptors with unusual properties. Exp Cell Res. 2011;317:674–84.CrossRefPubMedGoogle Scholar
  36. 36.
    Otero K, Vecchi A, Hirsch E, Kearley J, Vermi W, Del Prete A, et al. Nonredundant role of CCRL2 in lung dendritic cell trafficking. Blood. 2010;116:2942–9.CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Migeotte I, Franssen JD, Goriely S, Willems F, Parmentier M. Distribution and regulation of expression of the putative human chemokine receptor HCR in leukocyte populations. Eur J Immunol. 2002;32:494–501.CrossRefPubMedGoogle Scholar
  38. 38.
    Georges RB, Adwan H, Hamdi H, Hielscher T, Linnemann U, Berger MR. The insulin-like growth factor binding proteins 3 and 7 are associated with colorectal cancer and liver metastasis. Cancer Biol Ther. 2011;12:69–79.CrossRefPubMedGoogle Scholar
  39. 39.
    Georges R, Bergmann F, Hamdi H, Zepp M, Eyol E, Hielscher T, et al. Sequential biphasic changes in claudin1 and claudin4 expression are correlated to colorectal cancer progression and liver metastasis. J Cell Mol Med. 2012;16:260–72.CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Team RDC. R: a language and environment for statistical computing. Vienna, Austria R Foundation for Statistical Computing. 2011.Google Scholar
  41. 41.
    Muller A, Homey B, Soto H, Ge N, Catron D, Buchanan ME, et al. Involvement of chemokine receptors in breast cancer metastasis. Nature. 2001;410:50–6.CrossRefPubMedGoogle Scholar
  42. 42.
    Ghadjar P, Coupland SE, Na IK, Noutsias M, Letsch A, Stroux A, et al. Chemokine receptor CCR6 expression level and liver metastases in colorectal cancer. J Clin Oncol Off J Am Soc Clin Oncol. 2006;24:1910–6.CrossRefGoogle Scholar
  43. 43.
    Ghadjar P, Rubie C, Aebersold DM, Keilholz U. The chemokine CCL20 and its receptor CCR6 in human malignancy with focus on colorectal cancer. Int J Cancer J Int Cancer. 2009;125:741–5.CrossRefGoogle Scholar
  44. 44.
    Rubie C, Frick VO, Pfeil S, Wagner M, Kollmar O, Kopp B, et al. Correlation of IL-8 with induction, progression and metastatic potential of colorectal cancer. World J Gastroenterol WJG. 2007;13:4996–5002.CrossRefPubMedGoogle Scholar
  45. 45.
    Rubie C, Frick VO, Wagner M, Schuld J, Graber S, Brittner B, et al. ELR+ CXC chemokine expression in benign and malignant colorectal conditions. BMC Cancer. 2008;8:178.CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Rubie C, Frick VO, Ghadjar P, Wagner M, Justinger C, Faust SK, et al. Cxc receptor-4 mRNA silencing abrogates cxcl12-induced migration of colorectal cancer cells. J Transl Med. 2011;9:22.CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Brand S, Olszak T, Beigel F, Diebold J, Otte JM, Eichhorst ST, et al. Cell differentiation dependent expressed CCR6 mediates ERK-1/2, SAPK/JNK, and Akt signaling resulting in proliferation and migration of colorectal cancer cells. J Cell Biochem. 2006;97:709–23.CrossRefPubMedGoogle Scholar
  48. 48.
    Juremalm M, Nilsson G. Chemokine receptor expression by mast cells. Chem Immunol Allergy. 2005;87:130–44.CrossRefPubMedGoogle Scholar
  49. 49.
    Schober A, Zernecke A. Chemokines in vascular remodeling. Thromb Haemost. 2007;97:730–7.PubMedGoogle Scholar
  50. 50.
    Williams IR. Chemokine receptors and leukocyte trafficking in the mucosal immune system. Immunol Res. 2004;29:283–92.CrossRefPubMedGoogle Scholar
  51. 51.
    Vaday GG, Peehl DM, Kadam PA, Lawrence DM. Expression of CCL5 (RANTES) and CCR5 in prostate cancer. Prostate. 2006;66:124–34.CrossRefPubMedGoogle Scholar
  52. 52.
    Moller C, Stromberg T, Juremalm M, Nilsson K, Nilsson G. Expression and function of chemokine receptors in human multiple myeloma. Leukemia. 2003;17:203–10.CrossRefPubMedGoogle Scholar
  53. 53.
    Wu X, Fan J, Wang X, Zhou J, Qiu S, Yu Y, et al. Downregulation of CCR1 inhibits human hepatocellular carcinoma cell invasion. Biochem Biophys Res Commun. 2007;355:866–71.CrossRefPubMedGoogle Scholar
  54. 54.
    Shimada T, Matsumoto M, Tatsumi Y, Kanamaru A, Akira S. A novel lipopolysaccharide inducible C-C chemokine receptor related gene in murine macrophages. FEBS Lett. 1998;425:490–4.CrossRefPubMedGoogle Scholar
  55. 55.
    Brouwer N, Zuurman MW, Wei T, Ransohoff RM, Boddeke HW, Biber K. Induction of glial L-CCR mRNA expression in spinal cord and brain in experimental autoimmune encephalomyelitis. Glia. 2004;46:84–94.CrossRefPubMedGoogle Scholar
  56. 56.
    Zuurman MW, Heeroma J, Brouwer N, Boddeke HW, Biber K. LPS-induced expression of a novel chemokine receptor (L-CCR) in mouse glial cells in vitro and in vivo. Glia. 2003;41:327–36.CrossRefPubMedGoogle Scholar
  57. 57.
    Zabel BA, Nakae S, Zuniga L, Kim JY, Ohyama T, Alt C, et al. Mast cell-expressed orphan receptor CCRL2 binds chemerin and is required for optimal induction of IgE-mediated passive cutaneous anaphylaxis. J Exp Med. 2008;205:2207–20.CrossRefPubMedPubMedCentralGoogle Scholar
  58. 58.
    Oostendorp J, Hylkema MN, Luinge M, Geerlings M, Meurs H, Timens W, et al. Localization and enhanced mRNA expression of the orphan chemokine receptor L-CCR in the lung in a murine model of ovalbumin-induced airway inflammation. J Histochem Cytochem Off J Histochem Soc. 2004;52:401–10.CrossRefGoogle Scholar
  59. 59.
    Kitamura T, Kometani K, Hashida H, Matsunaga A, Miyoshi H, Hosogi H, et al. SMAD4-deficient intestinal tumors recruit CCR1+ myeloid cells that promote invasion. Nat Genet. 2007;39:467–75.CrossRefPubMedGoogle Scholar
  60. 60.
    Hirai H, Fujishita T, Kurimoto K, Miyachi H, Kitano S, Inamoto S, et al. CCR1-mediated accumulation of myeloid cells in the liver microenvironment promoting mouse colon cancer metastasis. Clin Exp Metastasis. 2014;31:977–89.CrossRefPubMedPubMedCentralGoogle Scholar
  61. 61.
    Reufsteck C, Lifshitz-Shovali R, Zepp M, Bauerle T, Kubler D, Golomb G, et al. Silencing of skeletal metastasis-associated genes impairs migration of breast cancer cells and reduces osteolytic bone lesions. Clin Exp Metastasis. 2012;29:441–56.CrossRefPubMedGoogle Scholar
  62. 62.
    Kovacheva M, Zepp M, Berger SM, Berger MR. Sustained conditional knockdown reveals intracellular bone sialoprotein as essential for breast cancer skeletal metastasis. Oncotarget. 2014;5:5510–22.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  • Israa G. Akram
    • 1
    • 2
  • Rania Georges
    • 1
    • 3
  • Thomas Hielscher
    • 4
  • Hassan Adwan
    • 1
    • 5
  • Martin R. Berger
    • 1
  1. 1.Toxicology and Chemotherapy UnitGerman Cancer Research Center (DKFZ)HeidelbergGermany
  2. 2.College of Science, Chemistry DepartmentAl-Mustansirya UniversityBaghdadIraq
  3. 3.Department of Pharmacology and ToxicologyAl-Baath UniversityHomsSyria
  4. 4.Department of BiostatisticsGerman Cancer Research Center (DKFZ)HeidelbergGermany
  5. 5.Department of Pharmacology and ToxicologyGerman University of CairoCairoEgypt

Personalised recommendations