A Pileri

Long-Term Follow-Up of Indolent Lymphoma Patients Treated With High-Dose Sequential Chemotherapy and Autografting: Evidence That Durable Molecular and Clinical Remission Frequently Can Be Attained Only in Follicular Subtypes

PURPOSE To evaluate the prognostic relevance of molecular monitoring of minimal residual disease in indolent lymphomas receiving high-dose sequential chemotherapy and autografting. PATIENTS, MATERIALS, AND METHODS A polymerase chain reaction- (PCR-)based strategy was used to evaluate the presence of residual tumor cells in a panel of 70 indolent lymphoma patients: 40 with follicular (FCL), 14 with small lymphocytic (SLL), and 16 with mantle-cell (MCL) lymphomas. They were treated either with first-line (n = 61) or second-line (n = 9) therapy with an intensified high-dose chemotherapy program followed by peripheral-blood progenitor cells autografting. The Bcl-1, Bcl-2, and immunoglobulin gene rearrangements were used as lymphoma-specific markers. Overall, a molecular marker was obtained from the diagnostic tissue in 60 of 70 patients (86%). Results The collection of PCR-negative cells and the achievement of posttransplantation molecular remission (MR) were common in patients with FCL subtype (54% and 70%, respectively), whereas they were not frequent among SLL and MCL (25% and 12.5%, respectively) patients. With a median molecular follow-up of 75 months, an 88% incidence of relapse was observed among patients never attaining MR. In contrast, relapse incidence was only 8% among patients attaining a durable MR (P <.005). At present, 26 patients (20 with FCL and six with non-FCL) are long-term survivors in absence of clinical and molecular disease. CONCLUSION Our results indicate that among indolent lymphomas, FCL and non-FCL subtypes show a significantly different behavior in terms of MR achievement, and MR after intensive chemotherapy and autografting is predictive for a prolonged disease-free survival, whereas persistent PCR positivity is associated with a high risk of relapse.

A novel nested-PCR strategy for the detection of rearranged immunoglobulin heavy-chain genes in B cell tumors.

Several methods have been developed for the detection of minimal residual disease (MRD) in B cell tumors. Chromosomal translocations or the rearrangement of the immunoglobulin heavy chain (IgH) and T cell receptor genes are generally employed. We report a novel PCR method to detect MRD using IgH genes. IgH rearranged variable region (VDJ) were amplified from tumor specimens using consensus primers for variable and joining region genes. Complementarity-determining regions (CDR) were identified and used to generate tumor-specific primers. Two-round amplifications using primers derived from CDRs and joining or constant regions were performed for MRD detection. IgH nested-PCR approach was tested on a panel of 75 B cell tumors including acute lymphoblastic and chronic lymphocytic leukemias, non-Hodgkin’s lymphomas and multiple myelomas. A VDJ sequence was obtained in 62 out of 75 cases (83%). Sensitivity using DNA or cDNA templates was 10−5 and −6, respectively. This method is specific and sensitive and provides a simple, non-radioactive approach for the evaluation of MRD in B cell tumors.

Maspin and mammaglobin genes are specific markers for RT-PCR detection of minimal residual disease in patients with breast cancer

BACKGROUND This study evaluates the specificity of some reverse-transcriptase polymerase chain reaction (RT-PCR) assays for the detection of residual tumor cells in breast cancer patients. The following markers have been analysed: carcinoembryonic antigen (CEA), cytokeratins (CK19 and CK20), polymorphic epithelial mucin (MUC-1), epidermal growth factor receptor (EGFR), maspin, and mammaglobin. RT-PCR was employed to detect breast cancer cells in peripheral blood (PB), bone marrow (BM), and stem cell leukoaphereses (PBPC). PATIENTS AND METHODS We evaluated the specificity of our RT-PCR assays on a panel of breast cancer specimens (n = 30), on PBPC in patients undergoing high-dose chemotherapy (n = 38), on BM (n = 7) and PB (n = 5) samples obtained from patients with breast cancer. Marrow cells, PB, and PBPC from normal subjects or hematological tumor patients were tested as negative controls. RESULTS Only maspin and mammaglobin met the criteria of sensitivity and specificity required for the detection of residual disease; they were expressed in 80% and 97% of breast cancer specimens, respectively, and not expressed in normal controls. CK19, CK20. EGFR, MUC-1, and CEA were sometimes expressed in normal blood cells and/or hematological tumors. CONCLUSIONS Our data support the notion that maspin and mammaglobin are useful markers for RT-PCR detection of minimal residual disease (MRD) in breast cancer patients, and that perspective clinical studies are needed to determine wether RT-PCR assays will be useful in assessing prognosis, tailoring therapy, or developing new strategies for ex vivo purging.

G-CSF administration following peripheral blood progenitor cell (PBPC) autograft in lymphoid malignancies: evidence for clinical benefits and reduction of treatment costs

Clinical value and costs of G-CSF administration following autograft with mobilized peripheral blood progenitor cells (PBPC) were evaluated in two sequential groups of 20 patients each, treated for lymphoid neoplasms in the period February 1993 to January 1996. One group was given G-CSF (Filgrastim) (5 μg/kg/day), starting on day +1 until ANC was >500/μl, the other received no G-CSF. All patients were conditioned with mitoxantrone 60 mg/m2 + L-PAM 180 mg/m2 and received large numbers of PBPC (median of 12 and 13 × 106 CD34+/kg, respectively). The median time to ANC >500/μl was 10 days in the G-CSF group vs 14 days in controls (P < 0.0001). g-csf was associated with a slightly faster platelet recovery (11 vs 13 days to plts >20 000/μl, P = 0.09). Median duration of fever (2.5 vs 5 days, P = 0.028), nonprophylactic antibiotics (8 vs 11 days, P = 0.019), and post-transplant hospitalization (13 vs 16 days, P = 0.0028) were also significantly reduced. The average cost per treatment in the G-CSF group amounted to about US

Long-term follow-up of idiotype vaccination in human myeloma as a maintenance therapy after high-dose chemotherapy

18 241 as compared to US

Overweight as an adverse prognostic factor for non-Hodgkin's lymphoma patients receiving high-dose chemotherapy and autograft

21 868 in the control group, implying a cost reduction of approximately 16%. Thus, G-CSF reduced morbidity with cost containment, supporting its use even if autograft is performed with large quantities of PBPC.

Allogeneic transplantation of unmanipulated peripheral blood stem cells in patients with multiple myeloma

The aim of this work was to evaluate the long-term immunological and clinical impact of idiotype (Id) vaccination in multiple myeloma (MM) patients in first remission after high-dose chemotherapy. A total of 15 patients received a series of subcutaneous (s.c.) injections of autologous Id, conjugated to keyhole limpet hemocyanin (KLH) and in association with low doses of GM-CSF. The median duration of follow-up was 110 months from diagnosis. The vaccine induced immune responses that lasted almost 2 years after the end of treatment. Antibody responses included anti-KLH IgM and IgG (90% of patients), anti-KLH IgE (30%), anti-GM-CSF IgG (20%), anti-Id IgG (20%), and anti-Id IgE (30%). Id-specific delayed type hypersensitivity skin tests were positive in 85% of tested patients. Following vaccination, a progressive recovery of T-cell receptor (TCR) diversity was observed and the loss of oligoclonality was significantly correlated with the remission duration. Although Id/KLH conjugates did not eliminate the residual tumor burden, the median progression-free survival, and overall survival were 40 and 82 months, respectively. A retrospective case-matched analysis showed similar results in patients treated with IFN-α alone or in association with steroids. This vaccine formulation can overcome Id-specific immune tolerance by inducing clinical responses that are worthy of further investigation.

Long-term follow-up of advanced-stage low-grade lymphoma patients treated upfront with high-dose sequential chemotherapy and autograft

Despite detailed evaluation of disease-associated prognostic factors, little is known about the impact of overweight in autograft programs for non-Hodgkins lymphoma (NHL) patients. In order to address this issue, 121 NHL patients were retrospectively evaluated. They had been upfront (92 patients) or in relapse (29 patients) and received high-dose sequential (HDS) chemotherapy including peripheral blood progenitor cell (PBPC) autograft. Body mass index (BMI) was calculated as weight in kilograms divided by the square of the height in meters; overweight was defined as BMI ⩾28. Univariate and multivariate analyses were used to determine the prognostic implication of overweight and other known prognostic indicators on overall (OS) and event-free (EFS) survival for the entire group and overweight and non-overweight (reference) subgroups. With a median follow-up of 3 years, the estimated 5-year OS and EFS for the entire group were 58% and 49%, respectively. Twenty-eight patients (23%) had BMI ⩾28. Their median OS and EFS were 2.2 and 1.4 years, respectively, whereas median OS and EFS for the reference group have not been reached, with a 5-year projection of 65 and 55%, respectively (P < 0.002). On multivariate analysis, the risk of death among overweight patients was 2.9 (CI, 1.3–6.2) times that of the reference group; using EFS as the end point, a similar association between overweight and survival was observed. In conclusion, in high-risk NHL patients undergoing intensive chemotherapy and PBPC autografting overweight is associated with a poorer outcome. Bone Marrow Transplantation (2000) 26, 1185–1191.

Clinical relevance of minimal residual disease monitoring in non-Hodgkin’s lymphomas: a critical reappraisal of molecular strategies

In multiple myeloma (MM), allogeneic bone marrow transplantation may produce complete and durable responses, but is accompanied by significant transplant-related mortality (TRM). To assess feasibility and possible advantages offered by the use of allogeneic, growth factor-primed PBSC instead of marrow, we analyzed the data of 10 patients with MM (IgG = 6, IgA = 1, BJ = 2, non-secreting = 1; stage II = 1, stage III = 8, plasma-cell leukemia = 1) who received an allogeneic transplant with PBSC. Their age ranged between 35 and 53 years (median 45). All were HLA-identical to their sibling donors. Prior to allograft, six patients received standard-dose chemotherapy (DAV or CY-Dexa) and four a sequential intensified scheme with autologous PBSC support. At the time of transplantation, three patients were in CR, three in PR, three had refractory disease, one progressive disease. Patients were conditioned with busulfan–melphalan (n = 9) or busulfan–cyclophosphamide (n = 1), and were allografted with unmanipulated PBSC obtained by apheresis after treatment with G-CSF alone (n = 6) or GM-CSF followed by G-CSF (n = 4). All patients engrafted, with 0.5 × 109/l PMN and 50 × 109/l platelets on (median) day 13. Four patients had ⩾grade II acute GVHD (grade II in 3, grade III in 1). Following allograft, CR was achieved in 71% patients. Eight are currently alive, with six in CR at a median of 18.5 months (range 7–28) from the transplant. Two patients died, 1 and 4 months from the allograft, respectively, and one is alive with progression. A PCR analysis of IgH rearrangement showed that residual disease was no more molecularly detectable in four out of seven evaluated patients following allograft. The results suggest that PBSC may improve the therapeutic efficacy of allogeneic transplant in MM, not only by a reduction of TRM but also by an improvement of rate and quality of response.

Negative immunomagnetic ex vivo purging combined with high-dose chemotherapy with peripheral blood progenitor cell autograft in follicular lymphoma patients: evidence for long-term clinical and molecular remissions.

Long-term outcome, after first line intensified high-dose sequential (i-HDS) chemotherapy, was evaluated in 46 patients, aged ⩽65 years, with advanced low-grade lymphoma. Seventeen patients had small lymphocytic lymphoma (SLL), 29 had follicular lymphoma (FL), 10 of them with histologic transformation. I-HDS included: (1) tumor debulkying, by 2 APO+2 DHAP courses; (2) sequential administration of high-dose (hd) etoposide, methotrexate, and cyclophosphamide, followed by peripheral blood progenitor cell (PBPC) harvest; (3) hd-mitoxantrone + melphalan with PBPC autograft. Ten FL patients had their PBPC immunologically purged ex vivo. There were two treatment-related deaths; five FL patients had short-lasting response followed by disease progression, five SLL reached a stable PR; overall, 34 patients (74%) reached CR. At a median follow-up of 4.3 years, the estimated 9-year OS and EFS were 84% and 45%, respectively. No significant differences were observed in the OS among patients at low, intermediate or high IPI score, with an estimated OS projection of 95%, 78%, and 75%, respectively. FL had longer survival without evidence of residual disease (9-year EFS: 59%) as compared to SLL patients (8.8-year EFS: 17%); however, both groups had prolonged survival and no need of salvage treatment, as shown by the time to disease progression curve, projected to 66% and 62% for SLL and FL, respectively. The results indicate that hd-approach in low-grade lymphoma: (1) is associated with longer progression-free survival as compared to conventional therapies; (2) may imply higher tumor mass reduction in FL as compared to SLL patients; (3) offers long life expectancy, with potential survival benefits at least for patients at intermediate/high IPI score.