Long-term survival of patients with resistant lymphoma treated with tandem stem cell transplant.

Dose-intensive chemotherapy with autologous stem cell support is commonly used in resistant/refractory cases of Hodgkin's disease (HD) and non-Hodgkin's lymphoma (NHL). The purpose of this study was to evaluate the role of tandem transplantation in these patients. We used non cross-resistant conditioning regimens with thiotepa, mitoxantrone and carboplatin (TMJ) followed by ifosphamide, carboplatin and etoposide (ICE) with autologous stem cell rescue in an attempt to maximize dose intensity and achieve long-term remission. Seventy-six patients were included in this study. Twenty-nine patients with HD and 47 with NHL underwent autologous stem cell transplant using TMJ as the conditioning regimen for the first transplant. Of these, 49 patients proceeded to the second transplant using ICE as the conditioning regimen. In 57 patients, only peripheral blood cells were used and in 11 patients both bone marrow and peripheral stem cells were used. Twelve patients died due to treatment-related toxicity. On an intent to treat basis, 32.14% of patients with HD refractory to initial or subsequent therapy survived long term as opposed to 12.76% of patients with NHL. With a median follow-up of 83 months (range 25 - 110 months), the median disease-free survival of patients with HD was 7 months, as opposed to 2 months for patients with NHL. Multivariate analysis identified that patients with HD had a superior outcome if they were less than 35 years of age and did not have B symptoms. Dose-intensive chemotherapy with tandem transplantation is an option in patients with resistant/refractory lymphoma who have very limited treatment options and poor prognosis.

High dose chemotherapy with thiotepa, mitoxantrone and carboplatin (TMJ) followed by autologous stem cell support in 100 consecutive lymphoma patients in a single centre: analysis of efficacy, toxicity and prognostic factors.

High dose chemotherapy with autologous stem cell transplant is often used in patients with Hodgkin's disease (HD) and non-Hodgkin's lymphoma (NHL) who either do not respond to, or relapse after conventional chemotherapy. There is no consensus on the "ideal" pretransplant conditioning regimen. In this study, we analyzed the results of 100 consecutive patients with HD and NHL who met our eligibility criteria and underwent autologous stem cell transplant at New York Medical College and Zalmen A. Arlin Cancer Institute. All patients received high dose chemotherapy with thiotepa, mitoxantrone and carboplatin (TMJ). One hundred patients, 37 with HD and 63 with NHL underwent autologous stem cell transplant using TMJ as a conditioning regimen. All patients with HD had chemo-sensitive relapse while 50 patients with NHL had chemo-sensitive relapse and 13 patients had first complete remission. The source of stem cells was bone marrow (18 patients), peripheral blood (50 patients) and both bone marrow and peripheral blood (32 patients). With a median follow up of 91 months (range 23-147 months), the median survival of patients with HD and NHL who underwent autologous stem cell transplant is 107 months and the 5 years disease free survival is 43%. Median survival of patients with HD and NHL is 87 and 107 months respectively. There were 4 transplant related deaths. Median survival of patients who had sensitive relapse at the time of transplant is 87 months while median survival has not been reached for patients who had first complete remission at the time of transplant. Multivariate analysis identified age>35 years (P=0.02) as a predictor for poor survival for the whole group as well as for patients with NHL (P=0.04). TMJ is a safe and effective regimen when used as a part of autologous stem cell transplant for patients with HD and NHL.

Sulforaphane and its metabolite mediate growth arrest and apoptosis in human prostate cancer cells.

The relation between the consumption of cruciferous vegetables and reduced prostate cancer occurrence has been documented, although the responsible phytochemicals are unknown. The effects of sulforaphane (SFN) which occurs as the precursor glucosinolate in broccoli and other cruciferous vegetables, and its metabolite N-acetylcysteine conjugate (SFN-NAC) on prostate cancer cells were investigated. SFN and SFN-NAC were analyzed with the androgen-dependent human prostate cancer LNCaP cell line model. Cell growth and apoptosis were determined with the expression of androgen receptor and prostate specific antigen, DNA synthesis, cell cycle progression, DNA strand breaks and caspase activation to ascertain the effects and mechanism. SFN and SFN-NAC were demonstrated for the first time to mediate a dose-dependent apoptosis and growth arrest in the prostate cancer cells. Caspases were activated and DNA strand breaks were detected in apoptotic cells. The expression of phosphorylated and dephosphorylated androgen receptors, and the production of prostate specific antigen were attenuated. The expression of cyclin D1 and DNA synthesis were inhibited along with G1 cell cycle block, causing decreased cell density and growth. SFN and its metabolite SFN-NAC have similar activities to induce growth arrest and apoptosis, indicating that the effects of SFN are maintained through the metabolic processes. SFN as a dietary component of cruciferous vegetables active in the prevention of prostate cancer is discussed.

Intravenous bolus topotecan in patients with myelodysplastic syndrome.

We treated 16 patients with myelodysplastic syndromes with 24 courses of bolus topotecan. Patients received topotecan as a daily 15 minute infusion for 5 days at 3 dose levels (4.0 mg/m2/d, 2.0 mg/m2/d or 2.5 mg/m2/d). There was one complete response and one partial response (overall response rate 12%). Toxicity included myelosuppression, diarrhea, ileus and mucositis. There were 3 treatment-related deaths. The results of this schedule of topotecan appeared to be inferior to that reported with infusional topotecan in patients with MDS.

Secondary acute myelogenous leukemia and myelodysplasia without abnormalities of chromosome 11q23 following treatment of acute leukemia with topoisomerase II-based chemotherapy.

Therapy-related MDS and AML are complications of intensive chemotherapy regimens. Traditionally, patients exposed to topoisomerase II inhibitors are reported to develop secondary AML with abnormalities of chromosome 11q23. We evaluated the long-term hematologic toxicity of topoisomerase II-intensive high-dose mitoxantrone-based chemotherapy in 163 newly diagnosed acute leukemia patients treated over an 8 year period. Nine (5.5%) patients developed new cytogenetic abnormalities. Four patients developed MDS with progression to AML, three patients developed new abnormalities at the time of relapse, and three patients (including one of the former patients) had changes that were not associated with hematologic disease. The abnormalities most frequently involved chromosomes 7q, 20q, 1q, and 13q. Despite the use of topoisomerase II-intensive treatment, no patient developed an abnormality involving chromosome 11q23. Spontaneous resolution of some changes and prolonged persistence of others in the absence of hematologic disease indicates that some cytogenetic changes are not sufficient to promote leukemogenesis.

Evaluation of topotecan and etoposide for non-Hodgkin lymphoma: correlation of topoisomerase-DNA complex formation with clinical response.

BACKGROUND: Topotecan, a topoisomerase I inhibitor, acts by stabilizing the topoisomerase DNA cleavage complex. Etoposide, a topoisomerase II inhibitor, mediates antitumor activity by stabilizing cleavage complex formed between topoisomerase II and DNA. These two agents have therapeutic activity in non-Hodgkin lymphoma. The authors report Phase I data of topotecan and etoposide combination for patients with recurrent or refractory non-Hodgkin lymphoma and correlation of topoisomerase-DNA complex formation to clinical response. METHODS: Twenty-two patients with recurrent or refractory aggressive non-Hodgkin lymphoma were treated at four dose levels of topotecan (1 mg/m(2)/day to 2.5 mg/m(2)/day). Topotecan was given at a 30-minute infusion daily with etoposide 150 mg/m(2)/day, both for 5 days. Topoisomerase-DNA covalent complex formation was measured using in vivo link assay, whereas topoisomerase I, IIalpha, and IIbeta in RNA expression levels were determined by reverse transcription-polymerase chain reaction in blood samples. The relation of these levels to clinical response was studied. RESULTS: The maximum tolerated dose of topotecan was 2.0 mg/m(2)/day for 5 days. Oropharyngeal mucositis was dose-limiting. Of 21 examinable patients, 3 patients achieved complete remission, and 5 patients achieved partial remission. Of six untreated patients who experienced a recurrence, three had complete remission, and the other three had partial remission. Drug-induced topoisomerase-DNA complex formation was observed throughout the treatment in blood samples of all the patients who responded. However, only 4 of 13 patients, who did not respond, formed covalent complex at all time points. This was statistically significant (P = 0.024). In all patients, expression levels of topoisomerase I and IIbeta mRNA remained similar to pretreatment levels, whereas topoisomerase IIalpha mRNA levels decreased dramatically by the third day. CONCLUSION: The recommended Phase II dose of topotecan with etoposide of 150 mg/m(2)/day for 5 days was 2.0 mg/m(2)/day for 5 days. Topoisomerase-DNA complex formation correlated with response to treatment.

Evaluation of solvent/detergent-treated plasma in patients with a prolonged prothrombin time.

OBJECTIVES: To compare the laboratory and clinical outcome of patients who received solvent/detergent-treated plasma (SDP) and fresh-frozen plasma (FFP). METHODS: A randomized, double-blinded study to assess the ability of SDP and FFP to reduce a prolonged prothrombin time (PT) to

Endothelin-1 from prostate cancer cells is enhanced by bone contact which blocks osteoclastic bone resorption.

The causes for the propensity of metastasized prostate cancer cells to grow in bone and to induce osteoblastic lesions remain unresolved. Co-culture of human prostate cancer cell lines with bone slices was determined to increase the level of endothelin-1 (ET-1) mRNA and its production. ET-1 is an ejaculate protein that also stimulates osteoblasts. Osteoclastic bone resorption was significantly blocked by the presence of androgen-independent prostate cancer cells in a dose-dependent manner as that of synthetic ET-1. The inhibition could be neutralized by specific ET-1 antibody, indicating the association of prostate cancer-derived ET-1 with inhibition of bone resorption. The combined ET-1 activity on osteoclasts and osteoblasts disrupts bone remodelling. ET-1 production is also elevated in the presence of prostate-specific antigen (PSA). ET-1 in turn enhances DNA synthesis of prostate cancer cells. Interactions among cancer cells, bone, ET-1 and PSA may be critical in cancer growth and lesions in bone.

High-dose chemotherapy and stem cell transplantation for patients with stage IV breast cancer without clinically evident disease: correlation of CD34+ selection to clinical outcome.

Forty-five patients with metastatic breast cancer without clinically evident disease were treated with thiotepa 750 mg/m2, mitoxantrone 40 mg/m2 and carboplatin 1000 mg/m2 followed by stem cell transplantation to determine the safety and efficacy of CD34+ selection of peripheral blood stem cells. Of these, 15 patients' (group I) stem cells were processed through Baxter Isolex 300 device for CD34+ selection, whereas 30 patients (group II) received unmanipulated stem cells. Toxicity, progression-free survival and survival were compared between these two groups. There was no difference in transfusion requirements, white cell count and platelet recovery and non-hematologic toxicity between the two groups. The survival of patients in group I was 27 months compared to 38 months in group II (P = 0.8). The progression-free survival was 12 months and 13.5 months for group I and group II patients, respectively (P = 0.6). Our results indicate that while there is no adverse effect, there is also no significant advantage of CD34+ selection in terms of progression-free survival and survival in patients with metastatic breast cancer without clinically evident disease. Bone Marrow Transplantation (2000).

Clinical and laboratory evaluation of all-trans retinoic acid modulation of chemotherapy in patients with acute myelogenous leukaemia.

All-trans retinoic acid (ATRA) is synergistic with chemotherapy in leukaemia cell lines. We treated 53 patients with newly diagnosed acute myelogenous leukaemia (AML) with high-dose cytarabine-based chemotherapy followed by ATRA. Peripheral blood and bone marrow samples were obtained to study the effect of in vitro exposure to ATRA and to measure apoptosis and bcl-2. The response rate was 72% for patients under age 60 years and 46% for patients aged 60 years or above. There was no difference in the percentage of responding patients, time to recurrence or overall survival for patients receiving chemotherapy with ATRA vs. historical controls receiving chemotherapy without ATRA. After in vitro exposure of day 3 bone marrow samples to ATRA, there was an increase in apoptotic cells in 25% of patient samples compared with samples not exposed to ATRA. Later date of peak apoptosis in peripheral blood and higher percentage of apoptotic cells in bone marrow on day 3 of treatment were associated with lack of clinical response to treatment. Increased bcl-2 in patient samples was associated with shorter time to recurrence and poor cytogenetic risk. The addition of ATRA to chemotherapy did not improve patient outcome. However, evidence of in vitro response to ATRA in 25% of patients suggests that retinoid pathways should be studied further in patients with AML.

Adenovirus and retrovirus mediated interferon alpha gene transfer into CD34+ cells maintains regeneration capacity and enhances adhesion molecules in K562 cells.

BACKGROUND: Systemic administration of interferon-alpha (IFN-alpha) results in cytogenetic remissions and enhanced survival in a significant percentage of patients with chronic mylogenous leukemia (CML) and lymphoma. However, this treatment is associated with deleterious toxic effects. Gene transfer of the IFN-alpha gene into hematopoietic progenitors represents a novel strategy to deliver high concentrations of IFN-alpha to a local area. METHODS: We compared the effect of the transfer of the IFN-alpha gene on the cell growth and differentiation of several CD34+ cells in culture and in a NOD/SCID animal model, using adenovirus and retrovirus constructs. RESULTS: Transient local expression of the IFN-alpha gene using an adenovirus vector was associated with normal proliferation of CD34+ progenitors as measured by a colony forming unit of granulocyte-macrophage (CFU-GM) growth. Flow cytometric determination revealed that there was no significant difference in viability of these cells for 24-hour transduction periods. Reverse transcriptase/polymerase chain reaction (RT-PCR) analysis of RNA from CD34+ harvested CFU-GM progenitors demonstrated expression of IFN-alpha mRNA; radioimmunoassay (RIA) revealed that transduced cells secreted substantial levels of IFN-alpha protein. Furthermore, we constructed a retroviral vector in which IFN-alpha cDNA was driven by a viral LTR promoter to evaluate the effect of permanent IFN-alpha gene expression on cell growth. Retroviral packaging cells PA317 with high titers of retrovirus were produced and used to infect CD34+ and K562 cells. RIA showed that IFN-alpha-transduced CD34+ cells (with the aid of fibronectin fragment CH-296) produced approximately 400 units of IFN-alpha protein compared to CD34+ cells, or cells transduced with empty vector. IFN-alpha transduced CD34+ generated similar numbers of CFU-GM colonies as compared to control CD34+ cells. Engraftment of CD34+ cells transduced with IFN-alpha gene in NOD/SCID mice was successful for the first 30 days. Additionally, we studied the effect of local IFN-alpha expression on the cellular adhesion molecules, VLA-4, Mac-1, ICAM-1, and L-selectin in K562 cells, and human umbilical endothelial vein cells. K562 cells transduced with the IFN-alpha gene expressed a significantly elevated level of VLA-4, Mac-1, and ICAM-1. CONCLUSIONS: We conclude that expression of the IFN-alpha gene using retrovirus vectors results in an adequate localized expression of IFN-alpha mRNA and protein.

Development of human prostate cancer cells to neuroendocrine-like cells by interleukin-1.

Growth and development of some prostate epithelial cells are androgen-dependent. Non-androgenic hormones and growth factors may also influence prostate cells and the effect of interleukin 1beta (IL-1beta) has been investigated with an androgen-dependent human prostate cancer cell line LNCaP. Exposure of LNCaP cells to IL-1beta at picomole ranges resulted in a dose-dependent and progressive differentiation to neuroendocrine-like cells evidenced by dendrite formation and the development of specific neuroendocrine cell markers. Quantification by computer-based image analysis after immunostaining revealed a two-fold increase of chromogranin A in 90% of the cells and a ten-fold increase in the remaining 10%. Additionally, serotonin was developed in all the cells with the staining intensity increased by five-fold. Significant increase in cytokeratin 8 and reduction of prostate specific antigen was also noted. Proliferation was reduced in parallel to the cellular development. The IL-1beta effect was irreversible after several days of IL-1beta incubation. IL-1beta is produced constitutively and its secreted level has an inversed relation during the exponential and plateau phases of cell growth. An IL-1 autocrine regulation in the growth and differentiation of prostate cells is discussed.

Regulation of growth and apoptosis of breast cancer cells by a 54 kDa lymphokine.

A normal human lymphocyte-derived 54 kDa polypeptide, capable of regulating cell growth has been identified as an isoform variant (abbreviated as NP54) of protein neuroleukin-phosphoglucose isomerase (PGI). Since distinct PGI variants undetectable in normal tissues had been identified in breast cancer tissues, the effect of NP54 on the growth of human breast cancer cells SK-Br-3 in cultures was analyzed. Exposure to NP54 caused a dose-dependent growth modulation. Approximately 40% reduction of cell density was detected at 40 pM of NP54, along with a blocking of G1 cells entering into S phase. The growth modulation was correlated with a significantly reduced expression of epidermal growth factor receptor (EGF-R) gene transcript, supporting the interpretation that the level of EGF-R expression and cell growth are related mechanisms. NP54 treatment also significantly increased cells with apoptotic morphological feature and fragmented DNA. Incubation with a monoclonal anti-NP54 antibody negated NP54 activity, confirming a regulatory activity in cell growth and apoptosis.

Enhancement of 5-fluorouracil cytotoxicity on human colon cancer cells by retrovirus-mediated interferon-alpha gene transfer.

Gene therapy has advantages in the treatment of a variety of disorders due to its selective expression within specific mammalian cells. Several reports documented the clinical effects of interferon-alpha (IFN-alpha) in management of patients with advanced colorectal carcinoma. We report for the first time, the successful transduction of human IFN-alpha gene into colon cancer cells, COLO 201 using a replication-defective retroviral vector. Retrovirus-containing supernatant from PA 317 packaging cells was used to infect colon cancer cells, COLO 201 and NIH 3T3 cells. Transient infection showed that cell proliferation and cell viability were significantly suppressed in colon cancer cells transduced with IFN-alpha gene. Moreover, IFN-alpha-transduced cells acquired less resistance to 5-FU induced apoptosis. These data demonstrate that IFN-alpha gene transfer may have a clinical application and can be combined with chemotherapy for treatment of advanced colorectal cancer.

Single and double autotransplants for relapsing/refractory Hodgkin's disease: results of two consecutive trials.

To evaluate a strategy of one cycle of dose-intensive chemotherapy for patients with Hodgkin's disease in sensitive relapse and two cycles for those with refractory disease, 122 patients received dose-intensive chemotherapy followed by autotransplant in two consecutive studies. Patients with refractory disease were offered a second transplant with different conditioning in the absence of progression or excessive toxicity. CR was present after treatment in 46% while 16% died in the peritransplant period. Of 41 patients with primary refractory disease and 42 with refractory relapse, 24 and 21 respectively received a second cycle. Of these 45 refractory patients, 12 were in CR and 11 in PR after the first cycle and 10 of these 11 in PR achieved a durable CR with the second transplant. The CR rate is 37% in patients with refractory relapse and 19% in those with primary refractory disease. At a median follow-up of 4 years, median survival is 45 months. Progression-free survival of the refractory patients who could receive a second cycle was similar to that of patients with sensitive disease. A sequential transplant strategy is feasible. A subgroup of patients with refractory disease can achieve long-term survival after sequential BMT.