Phase II study of a moderate-intensity preparative regimen with allogeneic peripheral blood stem cell transplantation for hematologic diseases: the Texas Transplant Consortium experience.

Conventional preparative regimens for allogeneic stem cell transplantation are associated with excessive regimen-related toxicity (RRT) in some patients because of underlying comorbidities, advanced age, or prior treatment. We studied a preparative regimen designed to reduce RRT, yet allow for adequate engraftment and development of a graft-versus-malignancy effect. Thirty patients (median age, 57 years) were entered on study. Twenty-nine patientsreceived stem cells from HLA-identical siblings and 1 from a sibling mismatched for 1 antigen at the A locus. Sixteen patients had received previous stem cell transplants (6 allogeneic and 10 autologous). The preparative regimen consisted of fludarabine 30 mg/M2 per day IV on day -10 to day -5, busulfan 1 mg/kg per dose PO (n = 6) or 0.8 mg/kg per dose IV (n = 24) for 8 doses every 6 hours on day -6 to day -5, and horse-derived antithymocyte globulin 5 mg/kg per day IV (n = 12) or 15 mg/kg per day IV (n = 18) on day -4 to day -1. GVHD prophylaxis consisted of cyclosporine (CYA) 3 mg/kg BID PO starting on day -3 (n = 13) or CYA and methotrexate 15 mg/m2 IV on day +1 and 10 mg/m2 IV on day +3 and day +6 (n = 17). The median number of CD34 cells transplanted was 3.19 x 10(6)/kg. All patients demonstrated recovery of hematopoietic function. Twenty-six (89%) of 29 evaluable patients achieved greater than 90% donor cell chimerism before day 100. Three patients never achieved greater than 90% donor chimerism, and another 3 patients subsequently lost donor chimerism. All 6 of these patients had autologous reconstitution with progressive disease. RRT was minimal; 7 patients had greater than grade II nonhematologic toxicity and there were no toxic deaths attributable to the conditioning regimen. Transplantation-related mortality was 7% (95% confidence interval [CI], 6%-8%) at 3 months and 28% (95% CI, 23%-34%) at 12 months after transplantation. Non-relapse-related mortality was most often due to infection. Grade II or greater GVHD developed in 56% of evaluable patients, and all patients with disease response developed GVHD. Actuarial estimates of overall and disease-free survival at 12 months were 52% (95% CI, 43%-63%) and 30% (95% CI, 24%-37%), respectively. Although this preparative regimen allowed adequate engraftment with minimal RRT, GVHD and infectious complications caused significant morbidity and mortality. Further study to define appropriate patient populations for this regimen, while limiting GVHD and infection risks, is needed.

A phase II trial evaluating the safety and effectiveness of the AastromReplicell system for augmentation of low-dose blood stem cell transplantation.

To reduce the number of apheresis procedures and maintain the usual rate of hematopoietic recovery in patients treated with high-dose chemotherapy, we studied the effect of adding a small volume of ex vivo expanded bone marrow to low doses of CD34(+) blood stem cells. Thirty-four patients with breast cancer received G-CSF (10 microg/kg/day) priming followed by a limited volume (50-100 ml) bone marrow aspiration and standard 10-liter aphereses. Marrow was expanded ex vivo using the AastromReplicell system and infused along with low doses of blood-derived CD34(+) cells, collected in one apheresis. Thirty-one evaluable patients received a median CD34(+) blood stem cell dose of 0.7 x 10(6)/kg (range, 0.2-2.5) and 4.7 x 10(7) nucleated cells/kg (range, 1.98-8.7) of ex vivo expanded marrow. All patients recovered with normal blood counts and engrafted 500 neutrophils/microl and 20 000 platelets/microl in a median of 10 and 13 days, respectively. Multivariate analysis revealed that, in addition to CD34(+) lineage negative cell quantity, the quantity of stromal progenitors contained in the ex vivo expanded product correlated with engraftment outcome (r = 0.551, P = 0.004). Our results indicate that ex vivo expanded bone marrow is capable of facilitating engraftment when combined with low doses of mobilized blood derived CD34(+) cells.

Graft failure in a patient with systemic lupus erythematosus (SLE) treated with high-dose immunosuppression and autologous stem cell rescue.

A 32-year-old female with WHO grade IV, dialysis dependent, lupus nephritis was treated with high-dose immunosuppression and autologous stem cell rescue. Stem cells were mobilized with cyclophosphamide (CY) and G-CSF, and 4.07 x 10(6) CD34+ cells/kg were obtained after CD34+ cell selection using the CellPro column. The preparative regimen consisted of CY, and antithymocyte globulin (ATG), with methylprednisolone. After apparent primary engraftment of neutrophils on day 9, the patient developed recurrent neutropenia on day 19. She showed no evidence of engraftment by day 35, and back-up unmanipulated stem cells were given without effect. Subsequently, she received unmanipulated peripheral stem cells (2 x 10(6) CD34+ cells/kg) from an HLA-identical sibling. The patient remained pancytopenic and expired on day 62 from disseminated fungal infection. An autopsy revealed no evidence of hematopoietic recovery. Progenitor cell assays were performed with the patient's stem cells, which were collected prior to transplantation, and serum collected day 27. Morphologic examination of the patient's cell colonies grown in the presence of her serum revealed abnormal shapes and non-adherent cells. There were significantly fewer BFU-e colonies and a trend toward fewer CFU-GM colonies with the patient's cells and serum compared to normal donor cells. We concluded that a substance present in her serum mediated graft failure and prevented engraftment after additional stem cell infusions.

Ig heavy chain CDR3 size diversities are similar after conventional peripheral blood and ex vivo expanded hematopoietic cell transplants.

It is largely unknown whether the immune repertoire can be reconstituted successfully after high-dose chemotherapy and transplantation using ex vivo expanded hematopoietic stem cell (HSC) grafts. It is critically important for the transplant outcome that immune repertoire reconstitution progresses after ex vivo expanded HSC graft transplants at least as efficiently as that seen after conventional HSC transplants. Previously, we showed that the T cell receptor V beta (TCRVB) third complementarity determining region (CDR3) diversification after ex vivo expanded bone marrow (BM) HSC graft transplants was similar to that seen after conventional peripheral blood stem cell transplants (PBSCTs). In the present study, the CDR3 diversity of the six immunoglobulin (Ig) heavy chain variable region gene (V(H)) families was examined in five breast cancer patients who were transplanted with ex vivo expanded BM HSCs as the only source of stem cells. For comparison, 12 healthy adults and four conventional PBSCT recipients were also studied. Using both CDR3 fingerprinting and single strand conformation polymorphism (SSCP) methodologies, it is shown that the contribution of the V(H) families to the overall repertoire among healthy adults is highly variable and not always proportional to V(H) family member size. After both ex vivo expanded HSC transplants and conventional PBSCTs, the V(H) CDR3 repertoires were limited in size diversity at 6 weeks post transplant. By 6 months, however, V(H) families displayed a repertoire diversity that was as complex as that seen in healthy adults. No difference was seen between ex vivo expanded HSC graft transplant recipients and conventional PBSCT recipients in V(H) repertoire diversity. In one patient there was a follow-up analysis 12 months after ex vivo expanded graft transplant, and the diversity of the V(H) families was maintained. In all patients, the amino acid size of the CDR3 regions fell within adult limits at all time points post transplant. These results indicate that B cell repertoire regeneration after ex vivo expanded hematopoietic cell graft transplants is similar to that seen after conventional PBSCT.

Ex vivo-expanded hematopoietic cell graft recipients exhibit T cell repertoire diversity similar to that seen after conventional stem cell transplants.

The feasibility of using ex vivo-expanded hematopoietic progenitor cells to reconstitute hematopoiesis after high-dose chemotherapy is presently being examined. Early studies have shown that myeloid and erythroid hematopoiesis can be successfully reconstituted after high-dose chemotherapy and ex vivo-expanded hematopoietic cell transplantation. The lymphoid reconstitution, however, has not been addressed previously. In this study, we examined the diversity of the T cell receptor V beta chain (TCRBV) repertoires in 5 breast cancer patients who were transplanted with ex vivo-expanded bone marrow mononuclear cells as the only source of hematopoietic graft. Using the TCRBV third complementarity determining region (CDR3) fingerprinting methodology, it is shown that CD4(+) and CD8(+) T cell subsets after ex vivo-expanded hematopoietic cell graft transplants exhibit TCRBV diversities that are similar in complexity when compared to those seen after conventional autologous peripheral blood stem cell transplants (PBSCT). No apparent difference in the extent of CDR3 diversity was found between ex vivo expanded and conventional autologous PBSCT recipients when the CD4(+) and CD8(+) subsets were further separated into CD45RA(+) "naïve" and CD45RO(+) "memory" subsets. The diversity of the CD45RA(+) naïve subsets was as complex as that of the CD45RO(+) memory subsets. These results indicate that T cell repertoire diversification is not further compromised when ex vivo-expanded hematopoietic cells are used instead of autologous peripheral blood stem cells as the only source of graft.

Infusional vinorelbine in relapsed or refractory lymphomas.

Vinorelbine (Navelbine is a semisynthetic vinca alkaloid devoid of serious neurotoxicity. When given weekly vinorelbine has documented activity against many tumors, including lymphomas. Since weekly schedules cannot be easily incorporated in combination regimens, we tested an infusional schedule of vinorelbine given every 21 days in adults with relapsed or refractory lymphoma. Patients with inadequate organ or bone marrow reserve, HIV or other serious infection, central nervous system disease, or prior stem cell or bone marrow transplantation were ineligible. In the phase I part, patients received a constant intravenous bolus of 8 mg/m(2), followed by intravenous continuous infusion over 24 hours daily for four days increasing from 10, 12, to 14 mg/m(2) /d in successive three-patient cohorts. Cycles were repeated every 21 days, and the daily continuous infusion dose was adjusted for toxicity. Dose-limiting mucositis and neutropenia were reached at the continuous dose of 14 mg/m(2) /d. Consequently, for the Phase II trial the starting continuous infusion dose was 12 mg/m(2) /d. After the first 19 patients were entered in the phase II study, the starting infusion dose was reduced to 10 mg/m(2) /d because of frequent grade (3/4) myelosuppression and mucositis. Forty-four patients were entered in the phase II study, of whom 41 are evaluable. Median age was 61 years, 23 were males, with clinically aggressive non-Hodgkin's lymphoma (NHL) in 22, indolent NHL in 18, and Hodgkin's Disease in one patient. The median number of prior regimens was 3 (range 1-11). The lymphoma was refractory to the initial regimen in nine patients, and to the regimen immediately before vinorelbine in 20 patients. Serum LDH was high in 2(1/4)1, and serum beta(2) -microglobulin > 3.0 mg / L in 16/31 patients. Responses were observed in four of 22 patients with aggressive NHL (18%, 95% confidence interval 5%-40%), and in six of 18 with indolent NHL (33%, 95% confidence interval 13%-59%). Median progression-free survival was 6 months for responders. During the Phase II trial 114 vinorelbine courses were administered. Neutrophil nadir was < 1000/microl in 65% and < 100/microl in 35% of courses, respectively. Platelet nadir was < 100,000/microl in 30% and < 20,000/microl in 8% of courses, respectively. Grade (3/4) mucositis was seen in 18% of courses, and neutropenic fever in 13%, and was complicated by death in one patient. We conclude that this dosage and schedule of vinorelbine has modest activity in patients with relapsed or refractory NHL. Myelosuppression is frequent but reversible, but there is no significant neurotoxicity. The role of vinorelbine in combination regimens for patients with relapsed lymphomas, particularly those of indolent histology, should be further investigated.

Ex-vivo expansion of bone marrow progenitor cells for hematopoietic reconstitution following high-dose chemotherapy for breast cancer.

The use of hematopoietic growth factors, stromal monolayers, and frequent medium exchange allows the expansion of hematopoietic progenitors ex-vivo. We evaluated the use of ex-vivo expanded progenitor cells for hematopoietic reconstitution following high dose chemotherapy (HDC) in breast cancer patients. Patients with high-risk Stage II or metastatic breast carcinoma underwent bone marrow aspirations using general anesthesia. A total of 675-1125 x 10(6) mononuclear cells (MNC) were seeded for ex-vivo expansion for 12 days in controlled perfusion bioreactors (Aastrom Biosciences, Inc.). The bone marrow cultures, which included the stromal cells collected with the aspirate, were supplemented with erythropoietin, granulocyte-macrophage-colony stimulating factor (GM-CSF)/IL-3 fusion protein (PIXY 321), and flt3 ligand. Stem cell transplant was performed with expanded cells after HDC. A median bone marrow volume of 52.9 mL (range 42-187 mL) was needed to inoculate the bioreactors. Median fold expansion of nucleated cells (NC) and colony forming unit granulocyte-macrophage (CFU-GM) was 4.9 and 9.5, respectively. The median fold expansion of CD34+lin- and long-term culture-initiating culture (LTC-IC) was 0.42 and 0.32, respectively. Five patients were transplanted with ex-vivo expanded NC. Median days to an absolute neutrophil count > 500/microL was 18 (range 15-22). Median days to a platelet count > 20,000/microl was 23 (range 19-39). All patients had sustained engraftment of both neutrophils and platelets. Immune reconstitution was similar to that seen after HDC and conventional stem cell transplantation. We conclude that ex-vivo expansion of progenitor cells from perfusion cultures of small volume bone marrow aspirates, allows hematopoietic reconstitution after HDC.

The presence of the Rb c-box peptide in the cytoplasm inhibits p210bcr-abl transforming function.

In order to test if the carboxyl terminal polypeptide of the Retinoblastoma (Rb) tumor suppressor protein, could be used to suppress the growth factor-independent growth phenotype of p210bcr-abl positive myeloid cells, we introduced a truncated form of the 3' end of the Rb cDNA encoding its last 173 amino acid residues (Rb C-box) which localize into the cytoplasm where the p210bcr-abl transforming protein is found, into myeloid cells (32D) which depends on the p210bcr-abl protein for IL3 growth factor-independent growth (32D-p210). The expression of the plasmid vectors carrying the Rb C-box cDNAs was shown to inhibit the abl tyrosine specific protein kinase activity of the p210(bcr-abl) oncoprotein and to suppress the IL3-independent growth phenotype of the 32D-p210 cells. The Rb C-box polypeptides did not suppress the growth of the untransfected 32D parental cell line in methylcellulose in the presence of IL3-conditioned medium. These results suggest that the cytoplasmic localization of the p210(bcr-abl) allows it to escape the effect of intranuclear proteins such as Rb which negatively regulate the p145(c-abl) kinase.

Hematopoietic retroviral gene marking in patients with follicular non-Hodgkin's lymphoma.

We conducted a double retroviral vector (RV) gene marking trial to test for the possible contribution to relapse of follicular non-Hodgkin's lymphoma (FNHL) cells present in bone marrow (BM) and peripheral blood (PB) grafts used for hematopoietic reconstitution of patients undergoing myelaoblative chemotherapy and autologous transplant. CD34 positive selection using the CellPro Ceprate CD34 column was performed on PB mononuclear cells obtained after cyclophosphamide/G-CSF mobilization. CD34 positive cells were exposed for 4-6 hours to the LNL6 or G1 Na RV in the absence of growth factors or stromal monolayers. One week later, BM mononuclear cells were similarly processed. Patients then received total body irradiation (TBI), cyclophosphamide, and etoposide followed by infusion of both PB and BM CD34 positive cells. Semiquantitative Southern blot analysis of DNA t(14;18) amplification products showed approximately a three log reduction in t(14;18) positive cells after CD34 positive selection. The first patient showed evidence of engraftment with RV positive BM and PB cells for 9 months. He relapsed one year after transplant. At relapse, one year after transplant, he had lost evidence of RV positive cells in ficolled mononuclear BM and PB cells as well as in CD19 positive cells. The second and third patients showed evidence of engraftment with RV positive cells up to 9 and 6 months post BMT respectively. The second and third patients are still in clinical remission. Our results demonstrate engraftment of RV transduced hematopoietic cells in the PB and BM for up to 9 months.

Paclitaxel plus high-dose cyclophosphamide with G-CSF support in patients with relapsed and refractory aggressive non-Hodgkin's lymphoma.

Based on the single-agent activity of both paclitaxel and cyclophosphamide in the treatment of non-Hodgkin's lymphoma (NHL), we conducted a phase II study to evaluate the efficacy of the combination of the two drugs in patients with refractory and relapsed aggressive NHL. All patients received 900 mg/m2 bolus of cyclophosphamide intravenously daily for 3 consecutive days with a concurrent infusion of 150 mg/m2 of paclitaxel over 72 h (50 mg/m2/d). 24 h after the completion of chemotherapy, patients received subcutaneous injections of 5 microg/kg of granulocyte-colony stimulating factor (G-CSF) daily until white cell count recovery. Treatment was repeated every 3 weeks. Patients who had at least a partial response (PR) after two courses continued to receive a maximum of four courses. Patients with responding disease were allowed to undergo high-dose chemotherapy followed by stem-cell/bone marrow transplantation if they were eligible. Of the 77 patients who were eligible for the study, 74 (96%) were evaluable for toxicity and treatment response. The overall response rate was 45% (95% CI 33-57%). Patients who received treatment after their disease relapsed from a complete response (CR) had an 81% response rate (38% CRs), whereas those with primary refractory disease had a 22% response rate. Toxicities of > grade 2 included alopecia (100%) and stomatitis (25%). Neutropenic fever of grade > 2 occurred after 18% of the courses, and platelet count of < or = 20 x 10(9)/l developed after 20% of the courses. Thus, the combination of paclitaxel plus high-dose cyclophosphamide is an effective new regimen in the treatment of refractory and relapsed NHL.

Chemotherapy immediately following autologous stem-cell transplantation in patients with advanced breast cancer.

Most patients relapse after high-dose chemotherapy (HDCT) with autologous stem-cell transplantation (ASCT) for metastatic breast cancer. Further chemotherapy immediately after hematopoietic recovery from ASCT is not given for fear of irreversibly damaging the newly engrafted stem cells. In a pilot chemoprotection trial, autologous CD34+ cells from patients with metastatic breast cancer were exposed to a replication-incompetent retroviral vector carrying MDR-1 cDNA and then reinfused after HDCT. Immediately on recovery, patients received multiple courses of escalating dose paclitaxel. All of the 10 patients tolerated reinfusion of modified cells without any toxicity and had myeloid engraftment within 12 days (range, 11-14). The bone marrow cells of three patients contained vector MDR-1-positive cells only at the time of the first course of posttransplant paclitaxel, indicating that the MDR-1 vector-modified cells had only short-term engrafting potential. A total of 83 courses of paclitaxel were administered starting at a median of 30 (range, 21-32) days from ASCT. The median dose of paclitaxel was 225 mg/m2 and the median interval between paclitaxel cycles of therapy was 21 (range, 20-41) days. Five of the six CR patients were able to receive all of the 12 courses of paclitaxel. Three patients who had achieved less than a complete response to the HDCT (2 patients) and partial response (1 patient) were converted to complete clinical responses during the 12 cycles of paclitaxel. No delayed toxicity or bone marrow failure was noted in these patients with a median follow-up of 2 years from ASCT. This is the first study of chemotherapy immediately after transplantation with autologous CD34+ cells. These data indicate that paclitaxel can be safely administered immediately after ASCT without any delayed toxicities. Paclitaxel given immediately after ASCT can further improve the response to pretransplant chemotherapy in patients with advanced breast cancer.

Peptide containing the BCR oligomerization domain (AA 1-160) reverses the transformed phenotype of p210bcr-abl positive 32D myeloid leukemia cells.

We first showed that the introduction of a bcr-abl transcription unit into the 32D murine myeloid cell line (P210bcrabl32D) converts this cell line from an IL3 dependent cell line to an IL3 growth independent cell line. We next cloned a fragment of the bcr-abl cDNA, which codes for the bcr oligomerization domain and neighboring regions. To test for a transformation inhibitory effect of this oligomerization inhibitory peptide transcription unit on the p210bcr-abl mediated IL3 independent growth of the P210bcrabl32D cell line, we transiently co-electroporated into the growth factor dependent 32D cells, mixtures of plasmids which contained varying ratios of the plasmid expression vectors for the bcr oligomerization inhibitory peptide along with a smaller amount of the plasmid expression vector for the full length p210bcr-abl. (The P210bcr-abl protein converts the 32D from a growth factor dependent into a growth factor independent cell line.) We then showed that the oligomerization domain containing fragment from the bcr and bcr-abl proteins, can be used to inhibit the IL3 independent growth of p210bcr-abl positive 32D cells. These studies may be of eventual interest for those investigators whose goal is to design molecular therapeutic approaches to CML based on the use of peptidomimetic chemical functionalities, which mimic the structure and the inhibitory binding properties of the oligomerization domain containing fragment so as to inhibit the transforming function of the P210bcr-abl oncoprotein.

Risk factors, treatment, and outcome of central nervous system recurrence in adults with intermediate-grade and immunoblastic lymphoma.

To evaluate the incidence, risk factors, and outcome of central nervous system (CNS) recurrence in adult patients with non-Hodgkin's lymphoma, we evaluated 605 newly diagnosed patients with large-cell and immunoblastic lymphoma who participated in prospective chemotherapy studies. The Kaplan-Meier estimate of probability of CNS recurrence at 1 year after diagnosis was 4.5% (95% confidence interval [CI], 4.4 to 4.6). Twenty-four patients developed CNS recurrence after a median of 6 months from diagnosis (range, 0 to 44 months). In univariate analysis, an increased risk for CNS recurrence was associated with an advanced disease stage (P = .0014), an increased LDH (P = .0000), the presence of B-symptoms (P = . 0037), involvement of more than one extranodal site (P = .0000), poor performance status (P = .0005), and B-cell phenotype (P = .008). Bone marrow involvement (P = .005), involvement of parenchymal organs (P = .03), and involvement of skin, subcutaneous tissue, and muscle (P = .002) were also associated with an increased risk for CNS disease. Multivariate logistic regression analysis identified only involvement of more than one extranodal site (P = .0005) and an increased LDH (P = .0008) as independent predictors of CNS recurrence. Established CNS recurrence had a poor prognosis. Only 1 of 24 patients remains alive and the Kaplan-Meier estimate of probability of survival at 1 year after the diagnosis of CNS recurrence is only 25.3% (95% CI, 6.9 to 43.7). Intrathecal treatment provided symptomatic benefit in only 1 of 6 patients. Radiation treatment provided symptomatic improvement in 6 of 9 patients treated. However, remissions were short and followed by systemic or CNS recurrence. Serum LDH and involvement of more than one extranodal site are independent risk factors for CNS recurrence in patients with large-cell lymphoma. The presence of both risk factors identifies a patient group at high risk for CNS recurrence. Established CNS recurrence can be rapidly fatal. Transient responses occur after radiation treatment.

Use of the international prognostic index and the tumor score to detect poor-risk patients with primary mediastinal large B-cell lymphoma: a study of 37 previously untreated patients.

We tested two prognostic models devised for intermediate-grade lymphomas, the age-adjusted international prognostic index and the tumor score, in 37 consecutive untreated patients treated for a diagnosis of primary mediastinal large B-cell lymphoma (PMLCL). Neither model selected for a group of patients with statistically significant differences in rates of complete response, failure-free survival (FFS) and overall survival (OS). Because the level of beta microglobulin (beta2m) is consistently low in the serum of patients with PMLCL despite bulky disease, we tested the median value of this continuous variable in the 37 patients and found it to be statistically significant for predicting FFS. A hypothetical tumor score model using the adjusted value for beta2m improved the prognostic accuracy for achievement of complete response (93% vs. 60%; P = 0.02), FFS (73% vs. 35%; P = 0.02), and OS (80% vs. 55%; P = 0.05). This hypothetical model merits further testing in a larger population of patients with PMLCL.

Ninety-six-hour paclitaxel infusion with mitoxantrone and ifosfamide/mesna and consolidation with ESHAP for refractory and relapsed non-Hodgkin's lymphoma.

A prospective phase II study was carried out in 48 patients with relapsed or refractory non-Hodgkin's lymphoma using paclitaxel 27.5 mg/M2 i.v. by continuous infusion over 24 hours daily on days 1, 2, 3, and 4 in combination with mitoxantrone 8 mg/M2 i.v. on day 1 and ifosfamide/mesna 1.33 grams/M2 i.v. daily on days 1, 2, and 3 (MINT). Responding patients completed four cycles of MINT and were consolidated with etoposide, solumedrol [methylprednisolone], high-dose cytarabine [Ara-C], and platinum (ESHAP). Forty-eight patients were entered in the study between 1994 and 1996 at The University of Texas M. D. Anderson Cancer Center. Overall response after the first four cycles of MINT was 67% (16% complete response [CR]+ 51% partial response [PR]) and after consolidation with ESHAP it was 49% (26% CR + 23% PR). Variables associated with an improved CR rate and better failure-free survival included the number of prior treatments and the response to prior treatment. A comparison with a similar group of patients treated with mesna, ifosfamide, mitoxantrone, and etoposide (MINE)-ESHAP revealed no major differences in outcome.

Phase I study of fludarabine and paclitaxel for the treatment of low-grade non-Hodgkin's lymphoma.

We conducted a phase I clinical trial of a new combination of fludarabine and paclitaxel in which 19 patients with histologically confirmed recurrent low-grade non-Hodgkin's lymphoma (NHL) were treated at five dose levels. Fludarabine was administered intravenously by bolus for 5 days and paclitaxel was given by intravenous (I.V.) continuous infusion for 96 or 72 hours starting day 1. Courses were repeated every 4 weeks. Patients whose disease responded received a maximum of six courses. All 19 patients received at least one course and could be evaluated for toxic effects, and 18 patients could be evaluated for response. The maximum tolerated dose (MTD) was 20 mg/m2/day I.V. bolus for 5 days of fludarabine plus 60 mg/m2/day I.V. of paclitaxel given as a continuous infusion over 72 hours. The limiting toxic effect was neutropenic fever, which was observed in five of the seven patients treated at the highest dose level. Grade 3 non-hematologic toxic effects of stomatitis (14%), neuropathy (14%), and hypotension (14%) were also observed at the highest dose level. No grade 4 non-hematologic toxic effects or treatment-related deaths occurred. One patient had herpes zoster infection of the skin 1 year after the completion of therapy. The overall response rate was 50%, with the two patients whose disease completely responded remaining disease free at 22 and 17 months. Patients with no prior exposure to either paclitaxel or fludarabine had 62% response rate. We conclude that the combination of fludarabine and paclitaxel appears to have promising activity for the treatment of recurrent low-grade NHL.

Paclitaxel activity for the treatment of non-Hodgkin's lymphoma: final report of a phase II trial.

In order to determine the activity of paclitaxel in patients with relapsed or refractory non-Hodgkin's lymphoma (NHL), we conducted a phase II clinical trial in which eligible patients received paclitaxel 200 mg/m2 intravenously over 3 h. Treatment was repeated every 3 weeks. Patients achieving complete or partial responses after two courses of paclitaxel continued to receive therapy for a maximum of eight courses, otherwise they were removed from the study. Of 96 evaluable patients, 45 (47%) had primary refractory disease, and 51 (53%) had relapsed lymphoma. The median number of prior treatment regimens was two (range one to 10 regimens). 45 patients had lowgrade, 44 had intermediate-grade, and seven had mantle cell lymphoma. 24/96 patients responded (10 complete and 14 partial remissions) for an overall response rate of 25% (95% CI 17-35%). Patients with relapsed lymphoma had a higher response rate than those with primary refractory disease (19/51 = 37% v 5/45 = 11%; P < 0.01), and patients with relapsed intermediate-grade lymphoma had a higher response than those with relapsed low-grade lymphoma (9/18 = 50% v 10/31 = 32%; P = 0.22). The treatment was very well tolerated with the most common side-effects being alopecia (100%), peripheral neuropathy (35% of > or = grade II), and arthralgia/myalgia (25% of > or = grade II). After the first course of paclitaxel, grade III/IV thrombocytopenia and neutropenia were observed in 21% and 23% of the patients respectively. 23 episodes of neutropenic fever developed after 250 courses of paclitaxel therapy (8%). We conclude that paclitaxel, at this dose and schedule, is an active new drug for the treatment of non-Hodgkin's lymphoma. The activity of paclitaxel combination programmes are currently under investigation.

Paclitaxel (Taxol) for the treatment of lymphoma.

Paclitaxel (Taxol) was recently tested in patients with relapsed and refractory lymphoma in two phase II clinical trials using two different infusion schedules. The first, reported from the NCI (USA), used a 96-hour intravenous continuous infusion schedule, and the second, from our group, used a 3-hour infusion. In the NCI trial, 29 evaluable patients were treated with 140 mg/m2 every three weeks, which achieved a 17% response rate (all PRs); while we treated 96 evaluable patients with 200 mg/m/ every three weeks, which achieved a 25% response rate (10 CRs and 14 PRs, 95% CI: 17%-35%). In our trial, patients with relapsed (not primary refractory) intermediate-grade lymphoma had a response rate of 50%, and those with relapsed low-grade lymphoma had a response rate of 31%. In a follow-up trial, 12 patients who failed to respond to 3-hour infusion of paclitaxel were crossed over to receive paclitaxel by 96-hour infusion. None of the 12 evaluable patients achieved a major clinical response. Similarly, of 25 patients treated with cyclosporine A and paclitaxel after failing therapy with single-agent paclitaxel, only one patient (4%) responded. We conclude that paclitaxel has a promising single-agent activity, most prominently in patients with relapsed intermediate-grade lymphoma. Paclitaxel-based combination programs are currently being evaluated in our institution.

Results of MDR-1 vector modification trial indicate that granulocyte/macrophage colony-forming unit cells do not contribute to posttransplant hematopoietic recovery following intensive systemic therapy.

To formally test the hypothesis that the granulocyte/macrophage colony-forming unit (GM-CFU) cells can contribute to early hematopoietic reconstitution immediately after transplant, the frequency of genetically modified GM-CFU after retroviral vector transduction was measured by a quantitative in situ polymerase chain reaction (PCR), which is specific for the multidrug resistance-1 (MDR-1) vector, and by a quantitative GM-CFU methylcellulose plating assay. The results of this analysis showed no difference between the transduction frequency in the products of two different transduction protocols: "suspension transduction" and "stromal growth factor transduction." However, when an analysis of the frequency of cells positive for the retroviral MDR-1 vector posttransplantation was carried out, 0 of 10 patients transplanted with cells transduced by the suspension method were positive for the vector MDR-1 posttransplant, whereas 5 of 8 patients transplanted with the cells transduced by the stromal growth factor method were positive for the MDR-1 vector transcription unit by in situ or in solution PCR assay (a difference that is significant at the P = 0.0065 level by the Fisher exact test). These data suggest that only very small subsets of the GM-CFU fraction of myeloid cells, if any, contribute to the repopulation of the hematopoietic tissues that occurs following intensive systemic therapy and transplantation of autologous hematopoietic cells.