Quantitative detection of t(14;18)-positive cells in patients with follicular lymphoma before and after autologous bone marrow transplantation

Hirt, Carsten; Dölken, Gottfried

doi:10.1038/sj.bmt.1702147

Cited by 43 publications

(26 citation statements)

References 35 publications

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“…8 Unlike CML, studies to determine the clinical significance of the tumor load in FL after SCT are very limited. 9 Hirt et al 10 have recently reported that relapses are associated with increasing numbers of circulating t(14;18)-positive cells, while continuous complete clinical remissions are associated with stable t(14;18)-positive cell counts in seven patients following autologous SCT. Mandigers et al 11 reported that levels of tumor load correlated with remission after using DLI to treat the relapse of FL following allogeneic SCT in one patient.…”

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confidence: 99%

Tumor load in patients with follicular lymphoma post stem cell transplantation may correlate with clinical course

Bredeson

Juckett

et al. 2003

Bone Marrow Transplant

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Summary:The purpose of this study was to evaluate if the tumor load, as determined by a real-time quantitative PCR (RQ-PCR) assay, correlated with the clinical course of follicular lymphoma patients after stem cell transplantation (SCT). Cryopreserved bone marrow and/or peripheral blood samples obtained at different time intervals after SCT from 11 patients (seven allogeneic, T-cell depleted/four autologous) were tested for tumor load, as defined by t(14;18) positive cells/total cells, using RQ-PCR. None of the six patients who remained in remission had samples with a tumor load 40.01% after SCT, although fluctuating tumor loads of p0.01% were observed in three of these patients. In contrast, four of the five patients (three allogeneic/two autologous) with relapsed/progressive disease had increasing tumor loads of 40.01% after SCT (0/6 vs 4/5, Po0.02, Fisher's exact). Our results suggest that RQ-PCR measurable tumor load 40.01% after SCT may correlate with relapsed/progressive disease. Prospective studies with greater numbers of cases are indicated to better determine the critical tumor load that predicts poor outcome after SCT with RQ-PCR.

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confidence: 99%

Tumor load in patients with follicular lymphoma post stem cell transplantation may correlate with clinical course

Bredeson

Juckett

et al. 2003

Bone Marrow Transplant

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“…Several studies have recently demonstrated that achievement of molecular complete remission is a desirable goal of new therapies because patients with molecular complete remission have longer disease-free status (16 -18). The clinical importance of achieving molecular response has been demonstrated in FL patients treated with several regimens, including rituximab (18,19) and stem cell therapy (3). Moreover, quantitative results may provide more prognostic information in the assessment of minimal residual disease detected by PCR analysis of peripheral blood, bone marrow, and stem-cell preparations used for transplantation after high-dose chemotherapy.…”

Section: Discussionmentioning

confidence: 99%

Real-Time t(14;18)(q32;q21) PCR Assay Combined with High-Resolution Capillary Electrophoresis: A Novel and Rapid Approach that Allows Accurate Quantitation and Size Determination of bcl-2/JH Fusion Sequences

et al. 2002

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Follicular lymphoma is characterized by the presence of the t(14;18)(q32;q21) chromosomal translocation that juxtaposes the bcl-2 gene at 18q21 with the immunoglobulin heavy chain (IgH) locus at 14q32. We have previously shown that accurate quantitation of t(14;18)-carrying cells in follicular lymphoma patients can be achieved by non-gelbased real-time TaqMan polymerase chain reaction (PCR; Applied Biosystems, Foster City, CA). Since our report, several studies have demonstrated that real-time PCR is highly sensitive and a reliable tool for evaluating treatment effectiveness and for following minimal residual disease in follicular lymphoma patients. Unfortunately, currently available real-time PCR methods do not determine the size of the amplification product, which is useful for excluding contamination and is commonly used as presumptive evidence of clonal identity or disparity when multiple samples from the same patient are analyzed. We describe a modified real-time PCR assay that rapidly allows accurate quantitation and precise determination of the size of the t(14;18) fusion sequence without the need for gel electrophoresis. In this assay, a consensus immunoglobulin heavy chain-joining region gene (JH) primer labeled at its 5' end with the fluorescent dye NED (Applied Biosystems) is included in the real-time PCR assay and thus is incorporated into the bcl-2/JH fusion product. The JH-NED primer did not interfere with the TaqMan probe fluorescent signal or target detection and allowed subsequent amplicon size determination by semiautomated highresolution capillary electrophoresis.KEY WORDS: Follicular lymphoma, High-resolution capillary electrophoresis, Real-time polymerase chain reaction, t(14;18). Mod Pathol 2002;15(4):448 -453Real-time polymerase chain reaction (PCR) for detection and quantitation of bcl-2/JH fusion sequences characteristic of t(14;18) (q32;q21) is being used increasingly to monitor tumor burden and the effect of therapy in patients with follicular lymphoma (FL; 1-5). Real-time PCR methods enable t(14;18) detection without the need to open amplification tubes and are target specific, highly sensitive, and reproducible. Thus, real-time PCR methods have several advantages over conventional PCR techniques that require gel electrophoresis-based amplicon detection and quantitation techniques. Currently available real-time PCR methods to detect the t(14;18) do not allow amplicon size determination. This is a major drawback for a few reasons. First, one cannot easily exclude contamination. Second, size of PCR products from multiple samples (either simultaneous or sequential) is a valuable means of determining clonal relatedness, as the size of bcl-2/JH fusion sequences generally vary from clone to clone (6 -8). Undoubtedly, sequencing of the fusion sequences is the most accepted method for determining clonal relatedness, but sequencing methods can be tedious and time consuming. Third, as t(14;18)-positive cells are detected in normal healthy individuals (9 -12), the ability to quantitate low-level...

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“…This supports the concept as previously expressed in a nontransplant setting that genetic abnormalities in addition to t(14;18) are required for lymphoma- 16 Similar to our observations, increasing quantitative levels of cells carrying t(14;18) seen with serial determinations has been shown to correlate with FL recurrence after autologous bone marrow transplant. 19 Our findings further denote that it may be essential to screen for the donor's t(14;18) status before using t(14;18) as the target for monitoring minimal residual disease (MRD) by RQ-PCR in FL patients after alloSCT to determine if there is a confounding donor's t(14;18) clone. We demonstrated that the significance of post transplant presence of t(14;18) in the recipients might be affected by whether the recipients had received grafts from donors carrying t(14;18) cells or not.…”

Section: Discussionmentioning

confidence: 99%

“…Our results indicate that the sensitivity, reproducibility, and linearity of this assay are the same, if not better, than other recently reported RQ-PCR assays for detecting t(14;18). [18][19][20][21] Patients' samples were tested in duplicate initially. If both runs were negative, an additional two runs were performed.…”

Section: Rq-pcrmentioning

confidence: 99%

The implication of follicular lymphoma patients receiving allogeneic stem cell transplantation from donors carrying t(14;18)-positive cells

Keever-Taylor

Bredeson

et al. 2005

Bone Marrow Transplant

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Summary:We performed real-time quantitative polymerase chain reaction (RQ-PCR) in peripheral blood (PB) and/or bone marrow (BM) samples collected pre-and post transplant from 23 recipient-donor pairs receiving allogeneic stem cell transplantation (allo-SCT) for follicular lymphoma (FL). Of 23 donors, 11 had a PB and/or BM sample positive for t(14;18) (BCL2/IGH fusion) at low levels (oone t(14;18) cell in 10K total cells). Recipients from donors with (n ¼ 11) and those without (n ¼ 12) detectable t(14:18) cells were similar in age, sex, and disease status pretransplant. No differences in the incidence of graft-versus-host-disease (GVHD), delayed engraftment, relapse rate, disease-free survival and overall survival were identified between the groups. Two recipients without detectable t(14;18) cells pretransplant showed detectable t(14;18) cells at 2 and 11 years after receiving grafts from donors with t(14:18) cells. Neither patient developed FL 1.5 and 2 years after the emergence of t(14;18) cells. Although the sample size is relatively small, our findings suggest that individuals carrying t(14;18) cells may not be excluded as donors given the lack of an association of t(14;18) detected in donors with adverse clinical outcome. It may be necessary to screen for the donor's t(14;18) status before using t(14;18) for monitoring minimal residual disease by RQ-PCR to exclude the possibility of confounding donor's t (14;18) [4][5][6][7][8][9][10][11] Allogeneic stem cell transplantation (allo-SCT) is of increasing interest as a therapeutic option for patients due to the potential of a graft-versus-lymphoma effect to provide long-term disease control. [12][13][14][15] We have recently reported that monitoring the tumor load of t(14;18) cells by real-time quantitative PCR (RQ-PCR) may predict the outcome of FL patients after allo-SCT. 16 Patients with tumor loads of greater than one t(14;18) cell per 10K total cells post transplantation have a higher risk of clinical relapse than those without. The monitoring of minimal residual disease (MRD) by RQ-PCR allows for the development of strategies, such as donor lymphocyte infusions to be employed at a stage of molecular relapse in patients whose RQ-PCR results suggest a high likelihood of subsequent clinical relapse.The utility of using RQ-PCR to monitor tumor load or MRD after allo-SCT, however, can be confounded by the presence of t(14;18) cells of donor origin. We recently described the disappearance of a patient's t(14;18) clone early post transplant with the concomitant emergence and long-term persistence of the donor's t(14;18) clone without development of FL. 17 Although it has not been well studied, donor carrying t(14;18) clones detectable by RQ-PCR could theoretically develop into FL in an immunosuppressed recipient who may have a genetic susceptibility to develop FL. In addition, the presence of t(14;18) clones in the donor could serve as a surrogate marker of a stem cell defect and thus affect transplant outcome in some other manner. Thus, the aims of the current ...

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Quantitative detection of t(14;18)-positive cells in patients with follicular lymphoma before and after autologous bone marrow transplantation

Cited by 43 publications

References 35 publications

Tumor load in patients with follicular lymphoma post stem cell transplantation may correlate with clinical course

Tumor load in patients with follicular lymphoma post stem cell transplantation may correlate with clinical course

Real-Time t(14;18)(q32;q21) PCR Assay Combined with High-Resolution Capillary Electrophoresis: A Novel and Rapid Approach that Allows Accurate Quantitation and Size Determination of bcl-2/JH Fusion Sequences

The implication of follicular lymphoma patients receiving allogeneic stem cell transplantation from donors carrying t(14;18)-positive cells

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