Advertisement

Molecular Medicine

, Volume 17, Issue 11–12, pp 1374–1382 | Cite as

Intraclonal Complexity in Chronic Lymphocytic Leukemia: Fractions Enriched in Recently Born/Divided and Older/Quiescent Cells

  • Carlo Calissano
  • Rajendra N. Damle
  • Sonia Marsilio
  • Xiao-Jie Yan
  • Sophia Yancopoulos
  • Gregory Hayes
  • Claire Emson
  • Elizabeth J. Murphy
  • Marc K. Hellerstein
  • Cristina Sison
  • Matthew S. Kaufman
  • Jonathan E. Kolitz
  • Steven L. Allen
  • Kanti R. Rai
  • Ivana Ivanovic
  • Igor M. Dozmorov
  • Sergio Roa
  • Matthew D. Scharff
  • Wentian Li
  • Nicholas Chiorazzi
Research Article

Abstract

The failure of chemotherapeutic regimens to eradicate cancers often results from the outgrowth of minor subclones with more dangerous genomic abnormalities or with self-renewing capacity. To explore such intratumor complexities in B-cell chronic lymphocytic leukemia (CLL), we measured B-cell kinetics in vivo by quantifying deuterium (2H)-labeled cells as an indicator of a cell that had divided. Separating CLL clones on the basis of reciprocal densities of chemokine (C-X-C motif) receptor 4 (CXCR4) and cluster designation 5 (CD5) revealed that the CXCR4dimCD5bright (proliferative) fraction contained more 2H-labeled DNA and hence divided cells than the CXCR4brightCD5dim (resting) fraction. This enrichment was confirmed by the relative expression of two cell cycle-associated molecules in the same fractions, Ki-67 and minichromosome maintenance protein 6 (MCM6). Comparisons of global gene expression between the CXCR4dimCD5bright and CXCR4brightCD5dim fractions indicated higher levels of pro-proliferation and antiapoptotic genes and genes involved in oxidative injury in the proliferative fraction. An extended immunophenotype was also defined, providing a wider range of surface molecules characteristic of each fraction. These intraclonal analyses suggest a model of CLL cell biology in which the leukemic clone contains a spectrum of cells from the proliferative fraction, enriched in recently divided robust cells that are lymphoid tissue emigrants, to the resting fraction enriched in older, less vital cells that need to immigrate to lymphoid tissue or die. The model also suggests several targets preferentially expressed in the two populations amenable for therapeutic attack. Finally, the study lays the groundwork for future analyses that might provide a more robust understanding of the development and clonal evolution of this currently incurable disease.

Notes

Acknowledgments

This work was supported in part by the National Cancer Institute/NIH (CA81554) and by philanthropic contributions from The Karches Foundation, Prince Family Foundation, Marks Foundation, Jerome Levy Foundation, Leon Levy Foundation, Andrew and Mona Albert Fund Inc., and Joseph Eletto Leukemia Research Fund. We thank Aarti Damle and The Feinstein Institute’s microarray core facility for gene expression analyses.

Supplementary material

10020_2011_17111374_MOESM1_ESM.pdf (5.7 mb)
Supplementary material, approximately 5878 KB.

References

  1. 1.
    Chiorazzi N, Rai KR, Ferrarini M. (2005) Chronic lymphocytic leukemia. N. Engl. J. Med. 352:804–15.CrossRefGoogle Scholar
  2. 2.
    Zenz T, Mertens D, Kuppers R, Dohner H, Stilgenbauer S. (2010) From pathogenesis to treatment of chronic lymphocytic leukaemia. Nat. Rev. Cancer. 10:37–50.CrossRefGoogle Scholar
  3. 3.
    Shanafelt TD, et al. (2008) Karyotype evolution on fluorescent in situ hybridization analysis is associated with short survival in patients with chronic lymphocytic leukemia and is related to CD49d expression. J. Clin. Oncol. 26:e5–6.CrossRefGoogle Scholar
  4. 4.
    Calissano C, et al. (2009) In vivo intraclonal and interclonal kinetic heterogeneity in B-cell chronic lymphocytic leukemia. Blood. 114:4832–42.CrossRefGoogle Scholar
  5. 5.
    Neese RA, et al. (2002) Measurement in vivo of proliferation rates of slow turnover cells by 2H2O labeling of the deoxyribose moiety of DNA. Proc. Natl. Acad. Sci. U. S. A. 99:15345–50.CrossRefGoogle Scholar
  6. 6.
    Messmer BT, et al. (2005) In vivo measurements document the dynamic cellular kinetics of chronic lymphocytic leukemia B cells. J. Clin. Invest. 115:755–64.CrossRefGoogle Scholar
  7. 7.
    Zupo S, et al. (1994) Expression of CD5 and CD38 by human CD5- B cells: requirement for special stimuli. Eur. J. Immunol. 24:1426–33.CrossRefGoogle Scholar
  8. 8.
    Stein JV, Nombela-Arrieta C. (2005) Chemokine control of lymphocyte trafficking: a general overview. Immunology. 116:1–12.CrossRefGoogle Scholar
  9. 9.
    van Gent R, et al. (2008) In vivo dynamics of stable chronic lymphocytic leukemia inversely correlate with somatic hypermutation levels and suggest no major leukemic turnover in bone marrow. Cancer Res. 68:10137–44.CrossRefGoogle Scholar
  10. 10.
    Defoiche J, et al. (2008) Reduction of B cell turnover in chronic lymphocytic leukaemia. Br. J. Haematol. 143:240–7.CrossRefGoogle Scholar
  11. 11.
    Herishanu Y, et al. (2010) The lymph node microenvironment promotes B-cell receptor signaling, NF-κB activation, and tumor proliferation in chronic lymphocytic leukemia. Blood. 117:563–74.CrossRefGoogle Scholar
  12. 12.
    Stehling-Sun S, Dade J, Nutt SL, DeKoter RP, Camargo FD. (2009) Regulation of lymphoid versus myeloid fate ‘choice’ by the transcription factor Mef2c. Nat. Immunol. 10:289–96.CrossRefGoogle Scholar
  13. 13.
    Yao X, et al. (2010) Promotion of self-renewal of embryonic stem cells by midkine. Acta. Pharmacol. Sin. 31:629–37.CrossRefGoogle Scholar
  14. 14.
    Ochiai K, Muto A, Tanaka H, Takahashi S, Igarashi K. (2008) Regulation of the plasma cell transcription factor Blimp-1 gene by Bach2 and Bcl6. Int. Immunol. 20:453–60.CrossRefGoogle Scholar
  15. 15.
    Basso K, Dalla-Favera R. BCL6: master regulator of the germinal center reaction and key oncogene in B cell lymphomagenesis. Adv. Immunol. 105:193-210.Google Scholar
  16. 16.
    Reif K, Cyster JG. (2000) RGS molecule expression in murine B lymphocytes and ability to down-regulate chemotaxis to lymphoid chemokines. J. Immunol. 164:4720–9.CrossRefGoogle Scholar
  17. 17.
    Wilker PR, et al. (2008) Transcription factor Mef2c is required for B cell proliferation and survival after antigen receptor stimulation. Nat. Immunol. 9:603–12.CrossRefGoogle Scholar
  18. 18.
    Novak AJ, et al. (2004) Expression of BLyS and its receptors in B-cell non-Hodgkin lymphoma: correlation with disease activity and patient outcome. Blood. 104:2247–53.CrossRefGoogle Scholar
  19. 19.
    Fu L, et al. (2009) BAFF-R promotes cell proliferation and survival through interaction with IKKβ and NF-κB/c-Rel in the nucleus of normal and neoplastic B-lymphoid cells. Blood. 113:4627–36.CrossRefGoogle Scholar
  20. 20.
    Zhai Y, et al. (1999) VEGI, a novel cytokine of the tumor necrosis factor family, is an angiogenesis inhibitor that suppresses the growth of colon carcinomas in vivo. FASEB J. 13:181–9.CrossRefGoogle Scholar
  21. 21.
    Coscia M, et al. (2011) IGHV unmutated CLL B cells are more prone to spontaneous apoptosis and subject to environmental prosurvival signals than mutated CLL B cells. Leukemia. 25:828–37.CrossRefGoogle Scholar
  22. 22.
    Porakishvili N, et al. (2011) CD180 functions in activation, survival and cycling of B chronic lymphocytic leukaemia cells. Br. J. Haematol. 153:486–98.CrossRefGoogle Scholar
  23. 23.
    Kay NE, et al. (2005) A recombinant IL-4-Pseudomonas exotoxin inhibits protein synthesis and overcomes apoptosis resistance in human CLL B cells. Leuk. Res. 29:1009–18.CrossRefGoogle Scholar
  24. 24.
    Nakamura M, Shimada K, Konishi N. (2008) The role of HRK gene in human cancer. Oncogene. 27 Suppl 1:S105–1CrossRefGoogle Scholar
  25. 25.
    Brosens JJ, Wilson MS, Lam EW. (2009) FOXO transcription factors: from cell fate decisions to regulation of human female reproduction. Adv. Exp. Med. Biol. 665:227–41.CrossRefGoogle Scholar
  26. 26.
    Samuel-Mendelsohn S, et al. (2011) Leptin signaling and apoptotic effects in human prostate cancer cell lines. Prostate. 71:929–45.CrossRefGoogle Scholar
  27. 27.
    Ma F, Zhang C, Prasad KV, Freeman GJ, Schlossman SF. (2001) Molecular cloning of Porimin, a novel cell surface receptor mediating oncotic cell death. Proc. Natl. Acad. Sci. U. S. A. 98:9778–83.CrossRefGoogle Scholar
  28. 28.
    Zhou Y, Hileman EO, Plunkett W, Keating MJ, Huang P. (2003) Free radical stress in chronic lymphocytic leukemia cells and its role in cellular sensitivity to ROS-generating anticancer agents. Blood. 101:4098–104.CrossRefGoogle Scholar
  29. 29.
    Parker CM, Cepek KL, Russell GJ, et al. (1992) A family of beta 7 integrins on human mucosal lymphocytes. Proc. Natl. Acad. Sci. U. S. A. 89:1924–8.CrossRefGoogle Scholar
  30. 30.
    Postigo AA, Sanchez-Mateos P, Lazarovits AI, Sanchez-Madrid F, de Landazuri MO. (1993) Alpha 4 beta 7 integrin mediates B cell binding to fibronectin and vascular cell adhesion molecule-1: expression and function of alpha 4 integrins on human B lymphocytes. J. Immunol. 151:2471–83.PubMedGoogle Scholar
  31. 31.
    Trentin L, et al. (1999) The chemokine receptor CXCR3 is expressed on malignant B cells and mediates chemotaxis. J. Clin. Invest. 104:115–21.CrossRefGoogle Scholar
  32. 32.
    Lopez-Giral S, et al. (2004) Chemokine receptors that mediate B cell homing to secondary lymphoid tissues are highly expressed in B cell chronic lymphocytic leukemia and non-Hodgkin lymphomas with widespread nodular dissemination. J. Leukoc. Biol. 76:462–71.CrossRefGoogle Scholar
  33. 33.
    Zlotnik A, Burkhardt AM, Homey B. (2011) Homeostatic chemokine receptors and organ-specific metastasis. Nat. Rev. Immunol. 11:597–606.CrossRefGoogle Scholar
  34. 34.
    Costantini JL, et al. (2009) TAPP2 links phosphoinositide 3-kinase signaling to B-cell adhesion through interaction with the cytoskeletal protein utrophin: expression of a novel cell adhesion-promoting complex in B-cell leukemia. Blood. 114:4703–12.CrossRefGoogle Scholar
  35. 35.
    Kappos L, et al. (2011) Natalizumab treatment for multiple sclerosis: updated recommendations for patient selection and monitoring. Lancet Neurol. 10:745–58.CrossRefGoogle Scholar
  36. 36.
    Wiernik PH, Adiga GU. (2011) Single-agent rituximab in treatment-refractory or poor prognosis patients with chronic lymphocytic leukemia. Curr. Med. Res. Opin. 27:1987–93.CrossRefGoogle Scholar
  37. 37.
    Nightingale G. (2011) Ofatumumab: a novel anti-CD20 monoclonal antibody for treatment of refractory chronic lymphocytic leukemia. Ann. Pharmacother. 45:1248–55.CrossRefGoogle Scholar
  38. 38.
    Rai KR, et al. (2002) Alemtuzumab in previously treated chronic lymphocytic leukemia patients who also had received fludarabine. J. Clin. Oncol. 20:3891–7.CrossRefGoogle Scholar
  39. 39.
    Calandra G, Bridger G, Fricker S. (2010) CXCR4 in clinical hematology. Curr. Top. Microbiol. Immunol. 341:173–91.PubMedGoogle Scholar
  40. 40.
    Cameron F, Whiteside G, Perry C. (2011) Ipilimumab: first global approval. Drugs. 71:1093–104.CrossRefGoogle Scholar
  41. 41.
    Callahan MK, Wolchok JD, Allison JP. (2010) Anti-CTLA-4 antibody therapy: immune monitoring during clinical development of a novel immunotherapy. Semin. Oncol. 37:473–84.CrossRefGoogle Scholar
  42. 42.
    Bajaj M, Heath EI. (2011) Conatumumab: a novel monoclonal antibody against death receptor 5 for the treatment of advanced malignancies in adults. Expert Opin. Biol. Ther. 11:1519–24.CrossRefGoogle Scholar
  43. 43.
    Byrd JC, et al. (2007) Phase 1 study of lumiliximab with detailed pharmacokinetic and pharmacodynamic measurements in patients with relapsed or refractory chronic lymphocytic leukemia. Clin. Cancer Res. 13:4448–55.CrossRefGoogle Scholar
  44. 44.
    Burger M, et al. (2005) Small peptide inhibitors of the CXCR4 chemokine receptor (CD184) antagonize the activation, migration and antiapoptotic responses of CXCL12 in chronic lymphocytic leukemia B cells. Blood. 106:1824–30.CrossRefGoogle Scholar
  45. 45.
    Puri S, et al. (2009) A review of studies on targeting interleukin 4 receptor for central nervous system malignancy. Curr. Mol. Med. 9:732–9.CrossRefGoogle Scholar
  46. 46.
    de Weers M, et al. (2011) Daratumumab, a novel therapeutic human CD38 monoclonal antibody, induces killing of multiple myeloma and other hematological tumors. J. Immunol. 186:1840–8.CrossRefGoogle Scholar
  47. 47.
    Pascual V, et al. (1994) Analysis of somatic mutation in five B cell subsets of human tonsil. J. Exp. Med. 180:329–39.CrossRefGoogle Scholar
  48. 48.
    Damle RN, et al. (1999) Ig V gene mutation status and CD38 expression as novel prognostic indicators in chronic lymphocytic leukemia. Blood. 94:1840–7.PubMedPubMedCentralGoogle Scholar
  49. 49.
    Quiroga MP, Burger JA. (2010) BCR-mediated decrease of CXCR4 and CD62L in CLL. Cancer Res. 70:5194; author reply 5195.CrossRefGoogle Scholar
  50. 50.
    Chiorazzi N, Ferrarini M. (2011) Cellular origin(s) of chronic lymphocytic leukemia: cautionary notes and additional considerations and possibilities. Blood. 117:1781–91.CrossRefGoogle Scholar
  51. 51.
    Griffin DO, Holodick NE, Rothstein TL. (2011) Human B1 cells in umbilical cord and adult peripheral blood express the novel phenotype CD20+ CD27+ CD43+ CD70. J. Exp. Med. 208:67–80.CrossRefGoogle Scholar
  52. 52.
    Fagarasan S, et al. (2000) Mechanism of B1 cell differentiation and migration in GALT. Curr. Top. Microbiol. Immunol. 252:221–9.PubMedGoogle Scholar
  53. 53.
    Dick JE. (2008) Stem cell concepts renew cancer research. Blood. 112:4793–807.CrossRefGoogle Scholar
  54. 54.
    Hayakawa K, Hardy RR, Herzenberg LA, Herzenberg LA. (1985) Progenitors for Ly-1 B cells are distinct from progenitors for other B cells. J. Exp. Med. 161:1554–68.CrossRefGoogle Scholar
  55. 55.
    Chiorazzi N. (2007) Cell proliferation and death: forgotten features of chronic lymphocytic leukemia B cells. Best Pract. Res. Clin. Haematol. 20:399–413.CrossRefGoogle Scholar
  56. 56.
    Burger JA, Ghia P, Rosenwald A, Caligaris-Cappio F. (2009) The microenvironment in mature B-cell malignancies: a target for new treatment strategies. Blood. 114:3367–75.CrossRefGoogle Scholar
  57. 57.
    Alfonso-Perez M, et al. (2006) Anti-CCR7 monoclonal antibodies as a novel tool for the treatment of chronic lymphocyte leukemia. J. Leukoc. Biol. 79:1157–65.CrossRefGoogle Scholar
  58. 58.
    Panayiotidis P, Ganeshaguru K, Jabbar SA, Hoffbrand AV. (1993) Interleukin-4 inhibits apoptotic cell death and loss of the bcl-2 protein in B-chronic lymphocytic leukaemia cells in vitro. Br. J. Haematol. 85:439–45.CrossRefGoogle Scholar
  59. 59.
    Yang JC, et al. (2007) Ipilimumab (anti-CTLA4 antibody) causes regression of metastatic renal cell cancer associated with enteritis and hypophysitis. J. Immunother. 30:825–30.CrossRefGoogle Scholar

Copyright information

© The Feinstein Institute for Medical Research 2011

Authors and Affiliations

  • Carlo Calissano
    • 1
  • Rajendra N. Damle
    • 1
    • 2
    • 10
  • Sonia Marsilio
    • 1
  • Xiao-Jie Yan
    • 1
  • Sophia Yancopoulos
    • 1
  • Gregory Hayes
    • 3
  • Claire Emson
    • 3
  • Elizabeth J. Murphy
    • 3
    • 4
  • Marc K. Hellerstein
    • 3
  • Cristina Sison
    • 5
  • Matthew S. Kaufman
    • 6
    • 10
  • Jonathan E. Kolitz
    • 1
    • 2
    • 10
  • Steven L. Allen
    • 1
    • 2
    • 10
  • Kanti R. Rai
    • 1
    • 6
    • 10
  • Ivana Ivanovic
    • 7
  • Igor M. Dozmorov
    • 7
  • Sergio Roa
    • 8
  • Matthew D. Scharff
    • 8
  • Wentian Li
    • 1
  • Nicholas Chiorazzi
    • 1
    • 2
    • 9
    • 10
  1. 1.The Feinstein Institute for Medical ResearchNorth Shore-LIJ Health SystemManhassetUSA
  2. 2.Department of Medicine, North Shore University HospitalNorth Shore-LIJ Health SystemManhassetUSA
  3. 3.KineMed, Inc.EmeryvilleUSA
  4. 4.Department of MedicineUniversity of CaliforniaSan FranciscoUSA
  5. 5.Biostatistics Unit, The Feinstein Institute for Medical ResearchNorth Shore-LIJ Health SystemManhassetUSA
  6. 6.Department of Medicine, Long Island Jewish Medical CenterNorth Shore-LIJ Health SystemNew Hyde ParkUSA
  7. 7.Oklahoma Medical Research FoundationOklahoma CityUSA
  8. 8.Department of Cell BiologyAlbert Einstein College of MedicineBronxUSA
  9. 9.Department of Molecular MedicineHofstra North Shore-LIJ School of MedicineHempsteadUSA
  10. 10.Hofstra North Shore-LIJ School of MedicineHempsteadUSA

Personalised recommendations