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Journal of Clinical Oncology, Vol 24, No 25 (September 1), 2006: pp. 4062-4068 © 2006 American Society of Clinical Oncology. DOI: 10.1200/JCO.2005.04.2739 Lymph Node Status and TS Gene Expression Are Prognostic Markers in Stage II/III Rectal Cancer After Neoadjuvant Fluorouracil-Based Chemoradiotherapy
From the Department of General Surgery, University Medical Center, Göttingen; Institute for Pathology, University of Technology, Dresden; Oncoscreen GmbH, Jena, Germany; and the Department of Biostatistics, Institute for Basic Medical Sciences, University of Oslo, Oslo, Norway. Address reprint requests to Torsten Liersch, MD, Department of General Surgery University Medical Center, Göttingen, Robert-Koch-Str 40, D-37075 Göttingen, Germany; e-mail: tliersc{at}gwdg.de
Purpose According to the CAO/ARO/AIO-94 trial of the German Rectal Cancer Study Group, preoperative combined fluorouracil (FU) -based long-term chemoradiotherapy (CT/RT) is recommended for patients with International Union Against Cancer (UICC) stage II/III rectal cancer. However, despite the local benefit of neoadjuvant treatment, the overall prognostic value remains uncertain in comparison with adjuvant CT/RT. Furthermore, the prognostic value of molecular biomarkers, such as thymidylate synthase (TS), thymidine phosphorylase (TP), and dihydropyrimidine dehydrogenase (DPD), all of which are involved in the FU metabolism, is unknown in neoadjuvant settings. We assessed the impact of standardized preoperative CT/RT and intratumoral TS, TP, and DPD levels on patient outcome. Patients and Methods Forty patients with rectal cancer pretherapeutic UICC stage II/III, receiving preoperative FU-based CT/RT (CAO/ARO/AIO-94 trial) followed by standardized surgery, including total mesorectal excision, were investigated. Downsizing, downstaging, tumor regression, as well as TS, TP, and DPD gene expression of post-treatment surgical specimens were correlated with disease-free survival (DFS) and overall survival (OS). Results Significant downsizing (P < .001) and downstaging (P = .001) were achieved with preoperative CT/RT. During a median follow-up of 49 months (95% CI, 43 to 58 months), the cancer recurrence rate was 28.2%. DFS and OS were significantly increased in patients with downstaging (P < .001 and P = .003, respectively), compared with patients without downstaging. All patients who developed cancer recurrence had a persistent positive lymph node status after preoperative CT/RT (P < .001) and a significantly higher TS gene expression (P = .035) compared with those patients without recurrence. Conclusion Persistent positive lymph node status and high intratumoral TS expression after preoperative CT/RT are predictive of an unfavorable prognosis in rectal cancer UICC stage II/III.
In Europe, rectal cancer is a major socioeconomic and health problem, with about 70,000 new cases diagnosed every year.1,2 In locally advanced disease, the local recurrence rate varies from 4% to 40%, and almost half of all patients will die as a result of progressive disease.3-5 In recent years, considerable progress in staging, surgical management (including total mesorectal excision), and cancer-related therapy of rectal cancer has resulted in an improvement in disease-free survival (DFS) and overall survival (OS).6-9 In the last decade, postoperative combined fluorouracil (FU) -based chemoradiotherapy (CT/RT) has been recommended as standard treatment for patients with rectal cancer International Union Against Cancer (UICC) stage II/III.10-14 Recently, the German Rectal Cancer Study Group published a randomized prospective trial that compared preoperative versus postoperative FU-based, long-term CT/RT in resectable rectal cancer pretherapeutic (c) UICC stage II/III (CAO/ARO/AIO-94 trial).15 The 5-year cumulative incidence of local cancer recurrence was 6% for patients assigned to preoperative CT/RT compared with 13% in the postoperatively treated group (P = .006).15 However, despite the local benefit of the preoperative treatment, no significant difference in the 5-year OS was achieved with preoperative versus postoperative CT/RT. According to these results, preoperative FU-based CT/RT in rectal cancer cUICC stage II/III is now recommended in Germany, parts of Europe, and the United States.16 As shown in several clinical studies, intratumoral thymidylate synthase (TS), thymidine phosphorylase (TP), and dihydropyrimidine dehydrogenase (DPD) gene expression may predict the response of advanced colorectal cancer to adjuvant FU-based therapy.17-23 However, most of the published studies are highly heterogeneous and therefore not comparable to each other.24 In this context, well-defined patient populations, standardized therapy schedules, and tissue collection seem to be mandatory to investigate the prognostic value of molecular markers. The aim of this study was to investigate the correlation between therapy-induced downsizing and downstaging on DFS and OS as well as the prognostic value of TS, TP, and DPD gene expression in rectal cancer patients treated within the preoperative CT/RT arm of the CAO/ARO/AIO-94 trial.
Eligibility During January 1998 to November 2001, 40 patients with primary rectal carcinoma (cUICC stage II/III) and a minimal follow-up of 36 months were enrolled onto long-term FU-based CT/RT in the neoadjuvant setting of the CAO/ARO/AIO-94 trial.15 All patients were treated at the Department of General Surgery, Medical Center of the University of Göttingen (Göttingen, Germany). Eligibility criteria included resectable adenocarcinoma of the rectum, located within 16 cm from the anal verge. Endorectal ultrasound (rES) and computed tomographic scans of the abdomen and pelvis, colonoscopy, chest x-ray, or thoracic computed tomographic scans were performed to exclude lower tumor stages or patients with evidence of distant metastases. Patients with previous cancer or previous cancer-related chemotherapy or radiotherapy were excluded. At the time of staging, none of the patients had contraindications to CT/RT, and all patients had an Eastern Cooperative Oncology Group performance status of 0 to 1. The trial was approved by the medical ethics committee of the University of Göttingen.
Treatment Surgery followed 4 to 6 weeks after completion of preoperative treatment and clinical restaging. Standardized surgery, including total mesorectal excision, was used, and in all patients a locally curative tumor resection was achieved. The surgical procedures included abdominoperineal resection in 15 patients (37.5%), low anterior resection in 23 patients (57.5%), and anterior resection in one patient (2.5%). One patient (2.5%) with a low-lying rectal cancer, initially cUICC stage II, refused the planned abdominoperineal resection, and therefore received a full rectal wall excision by transanal endoscopic microsurgery. Four weeks after surgery, four cycles of FU infusion 500 mg/m2/d on days 1 to 5, once per month, were started in an adjuvant setting.
Follow-Up
Histopathologic Staging and Assessment of Tumor Regression Downsizing was determined by comparison of pretreatment clinical T status assessed by rES and computed tomography compared with postoperative histopathologic T status. Downsizing or downstaging was present when the tumor regressed by at least one T stage and one UICC stage, respectively.
Microdissection, RNA Extraction, and Quantitation of Gene Expression Total RNA from paraffin-embedded tissue was extracted according to a previously described method.30,31 RNA was transcribed into cDNA using M-MLV reverse transcriptase (Invitrogen, Karlsruhe, Germany) and random hexamers (Amersham Pharmacia, Freiburg, Germany). Quantitation of gene expression was performed as described recently.32,33 Briefly, TS, TP, and DPD cDNA were quantified by TaqMan (Applied Biosystems, Foster City, CA) real-time polymerase chain reaction. cDNA was quantified in relation to beta-actin cDNA. Primer sequences were based on the GenBank accession numbers AB004047 (beta-actin), X02308 (TS), M63193 (TP), and U20938 (DPD). The primers and polymerase chain reaction conditions used in this study were described previously.32,33
Statistical Analysis
Tumor Staging and Histopathologic Tumor Regression After preoperative CT/RT, surgical tumor resection was performed in all 40 patients. One patient showed nonresectable liver metastases intraoperatively (R2 situation for distant metastases). The potential under-/overstaging rate of preoperative rES and computed tomography scans was investigated in a control group of patients receiving primary surgery (postoperatively treated control arm of the CAO/ARO/AIO-94 trial). The cUICC staging was correct in 15 of 16 patients (94%). The pretherapeutic and post-therapeutic histopathologic tumor stages (both UICC) are summarized in Table 1. Significant downsizing (P < .001) and downstaging (P = .001) were found after preoperative CT/RT. Downsizing was achieved in 19 patients (47.5%): 14 patients by one T stage, one patient by two T stages, three patients by three T stages, and one patient by four T stages. Downstaging was achieved in 18 patients (45%): seven patients by one UICC stage, 10 patients by two UICC stages, and one patient by three UICC stages. A distinct therapy-induced downsizing and staging was observed, especially in patients with cT3 and cUICC III, respectively. Downstaging from positive to negative lymph node status was achieved in 15 of 40 patients (37.5%). Eight (45%) of the patients with downstaging showed a TRG 3 or 4, six patients (33%) showed a TRG 2, and four patients (22%) showed a TRG 1, whereas 10 (53%) of the patients with downsizing showed a TRG 3 or 4, eight patients (42%) showed a TRG 2, and one patient (5%) showed a TRG 1.
Recurrence Rate, DFS, and OS After a median postoperative follow-up of 49 months (95% CI, 43 to 58 months), DFS was 72%. Three patients (7.5%) died as a result of a cause unrelated to cancer: one patient died on day 30 postoperatively as a result of pulmonary and cardiac failure (and was excluded for survival analysis); another patient remained without cancer recurrence (> 58 months postoperatively) until death caused by progressive dementia; the third patient died as a result of cardiac failure without cancer recurrence (49 months postoperatively). During the follow-up period, 11 patients (28.2%) developed cancer recurrence. The local recurrence rate was 7.7% (three of 39), and all were associated with the development of synchronous metastatic disease. As summarized in Table 2, two patients developed local recurrence associated with pulmonary metastases, and died as a result of the local recurrence. Another patient developed extraluminal cancer recurrence with extended peritoneal metastases 39 months postoperatively. To date, this patient (patient 11) is alive and has stable disease. Six patients developed cancer recurrence with distant metastases, and died despite induction of polychemotherapy. The remaining two patients (patients 8 and 10) with distant metastases are still alive. One patient received an R0 resection of his pulmonary metastases and is disease free at this time. The other patient with lymphatic and peritoneal metastases has progressive disease while receiving chemotherapy as of March 15, 2005.
Correlation Between Cancer Recurrence and Downstaging, Downsizing, and Tumor Regression After Preoperative CT/RT All patients who developed cancer recurrence belong to the patient subgroup in which no downstaging was achieved by preoperative CT/RT (Table 2). With the exception of one patient (patient 11), in all of these patients the local tumor showed a moderate to high CT/RT-induced TRG, but all patients had a persistent positive lymph node status. In this patient subgroup, no significant correlation between cancer recurrence and downsizing (P = .378) or TRG (P = .096) was found. Conversely, all patients with downstaging after preoperative CT/RT are disease free to date (P < .001), and also have a benefit in cancer-specific OS (P = .003; Fig 1). However, no significant benefit in DFS (P = .708) and OS (P = .515) was seen in patients with CT/RT-induced downsizing compared with patients without evidence of downsizing.
Correlation Between TS, TP, and DPD Gene Expression and Cancer Recurrence in Preoperatively Treated Patients There was a significant difference of TS gene expression between patients with and without cancer recurrence in the subgroup without CT/RT-induced downstaging after preoperative CT/RT (Fig 2). Patients who developed cancer recurrence had a significantly enhanced intratumoral TS gene expression of the post-therapeutic resection specimen (WMW test, P = .035; Table 3). No significant correlation was found between cancer recurrence and TP or DPD gene expression (online only appendix; Fig A1).
Influence of Preoperative CT/RT on Intratumoral TS, TP, and DPD Gene Expression In all surgical resection specimens, TS, TP, and DPD gene expression were analyzed after preoperative CT/RT. The mRNA levels were compared with intratumoral mRNA levels of rectal cancer patients (n = 16) treated with primary surgery in the postoperative (control) arm of the CAO/ARO/AIO-94 trial (randomized control arm). Intratumoral TS gene expression after preoperative CT/RT was significantly lower than that of the control group treated with primary surgery (P = .001; Table 4). In contrast, DPD gene expression was significantly higher in preoperatively treated tumors compared with the control group (P < .001). For TP gene expression, no tendency or significant difference between both patient groups was seen (Table 4).
The prognosis for recurrent disease in rectal cancer is poor and recurrent disease often is the cause of death. Accordingly, the prevention of early tumor recurrence is one of the main goals in rectal cancer treatment. The key message of the recently published CAO/ARO/AIO-94 trial of the German Rectal Cancer Study Group was that the local recurrence rate was significantly lower in the preoperatively treated patients than in the postoperatively treated patients.15 However, despite the local benefit of the preoperative treatment, no significant difference in the 5-year OS was achieved with preoperative versus postoperative CT/RT. Thus, the prognostic value of preoperative, long-term, FU-based CT/RT remains uncertain. Furthermore, the prognostic value of molecular biomarkers, such as TS, TP, and DPD, all of which are involved in FU metabolism, is unknown in preoperatively treated rectal cancer patients. To our knowledge, this patient cohort is the first study population with a standardized preoperative treatment lending itself to evaluate the impact of neoadjuvant CT/RT and intratumoral enzyme expression on patient outcome. There are two main observations in our study. First, all patients who developed cancer recurrence had a persistently positive lymph node status without downstaging after preoperative CT/RT, irrespective of the extent of downsizing or local tumor regression. In contrast, all patients with downstaging after preoperative CT/RT are disease free up to the follow-up examination (March 15, 2005; median, 49 months). Second, the patients in the subgroup without downstaging, who developed cancer recurrence, showed a significantly higher TS gene expression after preoperative CT/RT, compared with those who are disease free at present. The presence of downstaging as an important prognostic factor for cancer-specific survival has been shown by other investigators. Villafranca et al34 reported a 3-year DFS of 30% in patients without downstaging, compared with 88% in patients with downstaging, whereas Kaminsky-Forrett et al35 reported a 5-year cancer-specific survival of only 45% in patients without downstaging, compared with 100% in patients with downstaging. The fact that all patients with cancer recurrence had a positive lymph node status after therapy reflects the poor prognosis of a positive lymph node status, which is further emphasized by the development of synchronous peritoneal or hematogenous metastases. With these results in mind, the recommendation of local excision alone without lymphadenectomy, when preoperative CT/RT has achieved a high or total response of the local tumor,36-38 seems to be unjustified. The predictive value of high TS gene expression for poor prognosis has been shown in several other studies in advanced18-22 and adjuvant18-23 disease settings. A previous meta-analysis24 showed that these studies are heterogenous, both with respect to setup and results, and none of them has been standardized with respect to surgery. Therefore, a comparison of the study results is almost impossible. Furthermore, in contrast to other studies, we did not use tissue specimens collected prior to therapy; instead, tissue collected after CT/RT was used for measuring gene expression. This is another reason why our results cannot be compared with the findings of other studies. We gained biomarker data from the CAO/ARO/AIO-94 trial, which has been standardized comprehensively in a neoadjuvant setting. Apparently, the pretherapeutic clinical tumor stage, the postsurgical histomorphologic stage, and the characteristics of the surviving cells define additional clinical outcome. For the biomarker TS in residual tumor tissue after preoperative radiochemotherapy, we found that if there is no lymph node involvement, even tumors with relatively high residual TS do not have the capacity to recur. If there is lymph node involvement, however, only tumors with a particularly low TS gene expression level did not relapse. Microdissection should be mandatory for all molecular investigations on preoperatively treated specimens. As shown recently by our group,28 the results of those measurements are contaminated by the degree of tissue heterogeneity such as fibrosis, inflammation, or necrosis, which are marked features in neoadjuvantly treated tumor specimens. An interesting finding, although not directly related to the goal of this work, was that the absolute TS levels in tumors after neoadjuvant CT/RT were significantly lower than in tumors without neoadjuvant treatment. This observation may be explained by therapy-induced downregulation of TS, by selection processes that favor cells with low TS levels, or by a mixture of both. In any event, it has to be considered that the tumor tissue has been taken 4 to 6 weeks after completion of CT/RT. We believe that because rectal tumors are known to be regressing at the time point after CT/RT (our group's observation), the selection theory most likely is accurate. Intratumoral TP and DPD status had no impact on patient outcome in this study. We have not found any valid data in the literature about the role of the TP and DPD in neoadjuvant settings, and thus at present there is no hint for a prognostic value of these genes after neoadjuvant therapy. Interestingly, however, we observed an increase of average tumor DPD levels after CT/RT (data not shown). Again, this can be interpreted either as any kind of upregulation or positive selection. Given that DPD catalyzes the rate-limiting step of FU catabolism, it is tempting to speculate that cells with particularly high DPD expression are resistant to chemoradiotherapy and thus were selected. This would mean that neoadjuvant chemoradiotherapy confers resistance against FU and thus may hamper additional FU-based therapy of these patients. As a participant institution of the CAO/ARO/AIO-94 trial, we demonstrated herein that a high post-therapeutic TS level combined with a persistent positive lymph node status may identify patients at higher risk for cancer recurrence. We suggest that these patients may benefit more from an intensification of the adjuvant chemotherapy beyond FU monotherapy; for example, FU combined with oxaliplatin or irinotecan. According to the results of the CAO/ARO/AIO-94 trial, complete CT/RT-induced local tumor regression suggests an enhancement of DFS.39 Therefore, we would advocate an intensification of the preoperative CT/RT to prevent early cancer recurrence. As shown in phase I/II trials, local tumor response can be improved by using the oral fluoropyrimidine, capecitabine,40 or the combination of FU with oxaliplatin or irinotecan.41-44 Especially the combination of preoperative radiotherapy and capecitabine, which mimics the pharmacokinetics of continuous FU infusion, with oxaliplatin can achieve a doubling of the complete remission rate compared with FU monotherapy.43,45 In conclusion, persistently positive lymph nodes and high intratumoral TS gene expression after preoperative FU-based CT/RT are predictive of an unfavorable prognosis in patients with rectal cancer UICC stage II/III. Ongoing phase III trials will have to demonstrate the efficacy of dose-intensified treatment strategies in neoadjuvant and adjuvant settings, as well as the role of biomarkers, such as TS, TP, and DPD, in disease prognosis.
Fig A1
The authors indicated no potential conflicts of interest.
We thank David M. Goldenberg, ScD, MD, (Belleville, NJ), and Claus Rödel, MD, (Erlangen, Germany), for critical comments. Rolf Sauer, MD, (Erlangen, Germany), was the principal investigator of the CAO/ARO/AIO-94 trial of the German Rectal Cancer Study Group.
Authors' disclosures of potential conflicts of interest and author contributions are found at the end of this article.
1. Midgley R, Kerr D: Colorectal cancer. Lancet 353:391-399, 1999[CrossRef][Medline] 2. Landis SH, Murray T, Bolden S, et al: Cancer statistics, 1998. CA Cancer J Clin 48:6-29, 1998[Abstract] 3. Hermanek P Jr, Wiebelt H, Riedl S, et al: Long-term results of surgical therapy of colon cancer: Results of the Colorectal Cancer Study Group [German]. Chirurg 65:287-297, 1994[Medline] 4. Hermanek P, Wiebelt H, Staimmer D, et al: Prognostic factors of rectum carcinoma: Experience of the German Multicentre Study SGCRC—German Study Group Colo-Rectal Carcinoma. Tumori 81:60-64, 1995[Medline] 5. Abulafi AM, Williams NS: Local recurrence of colorectal cancer: The problem, mechanisms, management and adjuvant therapy. Br J Surg 81:7-19, 1994[Medline] 6. Moertel CG: Chemotherapy for colorectal cancer. N Engl J Med 330:1136-1142, 1994 7. O'Connell MJ, Martenson JA, Wieand HS, et al: Improving adjuvant therapy for rectal cancer by combining protracted-infusion fluorouracil with radiation therapy after curative surgery. N Engl J Med 331:502-507, 1994 8. Douglass HO Jr, Moertel CG, Mayer RJ, et al: Survival after postoperative combination treatment of rectal cancer. N Engl J Med 315:1294-1295, 1986[Medline] 9. Krook JE, Moertel CG, Gunderson LL, et al: Effective surgical adjuvant therapy for high-risk rectal carcinoma. N Engl J Med 324:709-715, 1991[Abstract] 10. Tepper JE, O'Connell M, Niedzwiecki D, et al: Adjuvant therapy in rectal cancer: Analysis of stage, sex, and local control—Final report of Intergroup 0114. J Clin Oncol 20:1744-1750, 2002 11. Tveit KM, Guldvog I, Hagen S, et al: Randomized controlled trial of postoperative radiotherapy and short-term time-scheduled 5-fluorouracil against surgery alone in the treatment of Dukes B and C rectal cancer: Norwegian Adjuvant Rectal Cancer Project Group. Br J Surg 84:1130-1135, 1997[CrossRef][Medline] 12. Wolmark N, Wieand HS, Hyams DM, et al: Randomized trial of postoperative adjuvant chemotherapy with or without radiotherapy for carcinoma of the rectum: National Surgical Adjuvant Breast and Bowel Project Protocol R-02. J Natl Cancer Inst 92:388-396, 2000 13. Rich TA, Shepard RC, Mosley ST: Four decades of continuing innovation with fluorouracil: Current and future approaches to fluorouracil chemoradiation therapy. J Clin Oncol 22:2214-2232, 2004 14. Benson AB III, Desch CE, Flynn PJ, et al: 2000 update of American Society of Clinical Oncology colorectal cancer surveillance guidelines. J Clin Oncol 18:3586-3588, 2000 15. Sauer R, Becker H, Hohenberger W, et al: Preoperative compared with postoperative chemoradiotherapy for locally advanced rectal cancer. N Engl J Med 351:1731-1740, 2004 16. Kachnic LA: The place of chemoradiotherapy in the management of rectal cancer. Am Soc Clin Oncol Ed Book 264-269, 2005 17. Beck A, Etienne MC, Cheradame S, et al: A role for dihydropyrimidine dehydrogenase and thymidylate synthase in tumour sensitivity to fluorouracil. Eur J Cancer 30A:1517-1522, 1994[CrossRef] 18. Ichikawa W, Uetake H, Shirota Y, et al: Combination of dihydropyrimidine dehydrogenase and thymidylate synthase gene expressions in primary tumors as predictive parameters for the efficacy of fluoropyrimidine-based chemotherapy for metastatic colorectal cancer. Clin Cancer Res 9:786-791, 2003 19. Johnston PG, Lenz HJ, Leichman CG, et al: Thymidylate synthase gene and protein expression correlate and are associated with response to 5-fluorouracil in human colorectal and gastric tumors. Cancer Res 55:1407-1412, 1995 20. Leichman CG, Lenz HJ, Leichman L, et al: Quantitation of intratumoral thymidylate synthase expression predicts for disseminated colorectal cancer response and resistance to protracted-infusion fluorouracil and weekly leucovorin. J Clin Oncol 15:3223-3229, 1997[Abstract] 21. Metzger R, Danenberg K, Leichman CG, et al: High basal level gene expression of thymidine phosphorylase (platelet-derived endothelial cell growth factor) in colorectal tumors is associated with nonresponse to 5-fluorouracil. Clin Cancer Res 4:2371-2376, 1998 22. Salonga D, Danenberg KD, Johnson M, et al: Colorectal tumors responding to 5-fluorouracil have low gene expression levels of dihydropyrimidine dehydrogenase, thymidylate synthase, and thymidine phosphorylase. Clin Cancer Res 6:1322-1327, 2000 23. Kornmann M, Schwabe W, Sander S, et al: Thymidylate synthase and dihydropyrimidine dehydrogenase mRNA expression levels: Predictors for survival in colorectal cancer patients receiving adjuvant 5-fluorouracil. Clin Cancer Res 9:4116-4124, 2003 24. Popat S, Matakidou A, Houlston RS: Thymidylate synthase expression and prognosis in colorectal cancer: A systematic review and meta-analysis. J Clin Oncol 22:529-536, 2004 25. Sobin LH, Wittekind C: UICC-TNM Classification of Malignant Tumors (ed 5). New York, NY, Wiley-Liss, 1997 26. Dworak O, Keilholz L, Hoffmann A: Pathological features of rectal cancer after preoperative radiochemotherapy. Int J Colorectal Dis 12:19-23, 1997[CrossRef][Medline] 27. Gavioli M, Bagni A, Piccagli I, et al: Usefulness of endorectal ultrasound after preoperative radiotherapy in rectal cancer: Comparison between sonographic and histopathologic changes. Dis Colon Rectum 43:1075-1083, 2000[CrossRef][Medline] 28. Jakob C, Aust DE, Meyer W, et al: Thymidylate synthase, thymidine phosphorylase, dihydropyrimidine dehydrogenase expression, and histological tumour regression after 5-FU-based neo-adjuvant chemoradiotherapy in rectal cancer. J Pathol 204:562-568, 2004[CrossRef][Medline] 29. Schutze K, Lahr G: Identification of expressed genes by laser-mediated manipulation of single cells. Nat Biotechnol 16:737-742, 1998[CrossRef][Medline] 30. Woodall CJ, Watt NJ, Clements GB: Simple technique for detecting RNA viruses by PCR in single sections of wax embedded tissue. J Clin Pathol 46:276-277, 1993 31. Chomczynski P, Sacchi N: Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156-159, 1987[Medline] 32. Kornmann M, Danenberg KD, Arber N, et al: Inhibition of cyclin D1 expression in human pancreatic cancer cells is associated with increased chemosensitivity and decreased expression of multiple chemoresistance genes. Cancer Res 59:3505-3511, 1999 33. Johnson MR, Wang K, Smith JB, et al: Quantitation of dihydropyrimidine dehydrogenase expression by real-time reverse transcription polymerase chain reaction. Anal Biochem 278:175-184, 2000[CrossRef][Medline] 34. Villafranca E, Okruzhnov Y, Dominguez MA, et al: Polymorphisms of the repeated sequences in the enhancer region of the thymidylate synthase gene promoter may predict downstaging after preoperative chemoradiation in rectal cancer. J Clin Oncol 19:1779-1786, 2001 35. Kaminsky-Forrett MC, Conroy T, Luporsi E, et al: Prognostic implications of downstaging following preoperative radiation therapy for operable T3–T4 rectal cancer. Int J Radiat Oncol Biol Phys 42:935-941, 1998[CrossRef][Medline] 36. Kim CJ, Yeatman TJ, Coppola D, et al: Local excision of T2 and T3 rectal cancers after downstaging chemoradiation. Ann Surg 234:352-359, 2001[CrossRef][Medline] 37. Schell SR, Zlotecki RA, Mendenhall WM, et al: Transanal excision of locally advanced rectal cancers downstaged using neoadjuvant chemoradiotherapy. J Am Coll Surg 194:584-591, 2002[CrossRef][Medline] 38. Bonnen M, Crane C, Vauthey JN, et al: Long-term results using local excision after preoperative chemoradiation among selected T3 rectal cancer patients. Int J Radiat Oncol Biol Phys 60:1098-1105, 2004[CrossRef][Medline] 39. Rodel C, Martus P, Papadoupolos T, et al: Prognostic significance of tumor regression after preoperative chemoradiotherapy for rectal cancer. J Clin Oncol 23:8688-8696, 2005 40. Dunst J, Reese T, Sutter T, et al: Phase I trial evaluating the concurrent combination of radiotherapy and capecitabine in rectal cancer. J Clin Oncol 20:3983-3991, 2002 41. Hofheinz RD, von Gerstenberg-Helldorf B, Wenz F, et al: Phase I trial of capecitabine and weekly irinotecan in combination with radiotherapy for neoadjuvant therapy of rectal cancer. J Clin Oncol 23:1350-1357, 2005 42. Gerard JP, Chapet O, Nemoz C, et al: Preoperative concurrent chemoradiotherapy in locally advanced rectal cancer with high-dose radiation and oxaliplatin-containing regimen: The Lyon R0-04 phase II trial. J Clin Oncol 21:1119-1124, 2003 43. Rodel C, Grabenbauer GG, Papadopoulos T, et al: Phase I/II trial of capecitabine, oxaliplatin, and radiation for rectal cancer. J Clin Oncol 21:3098-3104, 2003 44. Van Cutsem E, Dicato M, Wils J, et al: Adjuvant treatment of colorectal cancer (current expert opinion derived from the Third International Conference: Perspectives in Colorectal Cancer, Dublin, 2001). Eur J Cancer 38:1429-1436, 2002[CrossRef][Medline] 45. Schuller J, Cassidy J, Dumont E, et al: Preferential activation of capecitabine in tumor following oral administration to colorectal cancer patients. Cancer Chemother Pharmacol 45:291-297, 2000[CrossRef][Medline] Submitted September 16, 2005; accepted March 16, 2006.
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Copyright © 2006 by the American Society of Clinical Oncology, Online ISSN: 1527-7755. Print ISSN: 0732-183X
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ABSTRACT
INTRODUCTION
