Nonengraftment haploidentical cellular immunotherapy for refractory malignancies: tumor responses without chimerism.

Allogeneic bone marrow transplantation relies on immunosuppression, which controls graft-versus-host disease (GVHD) and allows engraftment at the expense of diminished graft versus-tumor (GVT) activity. Advances in hematologic transplantation have prompted the development of effective, less-toxic regimens that attempt to balance GVH and GVT immunoreactions. We analyzed the safety and efficacy of haploidentical transplantation in a Phase I/II nonimmunosuppressive, nonmyeloablative setting. A total of 41 patients with relapsed refractory cancer received 100 cGy of total body irradiation (TBI), along with an infusion of 1 x 10(6) to 2 x 10(8) CD3+ cells/kg; 29 patients received the highest dose. A postinfusional cellular graft rejection syndrome resembling engraftment syndrome was noted at the 2 highest CD3+ infusion cohorts. There were 26 patients with hematologic malignancies with 14 responses, 9 of which were major. Two of 6 patients with lymphoma remained free of disease at 76 months and 82 months, respectively; there were 5 durable complete responses and 4 partial responses in 13 patients with acute myelogenous leukemia (AML). All responses occurred outside of donor chimerism. TBI at 100 cGy followed by HLA-haploidentical immunotherapy is a biologically active therapy for patients with refractory AML and lymphoma. Possible mechanisms contributing to its effectiveness include initial GVT kill, breaking of host tolerance to tumor through cross-reactive alloreactive responses, persistent nondetectable microchimerism, or some combination of these.

Comparable outcomes in nonsecretory and secretory multiple myeloma after autologous stem cell transplantation.

Nonsecretory myeloma (NSM) accounts for <5% of cases of multiple myeloma (MM). The outcome of these patients following autologous stem cell transplantation (ASCT) has not been evaluated in clinical trials. We compared the outcomes after ASCT for patients with NSM reported to the Center for International Blood and Marrow Transplant Research (CIBMTR) between 1989 and 2003, to a matched group of 438 patients (4 controls for each patient) with secretory myeloma (SM). The patients were matched using propensity scores calculated using age, Durie-Salmon stage, sensitivity to pretransplant therapy, time from diagnosis to transplant, and year of transplant. Disease characteristics were similar in both groups at diagnosis and at transplant except higher risk of anemia, hypoalbuminemia, and marrow plasmacytosis (in SM) and plasmacytoma (more in NSM). Cumulative incidence of treatment-related mortality (TRM), relapse, progression-free survival (PFS), and overall survival (OS) were similar between the groups. In multivariate analysis, based on a Cox model stratified on matched pairs and adjusted for covariates not considered in the propensity score, we found no difference in outcome between the NSM and SM groups. In this large cohort of patients undergoing ASCT, we found no difference in outcomes of patients with NSM compared to those with SM.

ABO blood group barrier in allogeneic bone marrow transplantation revisited.

Reports have shown a worse outcome for donor-recipient pairs mismatched for ABO blood groups in bone marrow transplantation (BMT). These studies, however, included small and heterogeneous study populations, and not all considered bidirectional ABO incompatibility separately. Because the issue remains controversial, we analyzed the effect of ABO mismatch on the overall survival, transplant-related mortality, and occurrence of acute and chronic graft-versus-host disease (GVHD) in a large homogenous group of patients undergoing allogeneic BMT. A total of 3103 patients with early-stage leukemia who underwent transplantation between 1990 and 1998 with bone marrow from an HLA-identical sibling and who were reported to the Center for International Blood and Marrow Transplant Research were studied. The median follow-up was 54 months. A total of 2108 (67.9%) donor-recipient pairs were ABO identical, 451 (14.5%) had a minor mismatch, 430 (13.9%) had a major mismatch, and 114 (3.7%) had a bidirectional ABO mismatch. The groups did not differ significantly in patient or donor characteristics except for more female-to-male sex mismatch in the bidirectional ABO mismatch group (P = .017). In multivariate models of overall survival, transplant-related mortality, and grade II to IV acute GVHD, there were no significant differences among the 4 groups. Bidirectional ABO mismatch was associated with a significantly higher risk of grade III or IV acute GVHD (hazard ratio, 1.869; 95% confidence interval, 1.192-2.93; P = .006). Patients with major ABO mismatch received red blood cell transfusions (P = .001) for a longer timer after transplantation and had a slightly slower neutrophil recovery (P < .001). There was no evidence of a substantial effect of ABO blood group incompatibility on the outcome of conventional BMT among patients with leukemia.

Granulocyte-colony stimulating factor primed bone marrow and granulocyte-colony stimulating factor mobilized peripheral blood stem cells are equivalent for engraftment: which to choose?

The first reported bone marrow transplant was published in 1939, although it was deemed unsuccessful. Between 1957 and 1965, numerous reports of bone marrow transplants, many of which were successful, were published for patients with irradiation injury, aplastic anemia, leukemia, lymphoma, and myeloma. Sources of marrow were autologous, isologous, and homologous (often unrelated, including cadaveric) donors. Bone marrow infusion was shown to be safe. It was also demonstrated that an aliquot of marrow, removed (harvested) from the ileum, had sufficient hematopoietic stem cells (SC) to repopulate the marrow and restore blood counts after myeloablation. For about 20 yr, bone marrow was the only source of hematopoietic stem cells (HSC) for transplantation. The first reported autologous peripheral blood HSC transplant was recorded in 1981 using chemotherapy 'mobilized' SC collected by leukapheresis. Mobilization is defined for these purposes to be any treatment that enhances the number of HSC in the blood such that the collection contains sufficient HSC to repopulate the marrow and restore blood counts after myeloablation. Since the early 1990s, SCT using blood-derived stem cells has become very popular and very common. The principal reason is that mobilized (whether by H growth factor or during recovery after chemotherapy) blood-derived stem cells engraft more rapidly than do marrow-derived stem cells. On the one hand, bone marrow was always harvested in the resting, unperturbed state (steady state). On the other hand, blood stem cells (BSC) were virtually always collected after mobilization, usually with granulocyte-colony stimulating factor (G-CSF). There is one report of collection of steady state BSC used for transplantation, and slow engraftment was documented. Bone marrow was never harvested after either chemotherapy or growth factor (priming). It is the mobilization (most often with G-CSF alone or after chemotherapy) of BSC that produces more rapid engraftment than for steady state marrow stem cells (MSC). This contribution shows the data that changes the old paradigm to a new paradigm which states bone MSC and BSC engraft identically if collected after the same pretreatment of the donor with growth factor.

Anti-CD3 x anti-HER2 bispecific antibody effectively redirects armed T cells to inhibit tumor development and growth in hormone-refractory prostate cancer-bearing severe combined immunodeficient beige mice.

The bispecific antibody (BiAb) anti-CD3 x anti-Her2/neu (Her2Bi), combines Her2/neu targeting with nonmajor histocompatibility complex-restricted cytotoxicity mediated by activated T cells (ATCs). To evaluate this adaptive immunotherapeutic strategy for augmenting antitumor immune response toward hormone-refractory prostate cancer (HRPC), normal donor or patient T cells were activated with anti-CD3, expanded ex vivo in interleukin-2, and then armed with Her2Bi (5-500 ng per million ATCs). In vitro, arming ATCs with Her2Bi increased the percent specific cytotoxicity toward PC-3 prostate adenocarcinoma cells 2-3 fold and increased the secretion of Th1 cytokines granulocyte-macrophage colony-stimulating factor, tumor necrosis factor-alpha, and interferon-gamma when compared with unarmed ATCs or ATCs armed with an irrelevant BiAb. Her2Bi-armed ATCs administered with PC-3 (Winn Assay) or injected intratumorally prevented development or induced remissions, respectively, of PC-3 tumors in severe combined immunodeficient beige mice. Intravenously administered Her2Bi-armed ATCs localized to PC-3 xenografts mediated cytotoxicity toward tumor cells and produced significant tumor growth delay of PC-3 tumors, but not Her2/neu-negative LS174T colon adenocarcinoma xenografts. By flow cytometry analyses, Her2Bi-armed ATCs had a proliferative advantage over unarmed ATCs and persisted in the circulation and tumor tissues longer than unarmed ATCs. These findings suggest that Her2Bi-armed ATC therapy may be an effective, nontoxic, tumor-specific treatment for Her2-positive HRPC.

Primed marrow for autologous and allogeneic transplantation: a review comparing primed marrow to mobilized blood and steady-state marrow.

Mobilized peripheral blood collections, obtained following either chemotherapy (with or without granulocyte colony-stimulating factor (G-CSF)) or G-CSF administration alone, are rapidly replacing traditional bone marrow harvests as the source of cells for hematopoietic stem cell transplantation. According to the Autologous Blood and Marrow Transplant and the International Bone Marrow Transplant Registries, for the years 1998 through 2000, blood stem cell (BSC) transplants accounted for about 80% of autologous transplants in the pediatric age group and more than 90% of the autologous transplants among adults. In allogeneic transplantation, where the donor is a healthy family member or normal volunteer, G-CSF-mobilized BSC transplants are being used more and more frequently, accounting for about 20% of allogeneic transplants in the pediatric age range and more than 40% of allogeneic transplants among adults during the same time period. It is not, therefore, too great a stretch to imagine that BSC transplants will soon be, if not already, in the majority for allogeneic transplantation among adults. The principal reason why this is happening is the prevailing view that BSC engraft more rapidly than marrow stem cells (MSC). However, this view is based on comparisons between primed circulating blood cells (BSC) and unprimed resident marrow cells in the steady state (SS-MSC). If the reason why BSC engraft faster than SS-MSC were a consequence of G-CSF used for mobilization, then would priming of MSC by G-CSF (Prim-MSC) accelerate engraftment of marrow as well? We reviewed the literature of the last 10 years to see if there were enough data to answer this question.

Redirected T-cell cytotoxicity to epithelial cell adhesion molecule-overexpressing adenocarcinomas by a novel recombinant antibody, E3Bi, in vitro and in an animal model.

BACKGROUND: To redirect cytotoxic T cells to target a broad range of adenocarcinomas, the authors constructed a novel, recombinant, bispecific antibody, E3Bi, directed at the tumor-associated antigen, epithelial cell adhesion molecule (EpCAM), and the CD3 receptor on T cells. METHODS: T cells were prepared from healthy blood donors. The cytotoxicity of activated T cells (ATC) redirected to tumor cells by E3Bi was measured with in vitro (51)Cr release assays. In vivo studies were performed in a severe combined immunodeficient (SCID)/Beige mouse xenograft model. Tumor-bearing mice were treated with low doses (1 mg/kg) or high doses (10 mg/kg) of E3Bi along with ATC (2 x 10(9) cells/kg), and treatment efficacy was evaluated both by ex vivo tumor cell survival assay after in vivo treatments and by in vivo tumor growth delay studies. RESULTS: In vitro, targeting the EpCAM-overexpressing human tumor cell lines with E3Bi increased specific cytotoxicity of ATC by > 70% at an effector-to-target ratio of 2.5 (P < 0.001); this cytotoxicity was abolished competitively in the presence of an anti-EpCAM monoclonal antibody. In contrast, E3Bi did not enhance ATC cytotoxicity toward the low EpCAM-expressing tumor cell line. In ex vivo tumor cytotoxicity assays, a significant reduction in tumor cell survival (40% with low-dose E3Bi; 90% with high-dose E3Bi) was observed in E3Bi/ATC-treated mice compared with control mice that were treated with ATC only. In addition, SCID/Beige mice xenografted with LS174T tumors demonstrated a significant tumor growth delay (P = 0.0139) after receiving E3Bi/ATC/interleukin 2 (IL-2) compared with mice that received ATC/IL-2 alone. CONCLUSIONS: E3Bi specifically and very efficiently redirected T cells to destroy EpCAM-overexpressing tumors both in vitro and in an animal model. These results suggest a therapeutic utility for E3Bi in the treatment of adenocarcinomas.

Do G-CSF mobilized, peripheral blood-derived stem cells from healthy, HLA-identical donors really engraft more rapidly than do G-CSF primed, bone marrow-derived stem cells? No!

For more than a decade, the notion that peripheral blood-derived stem cells engraft more rapidly than bone marrow-derived stem cells after high-dose therapy has dominated our thinking. Recently, reports that granulocyte colony-stimulating factor (G-CSF) induces a proteolytic marrow microenvironment have provided mechanistic support for that belief, compelling us to review our own experience of 29 consecutive transplants with HLA-identical blood and marrow stem cells. In contrast to several reported randomized controlled trials, we found marrow stem cells engraft just as rapidly (median day 11 for granulocytes over 500/microl and median day 17 for platelets over 20,000/microl) as blood stem cells (median day 12 and median day 19, respectively) if the donor is treated with G-CSF in the same manner before marrow harvest as the donor is treated with G-CSF before leukapheresis. These observations with healthy HLA-identical donors confirm the results of our prior randomized autotransplant study. We propose the concept that the level of activation of the stem cells (induced by G-CSF) determines engraftment kinetics and not the anatomical site of derivation.

Novel approaches in the treatment of multiple myeloma.

Despite all the new advances, treatment of multiple myeloma remains very challenging. Table 6 summarizes the outcomes of some of the commonly used treatments from different clinical trails. Treatment with conventional chemotherapy alone resulted in median survival from 18 to 30 months. Aggressive treatments like VBMCP (vincristine, bleomycin, melphalan, cyclophosphamide, predisone) although resulting in higher response rates, resulted in higher treatment-related mortality and did not have any effect on median and 5 year survival. High dose chemotherapy with stem cell support increased the median survival to around 60 months and resulted in durable complete responses in a significant number of the patients. Some of the newer and promising treatments for multiple myeloma are summarized in Table 7.

The chronic leukemias: current and novel therapeutic approaches.

Recent developments with targeted therapies have expanded the therapeutic armamentarium (Table 1) for patients with both CLL and CML. Significant advances in allogeneic and autologous donor transplantation have increased the patient eligibility pool and reduced the toxicities while improving upon long term survival results.

The latest treatment advances for acute myelogenous leukemia.

The research into pathogenesis and mechanisms behind AML is advancing rapidly, but in general, translation into global application for the majority of patients is wanting. As more becomes known about the cytogenetic and molecular characteristics of leukemia cells and the pathways of leukemogenesis are further elucidated, it is hoped that future therapies will be directed more specifically toward the least toxic method to eradicate clonal malignant cells. HLA-haploidentical and alloBMT using KIR mismatch may dramatically improve survival for many more patients.

Shc proteins are strong, independent prognostic markers for both node-negative and node-positive primary breast cancer.

Activated Shc signaling proteins are implicated in many pathways associated with aggressive disease, and many breast cancer cell lines derived from highly aggressive tumors contain high levels of activated, tyrosine phosphorylated (PY)-Shc (the M(r) 46000 and M(r) 52000 isoforms) relative to levels of an inhibitory M(r) 66000 Shc isoform. It was, therefore, hypothesized that high amounts of PY-Shc relative to the M(r) 66000 Shc isoform would serve as a marker for aggressive neoplasms. Semiquantitative immunohistochemical analyses of PY-Shc and p66 Shc were performed on archival primary breast tumor specimens from 116 women, 17 of whom experienced relapse (6.1 years median follow-up of nonrelapsed patients). Consistent with our hypothesis, staining intensities demonstrated that increased amounts of PY-Shc (P = 0.01) and decreased expression of p66-Shc protein (P = 0.028) correlated with disease recurrence. Modeled as the ratio of PY-Shc to p66 Shc, the Shc ratio correlated strongly with nodal status (P = 0.003), tumor stage (P = 0.0025), and disease stage (P = 0.002) and was 2-fold higher in primary tumors of patients who subsequently relapsed (P < 0.001). Univariate Cox proportional hazards analysis of relapse-free survival demonstrated the prognostic value of PY-Shc (P = 0.01), p66 Shc (P = 0.04), and the Shc ratio (P = 0.004) as continuous variables, with a hazard ratio (HR) of 10 (P = 0.007) for the Shc ratio. Shc ratio cut points of <0.35 and >0.65 were identified and independently validated to maximize negative predictive value and positive predictive value. Patients with low Shc ratios (n = 36) had a 0.08 HR of relapse (P = 0.007) compared with patients with high Shc ratios, experiencing an 8-year cumulative 2.9% and 55% relapse hazard, respectively, compared with a 22% relapse hazard in the total cohort. The Shc ratio had similar prognostic value for disease-specific survival. In multivariate models, the Shc ratio, both as a continuous variable and as a cut point-categorized variable, was independent of all measured covariates (including nodal status, tumor stage, disease stage, grade, estrogen receptor status, and adjuvant therapy) and was a stronger prognostic marker than all but nodal status. All relapsed node-positive patients had very high Shc ratios (>0.80; P = 0.006) in their primary tumors. Furthermore, the Shc ratio was a strong, independent prognostic indicator in node-negative patients (79 patients, 10 recurrences), with a HR of 0.086 (P = 0.02) that was independent of clinical markers and adjuvant therapy. Patients with low and high Shc ratios experienced a 3.6% and 64% relapse hazard, respectively, compared with 20% in the total node-negative cohort.