Diseases of the Colon & Rectum

, Volume 40, Issue 4, pp 411–419 | Cite as

Flow cytometric DNA ploidy and S-phase fraction correlate with histopathologic indicators of tumor behavior in colorectal carcinoma

  • António E. Pinto
  • Paula Chaves
  • Paulo Fidalgo
  • António G. Oliveira
  • Carlos N. LeitÃo
  • Jorge Soares
Original Contributions


BACKGROUND: The clinical behavior of colorectal carcinoma is highly variable without reliable predictive biomarkers. Previous reports have shown that flow cytometric DNA analysis may provide valuable prognostic information in these tumors. PURPOSE AND METHODS: This study evaluates the DNA ploidy and the S-phase fraction (SPF) on frozen samples obtained from 61 patients with colorectal carcinoma by using flow cytometry, and it correlates the data with histopathologic features known to affect disease prognosis. Tumors were classified using the World Health Organization's histologic criteria and were staged according the American Joint Committee on Cancer's classification system. Grade of the neoplasm, vascular invasion, and perineural tumor spread were evaluated in every case. RESULTS: Fifty-nine percent of tumors were aneuploid and showed statistically significant higher S-phase values than diploid tumors (22.5vs.11.2 percent;P <0.00001). Mean SPF of the whole series was 17.9 (range, 4.2–44.2) percent. A statistically significant association was found between SPF values and histologic grade (P< 0.0016), nodal status (P<0.0007), distant metastasis(P <0.0001), tumor stage (P<0.0001), venous invasion (P< 0.0002), and lymphatic permeation (P< 0.01) but not with perineural growth and infiltration of the neoplasm through the bowel wall (T). DNA ploidy correlated positively with tumor stage (P<0.03), and the association between aneuploidy and advanced stages of the disease was statistically significant. CONCLUSIONS: These findings showed that flow cytometric DNA ploidy and SPF, evaluated in fresh samples, are potentially useful parameters to estimate colorectal carcinoma biopathology. Aneuploidy and high replicative neoplastic activity correlated with histopathologic features that are commonly associated with the prognosis of colorectal carcinoma, being SPF-related to disease dissemination and, therefore, an indicator of clinical relevance.

Key words

DNA ploidy S-phase fraction Flow cytometry Colorectal carcinoma Prognosis 


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  1. 1.
    Beahrs OH. Colorectal cancer staging as a prognostic feature. Cancer 1982;50:2615–7.PubMedGoogle Scholar
  2. 2.
    Chapuis P, Dent O, Fisher R,et al. A multivariate analysis of clinical and pathological variables in prognosis after resection of large bowel cancer. Br J Surg 1985;72:698–702.PubMedGoogle Scholar
  3. 3.
    Steinberg SM, Barkin JS, Kaplan RS, Stablein DM. Prognostic indicators of colon tumors: the Gastrointestinal Tumor Study Group experience. Cancer 1986;57:1866–70.PubMedGoogle Scholar
  4. 4.
    Wiggers T, Arends JW, Schutte B, Volovics L, Bosman FT. A multivariate analysis of pathologic prognostic indicators in large bowel cancer. Cancer 1988;61:386–95.PubMedGoogle Scholar
  5. 5.
    Ponz de Leon M, Sant M, Micheli A,et al. Clinical and pathologic prognostic indicators in colorectal cancer: a population-based study. Cancer 1992;69:626–35.PubMedGoogle Scholar
  6. 6.
    Wolley RC, Schreiber K, Koss LG, Karas M, Sherman A. DNA distribution in human colon carcinomas and its relationship to clinical behavior. J Natl Cancer Inst 1982;69:15–22.PubMedGoogle Scholar
  7. 7.
    Armitage NC, Robins RA, Evans DF, Turner DR, Baldwin RW, Hardcastle JD. The influence of tumor cell DNA abnormalities on survival in colorectal cancer. Br J Surg 1985;72:828–30.PubMedGoogle Scholar
  8. 8.
    Kokal W, Sheibani K, Terz J, Harada JR. Tumor DNA content in the prognosis of colorectal carcinoma. JAMA 1986;255:3123–7.CrossRefPubMedGoogle Scholar
  9. 9.
    Scott N, Wieand H, Moertel C, Cha S, Beart R, Lieber M. Colorectal cancer: Duke's stage, tumor site, preoperative plasma CEA level and patient prognosis related to tumor DNA ploidy pattern. Arch Surg 1987;122:1375–9.Google Scholar
  10. 10.
    Schutte B, Reynders MM, Wiggers T,et al. Retrospective analysis of the prognostic significance of DNA content and proliferative activity in large bowel carcinoma. Cancer Res 1987;47:5494–6.PubMedGoogle Scholar
  11. 11.
    Emdin SO, Stenling R, Roos G. Prognostic value of DNA content in colorectal carcinoma: a flow cytometric study with some methodologic aspects. Cancer 1987;60:1282–7.PubMedGoogle Scholar
  12. 12.
    Armitage NC, Ballantyne KC, Evans DF, Clarke P, Sheffield J, Hardcastle JD. The influence of tumor cell DNA content on survival in colorectal cancer: a detailed analysis. Br J Cancer 1990;62:852–6.PubMedGoogle Scholar
  13. 13.
    Kouri M, Pyrhonen S, Mecklin J,et al. The prognostic value of DNA ploidy in colorectal carcinoma: a prospective study. Br J Cancer 1990;62:976–81.PubMedGoogle Scholar
  14. 14.
    Dean PA, Vernava AM III. Flow cytometric analysis of DNA content in colorectal carcinoma. Dis Colon Rectum 1992;35:95–102.PubMedGoogle Scholar
  15. 15.
    Melamed MR, Enker WE, Banner P, Janov AJ, Kessler G, Darzynkiewicz Z. Flow cytometry of colorectal carcinoma with three-year follow-up. Dis Colon Rectum 1986;29:184–6.PubMedGoogle Scholar
  16. 16.
    Rognum TO, Thorud E, Lund E. Survival of large bowel carcinoma patients with different DNA ploidy. Br J Cancer 1987;56:633–6.PubMedGoogle Scholar
  17. 17.
    Jass JR, Mukawa K, Goh HS, Love SB, Capellaro D. Clinical importance of DNA content in rectal cancer measured by flow cytometry. J Clin Pathol 1989;42:254–9.PubMedGoogle Scholar
  18. 18.
    Hood DL, Petras RE, Edinger M, Fazio V, Tubbs RR. Deoxyribonucleic acid ploidy and cell cycle analysis of colorectal carcinoma by flow cytometry: a prospective study of 137 cases using fresh whole cell suspensions. Am J Clin Pathol 1990;93:615–20.PubMedGoogle Scholar
  19. 19.
    Russo A, Bazan V, Plaja S, Leonardi P, Bazan P. Patterns of DNA-ploidy in operable colorectal carcinoma: a prospective study of 100 cases. J Surg Oncol 1991;48:4–10.PubMedGoogle Scholar
  20. 20.
    Enker WE, Kimmel M, Cibas ES, Cranor ML, Melamed MR. DNA/RNA content and proliferative fractions of colorectal carcinomas: a five-year prospective study relating flow cytometry to survival. J Natl Cancer Inst 1991;83:701–7.PubMedGoogle Scholar
  21. 21.
    Bottger TC, Potratz D, Stockle M, Wellek S, Klupp J, Junginger T. Prognostic value of DNA analysis in colorectal carcinoma. Cancer 1993;72:3579–87.PubMedGoogle Scholar
  22. 22.
    Bauer KD, Bagwell CB, Giaretti W,et al. Consensus review of the clinical utility of DNA flow cytometry in colorectal cancer. Cytometry 1993;14:486–91.CrossRefPubMedGoogle Scholar
  23. 23.
    Merkel DE, McGuire WL. Ploidy, proliferative activity and prognosis: DNA flow cytometry of solid tumors. Cancer 1990;65:1194–205.PubMedGoogle Scholar
  24. 24.
    Morson BC, Sobin LH. Types histologiques des tumeurs intestinales. Genève: World Health Organization, 1976.Google Scholar
  25. 25.
    Beahrs OH, Henson DE, Hutter RV, Kennedy BJ, eds. Manual for staging of cancer. 4th ed. Philadelphia: JB Lippincott, 1992.Google Scholar
  26. 26.
    Deitch AD, Law H, White RD. A stable propidium iodide staining procedure for flow cytometry. J Histochem Cytochem 1982;30:967–72.PubMedGoogle Scholar
  27. 27.
    Vindelov LL, Christensson IJ, Nissen NI. Standardization of high-resolution flow cytometric DNA analyses by simultaneous use of chicken and trout red blood cells as internal reference standards. Cytometry 1983;3:328–31CrossRefPubMedGoogle Scholar
  28. 28.
    Dean PN, Jett JH. Mathematical analysis of DNA distributions derived from flow microfluorometry. J Cell Biol 1974;60:523–7.CrossRefPubMedGoogle Scholar
  29. 29.
    Hiddemann W, Schumann J, Andreeff M,et al. Convention on nomenclature for DNA cytometry. Cytometry 1984;5:445–6.CrossRefGoogle Scholar
  30. 30.
    Crissman JD, Zarbo RJ, Neibylski CD, Corbett T, Weaver D. Flow cytometric DNA analysis of colon adenocarcinoma: a comparative study of preparatory techniques. Mod Pathol 1988;1:198–204.PubMedGoogle Scholar
  31. 31.
    Frierson HF. Flow cytometric analysis of ploidy in solid neoplasms: comparison of fresh tissue with formalinfixed paraffin-embedded specimens. Hum Pathol 1988;19:290–4.PubMedGoogle Scholar
  32. 32.
    Bergers E, van Diest P, Baak J. Reproducibility of semiautomated cell cycle analysis of flow cytometric DNA histograms of fresh breast cancer material. Anal Cell Pathol 1995;8:1–13.PubMedGoogle Scholar
  33. 33.
    Albe X, Vassilakos P, Helfer-Guarnori K,et al. Independent prognostic value of ploidy in colorectal cancer. Cancer 1990;66:1168–75.PubMedGoogle Scholar
  34. 34.
    Remvikos Y, Muleris M, Vieth P, Salmon R, Dutrilaux B. DNA content and genetic evolution of human colorectal adenocarcinoma: a study by flow cytometry and cytogenetic analysis. Int J Cancer 1988;42:539–43.PubMedGoogle Scholar
  35. 35.
    Shankey TV, Rabinovitch PS, Bagwell B,et al. Guidelines for implementation of clinical DNA cytometry. Cytometry 1993;14:472–7.CrossRefPubMedGoogle Scholar
  36. 36.
    Wersto RP, Liblit RL, Koss LG. Flow cytometric DNA analysis of human solid tumors: a review of the interpretation of DNA histograms. Hum Pathol 1991;22:1085–98.CrossRefPubMedGoogle Scholar
  37. 37.
    Frierson HR. The need for improvement in flow cytometric analysis of ploidy and S-phase fraction. Am J Clin Pathol 1991;95:439–41.PubMedGoogle Scholar
  38. 38.
    Wersto RP, Stetler-Stevenson M. Debris compensation of DNA histograms and its effect on S-phase analysis. Cytometry 1995;20:43–52.CrossRefPubMedGoogle Scholar
  39. 39.
    Baldetorp B, Bendahl P, Ferno M,et al. Reproducibility in DNA flow cytometric analysis of breast cancer: comparison of 12 laboratories results for 67 samples homogenates. Cytometry 1995;22:115–27.CrossRefPubMedGoogle Scholar
  40. 40.
    Weaver DL, Bagwell CB, Hitchcox SA,et al. Improved flow cytometric determination of proliferative activity (S-phase fraction) from paraffin-embedded tissue. Am J Clin Pathol 1990;94:576–84.PubMedGoogle Scholar
  41. 41.
    Scott N, Cross D, Plumb MI, Dixon MF, Quirke P. An investigation of different methods of cell cycle analysis by flow cytometry in rectal cancer. Br J Cancer 1992;65:8–10.PubMedGoogle Scholar
  42. 42.
    Silvestrini R, and the SICCAB Group for Quality Control of Cell Kinetic Determinations. Quality control for evaluation of the S-phase fraction by flow cytometry: a multicentric study. Cytometry 1994;18:11–6.CrossRefPubMedGoogle Scholar
  43. 43.
    Bauer KD, Lincoln ST, Vera-Roman JM,et al. Prognostic implications of proliferative activity and DNA aneuploidy in colonic adenocarcinomas. Lab Invest 1987;57:329–35.PubMedGoogle Scholar
  44. 44.
    Quirke P, Dixon MF, Clayden AD,et al. Prognostic significance of DNA aneuploidy and cell proliferation in rectal adenocarcinomas. J Pathol 1987;151:285–91.CrossRefPubMedGoogle Scholar
  45. 45.
    Witzig TE, Loprinzi CL, Gonchoroff NJ,et al. DNA ploidy and cell kinetic measurements as predictors of recurrence and survival in stages B2 and C colorectal adenocarcinoma. Cancer 1991;68:879–88.PubMedGoogle Scholar
  46. 46.
    Bosari S, Lee A, Wiley B, Heatley G, Silverman M. Flow cytometric and image analyses of colorectal adenocarcinomas: a comparative study with clinical correlations. Am J Clin Pathol 1993;99:187–94.PubMedGoogle Scholar
  47. 47.
    Linden MD, Ma CK, Kubus J, Brown RD, Zarbo RJ. Ki-67 and proliferating cell nuclear antigen tumor proliferatives indices in DNA diploid colorectal adenocarcinomas: correlation with histopathologic characteristics and cell cycle analysis with two-color DNA flow cytometry. Am J Clin Pathol 1993;100:206–12.PubMedGoogle Scholar
  48. 48.
    Ngoi SS, Staiano-Coico L, Godwin TA, Wong RJ, De-Cosse JJ. Abnormal DNA ploidy and proliferative patterns in superficial colonic epithelium adjacent to colorectal cancer. Cancer 1990;66:953–9.PubMedGoogle Scholar
  49. 49.
    Sahin AA, Ro JY, Brown RW,et al. Assessment of Ki-67 derived tumor proliferative activity in colorectal carcinomas. Mod Pathol 1994;7:17–22.PubMedGoogle Scholar
  50. 50.
    Streffer C, van Beuningen D, Gross E, Schabronath J, Eigler F, Rebmann A. Predictive assays for the therapy of rectum carcinoma. Radiother Oncol 1986;5:303–10.PubMedGoogle Scholar
  51. 51.
    Lenner P, Roos G, Johansson H, Lindh J, Dige U. NonHodgkin lymphoma: multivariate analysis of prognostic factors including fraction of S-phase cells. Acta Oncol 1987;26:179–83.PubMedGoogle Scholar
  52. 52.
    Christensson B, Lindemalm C, Johansson B, Mellstedt H, Tribukait B, Biberfeld P. Flow cytometric analysis: a prognostic tool in non-Hodgkin's lymphoma. Leuk Res 1989;13:307–14.CrossRefPubMedGoogle Scholar
  53. 53.
    Gansler T, Chatten J, Varello M, Bunin GR, Atkinson B. Flow cytometric DNA analysis of neuroblastoma: correlation with histology and clinical outcome. Cancer 1986;58:2453–8.PubMedGoogle Scholar
  54. 54.
    Tubiana M, Courdi A. Cell proliferation kinetics in human solid tumors: relation to probability of metastatic dissemination and long-term survival. Radiother Oncol 1989;15:1–18.CrossRefPubMedGoogle Scholar
  55. 55.
    Riley RS. Cellular proliferation markers in the evaluation of human cancer. Clin Lab Med 1992;12:163–99.PubMedGoogle Scholar
  56. 56.
    Silvestrini R, Daidone M, Costa A. Cell kinetics of solid tumors with time and its clinical implication. Tumori 1989;75:367–72.PubMedGoogle Scholar
  57. 57.
    Sun XF, Wingren S, Carstensen JM,et al. ras p21 expression in relation to DNA ploidy, S-phase fraction and prognosis in colorectal adenocarcinoma. Eur J Cancer 1991;27:1646–9.PubMedGoogle Scholar
  58. 58.
    Steiner MG, Harlow SP, Colombo E, Bauer KD. Chromosomes 8,12, and 17 copy number in Astler-Coller stage C colon cancer in relation to proliferative activity and DNA ploidy. Cancer Res 1993;53:681–6.PubMedGoogle Scholar

Copyright information

© American Society of Colon and Rectal Surgeons 1997

Authors and Affiliations

  • António E. Pinto
    • 1
  • Paula Chaves
    • 1
  • Paulo Fidalgo
    • 2
  • António G. Oliveira
    • 2
  • Carlos N. LeitÃo
    • 2
  • Jorge Soares
    • 1
  1. 1.Departamento de Patologia MorfológicaInstituto PortuguÊs de OncologiaLisboa CodexPortugal
  2. 2.ServiÇo de GastroenterologiaInstituto PortuguÊs de Oncologia de Francisco GentilLisboaPortugal

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