Mechanisms of donor-specific tolerance in recipients of haploidentical combined bone marrow/kidney transplantation.

We recently reported long-term organ allograft survival without ongoing immunosuppression in four of five patients receiving combined kidney and bone marrow transplantation from haploidentical donors following nonmyeloablative conditioning. In vitro assays up to 18 months revealed donor-specific unresponsiveness. We now demonstrate that T cell recovery is gradual and is characterized by memory-type cell predominance and an increased proportion of CD4⁺ CD25⁺ CD127⁻ FOXP3⁺ Treg during the lymphopenic period. Complete donor-specific unresponsiveness in proliferative and cytotoxic assays, and in limiting dilution analyses of IL-2-producing and cytotoxic cells, developed and persisted for the 3-year follow-up in all patients, and extended to donor renal tubular epithelial cells. Assays in two of four patients were consistent with a role for a suppressive tolerance mechanism at 6 months to 1 year, but later (≥ 18 months) studies on all four patients provided no evidence for a suppressive mechanism. Our studies demonstrate, for the first time, long-term, systemic donor-specific unresponsiveness in patients with HLA-mismatched allograft tolerance. While regulatory cells may play an early role, long-term tolerance appears to be maintained by a deletion or anergy mechanism.

Long-term outcome and prognostic factors of second allogeneic hematopoietic stem cell transplant for acute leukemia in patients with a median follow-up of ⩾10 years.

Second allogeneic hematopoietic stem cell transplantation (HSCT2) is a frequently used treatment option for relapse of acute leukemia after first allogeneic transplantation. Remission can be induced in selected patients, but data on long-term outcome and finally cure are limited. To estimate the long-term results of HSCT2, we retrospectively analyzed the course of 286 patients receiving myeloablative HSCT2 between 1985 and 2000, with a median follow-up of 11.3 years. Overall survival (OS) and leukemia-free survival (LFS) at 10 years from HSCT2 were 10±2 and 7±2%, respectively. Cumulative 10-year incidence of relapse and non-relapse mortality were 58±3% and 35±3%, respectively. CR at HSCT2, an interval from first transplant to relapse >10 months and TBI as part of the conditioning for HSCT2 favorably influenced LFS and OS. Patients with all three favorable factors had a 10-year OS of 36±10% and LFS of 25±9%, whereas patients showing no favorable factor had all died before year 5. Although retrospective, the long follow-up of this analysis supports the curative potential of alloHSCT2 in selected patients, who might be identified in advance, based on prognostic factors.

Cytokines in cancer therapy.

Cytokines are crucial factors in the activation and development of immune response, including responses against tumor cells. Interleukin (IL)-2, a T-cell growth factor, has been largely used to activate T and NK cells in vivo and to maintain such an activation for therapeutic purposes. When given to patients, IL-2 was shown to cause clinical responses, especially in metastatic melanoma and renal cancer patients, though its mechanism of action could not be completely elucidated. Cytokines (IL-2, IL-12, GM-CSF) are also used as natural adjuvants of vaccines of various formulation to help in activating and maintaining an antitumor immune response. This review summarizes findings deriving from the use of cytokines in cancer therapy and provides insights into future approaches when a more appropriate use of cytokines, together with new vaccines, is likely to improve clinical outcome.

Rationale for the use of gemcitabine in breast cancer (Review).

Many active cytotoxic drugs and several regimens exist for breast cancer therapy. However, these conventional treatments have not changed the outcome of patients with locally advanced and metastatic disease. As a consequence, the dynamic balance between chemotherapy-induced side effects and benefits attributable to relief of cancer-related symptoms must be carefully considered in this setting. Gemcitabine is a pyrimidine nucleoside antimetabolite that has shown activity in a variety of solid tumors, a good toxicity profile, and non-overlapping toxicity with other chemotherapeutic drugs. As a single agent, gemcitabine yields response rates ranging from 14 to 37% as first-line treatment for advanced breast cancer and 12-30% as salvage therapy for patients previously treated with anthracycline and/or taxane treatment. Combined with vinorelbine, platinum, anthracyclines, and taxanes as doublets or triplets, response rates of 50 to 80% have been reported in phase II clinical studies. Gemcitabine in combination with anthracyclines and taxanes has been evaluated in the neoadjuvant setting in patients with early-stage breast cancer with interesting clinical and pathological response rates. Preliminary results of gemcitabine in combination with the biologic agent, trastuzumab, are encouraging. Phase III trials of gemcitabine combinations compared to standard regimens are ongoing with the aim to assess the independent contribution of gemcitabine.

Filgrastim XM02 (Tevagrastim®) after autologous stem cell transplantation compared to lenograstim: favourable cost-efficacy analysis.

PURPOSE: Granulocyte colony-stimulating factors (G-CSFs), filgrastim and lenograstim, are recognised to be useful in accelerating engraftment after autologous stem cell transplantation. Several forms of biosimilar non-glycosylated G-CSF have been approved by the European Medicines Agency, with limited published data supporting the clinical equivalence in peripheral blood stem cell mobilisation and recovery after autologous stem cell transplantation. METHOD: With the aim of comparing cost-effective strategies in the use of G-CSF after autologous stem cell transplantation, we retrospectively evaluated 32 patients consecutively treated with biosimilar filgrastim XM02 (Tevagrastim) and 26 with lenograstim. All patients received G-CSF (biosimilar or lenograstim) at a dosage of 5 mcg/kg/day subcutaneously from day 5 to absolute neutrophil count of 1500/mmc for three days. RESULTS: The median time to absolute neutrophil count engraftment was 11 days for the filgrastim XM02 group and 12 days for the lenograstim group. As for platelets recovery, the median time was 12 days in both groups. The median number of G-CSF vials used for patients was 9.5 for Tevagrastim and 10.5 for lenograstim, reflecting a mean estimated cost of about 556.1 euros for Tevagrastim versus 932.2 euros for lenograstim (p< 0.001). The median days of febrile neutropenia were 1.5 and 1 for filgrastim XM02 and lenograstim, respectively. No adverse event related to the use of XM02 filgrastim was recorded. CONCLUSION: In our experience, filgrastim XM02 and lenograstim showed comparable efficacy in shortening the period of neutropenia after cytoreduction and autologous stem cell transplantation, with a favourable cost effect for filgrastim XM02.

Rituximab and subcutaneous 2-chloro-2'-deoxyadenosine as therapy in untreated and relapsed Waldenström's macroglobulinemia.

With the aim to assess the efficacy of subcutaneous cladribine in combination with rituximab, 29 newly diagnosed/pretreated WM patients were enrolled in a multicenter phase II trial. Intended therapy consisted of rituximab on day 1 followed by s.c. cladribine 0.1 mg/kg for 5 consecutive days, administered monthly for 4 cycles. The expression of genes involved in cladribine metabolism was evaluated. With a median follow-up of 50 months the ORR rate observed was 89.6% without any difference between newly or pretreated patients (P=.522). The therapy was well tolerated; no major infections were observed, no patients developed transformation to high-grade NHL nor myelodysplasia. Low expression levels of hCNT1 correlated with the failure to achieve a CR (P=.024). The combination of rituximab/cladribine is safe and effective in WM patients requiring treatment. The pharmacogenomic analysis suggests that hCNT1 might be beneficial in predicting clinical response to such a combination treatment.

Tissue microarrays in diffuse large B-cell lymphomas: are they really able to identify distinct prognostic groups in lymphomas of both nodal and extranodal origin?

AIMS: Diffuse large B-cell lymphomas (DLBCL) can be divided into different subgroups (germinal center B-cell-like [GCB] and non-GCB) according to their gene expression profiles. Immunohistochemistry has been proposed as a surrogate for identifying these subgroups, but data about its efficacy in providing prognostic information are conflicting. METHODS AND RESULTS: This study retrospectively analyzed a series of 105 DLBCL, defined as GCB and non-GCB according to CD10, bcl-6, and MUM1 expression. All patients received a first-line anthracycline-based (CHOP-like) chemotherapy. A total of 50 patients (48%) were identified as GCB and 55 (52%) as non-GCB. The overall response rate was 89% (94/105), with 62 (59%) complete response. Disease progressions were equally distributed between the 2 subgroups and were not significantly different (P = .756) considering the primary site of involvement (nodal or extranodal). The median follow-up was 62 months (range 5-126 months). Overall survival at 5 years was not significantly different between the groups (P = .3468) and was 72.3% and 66.6% for GCB and non-GCB, respectively. CONCLUSION: The results do not support the prognostic value of GCB and non-GCB immunohistochemical categories in DLBCL of both nodal and extranodal origin. Furthermore, a limited number of antigens may be not sufficient to identify the same patterns defined by cDNA microarray. Prospective studies are warranted to address this issue.

Expression of the human concentrative nucleotide transporter 1 (hCNT1) gene correlates with clinical response in patients affected by Waldenström's Macroglobulinemia (WM) and small lymphocytic lymphoma (SLL) undergoing a combination treatment with 2-chloro-2'-deoxyadenosine (2-CdA) and Rituximab.

PURPOSE: Resistance to nucleoside analogues agents is likely to be multifactorial and could involve a number of mechanisms affecting drug penetration, metabolism and targeting. In vitro studies of resistant human cell lines have confirmed that human concentrative nucleoside transporter 1 (hCNT1)-deficient cells display resistance. EXPERIMENTAL DESIGN: We applied real-time PCR method to assess the mRNA expression of equilibrative and concentrative nucleoside transporter (hENT1, hCNT1), deoxycytidine and deoxyguanosine kinase (dCK, dGK), 5'-nucleotidase (5'-NT), ribonucleotide reductase catalytic and regulatory (RR1, RR2) subunits in bone marrow cells from 32 patients with Waldenström's Macroglobulinemia (WM) and small lymphocytic lymphoma (SLL) who received 2CdA-based chemotherapy. Responses to chemotherapy, were then correlated to the expression of these markers. RESULTS: All 32 patients enrolled expressed lower levels of hCNT1 as compared to healthy donors. In univariate analysis, lower expression level of hCNT1 (p=0.0021) and RR2 (p=0.02) correlated with response to chemotherapy. In particular, patients with low levels of hCNT1 achieved inferior clinical response. No significant correlation between these genes expression and age, stage of disease was found. This study suggests that nucleotidase expression levels can be used to identify subgroups of WM and SLL patients who will likely respond differently to a 2CdA-based therapy.

T-cell recognition of melanoma-associated antigens.

In this review, we summarize the significant progress that has been made in the identification of melanoma-associated antigens (MAA) recognized by cytotoxic T-lymphocytes (CTL). These antigens belong to three main groups: tumor-associated testis-specific antigens (e.g. , MAGE, BAGE, and GAGE); melanocyte differentiation antigens (e.g., tyrosinase, Melan-A/MART-1); and mutated or aberrantly expressed molecules (e.g, CDK4, MUM-1, beta-catenin). Although strong CTL activity may be induced ex vivo against most of these antigens, often in the presence of excess cytokines and antigen, a clear understanding of the functional status of CTL in vivo and their impact on tumor growth, is still lacking. Several mechanisms are described that potentially contribute to tumor cell evasion of the immune response, suggesting that any antitumor efficacy achieved by immune effectors may be offset by factors that result ultimately in tumor progression. Nevertheless, most of these MAA are currently being investigated as immunizing agents in clinical studies, the conflicting results of which are reviewed. Indeed, the therapeutic potential of MAA has still to be fully exploited and new strategies have to be found in order to achieve an effective and long-lasting in vivo immune control of melanoma growth and progression.