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© 2001 American Society for Clinical Oncology Clinical Impact of Germ Cell Tumor Cells in Apheresis Products of Patients Receiving High-Dose ChemotherapyFrom the Department of Hematology/Oncology, University of Tuebingen, Tuebingen; Städt Kliniken Oldenburg, Med Klinik II, Abt Hämatologie und Onkologie; Westdeutsches Tumorzentrum, Innere Klinik und Poliklinik, Universitätsklinikum Essen, Essen; Katharinenhospital Stuttgart, Hämatologie und Intern Onkologie, Stuttgart, Germany; and Pathology/Laboratory for Experimental Patho-Oncology, Academic Hospital Rotterdam/Daniel Josephine Nefkens Institute, Erasmus University, Rotterdam, the Netherlands. Address reprint requests to C. Bokemeyer, MD, Department of Hematology/Oncology, University of Tuebingen, Otfried-Mueller-Str 10, 72076, Tuebingen, Germany; email: carsten.bokemeyer{at}med.uni-tuebingen.de
PURPOSE: High-dose chemotherapy (HD-Ctx) followed by autologous peripheral-blood stem-cell (PBSC) transplantation is currently investigated in patients with poor prognosis or relapsed metastatic germ cell tumor (GCTs). This study analyzed the presence and the clinical importance of contaminating tumor cells in PBSC preparations used to support HD-Ctx in GCT patients. PATIENTS AND METHODS: Seven targets for reverse transcription polymerase chain reaction (RT-PCR)-based detection of GCT cells were able to detect seminomatous and different histologic variants of nonseminomatous tumor cells. PBSC preparations from 57 patients were investigated for the presence of contaminating tumor cells using this set of targets, including beta human chorionic gonadotropin (ß-hCG), fibronectin (EDB variant), epidermal growth factor receptor (EGFR), CD44 (v8 to 10 variant), germ cell and placental alkaline phosphatase (AP), human endogenous retrovirus type K (ENV and GAG), and XIST. Samples of PBSC preparations from four healthy donors for allogenic transplantations as well as blood specimens from 10 healthy volunteers served as negative controls. RESULTS: Fifty patients (43 first-line and seven second-line Ctx) were assessable. Combining all RT-PCR results, 29 PBSC preparations (58%) were positive for tumor-specific amplification products (HERV-K 0, fibronectin 4, XIST 14, ß-hCG 19, AP 19, CD44 24, EGFR 26). Ten (35%) of 29 patients who underwent transplantation with positive PBSC preparations and seven (33%) of 21 patients with negative PBSC preparations have suffered relapse or progression (not significant [ns]). With a median follow-up of 22 months (2 to 66) post–HD-Ctx projected 3-year survival rates are 68% (RT-PCR+) and 58% (RT-PCR-) (ns). None of the 10 control peripheral-blood samples showed positivity for any of the targets studied. CONCLUSION: GCT cells can be detected in more than 50% of PBSC preparations using a RT-PCR approach with multiple targets. Despite the presence of tumor cells, retransplantation of the PBSC products did not effect long-term outcome. Factors such as responsiveness to chemotherapy and tumor mass seem to overcome the importance of potentially re-infused tumor cells.
MALIGNANT GERM cell tumors (GCTs) are the most common solid tumors in young men aged 20 to 40 years. With the introduction of modern cisplatin-based combination chemotherapy in the 1970s, this disease has become curable in many patients (up to 90% to 95%).1 There are, however, two subsets of patients for whom the results of conventional-dose chemotherapy are not satisfactory: patients with poor prognosis metastatic disease according to International Germ Cell Cancer Classification Group criteria,2 who will achieve remissions only in 50% of cases because of their widespread metastatic disease. The other group consists of patients relapsing after cisplatin-based first-line treatment who will only achieve long-term cure rates of 20% to 25%.1,3 For these two specific patient subgroups, high-dose chemotherapy followed by autologous peripheral-blood stem-cell (PBSC) support has been widely investigated during the last 10 years in order to improve their prognosis.3,4 High-dose strategies have indeed been shown to be successful in patients with disease responsive to chemotherapy, whereas patients with primary mediastinal GCTs and those with cisplatin-refractory disease (progression within 4 weeks of last cisplatin-based regimen) are less likely to benefit from this type of therapy.5,6,7 Patients receiving high-dose treatment followed by autologous PBSC transplantation usually present with a high tumor burden at the time of procedure initiation. Resistance to chemotherapeutic drugs is one of the most important reasons for relapse after high-dose therapy. In addition, the possible reinfusion of GCT cells in apheresis products collected at the time of stem-cell mobilization may also be a potential cause of treatment failure. However, both frequency and clinical impact of potentially contaminating tumor cells in leukapheresis products have not yet been widely investigated in GCT patients. Notably, in breast cancer patients, the presence of tumor-cell contamination of bone marrow and stem-cell products has been a rather frequent finding.8 While the presence of microscopic bone marrow cells has been demonstrated to be of prognostic value, the ability of reinfused breast cancer cells after high-dose chemotherapy to cause disease relapse has not yet been definitely demonstrated.9,10 GCTs can be composed of a variety of histologic components, ie, seminoma and different variants of nonseminomas: embryonal carcinoma, teratoma, yolk sac tumor, and choriocarcinoma.11 In the present analysis, we explored the applicability of several targets for reverse transcription polymerase chain reaction (RT-PCR) to detect the various histologic elements of GCTs and their usefulness to identify contaminating tumor cells in PBSC preparations. The RT-PCR results in leukapheresis products were pooled and the presence of GCT cells was correlated to the frequency of relapse and overall survival of patients receiving high-dose chemotherapy.
Patients Patients’ characteristics are given in Table 1. For the analysis of contaminating tumor cells, the leukapheresis products of 57 patients scheduled to receive high-dose chemotherapy either as intensified first-line treatment (n = 48) or as salvage treatment (n = 9) at relapse were available. In total the results from 50 analyses were assessable, since four patients did not receive high-dose chemotherapy because of disease progression during induction treatment, two patients’ samples for RNA analysis for PCR failed, and one patient was lost to follow-up immediately after high-dose treatment. The median follow-up for the 50 assessable patients was 22 months (2 to 66 months). All patients were treated within prospective clinical trials on high-dose chemotherapy at one of four participating centers (Oldenburg, Essen, Stuttgart, and Tübingen) in Germany. Before study entry, the diagnosis of GCT had to be unequivocally demonstrated in all patients by the presence of elevated tumor markers (beta human chorionic gonadotropin [ßhCG], alpha-fetoprotein [AFP], lactate dehydrogenase [LDH]) and/or histopathologic evaluation. The clinical study protocols were approved by each institution’s ethical committee, and all patients gave informed consent.
Study Protocols Patients with relapsed disease were treated by three cycles of conventionally dosed chemotherapy with paclitaxel, ifosfamide, and cisplatin (TIP) at 3-week intervals. This conventional-dose induction treatment was used to achieve tumor control before high-dose chemotherapy, to test chemotherapy sensitivity, and to allow the mobilization of PBSCs. Apheresis was performed after the first cycle of TIP chemotherapy using granulocyte colony-stimulating factor (G-CSF) 5 to 10 µg/kg subcutaneously daily, starting immediately after chemotherapy until completion of leukapheresis. The subsequently scheduled high-dose chemotherapy cycle consisted of thiotepa, etoposide, and carboplatin (TEC) followed by autologous PBSC rescue with at least 2 x 106 CD34+ cells/kg body weight.12 The first-line treatment protocol for patients with poor prognosis according to International Germ Cell Cancer Classification Group criteria consisted of one cycle of standard-dose cisplatin, etoposide, and ifosfamide (PEI) chemotherapy followed by stem-cell leukapheresis. G-CSF was used as in patients with relapsed disease. All patients went on to receive three consecutive cycles of high-dose PEI with intensification of etoposide and ifosfamide doses. Each high-dose PEI cycle was repeated at 3-week intervals and supported by PBSC transplantation with at least 2 x 106 CD34+ cells/kg body weight. Both treatment protocols have been previously described in detail.4,12 Response to high-dose chemotherapy was classified according to World Health Organization criteria. Additionally, patients with normalization of previously elevated GCT serum markers with radiologic evidence of residual disease were considered to be in marker-negative partial remission. Surgical resection of all residual masses was attempted in these cases.
Selection of Markers for Tumor-Cell Detection
The primer sequences used for these markers and the RT-PCR conditions are given in Table 2.
RNA Isolation and PCR RNA was isolated from 1 mL leukapheresis samples that had been stored in liquid nitrogen since the day of stem-cell collection. These cells were thawed in a 37°C water bath, washed with 100% fetal bovine serum, and resuspended in serum-free culture medium. Total RNA was isolated using RNA-STAT-60 according to the supplier’s protocol (Tel-Test "B" Inc; Friendswood, TX) The expression of the various genes in tissue samples, apheresis products, and cell lines was determined by PCR amplification of the generated cDNA using gene or transcript-specific primer combinations spanning the particular intron sequences. When no introns were present, the samples were pretreated with DNase. HPRT was used as quality-control of the cDNAs tested. A negative control, in which cDNA was omitted, and a positive control were used for every PCR assay. Initially the PCR assay was checked on the positive controls. Subsequently, normal peripheral-blood samples were analyzed. When found to be negative, they were studied regarding their expression on seminomatous and nonseminomatous GCTs, as well as five GCT-derived cell lines (NTera2, TERA1, 2102Ep, NCCIT, and 833KE). Subsequently, the four control leukapheresis samples were investigated. Cytokeratin 19 PCR was applied on three consecutive peripheral-blood samples, to exclude possible contamination with skin cells.
Sensitivity of the Amplification Method
Statistical Analysis
Informativity of the Various Targets to Detect the Histologic Variants of GCT The possible application of detection of circulating GCT cells by analyses of the presence of specific transcripts by RT-PCR has been poorly investigated. The targets included in this study are indicated in Table 3, of which the conditions are described in Patients and Methods and in Table 2. We have reported previously that a specific transcript of 1.5 kb of the platelet-derived growth factor receptor is suitable to detect GCT cells.21,22 However, we recently found that a subpopulation of B cells, present in normal peripheral blood, results in positivity in peripheral blood, and therefore this marker is not suitable for detection of contaminated tumor cells in apheresis products (Palumba et al, manuscript in preparation). Similarly, transcripts specific for cytokeratin 19, the standard form and v10 variant of CD44, GDF3, the EDA variant of fibronectin, and the GAG gene of HERV-K show positivity in normal peripheral-blood samples and were therefore excluded from this study (Table 3). In contrast, seven transcripts were negative in all peripheral-blood samples and were studied in more detail. These targets were tested on a series of invasive GCTs of different histologic composition, of which the results are also listed in Table 3. All seminomas, embryonal carcinomas, teratomas yolk sac tumors, and mixed nonseminomas were positive for ß-hCG, CD44 v8 to 10, XIST, and EGFR. In addition, all seminomas were positive for HERV-K ENV, and AP. None of the seminomas showed expression of the EDB variant of fibronectin. In contrast, this variant was present in pure yolk sac tumors and most mixed nonseminomas containing a significant yolk sac tumor element. In conclusion, the EDB variant seems to be a specific marker for yolk sac tumor. No expression of the HERV-K gene was found in pure mature teratomas or in two mixed nonseminomas mainly containing such a histologic composition. This is in agreement with our previous results.17 AP positivity was detected in the majority of nonseminomas, with the exception of mature teratoma. Moreover, we found that the five GCT-derived cell lines all showed positivity for the targets studied (Table 3).
Detection of Tumor Cell Contamination In order to determine the sensitivity of our RT-PCR method to detect the different transcripts found to be present in GCTs, both for seminomas and nonseminomas (see above), serial dilutions were prepared. Under standard PCR conditions (see Patients and Methods for details), roughly 10 transcripts could be detected for the different targets within a single amplification experiment (performed on cDNA derived from 125 ng of RNA). In addition, the dilution experiments using the NTera2 cells in peripheral blood indicate that this corresponded with about one tumor cell per 1,000 host cells (not shown). The results of detection of GCT tumor cells in apheresis products with the targets used are indicated in Table 4. Note that they remained negative in multiple-control PBSC preparations, even after a booster amplification of another 12 cycles (not shown). The results of the analysis of the apheresis samples and the number of patients with relapse are given in Table 4. An example of the PCR results for ß-hCG is shown in Fig 1. The frequency of positive signals ranged from 0% up to 52% using the different markers, eg, HERV-K ENV and EGFR.
When considering an apheresis product as contaminated by tumor cells in case of a positive signal for any of the markers, this combined analysis results in a tumor-cell contamination rate of 58% (29 of 50 samples). Among these 29 patients, 10 (35%) have subsequently relapsed or progressed. Comparably, treatment has also failed for seven (34%) of 21 patients with complete negativity for all markers. The comparison of patients’ characteristics depending on the positivity or negativity for PCR signals showed no statistical differences between those two groups. The median age in both groups (34 and 32 years) was comparable, as were the median follow-up times. When we compared the results of the PCR on individual patients, both AP and ß-hCG were found to detect the same 19 patients’ samples as positive. When analyzing the combination of markers that yielded the same frequency of positive results as the combination of all markers, positivity for CD44 and EGFR was sufficient to detect all 29 positive samples. The relapse rate among patients with negative signals was not significantly different from that of patients with positive signals for the markers analyzed separately for contaminating cells (Table 4).
Correlation to Clinical Results
The overall pattern of relapse after high-dose chemotherapy in this cohort of patients was not unusual. In most patients the sites at relapse were the same as those that were involved during the course of disease before high-dose therapy. Three patients had liver involvement for the first time at relapse after high-dose chemotherapy, and three different patients eventually developed brain metastases (one as the only site of disease, the other two in addition to further sites). Only two of these three patients with brain metastases had received PBSC support with detectable tumor-cell contamination. A separate analysis with respect to the presence of contaminating tumor cells in the seven patients receiving salvage high-dose chemotherapy is listed in Table 5. Again, no correlation to clinical outcome was found.
Among the 29 patients who received tumor-marker–positive apheresis products, 10 had disease progression and/or died, and 19 are still in complete (n = 10) or tumor-marker–negative partial remission (n = 9). Seventeen patients relapsed after a median time period of 7 months (range, 2 to 18 months). Among these, nine patients died of disease, five are alive with progressive disease, and three have responded favorably to subsequent salvage chemotherapy and are again in complete or partial remission at the time of follow-up. Ten of these 17 patients had received transplants of PCR-marker–positive PBSC products, while seven patients had received PCR-signal–negative stem-cell preparations.
In the present study, patients with metastatic germ-cell tumors undergoing autologous high-dose chemotherapy plus PBSC transplantation were found to have contaminating tumor cells at the time of apheresis in more than 50% of cases. For the detection of GCT cells, a panel of seven targets for PCR amplification was used, yielding positive signals in 29 (58%) of 50 evaluated apheresis samples. The markers used are able to identify the different histologic variants of GCT, including seminoma and various elements of nonseminomas. Interestingly, four of the markers were suitable to detect all histologies (ß-hCG, CD44 v8 to 10, XIST and EGFR), while fibronectin EDB was found to be specific for yolk-sac tumor. XIST could be one of the most promising targets to detect contaminating GCT cells, because expression of this gene is restricted to female cells and to cells containing extra copies of the X chromosome, like GCT cells.20 Theoretically, the other genes used as targets to detect GCT cells in leukapheresis samples might be expressed in host cells of the patient, although our data do not indicate that this is the case in our series. The frequency of contaminating tumor cells in this study is even higher than among those patients reported by Fan et al.23 Among their 20 patients with ß-hCG–secreting tumors, eight apheresis products (40%) were found positive. However, the detection of contaminating tumor cells was only addressed by a PCR test for this marker, so that GCT cells negative for this target would remain undetected. In fact, our data indicate that, using ß-hCG as target, 19 out of the 50 cases were positive, accounting for 38%, which agreed with the data obtained by Fan et al. In most patients of the current series, PBSC separation was performed after one standard-dose chemotherapy cycle of PEI. In the subset of patients with relapsed disease included in this study, a similar pattern of results was found, possibly due to a similar frequency of contaminating tumor cells. The PBSCs of these patients were collected after one cycle of TIP chemotherapy. In all cases, G-CSF was used to enhance the stem-cell mobilization.12 Circulating tumor cells have also been investigated in other diseases, eg, in patients with breast cancer and with multiple myeloma following induction chemotherapy and growth factor application.9,24 In a study of myeloma patients that compared the frequency of contaminating tumor cells in apheresis samples gained either after the first or the second cycle of intensified chemotherapy plus G-SCF, no differences in the quantity of tumor cells per number of CD34-positive cells were found.25 This is in accordance with data from patients with breast cancer receiving sequential high-dose chemotherapy.26 Thus it appears unlikely that the high frequency of contaminating tumor cells in our series of patients with GCT would have been substantially reduced if apheresis would have been performed after the second or third cycle of induction chemotherapy. High-dose chemotherapy has been used in the treatment of metastatic GCTs because this is a chemosensitive malignancy with potentially high cure rates. Prospective studies are currently investigating the role of high-dose chemotherapy plus PBSC transplantation in patients with "poor prognosis" characteristics at the time of presentation and in patients with relapsed disease after standard cisplatin-based combination chemotherapy.1,6,12 Several prognostic factors have been identified to determine the chances of response to and long-term survival after high-dose chemotherapy in these patient groups. Particularly in patients with relapsed disease, a mediastinal primary tumor, elevated ß-hCG levels before high-dose therapy, and disease refractory to standard cisplatin-based chemotherapy are all considered unfavorable characteristics resulting in a poor outcome.4 It has not yet been determined whether the presence of contaminating tumor cells in the transplanted PBSC apheresis products may also contribute to the risk of relapse after high-dose chemotherapy. In the present series of 50 patients undergoing high-dose chemotherapy, the presence of contaminating tumor cells had neither an impact on overall survival nor did it effect event-free survival in the 32 patients with a favorable response to high-dose chemotherapy. Since no significant influence of the presence of contaminating tumor cells was found by univariate analysis, a multivariate analysis comparing the presence of these cells to established prognostic factors was not performed. These data clearly indicate that other prognostic factors, such as tumor mass, response to chemotherapy, and presence of residual disease after chemotherapy, may be more important in determining the overall prognosis of the patient than contaminating tumor cells retransplanted after high-dose chemotherapy. Since 19 of the patients who received transplants of GCT-positive apheresis products are still in complete or tumor-marker–negative partial remission, this finding might also indicate that the GCT cells transplanted are not clonogenic and might not have had the biologic potential to cause relapse. Hypothetically, the cells mobilized had already been damaged by the previous chemotherapy and had then undergone apoptosis after retransplantation. The series presented here with 50 patients followed for 2 years after high-dose chemotherapy and with 63% of patients remaining in remission and with a tumor cell contamination rate of more than 50% is clearly large enough to statistically detect a possible effect of these tumor cells on the patient’s prognosis. However, it seems that contaminating tumor cells may not be of large clinical relevance in GCT patients undergoing autologous transplantation. It may still be of interest to use the proposed marker panel in patients at the time of diagnosis and screen blood samples for the presence of circulating tumor cells. At that time, these cells may still be viable. Particularly in patients in low stages of disease, circulating tumor cells may identify a subset of patients with a less favorable prognosis. In this setting, it would also be worthwhile to use the marker panel shown here in order to detect certain biologic characteristics of the tumor cells in circulation and correlate the results to the histological features of the primary tumor. It could be of interest to include target selections so that minimal numbers of tumor cells will not be missed by the approach applied. In summary, in patients with advanced metastatic disease, circulating tumor cells may be frequently found in apheresis samples but do not seem to adversely affect the prognosis when retransplanted after high-dose chemotherapy. Thus, the attempt to deplete these cells in stem-cell apheresis products does not seem necessary.
Supported by grant no. 505 U from the Fortuene-Program for Research at Tuebingen University.
Presented in part at the Thirty-Sixth Annual Meeting of the American Society of Oncology, New Orleans, LA, May 20-23, 2000.
1. Bosl GJ, Motzer RJ: Testicular germ-cell cancer. N Engl J Med 337: 242-253, 1997
2.
Mead GM, Stenning SP, Cook P: International germ cell consensus classification: A prognostic factor-erased staging system for metastatic germ cell cancers. J Clin Oncol 15: 594-603, 1997
3.
Siegert W, Beyer J, Strohscheer I, et al: High-dose treatment with carboplatin, etoposide, and ifosfamide followed by autologous stem-cell transplantation in relapsed or refractory germ cell cancer: A phase I/II study—The German Testicular Cancer Cooperative Study Group. J Clin Oncol 12: 1223-1231, 1994 4. Bokemeyer C, Harstrick A, Beyer J, et al: The use of dose-intensified chemotherapy in the treatment of metastatic nonseminomatous testicular germ cell tumors: German Testicular Cancer Study Group. Semin Oncol 25: 24-32, 1998 (suppl 4)
5.
Beyer J, Kramar A, Mandanas R, et al: High-dose chemotherapy as salvage treatment in germ cell tumors: A multivariate analysis of prognostic variables. J Clin Oncol 14: 2638-2645, 1996
6.
Bokemeyer C, Kollmannsberger C, Meisner C, et al: First-line high-dose chemotherapy compared with standard-dose PEB/VIP chemotherapy in patients with advanced germ cell tumors: A multivariate and matched-pair analysis. J Clin Oncol 17: 3450-3456, 1999 7. Hartmann JT, Kanz L, Bokemeyer C: Diagnosis and treatment of patients with testicular germ cell cancer. Drugs 58: 257-281, 1999[Medline]
8.
Diel IJ, Kaufmann M, Costa SD, et al: Micrometastatic breast cancer cells in bone marrow at primary surgery: Prognostic value in comparison with nodal status. J Natl Cancer Inst 88: 1652-1658, 1996
9.
Brugger W, Bross KJ, Glatt M, et al: Mobilization of tumor cells and hematopoietic progenitor cells into peripheral blood of patients with solid tumors. Blood 83: 636-640, 1994 10. Hoon DS, Sarantou T, Doi F, et al: Detection of metastatic breast cancer by beta-hCG polymerase chain reaction. Int J Cancer 69: 369-374, 1996[Medline] 11. Mostofi FK, Sesterhenn IA, Davis CJ: Immunopathology of germ cell tumors of the testis. Semin Diagn Pathol 4: 320-341, 1987[Medline]
12.
Rick O, Bokemeyer C, Beyer J, et al: Salvage treatment with paclitaxel, ifosfamide, and cisplatin plus high-dose carboplatin, etoposide, and thiotepa followed by autologous stem-cell rescue in patients with relapsed or refractory germ cell cancer. J Clin Oncol 19: 81-88, 2001 13. Hildebrandt MO, Blaser F, Beyer J, et al: Detection of tumor cells in peripheral blood samples from patients with germ cell tumors using immunocytochemical and reverse transcriptase-polymerase chain reaction techniques. Bone Marrow Transplant 22: 771-775, 1998[Medline] 14. Miyake H, Hara I, Yamanaka K, et al: Expression patterns of CD44 adhesion molecule in testicular germ cell tumors and normal testes. Am J Pathol 152: 1157-1160, 1998[Abstract] 15. Caricasole AA, van Schaik RH, Zeinstra LM, et al: Human growth-differentiation factor 3 (hGDF3): Developmental regulation in human teratocarcinoma cell lines and expression in primary testicular germ cell tumours. Oncogene 16: 95-103, 1998[Medline] 16. Mighell AJ, Thompson J, Hume WJ, et al: RT-PCR investigation of fibronectin mRNA isoforms in malignant, normal and reactive oral mucosa. Oral Oncol 33: 155-162, 1997[Medline]
17.
Roelofs H, van Gurp RJ, Oosterhuis JW, et al: Detection of human endogenous retrovirus type K-specific transcripts in testicular parenchyma and testicular germ cell tumors of adolescents and adults: Clinical and biological implications. Am J Pathol 153: 1277-1282, 1998
18.
Schar BK, Otto VI, Hanseler E: Simultaneous detection of all four alkaline phosphatase isoenzymes in human germ cell tumors using reverse transcription-PCR. Cancer Res 57: 3841-3846, 1997 19. Wells A: EGF receptor. Int J Biochem Cell Biol 31: 637-643, 1999[Medline] 20. Looijenga LH, Gillis AJ, van Gurp RJ, et al: X inactivation in human testicular tumors: XIST expression and androgen receptor methylation status. Am J Pathol 151: 581-590, 1997[Abstract]
21.
Mosselman S, Looijenga LH, Gillis AJ, et al: Aberrant platelet-derived growth factor alpha-receptor transcript as a diagnostic marker for early human germ cell tumors of the adult testis. Proc Natl Acad Sci U S A 93: 2884-2888, 1996 22. Oosterhuis JW, Gillis AJ, van Roozendaal CE, et al: The platelet-derived growth factor alpha-receptor 1.5 kb transcript: Target for molecular detection of testicular germ cell tumours of adolescents and adults. APMIS 106: 207-213, 1998[Medline] 23. Fan Y, Einhorn L, Saxman S, et al: Detection of germ cell tumor cells in apheresis products using polymerase chain reaction. Clin Cancer Res 4: 93-98, 1998[Abstract] 24. Kiel K, Cremer FW, Rottenburger C, et al: Analysis of circulating tumor cells in patients with multiple myeloma during the course of high-dose therapy with peripheral blood stem cell transplantation. Bone Marrow Transplant 23: 1019-1027, 1999[Medline] 25. Kiel K, Cremer FW, Ehrbrecht E, et al: First and second apheresis in patients with multiple myeloma: No differences in tumor load and hematopoietic stem cell yield. Bone Marrow Transplant 21: 1109-1115, 1998[Medline]
26.
Basser RL, To LB, Collins JP, et al: Multicycle high-dose chemotherapy and filgrastim-mobilized peripheral-blood progenitor cells in women with high-risk stage II or III breast cancer: Five-year follow-up. J Clin Oncol 17: 82-92, 1999 Submitted August 4, 2001; accepted March 15, 2001. This article has been cited by other articles:
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Copyright © 2001 by the American Society of Clinical Oncology, Online ISSN: 1527-7755. Print ISSN: 0732-183X
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ABSTRACT
INTRODUCTION
