Prolonged clinical remissions in patients with relapsed or refractory follicular lymphoma treated with autologous stem cell transplantation incorporating rituximab.

Three sequential phase II trials were conducted with different immunotherapy approaches to enhance the outcome of autologous transplant (high-dose therapy and autologous stem cell transplantation (HDT/ASCT)) for recurrent follicular lymphoma. Seventy-three patients were enrolled from 1996 to 2009. Patients received HDT/ASCT combined with (1) interferon-α 3 MU/m(2) subcutaneously (SC) three times per week (TIW) for 2 years post-ASCT, (2) rituximab (R) 375 mg/m(2) for in vivo purging 3-5 days pre-stem cell collection and 2 × 4 weekly R at 2 and 6 months post-ASCT, respectively, or (3) three infusions of R pre-stem cell collection followed by 6× R weekly and interferon-α 3 MU/m(2) SC TIW. Although not statistically significant, progression-free survival (PFS) for patients who received rituximab was 56.4 and 49.1% at 5 and 10 years compared to 36 and 21% in those who did not receive rituximab. Molecular relapse post-HDT/ASCT was the strongest predictor of PFS in a multivariate analysis. Molecular relapse was coincident with or preceded clinical relapses in 84% of patients who relapsed­median of 12 months (range 0-129 months). Adverse events included secondary malignancy, transformation to diffuse large B cell lymphoma, prolonged mostly asymptomatic hypogammaglobulinemia, and pulmonary fibrosis. The long-term toxicity profile must be considered when selecting patients for this treatment.

Earliness per se QTLs and their interaction with the photoperiod insensitive allele Ppd-D1a in the Cutler × AC Barrie spring wheat population.

Earliness per se regulates flowering time independent of environmental signals and helps to fine tune the time of flowering and maturity. In this study, we aimed to map earliness per se quantitative trait loci (QTLs) affecting days to flowering and maturity in a population developed by crossing two spring wheat cultivars, Cutler and AC Barrie. The population of 177 recombinant inbred lines (RILs) was genotyped for a total of 488 SSR and DArT polymorphic markers on all 21 chromosomes. Three QTLs of earliness per se affecting days to flowering and maturity were mapped on chromosomes 1B (QEps.dms-1B1 and QEps.dms-1B2) and 5B (QEps.dms-5B1), in individual environments and when all the environments were combined. A QTL affecting flowering time (QFlt.dms-4A1) was identified on chromosome 4A. Two grain yield QTLs were mapped on chromosome 5B, while one QTL was mapped on chromosome 1D. The population segregated for the photoperiod insensitive gene, Ppd-D1a, and it induced earlier flowering by 0.69 days and maturity by 1.28 days. The photoperiod insensitive allele Ppd-D1a interacted in an additive fashion with QTLs for flowering and maturity times. The earliness per se QTL QFlt.dms-5B.1 inducing earlier flowering could help to elongate grain filling duration for higher grain yield. Hence, chromosome 5B possesses promising genomic regions that may be introgressed for higher grain yield with earlier maturity through marker-assisted selection in bread wheat.

Differential immune responses mediated by adenovirus- and lentivirus-transduced DCs in a HER-2/neu overexpressing tumor model.

Recent investigations have demonstrated that adenoviral and lentiviral vectors encoding HER-2 can be utilized in cancer immunotherapy. However, it is not known whether both viral systems elicit a similar immune response. Here, we compare the immune response in mice induced by dendritic cells (DCs) infected with either recombinant adenovirus or lentivirus encoding rat HER-2 (rHER-2). Both vaccine types yielded similar control of tumor growth, but we found clear differences in their immune responses 10 days after DC immunization. Adenovirus rHER-2-transduced DCs elicited locally and systemically high frequencies of CD4+ and CD8+ T cells, while lentivirus rHER-2-transduced DCs predominantly led to CD4+ T-cell infiltration at the tumor site. Splenocytes from mice immunized with lentivirus rHER-2-transduced DCs secreted higher levels of interferon (IFN)-γ, mainly by CD4+ T cells, following stimulation by RM-1-mHER-2 tumors. In contrast, the adenovirus vaccinated group exhibited CD4+ and CD8+ T cells that both contributed to IFN-γ production. Besides an established cellular immune response, the rHER-2/DC vaccine elicited a significant humoral response that was highest in the adenovirus group. DC subsets and regulatory T cells in the spleen were also differentially modulated in the two vaccine systems. Finally, adoptive transfer of splenocytes from both groups of immunized mice strongly inhibited in vivo tumor growth. Our results suggest that not only the target antigen but also the virus system may determine the nature and magnitude of antitumor immunity by DC vaccination.

Prolonging microtubule dysruption enhances the immunogenicity of chronic lymphocytic leukaemia cells.

Cytotoxic chemotherapies do not usually mediate the expression of an immunogenic gene programme in tumours, despite activating many of the signalling pathways employed by highly immunogenic cells. Concomitant use of agents that modulate and complement stress-signalling pathways activated by chemotherapeutic agents may then enhance the immunogenicity of cancer cells, increase their susceptibility to T cell-mediated controls and lead to higher clinical remission rates. Consistent with this hypothesis, the microtubule inhibitor, vincristine, caused chronic lymphocytic leukaemia (CLL) cells to die rapidly, without increasing their immunogenicity. Protein kinase C (PKC) agonists (such as bryostatin) delayed the death of vincristine-treated CLL cells and made them highly immunogenic, with increased stimulatory abilities in mixed lymphocyte responses, production of proinflammatory cytokines, expression of co-stimulatory molecules and activation of c-Jun N-terminal kinase (JNK), p38 and nuclear factor kappa B (NF-kappaB) signalling pathways. This phenotype was similar to the result of activating CLL cells through Toll-like receptors (TLRs), which communicate 'danger' signals from infectious pathogens. Use of PKC agonists and microtubule inhibitors to mimic TLR-signalling, and increase the immunogenicity of CLL cells, has implications for the design of chemo-immunotherapeutic strategies.

PPARγ inhibits breast cancer progression by upregulating PTPRF expression.

OBJECTIVE: Peroxisome proliferator-activated receptor γ (PPARγ) regulates fatty acid storage and glucose metabolism. Recently, PPARγ has been reported to be involved in cancer. The present study reported a PPARγ consensus binding site (AGGTCA) in the ptprf promoter and identified a strong association between PPARγ and PTPRF expression, as well as their tumor suppressor roles in a v-Ha-Ras-induced model of breast cancer. MATERIALS AND METHODS: The prognostic potential of PPARγ was assessed with a KM analysis of raw data from 3,951 breast cancer patients. The expression of PPARγ and PTPRF in the rat breast cancer cell lines was detected by Western blot and qPCR. The impact of PPARγ on cancer cell migration, invasion, and growth was confirmed using cell migration assay, transwell cell invasion assay, tri-dimensional soft agar culture, respectively. The binding of PPARγ with the ptprf promoter was then examined using electrophoretic mobility shift assay. The inhibitory effect of PPARγ on tumor growth was then examined in mouse tumor model in vivo. RESULTS: It was identified that PPARγ expression is lost in the aggressive v-Ha-Ras-induced breast cancer cell line FE1.2 but highly expressed in less malignant FE1.3 cells. Exogenous expression of PPARγ in FE1.2 cells (FE1.2-PPARγhi) resulted in a marked inhibition of proliferation compared with that in FE1.2-Vector control group. FE1.2-PPARγhi cells also exhibited reduced migration, invasion, and colony formation abilities compared with those of the controls. The PPARγ agonist rosiglitazone also suppressed the malignant properties of FE1.2 cells. Protein tyrosine phosphatase receptor F (PTPRF), a downstream target of PPARγ, was markedly induced in FE1.2-PPARγhi cells. A PPARγ consensus binding site (AGGTCA) was identified in the ptprf promoter, and an electrophoretic mobility shift assay confirmed that PPARγ bind to this promoter. Similar to the effect of vector-mediated overexpression of PPARγ, ectopic overexpression of PTPRF in FE1.2 cells led to reduced proliferation. Furthermore, a PPARγ antagonist (GW9662) and PTP inhibitor (NSC87877) abrogated the suppressive function of PPARγ and PTPRF in FE1.2 cells, respectively. PPARγ overexpression or activation suppressed the progression and distant organ metastasis of breast cancer cells in a NOD/SCID mouse model. CONCLUSIONS: These results suggest that PPARγ inhibits tumor cell proliferation, at least in part, through direct regulation of the ptprf gene and that PPARγ is a potential target for breast cancer treatment.

Toll-like receptor agonists in the treatment of chronic lymphocytic leukemia.

Advances in our understanding of the Toll-like receptors (TLRs) have led to the identification of several agonists that are suitable for clinical development. Chronic lymphocytic leukemia (CLL) may be especially amenable to TLR agonists because it is an immunologically susceptible tumor with strong expression of several TLRs, particularly TLR-7 and TLR-9. TLR agonists may indirectly clear CLL cells by enhancing the activity of natural killer and tumor-reactive T cells, or by altering the tumor microenvironment and inhibiting angiogenesis. However, signaling pathways can be activated directly in CLL cells by TLR-7 and TLR-9 agonists, leading to the production of cytokines and costimulatory molecules in a manner that is dependent on the underlying cytogenetic abnormalities, but rendering the tumor cells more sensitive to killing by cytotoxic T cells, immunotoxins and some chemotherapeutic drugs. Imidazoquinolines are TLR-7 agonists with strong local activity against CLL, and phase I trials of systemically administered imidazoquinolines (and also cytosine-phosphate-guanosine oligonucleotides that are TLR-9 agonists) are currently ongoing at different centers. The potential importance of these TLR agonists in the treatment of CLL is suggested by their ability to sensitize tumor cells to cytotoxic agents, and their future probably lies in combination with radiotherapies, chemotherapies, monoclonal antibodies and cancer vaccines.

PPAR-delta modulates membrane cholesterol and cytokine signaling in malignant B cells.

A deeper understanding of the mechanisms that underlie aberrant signal transduction in B-cell cancers such as chronic lymphocytic leukemia (CLL) may reveal new treatment strategies. The lipid-activated nuclear receptor peroxisome proliferator-activated receptor delta (PPARδ) accounts for a number of properties of aggressive cancers and was found to enhance Janus kinase (JAK)-mediated phosphorylation of signal transducer and activator of transcription (STAT) proteins in B lymphoma cell lines and primary CLL cells. Autocrine production of cytokines such as IL10 and interferon-beta was not increased by PPARδ but signaling responses to these cytokines were amplified and associated with increased cholesterol biosynthesis and plasma membrane levels. Plasmalemmal cholesterol and STAT phosphorylation from type 1 interferons (IFNs) were increased by PPARδ agonists, transgenes and exogenous cholesterol, and decreased by cyclodextrin, PPARD deletion and chemical PPARδ inhibitors. Functional consequences of PPARδ-mediated perturbation of IFN signaling included impaired upregulation of co-stimulatory molecules. These observations suggest PPARδ modulates signaling processes in malignant B cells in part by altering cholesterol metabolism and changes the outcomes of signaling from cytokines such as IFNs. PPARδ antagonists may have therapeutic activity as anti-leukemic signal transduction modulators.

Immunomodulatory effects of Toll-like receptor-7 activation on chronic lymphocytic leukemia cells.

Weak immunogenicity of chronic lymphocytic leukemia (CLL) cells may contribute to disease progression and inhibit effective immunotherapy. Accordingly, agents that enhance the immunogenicity of CLL cells may be useful in immunotherapeutic approaches to this disease. Since Toll-like receptors (TLRs) are major regulators of innate immunity and initiation of adaptive immunity, we studied the effects of viral pathogen associated molecular pattern agonists (that are recognized by TLRs) on the costimulatory phenotype and function of CLL cells. CLL cells (especially those with high endogenous expression of CD38) responded to TLR7-activating imidazoquinolines and guanosine analogs by increasing costimulatory molecule expression, producing inflammatory cytokines, and becoming more sensitive to killing by cytotoxic effectors. Additional activation of protein kinase C pathways increased the ability to stimulate T-cell proliferation, blocked phosphorylation of the transcription factor, signal transducer and activator of transcription (STAT)3, and resulted in the acquisition of a dendritic cell surface phenotype by TLR7-activated CLL cells. Normal B cells also responded to TLR7 activation by increasing costimulatory molecule expression and cytokine production. These findings suggest a potential role for TLR7 agonists in CLL immunotherapy.

PPAR-delta promotes survival of chronic lymphocytic leukemia cells in energetically unfavorable conditions.

Targeting the mechanisms that allow chronic lymphocytic leukemia (CLL) cells to survive in harsh cancer microenvironments should improve patient outcomes. The nuclear receptor peroxisome proliferator activated receptor delta (PPARδ) sustains other cancers, and in silico analysis showed higher PPARD expression in CLL cells than normal lymphocytes and other hematologic cancers. A direct association was found between PPARδ protein levels in CLL cells and clinical score. Transgenic expression of PPARδ increased the growth and survival of CD5+ Daudi cells and primary CLL cells in stressful conditions including exhausted tissue culture media, low extracellular glucose, hypoxia and exposure to cytotoxic drugs. Glucocorticoids and synthetic PPARδ agonists up-regulated PPARD expression and also protected Daudi and primary CLL cells from metabolic stressors. Survival in low glucose was related to increased antioxidant expression, substrate utilization and mitochondrial performance, and was reversed by genetic deletion and synthetic PPARδ antagonists. These findings suggest PPARδ conditions CLL cells to survive in harsh microenvironmental conditions by reducing oxidative stress and increasing metabolic efficiency. Targeting PPARδ may be beneficial in the treatment of CLL.

PPAR-delta promotes survival of breast cancer cells in harsh metabolic conditions.

Expression of the nuclear receptor peroxisome proliferator activated receptor delta (PPARδ) in breast cancer cells is negatively associated with patient survival, but the underlying mechanisms are not clear. High PPARδ protein levels in rat breast adenocarcinomas were found to be associated with increased growth in soft agar and mice. Transgenic expression of PPARδ increased the ability of human breast cancer cell lines to migrate in vitro and form lung metastases in mice. PPARδ also conferred the ability to grow in exhausted tissue culture media and survive in low-glucose and other endoplasmic reticulum stress conditions such as hypoxia. Upregulation of PPARδ by glucocorticoids or synthetic agonists also protected human breast cancer cells from low glucose. Survival in low glucose was related to increased antioxidant defenses mediated in part by catalase and also to late AKT phosphorylation, which is associated with the prolonged glucose-deprivation response. Synthetic antagonists reversed the survival benefits conferred by PPARδ in vitro. These findings suggest that PPARδ conditions breast cancer cells to survive in harsh microenvironmental conditions by reducing oxidative stress and enhancing survival signaling responses. Drugs that target PPARδ may have a role in the treatment of breast cancer.

Phosphorylation of p53 protein in response to ionizing radiation occurs at multiple sites in both normal and DNA-PK deficient cells.

The tumour suppressor gene product, p53, is involved in mediating cellular responses to DNA damage including growth arrest and/or apoptosis. The mechanism by which p53 protein senses the presence of damaged DNA is not understood. The possibility that p53 may be post-translationally modified by enzymes that are activated in response to DNA damage including DNA-dependent protein kinase (DNA-PK), poly(ADP-ribose) polymerase and stress activated protein kinase has received considerable attention. Recent studies have indicated that DNA-PK is not required for the transactivation or apoptosis-promoting activities of p53 protein. However, the possibility that other functions of p53 may be dependent on phosphorylation by DNA-PK has not been explored. Here we describe a series of experiments that compares the expression, function and phosphorylation status of p53 protein in normal and DNA-PK-deficient scid cells. While several novel p53 phosphoforms are generated in response to DNA damage in normal cells, the same phosphoforms are observed in scid cells.

Immunogenicity of whole-cell tumor preparations infected with the ALVAC viral vector.

The immunogenicity of recombinant canarypox (ALVAC) viral vectors within murine whole-cell tumor vaccines was evaluated using the T cell thymic lymphoma STF10 and the B16 melanoma. Tumor cells were modified with the recombinant ALVAC vectors and injected into syngeneic mice. Control mice receiving cells alone all developed tumors, while mice injected with tumor variants bearing parental and recombinant vectors either completely rejected their tumors, or exhibited a significant delay in tumor formation. Rechallenge of mice receiving STF10-variant vaccines yielded a protective effect against parental tumor cells only when a modified regimen incorporating two vaccinations was utilized. Notably, the parental ALVAC virus was equivalent to all other recombinant ALVAC viruses in conferring antitumor immunity when using a prime-and-boost protocol. Tumorigenicity experiments in nude mice revealed that the effector mechanism mediating rejection of tumor cells bearing ALVAC vectors is multifactorial, in that the immunogenicity of STF10/ALVAC vaccines is reduced, but not completely abolished in these mice. Finally, in vitro experiments revealed that cytotoxic T cells specific for parental STF10 cells could be generated as a result of in vivo immunization with STF10/ALVAC vaccines.

Stem cell function and engraftment is not affected by "in vivo purging" with rituximab for autologous stem cell treatment for patients with low-grade non-Hodgkin's lymphoma.

The chimeric anti-CD20 monoclonal antibody rituximab (Rituxan; IDEC Pharmaceuticals, San Diego, CA, and Genentech, Inc, San Francisco, CA) has recently been approved by the US Food and Drug Administration as single-agent treatment of relapsed/refractory low-grade or follicular non-Hodgkin's lymphoma. Initial results from the pivotal clinical trial revealed that response rates to rituximab were higher in patients who previously had high-dose therapy and autologous stem cell transplantation. We have initiated a clinical trial that combines the use of rituximab with high-dose chemotherapy followed by autologous stem cell transplantation for patients with chemosensitive relapsed follicular small cleaved or mantle cell lymphoma. A unique feature of this study is that in addition to eight maintenance infusions of rituximab after autologous stem cell transplantation, patients also received rituximab 375 mg/m2 2 days before a granulocyte colony-stimulating factor-mobilized stem cell collection as "in vivo purge." We report on preliminary results demonstrating the safety and efficacy of the in vivo purge on 10 patients undergoing stem cell mobilization, nine of whom have already undergone transplantation. The peripheral blood CD34+ counts were 14.92 and 20 x 10(6)/L on day 4 and day 5, respectively, of the stem cell mobilization with granulocyte colony-stimulating factor. This compares with 11.7 and 11.8 x 10(6)/L, respectively, for the control population. The median CD34 stem cell yield in the graft collection was 3.7 x 10(6)/kg in patients receiving rituximab in vivo purge compared with 3.1 x 10(6)/kg in the control population. The target stem cell collection was successfully collected in six of 10 patients in a 1-day single large-volume leukapheresis collection, while two patients required 2 days and the last two patients required 3 days. Functional assays revealed the stem cell colony-forming unit-granulocyte monocyte and burst-forming unit-erythrocyte to be 55 and 44 colonies per plate, respectively, for the patients receiving the in vivo rituximab purge. This compares favorably with 37 and 38.5 colonies per plate, respectively, for the control population. Neutrophil engraftment took a median of 11 days for both cohorts; platelet independence was achieved in 8 days compared with 10 days for the control population. The median number of platelet transfusions was two for patients receiving rituximab and 2.5 for the control group. Assessment of serum cytokines immediately before the rituximab infusion during the stem cell mobilization and immediately after revealed a twofold to sevenfold increase in interleukin-1beta, tumor necrosis factor-alpha, and interleukin-6. The polymerase chain reaction analysis for minimal residual disease in stem cell collections and in peripheral blood and bone marrow samples of these patients will help to determine the efficacy of rituximab in vivo purge on disease progression.

Characterization of graft-versus-host disease in SCID mice and prevention by physicochemical stressors.

BACKGROUND: Graft versus host disease (GVHD) prevents potentially curative allogeneic stem cell transplantation from being offered to cancer patients who lack a suitably matched donor. New methods to prevent GVHD are required to allow successful transplants across major histocompatibility complex barriers. METHODS: A model of GVHD in C.B-17 SCID mice was developed to allow the study of allo-activated donor T cells without confounding effects of host lymphocytes. The abilities of cyclosporin-A, anticytokine antibodies, and oxidative stress to prevent GVHD in this model was studied. RESULTS: T cells from major histocompatibility-mismatched donor mice caused severe GVHD in sublethally irradiated SCID hosts that could be ameliorated by coadministration of donor bone marrow but not by cyclosporine-A or anticytokine antibodies. In contrast, three-log more T cells could be injected without clinical consequences if they had been pretreated with a combination of heat, ultraviolet light, and oxygenation. The effect was not the trivial result of donor T cell destruction because T cell reconstitution, although delayed, recovered to normal levels within 2 weeks. Protection from GVHD required oxygenation and was associated with normalization of the CD4/CD8 donor T cell ratio, recovery of host hematopoiesis, and decreased inflammatory cytokine production. CONCLUSION: Pretreatment of donor T cells with a combination of physicochemical stressors effectively prevents GVHD caused by major major histocompatibility disparities and may facilitate the safe transplantation of patients without HLA-identical donors.

Acute intestinal graft-versus-host disease in a syngeneic bone marrow transplant recipient.

BACKGROUND: We report a case of intestinal graft-versus-host disease (GVHD) in a syngeneic bone marrow transplant patient. METHODS: Several days after receiving a bone marrow transplant from his identical twin for treatment of non-Hodgkin's lymphoma, a 47-year-old man developed a skin rash and diarrhea. RESULTS: A colonic biopsy on day +15 revealed characteristic changes of acute intestinal GVHD. Molecular studies (microsatellite DNA and HLA sequence-specific primer polymerase chain reaction analyses) confirmed the genotypic identity of donor and host and the improbability of transfusion-associated GVHD. CONCLUSION: This case illustrates that pathological evidence of GVHD does not absolutely require the presence of genetic differences between host and donor and questions existing concepts about the nature of cyclosporine-induced GVHD.

Abnormal thyroid stimulating hormone (TSH) levels in adults following allogeneic bone marrow transplants.

Thyroid function abnormalities in 270 adult patients post-BMT are described. Various conditioning regimens were used and the effects of three TBI and one chemotherapy only based regimens are compared. The overall incidence of elevated TSH is 8.9; 3.8, 7.2 and 16.7% in those patients who received 300, 500 and 1200 cGy respectively and 11.7% in those who received BuCy conditioning. Three cases (1.1%) of clinial hypothyroidism were observed. Compensated hypothyroidism defined as an elevated TSH in the presence of normal T3, T4 levels and transient in some cases, was the most common finding. All but four cases occurred in the first 2 years after BMT. In the remaining four, three occurred in patients with chronic GVHD. The results reported here show a lower prevalence than observed in most other reviews, particularly for children. A trend was observed with increasing radiation doses. The results are not significantly different from those we observed in the BuCy regimen.

Bone marrow aspirates as part of routine donor assessment for allogeneic blood and marrow transplantation can reveal presence of occult hematological malignancies in otherwise asymptomatic individuals.

Pre transplant screening work-up of donors for allogeneic blood and marrow transplantation is essential in an effort to minimize risks to the recipient and protect the donor. At Princess Margaret Hospital, every potential donor is screened with a bone marrow aspirate. The case histories of three asymptomatic potential donors who presented within 1 year with normal complete blood counts, history and physical examination are presented. A 65-year-old male patient was diagnosed with smouldering multiple myeloma, a 72-year-old male patient with chronic lymphocytic leukemia and a 42-year-old male patient with myelodysplastic syndrome. Bone marrow examination led to the diagnosis in each one of these cases. Of note is that each of the potential donors was discovered to have the same disease as the transplant recipient. In vitro clonogenic hemopoietic progenitor assays were compared to those of 20 normal volunteers. Inferior growth of hemopoietic progenitor colonies in all three was noted. In conclusion, particularly in older donors and donors with potential for familial malignancies, more screening investigations including bone marrow aspiration may be reasonable to investigate for occult hematological malignancies prior to stem cell donation. Clonogenic assays can contribute to detect hemopoietic abnormalities pre transplant.

Clinical and economic analysis of allogeneic peripheral blood progenitor cell transplants: a Canadian perspective.

Allogeneic peripheral blood progenitor cell (PBPC) transplants are an alternative to BMT, although G-CSF mobilization dose, timing of pheresis and risk of GVHD are not well defined. We compared harvest characteristics, donor and recipient outcomes and costs of two PBPC transplant strategies with historical controls who received BMT. Twenty donors mobilized with four daily s.c. G-CSF doses (5 microg/kg/day) (group 1) and 20 mobilized with 10 microg/kg/day G-CSF (group 2) were compared with 20 BM controls (group 3). G-CSF and phereses were well tolerated. Four of 40 PBPC donors required femoral catheter placement. At least 2.5 x 10(6) CD34+/kg recipient weight were collected with two phereses in 19/20 donors (group 1) and 18/20 donors (group 2). Time to neutrophil (18 vs 20 vs 22 days, P = 0.02) and platelet (21 vs 24 vs 27 days, P = 0.005) engraftment was shorter in the PBPC groups (group 2 vs group 1 vs group 3) but secondary engraftment outcomes were not different. The incidence of grade 2-4 aGVHD was higher in the low-dose G-CSF group (group 1) but there was no difference in cGVHD, 100-day or 1-year survival. The mean PBPC transplant cost (group 1) at first hospital discharge was less than BM (group 3) ($34,643 vs $37,354) but the mean overall cost for both groups was similar at 100 days ($46,334 vs $46,083). Allogeneic PBPC transplant with short course, low-dose G-CSF mobilization is safe, feasible and cost equivalent to allogeneic BMT.

Bcl-2 clearance: optimising outcomes in follicular non-Hodgkin's lymphoma.

The long median survival time of patients with follicular non-Hodgkin's lymphoma (NHL), means that the efficacy of new treatments are difficult to assess in the short term. Bcl-2 is an inhibitor of apoptosis and overexpression of the bcl-2 gene in the blood or bone marrow is a feature in up to 85% of patients with follicular NHL. Levels of bcl-2(+) cells in the peripheral blood or bone marrow therefore are a useful measure of disease status in such patients and can be detected by polymerase chain reaction (PCR). Complete bcl-2 clearance from the bone marrow (molecular remission) following autologous stem cell transplant (ASCT) for follicular NHL is considered to be an important prognostic factor for disease-free survival. Tumour cell contamination of the stem cell grafts used in ASCT is commonly associated with relapse. This can be addressed by purging the stem cell harvest prior to transplantation. Various methods of in vitro purging after stem cell collection have been shown to reduce the level of contamination but yield is invariably reduced and grafts remain bcl-2 positive. However, in vivo purging with rituximab during the process of collection has been used to obtain bcl-2-negative stem cell harvests without compromising the yield. Rituximab is a monoclonal antibody licensed for treatment of relapsed and refractory low-grade or follicular NHL. Rituximab targets the CD20 antigen, which is found on cells of the B cell lineage. When used for in vivo purging it depletes the peripheral blood of CD20-positive cells and prevents contamination by lymphoma cells. Molecular remission, as measured by bone-marrow bcl-2 clearance, has been achieved in 7/7 patients with follicular NHL at 1 year after treatment with ASCT using rituximab as an 'in vivopurse', followed by rituximab maintenance. Early clinical outcomes are also encouraging.

Prolonged molecular and clinical remission after treatment of a patient with follicular lymphoma with rituximab.

Rituximab is a chimeric anti-CD20 monoclonal antibody that has approval for single agent therapy in the treatment of relapsed/refractory low grade or follicular non-Hodgkin's Lymphoma. In published phase II trials, molecular remissions of PCR detectable t(14;18) disease in the peripheral blood have been reported in up to 62% of patients by three months. We report a case of a patient who achieved prolonged clinical and molecular remission following a single four week course of Rituximab that has exceeded any previous remission achieved with chemo-radiotherapy. The implications of molecular remission as a surrogate of clinical remission and molecular relapse as a harbinger of clinical relapse are reviewed and discussed.