Re: “Detection and prognostic relevance of cytokeratin 20 in differentiated and anaplastic thyroid carcinomas by RT-PCR”

Letters to the Editors 123

Surgery Volume 137, Number 1

2. Rha SE, Ha HK, Lee SH, Kim JH, Kim JK, Kim PN, et al. CT and MR imaging findings of bowel ischemia from various primary causes. RadioGraphics 2000;20:29-42. 3. Nyman U, Ivancev K, Lindh M, Uher P. Endovascular treatment of chronic mesenteric ischemia: results of five cases. Cardiovasc Intervent Radiol 1998;21:305-13. 4. Spittler C, Chari V, Husni E, Patyakis J, Li P, Yelis J, et al. Second-look laparoscopy for visceral ischemia facilitated by preinstalled ports. Am Surg 1997;63:732-4. 5. Ogihara S, Yamamura S, Tomono H, Iwabuchi H, Ebihara T, Minagawa Y. Superior mesenteric arterial embolism: treatment by trans-catheter trombo-aspiration. J Gastroenterol 2003;38:272-7. doi:10.1016/j.surg.2004.05.053

Re: ‘‘Detection and prognostic relevance of cytokeratin 20 in differentiated and anaplastic thyroid carcinomas by RT-PCR’’ To the Editors: We read with interest the article by Schmitz-Winnenthal et al1 describing the prognostic relevance of CK20 reverse transcription polymerase chain reaction (RTPCR) assay in thyroid carcinomas. The authors demonstrated that CK20 is a valuable differentiation marker in thyroid carcinomas. On the one hand, CK20 expression in thyroid carcinoma tissue is correlated with well-differentiated tumors and a better prognosis. On the other hand, CK20 expression in circulating cells detected by RT-PCR in the blood of thyroid carcinoma patients is associated with a worse prognosis. We would like to highlight the technical refinements that the authors used to obtain such valuable results. They controlled the illegitimate transcription of CK20 in granulocytes by extracting mononuclear cells before performing RNA extraction. Previous studies demonstrated that CK20 may be of major interest in the detection of circulating epithelial cells in the blood of cancer patients.2 But illegitimate transcription of CK20 in granulocytes3 dampened the enthusiasm for the preliminary results on the prognostic relevance of CK20 in several epithelial cancers, notably in colon cancer.2 We personally studied the illegitimate transcription of CK20 in the blood of 31 colorectal cancer patients and 132 patients without any epithelial cancer, and found that illegitimate transcription was present in about 30% of the healthy controls. Because cutaneous granulocyte cells (Merckel cells) could have contaminated blood at the time of venipuncture, we also analyzed 27 patients with venous central catheterism, and found the same ratio of falsepositivity in this population. These results pinpoint the illegitimate transcription of CK20 in granulocytes as the sole explanation for such discrepant results (Table I). Several techniques are available to avoid the illegitimate transcription of CK20 in granulocytes, including (1) negative selection by withdrawing blood mononuclear cells before performing RNA extraction using either antibody-coupling beads4 or the technique of Schmitz-Winnenthal et al;1 (2) positive selection of circulating tumor cells using specific antiepithelial anti-

Table I. Blood CK20 RT-PCR positivity in different populations Population tested Healthy controls ICU patients Colon cancer patients Stage II Stage III Stage IV

Blood sampling location

Number of positive Number Percent tests of patients positivity

Antecubital

42

132

32

Central catheter Antecubital

8

27

30

15

31

48

Antecubital Antecubital Antecubital

2 3 10

6 8 17

33 38 59

bodies;4 and (3) avoiding selection and studying CK20 RNA quantitative expression, as reported by Schuster et al.5 In quantitative techniques, background noise represented by illegitimate transcription of CK20 in granulocytes might be measured and used to define a clear cutoff level distinguishing healthy people from cancer patients. Above this threshold, circulating CK20-expressing tumor cells might be effectively detected in the blood of epithelial cancer patients. We are therefore actually studying the quantitative technique combined with the sampling method of Schmitz-Winnenthal et al1 to resume our studies of the clinical relevance of CK20 RT-PCR assay in the blood of colorectal cancer patients. Richard Douard, MDa Ste´phane Moutereau, MDb Philippe Wind, MDc Anne Bergera Paul-Henri Cugenec, MDa Sylvan Loric, MD, PhDb General Surgery Unit, AP-HP European Georges Pompidou University Hospital, 20 Rue Leblanc, 75908 Paris, Francea Clinical Biochemistry Laboratory, AP-HP Henri Mondor University Hospital, Cre´teil, Franceb General Surgery Unit, AP-HP Avicenne Hospital, Bobigny, Francec

References 1. Schmitz-Winnenthal FH, Weckauf H, Haufe S, Hinz U, Z’graggen K, Klar E, et al. Detection and prognostic relevance of cytokeratin 20 in differentiated and anaplastic thyroid carcinomas by RT-PCR. Surgery 2003;134:964-72. 2. Funaki NO, Tanaka J, Ohshio G, Onodera H, Maetani S, Imamura M. Cytokeratin 20 mRNA in peripheral venous blood of colorectal carcinoma patients. Br J Cancer 1998;77:1327-32. 3. Jung R, Petersen K, Kruger W, Wolf M, Wagener C, Zander A, et al. Detection of micrometastasis by cytokeratin 20 RT-PCR is limited due to stable background transcription in granulocytes. Br J Cancer 1999;81:870-3. 4. Ellis WJ, Pfitzenmaier J, Colli J, Arfman E, Lange PH, Vessella RL. Detection and isolation of prostate cancer cells from peripheral blood and bone marrow. Urology 2003;61:277-81. 5. Schuster R, Max N, Mann B, Heufelder K, Thilo F, Grone J, et al. Quantitative real-time RT-PCR for detection of disseminated tumor cells in peripheral blood of patients with colorectal cancer using different mRNA markers. Int J Cancer 2004;108:219-27. doi:10.1016/j.surg.2004.07.017