bluuebunny/crossref_metadata_2025 huggingface dataset:
First Report of the HOVON 45: A Phase II Study with Rituximab, High Dose Ara-C and Autologous Stem Cell Transplantation in the Primary Treatment of Mantle Cell Lymphoma.
Authors: Van’t Veer Mars B. , Notenboom Annelise , McKenzie Marius , Kluin-Nelemans Hanneke , van Oers Rien , Zijlstra Jose , de Jong Daphne , Sonneveld Pieter , Hagenbeek Anton
Published: October 2019

The Dutch Hemato-Oncology Study Group (HOVON) evaluated in a phase II trial the efficacy of the addition of rituximab, high dose Ara-C and autologous stem cell transplantation after BEAM conditioning to conventional CHOP in 88 previously untreated patients with mantle cell lymphoma, with respect to remission rate, failure free survival (FFS) and overall survival (OS). From 2002 to 2005, 86 eligible patients were included and evaluated. The male : female ratio was: 4 and the median age 55 years (32–66). Median follow-up of all patients still alive was 25.5 months. Patients were treated with R-CHOP21 x 3 (rituximab 375 mg/m2, cyclophosphamide 750 mg/m2, doxorubicin 50 mg/m2, vincristine 1.4 mg/m2 day 1 and prednisone 100 mg days 1 – 5). Stem cells were harvested after high dose Ara-C x 1 (2g/m2 BID days 1 – 4), followed by rituximab on day 11 for in vivo purging and G- CSF. After haematological recovery BEAM conditioning (carmustine 300mg/m2 day 1, Ara-C 200mg/m2 and etoposide 200 mg/m2 days 2 – 5 and melphalan 2 x 100 mg/m2 day 6) was given with autologous stem cell support day 8). After 3 x R-CHOP 17 patients did not reach PR and went off study, according to the protocol, and 5 patients were excluded for other reasons. Sixty four patients received HD Ara-C and 63 patients proceeded to BEAM. Haematological toxicity was as follows: the median time of recovery of leukocytes (> 1,0 x 109/l) from start of high dose Ara-C was 17 days (range: 0 – 59 days), from start of BEAM 26 days (range: 17 – 55). For platelets the median time to recovery (> 50 x 109/l) from start of high dose Ara-C was 23 days (range: 0 – 44 days) and from start of BEAM 25 days (range: 16 – 364). Non-hematological toxicity grade 3–4 was seen during CHOP in 13%, after HD Ara-C in 20%, mainly gastro-intestinal, and after BEAM in 51%, mainly gastro-intestinal and liver, of the patients. Grade 3–4 infections were seen during CHOP in 17%, after HD Ara-C in 30% and after BEAM in 59% of the patients. Responses to treatment are summarised in Table 1. At two years FFS was 67%, OS was 81%. An analysis restricted to the 63 patients who completed the protocol treatment showed FFS 90% and OS 98% at two years.

We conclude that intensification of first line treatment with rituximab, HD Ara-C and BEAM is beneficial with respect to FFS and OS in younger MCL patients and that this regimen is well tolerated.

Table 1. Response to treatment N CR PR NR/PD Not yet known after 3x CHOP 83 13 (16%) 51 (61%) 17 (20%) 2 after HD Ara-C 64 22 (34%) 28 (44%) 14 after BEAM 63 43 (68%) 13 (21%) 7 died early 1
Phase II Study of Rituximab and Cladribine (2-CDA) in Newly Diagnosed Mantle Cell Lymphoma (MCL) (N0189).
Authors: Inwards David James , Hillman David W. , Fishkin Paul A. , White William L. , Morton Roscoe F. , Dakhil Shaker R. , Nikcevich Daniel A. , Wender Donald B. , Fitch Tom R. , Kurtin Paul J.
Published: October 2019

Background: A previous trial of 2-CDA as a single agent for therapy of mantle cell lymphoma demonstrated this agent to be efficacious with an overall response rate of 81% (31% complete responses) (Blood 1999 Nov 15; 94:660a). A phase II study of the addition of rituximab to 2-CDA was conducted by the North Central Cancer Treatment Group based on improved outcomes achieved by the addition of rituximab to other regimens active in MCL.

Methods: This one-stage phase II study was designed to determine the complete response (CR) or complete response/unconfirmed (CRu) rate. Central pathology confirmation of cyclin D1 positive mantle cell lymphoma was required. No previous therapy for lymphoma was allowed, with the exception of splenectomy. The schedule was rituximab 375 mg/m2 IV day 1; 2-CDA 5 mg/m2/d IV days 1–5 of a 4-week cycle. After 2 of the first 6 patients developed grade 4 neutropenia, subsequent patients received either pegfilgrastim or filgrastim support. Patients received 2–6 cycles of therapy, depending on response. Patients were required to achieve at least a PR after 2 cycles of therapy to continue on protocol therapy.

Results: Patient characteristics of all 29 eligible pts: median age: 70 (range: 41–86); 21 male, 8 female; PS 0 (55.2%), PS 1 (41.4%), PS 2 (3.5%); stage II (6.9%), stage III (3.5%), stage IV (89.7%); prior splenectomy (20.7%). The only grade 4 adverse events occurring more than once were neutropenia (24.1%) and leucopenia (6.9%). One patient died of cerebral ischemia in the setting of pneumonia without neutropenia. Fifteen (51.7%; 95% CI: 32.5–70.6%) pts achieved a CR; only one has relapsed to date (665 days after starting therapy). Four additional pts achieved a PR. Ten pts have progressed with 4 pts progressing early at 17, 45, 46, and 71 days (two of whom have died). Ten pts (34.0%) went on to receive further therapy off study, 5 in less than a PR after 2 cycles, 2 in PR after study therapy, and 1 who went off study for a rash. Fourteen pts (48.3%) have progressed or gone on to receive additional therapy off study. At last contact, 26 (89.7%) were alive (median follow-up 14.3 months; range: 6–34). One year survival rate is 89.3% (95% CI: 78.5–100).

Conclusions: Rituximab and cladribine were well tolerated for the treatment of MCL in a group of elderly patients. The response rate may have been underestimated due to the study design, which required at least a PR after 2 cycles to continue therapy. Despite this, 52% achieved a complete remission. Complete remissions attained with this regimen appear to be durable, with a single relapse to date among 15 patients achieving CR.
Blastoid Mantle Cell Lymphoma (MCL): 5-yr Failure-Free Survival (FFS) Rate of 50%, without Failures after 2.5 Years Following Treatment with Rituximab (R)-HyperCVAD Alternating with R-Methotrexate/Cytarabine (M/A).
Authors: Romaguera Jorge Enrique , Fayad Luis , Rodriguez Maria Alma , Hagemeister F.B. , Pro Barbara , McLaughlin Peter , Younes Anas , Samaniego Felipe , Goy Andre , Sarris Andreas H. , Dang Nam H. , Wang Luhua , Kantarjian Hagop M. , Beasley Virginia , Medeiros L. Jeffrey , Katz Ruth L. , Cabanillas Fernando
Published: October 2019

Blastoid MCL has a very poor prognosis, with a median survival of 16–20 months after treatment with CHOP-like chemotherapy regimens. We recently described an intense regimen where R-HyperCVAD is alternated with R-M/A for 6–8 cycles, capable of achieving 87% complete remission (CR) rates and an overall 3-year FFS rate of 64% in a group of 97 patients with newly diagnosed aggressive MCL treated under an institutionally approved clinical trial from March 1999 to March 2001 (J Clin Oncol 23:7013–7023).

Fourteen of these patients presented with the blastoid cytologic variant and the following clinical features were compared with those without a blastoid cytology: age > 65 years, beta 2 microglobulin ≥ 3 gm/dL, elevated serum lactic dehydrogenase (LDH), and high international prognostic score (IPI). Only serum LDH was significantly different, being higher for those with blastoid presentation (p = 0.03). Rates of complete remission were lower for blastoid when compared to non-blastoid cytology (79% vs 89%, respectively) but not statistically significant (p = 0.72). With a median follow up of 57 months, there is a plateau in the FFS curve for blastoid MCL as shown below. At 57 months the median overall survival has not been reached. Contrary to what we had expected, the improved outcome in this group of patients with blastoid cytology suggests a potential for long term remission after intense, non-myeloablative chemotherapy.

Figure Figure
Long Term Efficacy of Rituximab in B-Cell Post Transplantation Lymphoproliferative Disorders (B-PTLD): Update of the Multicenter, Open Label, Phase II Trial (M39037 TRIAL).
Authors: Choquet Sylvain , Herbrecht Raoul , Stoppa Anne-marie , Vandenberghe Peter , Feremans Walter , Morschhauser Franck , Salles Gilles Andre , Jaccard Arnaud , Lamy Thierry , Lebranchu Yves , Offner Philippe , Oksenhendler Eric , Raphael Martine , Leblond Veronique , Milpied Noel-Jean
Published: October 2019

Background:

B-PTLD is a rare but severe complication observed after organ transplantation. There is no consensus on treatment modalities in this setting but Rituximab monotherapy has been presented as an effective and well tolerated treatment. Long term efficacy is unknown.

Aim:

We recently published results of the first prospective study on PTLD treatment, using four weekly injection of Rituximab in progressive or non responding tumours after immunosuppression diminution (Blood2006; 107; 3053–7). We present here update results up to 6 years after inclusion.

Methods:

From may 2000 to December 2001, 43 PTLD after solid organ transplantation have been enrolled in M39037, a franco-belgian multicentric prospective trial. The primary end point was day 80 response rate and one year results were also presented. Participating centres were contacted in order to obtain update data on surviving patients at one year, especially for their tumour status (complete response (CR), partial response (PR) or progressive disease (PD)), date of last information, eventual death and cause of death.

Results:

At one year, on 43 included patients, 26 were still alive, 12 in CR or Cru, one in PR, 10 in PD and two with insufficient information. At this time, update data are available on eight patients, six were in CR or Cru at one year and two in PD. Five patients died by organ failure (n=1), graft rejection (n=1), sudden death (n=2) or heart biopsy complication (n=1); all but one were in CR at this time. Three patients are still alive and in CR. No relapse has been diagnosed and median survival is 1188 days (with a median follow up of 1689 days for surviving patients). Final analysis on all patients will be presented at the ASH meeting.

Conclusion:

Rituximab in monotherapy seems to have a durable efficacy on PTLD after solid organ transplantation with a follow up of more than 4,5 years. Complete results will be presented at the ASH Meeting.
Treatment of 22 Patients with Amyothrophyc Lateral Sclerosis, Grafting Stem Cells through Neuroendoscopy.
Authors: De Bellis Roberto , Cordoba Alvaro , Muxi Pablo J. , Bello Laura , Caneiro Ada
Published: October 2019

Introduction: experimental and clinical research have shown the plasticity of Stem Cell in varied conditions of tissular damage. Amyothrophyc Lateral Sclerosis (ALS) is a degenerative disorder, culminating in a respiratory insufficiency and death. We have designed a trial in order to implant stem cells, in the spinal cord and in the cerebrospinal fluid (CSF), through a neuroendoscopic procedure minimally invasive.

Material and Methods: 22 patients with ALS met the inclusion criteria for this study. Spirometry with forced expiratory volume and forced vital capacity ratio not inferior to 60% was required. All patients were measured with ALS function scale to settle an individual initial score. Peripheral blood stem cells were collected through a central venous catheter by a continuous flow apheresis equipment. CD34+ cells were tested and selected. In the operating room, the first step was to insert a spinal needle in the vertebral space between L4 – L5, to obtain 4 milliliters (ml) of CSF and to suspend 2 ml of stem cells in it, injecting the mixture in the subaracnoidal space. The second step was to locate the intervertebral space between D8 – D9 and to introduce percutaneously a flexible neuroendoscope to find the spinal posterior sulcus. Once 1 millimeter (mm) deep in the posterior side of the spine, 1 ml of concentrated stem cells were transplanted. After checking through the neuroendoscope, the graft is firmly attached, 2 ml of concentrated stem cells were injected in the perispinal space and the procedure was finished.

Results: 22 patients with ALS (17 males, 5 females); with an average of 57 years old (38 – 75); 27 months after diagnosis (8 – 36) were treated. The amount of CD34+ cells employed was 7.0×10–6 per kilogram (3.3 – 17.8). Periodically assessment of the evolution of the score points of the international ALS scale, revealed that 9 improved their initial scores, 11 kept the same condition with no progressive deterioration of their neurological performance and 2 decrease their neuromuscular status. In all cases a remarkable decrease of the neuromuscular fasciculations was observed.

Conclusions: The procedure was safe, feasible and easy to reproduce. No complications or morbility was observed. Improvement of the neurological condition was registered in 9 patients and stabilization of the progressive disease in other 11. In spite of the short follow up, we think it is an encouraging new approach for an up to now, always deadly disease.
Overexpression of Lipases Enables Specific Cytotoxicity by the Lipase Inhibitor Orlistat in Chronic Lymphocytic Leukemia Cells.
Authors: Pallasch Christian P. , Schwamb Janine , Schulz Alexandra , Debey Svenja , Kofler David , Schultze Joachim L. , Hallek Michael , Ultsch Alfred , Wendtner Clemens M.
Published: October 2019

Genome-wide gene expression profiling of chronic lymphocytic leukemia (CLL) cells in comparison to healthy donor CD5-positive B-cells revealed deregulated expression of lipase-associated genes. A set of 19 lipase activity defined genes, e.g. LPL, phospholipases A1, −A2, −C and −D2 family members and other lipase-associated genes were overexpressed in CLL. Recently lipoprotein lipase (LPL) was identified as prognostic factor in CLL. Here we show that the expression of LPL in CLL is induced by B-cell receptor (BCR) stimulus both in mutated and unmutated CLL samples. In native and BCR-stimulated CD5+ healthy B-cells no LPL-expression was detected. Antigenic stimulation via the BCR is thought to be functionally and prognostically relevant in CLL pathogenesis, LPL expression in CLL cells could reflect a permanent BCR-stimulus due to autoantigens including also unmutated IgVH cases.

We hypothesized that the overexpression of lipases and especially of LPL reveals a putative therapeutic target by lipase inhibition through the the FDA-approved obesity drug and lipase inhibitor orlistat (tetrahydrolipstatin), which is known to inhibit LPL as well. Treatment of CLL cells with orlistat in vitro revealed significant cytotoxicity and induction of apoptosis in primary CLL cells with an IC50 of 5.48 μM (n=18). In comparison, no significant cytotoxicity was seen with healthy PBMC’s (n=12; p<0.001), even using high doses of orlistat up to 100 μM. Induction of apoptosis induction was observed both in low-risk (ZAP70 negative, n=15) and high-risk (ZAP70-positive, n=10) CLL samples. Orlistat mediated cytotoxicity was slightly decreased by BCR stimulus while additive cytotoxic effects where observed in combination with fludarabine treatment. Susceptibility to orlistat treatment was not dependent on Binet stage.

In summary, we provide in vitro data for a criticial role of fatty acid metabolism in CLL pathogenesis and suggest a therapeutic potential of the lipase inhibitor orlistat. No severe systemic side effects of orlistat have been observed in several animal models and in the current clinical application as anti-obesity drug. Orlistat seems to be a promising candidate for an anti-leukemic therapy in CLL, ongoing in vivo experiments applying the TCL1 mouse model will further elucidate the therapeutic potential of orlistat.
Polyreactive Monoclonal Antibodies Synthesized by Some B-CLL Cells Recognize Specific Antigens on Viable and Apoptotic T Cells.
Authors: Catera Rosa , Hatzi Katerina , Chu Charles C. , Herve’ Maxime , Meffre Eric , Ferrarini Manlio , Oscier David Graham , Davis Zadie , Allen Steven L. , Rai Kanti R. , Chiorazzi Nicholas
Published: October 2019

B-cell chronic lymphocytic leukemia (CLL) is a disease characterized by uncontrolled clonal expansion of a CD5+/CD19+ B cell. Clones from different patients often express receptors/mAbs with remarkably similar VHDJH rearrangements, suggesting that the same antigen(s) or epitope(s) led to the initial clonal expansion. Although the specific antigens and epitopes remain to be identified, recent data indicate that B-CLL mAbs often bind autoantigens. In certain systemic autoimmune diseases, autoantibodies can react with cell surface molecules of viable lymphocytes (anti-lymphocyte mAbs) and with intracellular molecules of apoptotic cells that are transported to the cell membrane and function as autoantigens. Therefore, we expressed recombinant antibodies from B-CLL cells as IgG1 mAbs and tested them by flow cytometry for immunoreactivity with surface molecules on irradiated (apoptotic) and non-irradiated Jurkat cells, a human T-cell line. We chose to use T cells as our target for analysis to avoid difficulties that might arise from binding to FcgRIIb on B lymphocytes.

Initial screening indicated that 5 of 25 mAbs showed intense binding. Antibodies from CLL Nos. 141 and DO8 bound to live but not apoptotic cells; in contrast, CLL Nos. 014, 114, and DO13 bound to apoptotic but not live cells. These five antibodies express unmutated VH genes. CLL 141 and DO8 express VH4-34. CLL 014 expresses VH1-69, CLL 114 VH4-39 and DO13 VH1-02. Other antibodies expressing VH1-69 in its unmutated form also bound Jurkat cells, albeit weaker than CLL 014, whereas mAbs expressing different unmutated genes from the VH1 family did not react. Among the mAbs expressing VH1-02 that were tested, only DO13 showed reactivity. This indicates that the VH1-02 gene was not solely responsible for the observed binding, and that DJ and/or VLJL segments were also involved. Remarkably, none of the tested mAbs belonging to VH3 family recognize antigens on Jurkat cells; all of these VH3 mAbs were mutated. Only CLL 114 belongs to a previously identified stereotypic set. Of interest, although CLL 014 and 141 are polyreactive, binding ssDNA, dsDNA, insulin, LPS, and HEp-2 cell lysates, their reactivities with surface antigens were specific in that CLL 141 bound only live cells and CLL 014 reacted solely with apoptotic cells.

Our results indicate that, despite their inherent polyreactivity, certain B-CLL mAb bind specifically to antigenic epitopes on the surface of live as well as apoptotic Jurkat cells. The identity of these autoantigens is currently being investigated. These data highlight the importance of autoreactivity in the promotion and evolution of B-CLL.
Randomized Comparison of Cladribine Plus Cyclophsophamide with Fludarabine Plus Cyclophosphamide in Progressive Chronic Lymphocytic Leukemia: An Updated Report of Prospective PALG-CLL3 Study.
Authors: Robak Tadeusz , Blonski Jerzy Z. , Gora-Tybor Joanna , Jamroziak Krzysztof , Ceglarek Bernadetta , Konopka Lech , Calbecka Malgorzata , Kostyra Aleksandra , Stella-Holowiecka Beata , Kloczko Janusz , Warzocha Krzysztof , Seferynska Ilona , Dmoszynska Anna , Kowal Malgorzata , Lewandowski Krzysztof , Dwilewicz-Trojaczek Jadwiga , Wiater Elzbieta , Potoczek Stanislaw , Hellmann Andrzej , Skotnicki Aleksander , Sulek Kazimierz , Dybowicz Jacek , Zawilska Krystyna
Published: October 2019

Purine analogues, Cladribine (2-CdA) and Fludarabine (FA), are highly effective in treatment of chronic lymphocytic leukemia (CLL). This prospective randomized phase 3 trial was designed to compare the efficacy and toxicity of 2-CdA and cyclophsophamide (CC regimen) with FA and cyclophsophamide (FC regimen) in previously untreated progressive CLL. The primary end points of the study were complete response (CR) and overall response (OR) after completion of the therapy. The secondary end points were progression free survival (PFS), overall survival (OS) and treatment related toxicity. Eligible patients were assigned to receive 6 courses of either 2-CdA 0.12 mg/kg/d i.v. with cyclophosphamide 250 mg/m2/d i.v. for 3 consecutive days or FA 25 mg/m2/d i.v. with cyclophosphamide 250 mg/m2/d i.v. for 3 consecutive days administered at 28 day intervals. The treatment response and toxicity were evaluated according to NCI-SWOG guidelines. The study started in January 2004, and here we present updated results from the interim analysis of 227 evaluated patients performed in July 2006. As shown in the table, there were no significant differences in the rates of CR and OR between the treatment groups. Moreover, the treatment related toxicity, including grade ¾ thrombocytopenia, neutroepenia and infections was similar in patients receiving FC or CC regimens. Median time of progression free survival did not differ between both groups (11.0 months in CC arm and 11.2 months in FC arm, p=0.41). There were 5 deaths (3.2%) observed in CC arm and 15 (8.1%) in FC arm. In conclusion, the preliminary results of our study indicate that CC and FC protocols have similar activity and toxicity in previously untreated patients with CLL.

Characteristic CC arm FC arm p value Pts enrolled 158 174 Pts evaluated 105 122 OR (%) 88 (83.8) 99 (81.2) 0.60 CR (%) 41 (39.0) 49 (40.2) 0.86 Thrombocytopenia gr 3/4 (%) 12 (10.7) 17 (13.4) 0.53 Neutropenia gr 3/4 (%) 21 (18.8) 30 (23.6) 0.36 Infections gr 3/4 (%) 34 (30.4) 37 (29.1) 0.84
Significant Correlation between Survival Endpoints and Exposure to Ofatumumab (HuMax-CD20) in Chronic Lymphocytic Leukemia.
Authors: Coiffier Bertrand , Tilly H. , Pedersen L.M. , Plesner T. , Frederiksen H. , van Oers M.H.J. , Wooldridge J. , Kloczko J.S. , Holowiecki J. , Hellmann A. , Walewski J. , Flensburg M. , Petersen J. , Robak T.
Published: October 2019

Background: The fully human monoclonal IgG1 antibody HuMax-CD20 targets a novel epitope of the CD20 molecule on B-cells. HuMax-CD20 stops growth of engrafted B-cell tumors in SCID mice more efficiently than Rituximab and i.v. infusion of HuMax-CD20 in cynomolgus monkeys leads to profound, long lasting, dose-dependent B-cell depletion.

Aim: The objective of the present trial was to establish the safety, efficacy and the pharmacokinetics of HuMax-CD20 in patients with chronic lymphocytic leukemia.

Methods: Data are presented from an open label, dose-escalation, multicenter phase I/II clinical trial. 3 cohorts of 3 (A), 3 (B) and an extended cohort of 27 (C) patients with relapsed or refractory chronic lymphocytic leukemia (B-CLL) received 4 weekly i.v. infusions of HuMax-CD20 and were followed for 12 months. The first infusion was 100 mg, 300 mg and 500 mg in cohort A, B and C and the following 3 infusions were of 500, 1000 and 2000 mg, respectively. The endpoints were B-cell depletion, adverse events, objective response according to the NCI working group guidelines for CLL, time to progression, duration of response, time to next anti-CLL treatment, and pharmacokinetics. Data are presented from group C.

Results: Median age was 63 years (range, 27–82); median time since diagnosis was 6.1 years (range, 1–14). Maximum tolerated dose was not reached and none of the patients developed human anti human antibodies. 13 of 27 patients (48%) reported infections and infestations most frequently CTC grade 1 or 2 events of nasopharyngitis, gastroenteritis, sinusitis and upper respiratory tract infection without causal relation to ofatumumab. All patients had pronounced reduction of the leukemic CD19+CD5+ cell counts. Objective response rate was 46% (12 of 26 evaluable patients) with 1 nPR and 11 PR. Median time to progression (TTP) was 133 days (95 % CI 105–158) in the full analysis population and 161 days (95 % CI 140–220) in the subgroup of responders. The duration of response was 112 days (95 %CI 100–133). Time to next anti-CLL therapy was 366 days (95 % CI 258-N.A.). AUC (median 344697 μg/mL*h; range 62709–1362933) correlated significantly to both TTP and time to next anti-CLL therapy to which parameters the clearance (Cl) of ofatumumab (median 10 mL/h; range 3–42) correlated inversely.

Correlation between survival endpoints and PK Time to progression Time to next anti-CLL therapy ρ p-value n ρ p-value n ρ: Spearman correlation coefficient, n: number of observations AUC,μg/mL*h 0.55 0.005 24 0.62 0.001 24 Cl, mL/h −0.59 0.003 23 −0.58 0.003 23

Conclusion: This analysis of data from the first study of ofatumumab in CLL patients provides an indication of clinical benefit associated with high exposure and low clearance.
Respiratory Syncytial Virus Infections (RVS) after Allogeneic Hematopoietic Stem Cell Transplantation (HSCT) in the Palivizumab (P) Era.
Authors: de Fontbrune Flore Sicre , Robin Marie , Scieux Catherine , de Latour Régis Peffault , Ferry Christèle , Rocha Vanderson , Boudjedir Karim , Devergie Agnès , Gluckman Eliane , Socié Gérard , Ribaud Patricia
Published: October 2019

Respiratory syncytial virus infections after HSCT can lead to severe respiratory failure and is associated with a fatal issue in a substantial number of patients (pts). P has been used to treat RSV infections in HSCT recipients, however its impact on outcome is not well established. From January 1999 to March 2006, all pts with RSV infections after HSCT at Saint Louis Hospital were retrospectively reviewed in order to determine cumulative incidence of RSV-related death, risk factors for 4-month transplant-related-mortality post RSV diagnosis (early TRM), and eventual impact of P. Forty pts with RSV infections were identified, RSV occurred in a median of 177 days (d) (range: −15 d to 987 d ) after HSCT in 3 to 65 year-old recipients (median age: 16 years). 30 pts had received a graft from an unrelated donor. Characteristics at diagnosis were: pneumonia in 16, bronchitis or bronchiolitis in 8 and upper respiratory disease in 16 pts; 16 pts had hypoxemia. Among the 40 pts, 18 received P at diagnosis (15mg/kg - intravenous course - 1 to 3 monthly injections). The groups of pts with or without P were significantly unbalanced for age, source of stem cells, neutrophil and lymphocyte counts, median time from to transplantation to diagnosis (younger age, more cord blood transplantations, lower neutrophil and lymphocyte counts, and shorter duration between transplantation and infection in the P group). Only one patient (treated with P) died from RSV pneumonia alone, giving a 2.6% cumulative incidence of RSV-related death (95% confidence interval (CI): 0–7.7). Seven other pts died during the 4 months following RSV diagnosis. All of them died from respiratory failure of multiple causes (RSV +: n = 4; RSV−: n = 3). Early TRM incidence was 22% (95%CI: 17–27) and 3-year overall survival was 59% (95%CI: 43–83). Risk factors for early TRM in a multiple Cox model were stem cell source (unrelated cord blood, hazard ratio (HR): 4.9, 95%CI: 1.1–22, p=0.039) and hypoxemia at RSV diagnosis (HR: 7.31, 95%CI: 1.8–49). As P was predominantly used in pts with poorer prognosis, we cannot easily test its impact on early TRM: 23% (95%CI: 0–50) and 9% (95%CI:0–22) with or without P, respectively. Nevertheless, pts who received an unrelated graft and who were receiving ≥ 0.5 mg/kg/d prednisolone equivalent at RSV diagnosis were at higher risk of early death (HR: 8.15, 95%CI: 1.56–42). Interestingly, cumulative incidence of death in this group did not change with or without P (37% versus 40%). In contrast, there was no death in pts grafted from a related donor if RSV was diagnosed after engraftment. Overall survival of HSCT pts with RSV infection appeared better than previously reported, and RSV was rarely the direct cause of death after transplantation. The impact of P remains to be defined.
Alemtuzumab but Not ATG Decreases Natural Killer (NK) Cell Activity towards Tumor Targets in the Early Phase after Allogeneic Stem Cell Transplantation.
Authors: Penack Olaf , Fischer Lars , Stroux Andrea , Gentilini Chiara , Nogai Axel , Muessig Arne , Ganepola Susanne , Thiel Eckhard , Uharek Lutz
Published: October 2019

Background: In vivo T cell depletion with ATG or Alemtuzumab is effective to reduce the incidence of graft-versus host disease (GVHD) caused by alloreactive T cells. However, there is also a potential impact of these substances on the function of natural killer (NK) cells who are the predominant cells in peripheral blood in the early phase after hematopoietic stem cell transplantation (HSCT) and mediate beneficial graft-versus-tumor activity. Using a novel flow cytometric assay, which detects the lytic granule membrane protein CD107a as a marker for NK cell degranulation, we investigated the effect of T cell depletion with ATG and Alemtuzumab on NK cell function in the early phase after HSCT.

Methods: PBMCs of 34 patients (pts) at day +30 after allogeneic HSCT and of 16 healthy donors were coincubated at 37°C for 3 h with the NK sensitive cell line HL60. In each tube, containing 400μl effector/target suspension (2x106 cells), 20μl of PE-Cy5 conjugated anti-CD107a monoclonal antibody was added prior to incubation. After the first 1 h 10μl of the secretion inhibitor 2 mM monensin was added. At the end of coincubation cells were stained with mAbs (CD56, CD3) for flow cytometry. The percentage of CD107a expressing NK cells was assessed and the absolute number of degranulating NK cells/μl was calculated.

Results: Treatment Characteristics: Fourteen pts received ATG, ten pts were treated with Alemtuzumab and ten patients did not receive T cell depletion. The source of donor was: MRD 12 and MUD 22. NK cell count: The median NK cell count was: 250/μl in healthy individuals, 250/μl in pts without T cell depletion, 400/μl in pts with ATG and 100/μl in pts receiving Alemtuzumab (p<0.0005; Kruskal-Wallis test). NK cell activity: The median percentage of degranulating NK cells was 5.4% in healthy donors, 4,4% without T cell depletion, 2,8% when ATG was used and 0,8% when Alemtuzumab was given (p<0.0005). The absolute number of CD107a+ NK cells in response to the standardized tumor targets accounted for 13,4/μl (median) in normal donors, 12,9/μl in pts without T cell depletion, 7,6/μl in pts with ATG and 0,9/μl in pts with Alemtuzumab (p=0.001). The percentage and absolute number of CD107a+ NK cells were not significantly different between patients receiving ATG and patients not receiving T cell depletion (p=NS).

Conclusion: With a new and feasible method we were able to quantify and characterize tumor reactive NK cells after HSCT. We found that NK cell mediated cytotoxicity towards tumor targets is influenced by the type of T cell depletion: The NK cell activity in patients receiving Alemtuzumab was considerably reduced whereas ATG had only moderate impact on the NK cell activity in the early phase after HSCT.
Rituximab Reduces the Incidence of Acute Graft Versus Host Disease in an Unselected Series of Twenty-One Patients Undergoing Allogeneic Stem Cell Transplantation at a Single Center.
Authors: Christopeit M. , Mueller L. , Grothe W. , Weber T. , Muetherig A. , Theurich S. , Behre G.
Published: October 2019

Recent evidence highlights an implication of B lymphocytes in the development and sustainment of chronic graft versus host disease (cGvH). The monoclonal anti-CD20-antibody rituximab has proven effective against steroid refractory cGvH. Data on the role of B lymphocytes in acute GvH (aGvH) is sparse. We report on a reduction of the incidence of aGvH in patients receiving rituximab as an element of conditioning as compared to patients conditioned without rituximab. All 6 patients receiving rituximab suffered from CD20 positive Non-Hodgkin-Lymphoma (NHL) and received cyclosporine and mycophenolate immunosuppression. Immunosuppression in the other 15 patients was achieved either with cyclosporine and mycophenolate (N=11) or cyclosporine and methotrexate (N=4). All patients except for one received dose reduced conditioning. Only patients beyond day 25 are included in the analysis; as of july 20th, 2006 median follow up is 151 days (range 27 to 322 days). Out of six patients receiving rituximab as an element of conditioning, one (16.7 %) developed moderate aGvH. Out of 15 patients who did not receive rituximab during conditioning, 10 (66.7 %) developed moderate to severe aGvH. Two patients developed severe acute GvH after early cessation of immunosuppression and/or donor lymphocyte infusion for progressive disease. If removed from this analysis, 8 out of 13 patients (61.5 %) developed aGvH after conditioning without rituximab. Taken together, these data present a trend that in patients conditioned with rituximab acute GvH occurs less frequently. These are amongst the first data indicating that rituximab might suppress the development of acute GvH. They are in accordance with work published before yet this is the first study systematically comparing two groups of patients. At present, no data exploring the influence of CD20 positive B lymphocytes or the effect of rituximab on graft-versus-tumor effects exist. It therefore remains to be investigated whether the suggested suppression of GvH by rituximab benefits overall survival by reducing transplantation related mortality or whether it is associated with a higher relapse rate due to an abrogated graft versus tumor effect. Furthermore, it remains to be elucidated whether the administration of rituximab before transplantation or during the restoration of the allogeneic immune system exerts prophylactic effects against cGvH and whether this influences both graft versus tumor effects and survival. Finally, we conclude that the addition of rituximab to the treatment of severe steroid refractory acute graft versus host reactions has to be tested in randomized trials.
Chemokine Profiling in Patients with Graft Versus Host Disease (GVHD) Following Allogeneic Blood Stem Cell Transplantation.
Authors: Ziegler Judy A. , Lokshin Anna , Lentzsch Suzanne , Agha Mounzer , Arnold Renate , Mapara Markus Y.
Published: October 2019

Development of GVHD following allogeneic blood stem cell or bone marrow transplantation (BMT) is dependent on the immunogenetic disparity between donor and host, subsequent alloactivation and the inflammatory reaction occurring secondary to conditioning regimen-induced tissue damage. The inflammatory response occurring early following conditioning therapy is thought to be particularly crucial for the subsequent development of GVHD as it occurs in response to tissue damage and is amplified by the alloresponse. Chemokines are predominantly small molecules (8–14 kd) that bind to a family of heterotrimeric G-protein-coupled receptors with a seven-transmembrane-spanning serpentine structure and play an important role in leukocyte trafficking and are thought to be involved in tissue infiltration during GVHD. Reports from several investigators using murine model have demonstrated the relevance of distinct chemokines in the pathogenesis of GVHD. Furthermore, we have recently demonstrated that conditioning intensity and genetic host factors influence the tissue and systemic expression of chemokines during the early course of GVHD in rodent BMT models. However, the pathobiologic relevance of chemokines expression at later time points during GVHD and in human GVHD is so far unclear.

Using luminex multiplex assays we studied the sera of patients, who had undergone peripheral blood stem transplantation from an allogeneic HLA matched sibling or matched unrelated donor in order to determine the chemokine profiles during GVHD. In addition to the most relevant cytokines/cytokine receptors (IL-1a, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IFN-a, IFN-g, GM-CSF, IL-12 (p40), IL-13, IL-15, IL17, TNF-alpha, TNF-RI, TNF-RII, DR5, FGF-b, VEGF, HGF, IL-1Ra, sIL-2R and sIL-6R), we analyzed the serum expression of CCL2, CCL3, CCL4, CCL5, CCL11, CXCL9, and CXCL10. Patients with acute GVHD of the gut (n=8), patients without GVHD (n = 4) and patients with chronic GVHD (n=4) were studied. Patients with acute/chronic GVHD had significantly elevated levels of CCL3 (90 pg/ml vs. 58 pg/ml, p= 0.004) and CCL11 (167pg/ml vs. 93pg/ml) compared to controls patients without GVHD. Patients with acute GVHD of the gut had also markedly elevated CXCL10, CCL4 levels although not reaching statistical significance. In addition, patients with acute or chronic GVHD were found to have significantly elevated serum levels of HGF, IL-1Ra, and TNF-RI.

In conclusion we are able to demonstrate that CCL3 appears to be the most prominent chemokine to be elevated in the serum of patients with a/cGVHD supporting results obtained in preclinical BMT models. Given the complex and frequently contradictory preclinical results, further studies are warranted to confirm our findings in larger cohorts of patients and to delineate whether targeting CCL3 or one of its receptor (CCR1 or CCR5) might prove beneficial for the treatment of GVHD.
Early Reconstitution of Natural Killer Cells Following Non-Myeloablative Hematopoietic Stem Cell Transplantation Predicts the Development of Acute Graft-Versus-Host Disease.
Authors: Boyiadzis Michael , Hakim Frances T. , Memon Sarfraz A. , Dean Robert , Fowler Daniel , Gress Ronald E. , Bishop Michael R.
Published: October 2019

Natural killer (NK) cells are among the first lymphocytes to recover following allogeneic hematopoietic stem cell transplantation (HSCT). Their levels may reflect both homeostatic and inflammatory processes. In this study we analyzed the early reconstitution of NK cells in fifty-one patients with hematologic malignancies after a reduced-intensity regimen consisting of fludarabine and cyclophosphamide and its relationship to the development of acute graft-versus-host disease (aGVHD) and cytomegalovirus (CMV) reactivation.

NK cells were severely depleted by the fludarabine-cyclophosphamide conditioning regimen. On the day of the transplant, the NK levels had been reduced in all patients to 0 cells/microliter. Subsequent NK cell recovery was rapid. By two weeks post-transplant NK cells had recovered to median levels comparable those in the original hosts (median= 119 cells/microliter; range, 0–816) and by four weeks increased to a median of 161 cell/microliter (range, 6–701; p< 0.0001) as compared to pre-transplant levels. By the second month post-transplant the median levels decreased and were maintained at pre-transplant levels.

CMV antigenemia developed in 23 of the 51 patients at a median of 30 days post transplant (range, 20–213 days). The median NK cell reconstitution at 2 weeks in the patients who subsequently developed CMV antigenemia was 158 cells/microliter (range, 2–816) and was higher than that among patients without CMV antigenemia (median 71 NK cells/microliter, range 10–228;Wilcoxon rank sum test p = 0.05).

Grade I to III aGVHD developed in 31 out of 51 patients (median time = 27 days; range, 11–92 days). The NK cell levels in the patients who developed aGVHD were significantly higher at two weeks post-transplant (median NK = 193 cells/microliter; range, 0–816) than those in the patients who did not (median NK = 66 cells/microliter; range 2–215) (p = 0.005 by Wilcoxon rank sum test). Furthermore, based on a Kaplan-Meier analysis of time to aGVHD as a function of the NK levels at two weeks post-transplant, patients who had NK cell counts greater than 195 cells/microliter at two weeks post transplant had a significantly higher probability of developing aGVHD (p = 0.0036 by log-rank test) Figure 1.

These results suggest that the early NK cell reconstitution may serve as a biomarker of transplant outcomes, especially in predicting the development of acute GVHD.

Figure Figure
Transplantation of Positive Selected Peripheral Stem Cells with Add-Back of T Cells from Unrelated Donors in Children: Favourable Survival and Low Incidence of GvHD.
Authors: Lang Peter J. , Greil Johann , Bader Peter , Mura Ulrike , Schumm Michael , Feuchtinger Tobias , Pfeiffer Matthias , Handgretinger Rupert
Published: October 2019

There is still a significant risk of GvHD in transplantation of peripheral stem cells from matched unrelated donors, despite the use of ATG and pharmacological prophylaxis. Especially in the case of young pediatric patients and adult donors, grafts with high numbers of stem cells but also with extreme amounts of T cells can be obtained. We present an update of a clinical study with T cell reduced grafts in 22 patients, which offer both large stem cell numbers and clearly defined amounts of T cells. Positive selection of peripheral stem cells was done with CD34 or CD133 coated magnetic microbeads and the CliniMACS™ device. Afterwards, an unselected aliquot containing 10x106/kg T cells was added to the purified stem cells and infused on day 0, resulting in a median T cell depletion of 2 log. The diagnoses were: acute lymphatic leukemia (n=9; CR1=4, CR2=5), NHL (n=1) acute (n=5; CR1=1, CR2=4) and chronic myeloic leukemias (n=3), MDS (n=3) and Wiskott-Aldrich (n=1). Median age was 10 years (0.5–18). The donors were matched for HLA-A/B (medium resolution typing) and DRB1/DQB1 (high resolution). A short course of MTX (2–3x10 mg/m2) and CSA (2–3 mg/kg, adjusted to blood levels) until day 100 were given. 21/22 patients had primary engraftment with a median time to ANC >500 of 18 days (12–60). No G-CSF was given. One patient rejected his graft and was successfully retransplanted from another MUD. Platelet recovery was fast (median time to reach independence from substitution: 20 days). Mean numbers of CD3+, CD3+CD4+ and CD3+CD8+ on day 180 (365) were 259/μl (868/μl), 100/μl (400/μl) and 146/μl (456/μl). 19/22 patients had GvHD grade 0–I (86%), 1 and 2 patients had grade II and III, (5%, 10%) respectively. Chronic GvHD occurred in 2 patients (10%). 15/22 patients are alive with a median follow up of 3 years (1.5–5). 3 year EFS was 64% (all patients), 50% (ALL/NHL) and 73% (myeloic leukemias/MDS). Relapse probability at 2 years was 25%, probability of TRM at 2 years was 15%.Causes of death were relapse (n=4) and infections (n=3). Thus, stable and favourable survival rates with a low incidence of GvHD were achieved. The method allows to administer clearly defined T cell numbers independent from the size of the grafts. This may be of advantage in particular in small children and if the donor does not accept a bone marrow harvest: all available stem cells can be infused without the limitation of unacceptable high T cell numbers.
Tacrolimus Concentrations in Hematopoetic Stem Cell Transplantation Patients Are Related to Cytochrome P450 3A5 Gene Polymorphisms.
Authors: Kobayashi Koichiro , Ogasawara Masahiro , Kikuchi Misato , Kimura Satoru , Miyazono Takayoshi , Nakata Masanobu , Imai Kiyotoshi , Hirano Teiichi , Kobayashi Naoki , Kiyama Yoshio , Tanaka Junji , Imamura Masahiro , Tanimoto Mitsune , Kasai Masaharu
Published: October 2019

Objective: Tacrolimus, a commonly used potent immunosuppressive drug in hematopoetic stem cell transplantation (SCT), has large individual variation in concentrations and a relatively narrow therapeutic range. It is metabolized mainly by cytochrome P450 (CYP) 3A which has genetic polymorphisms influencing enzymatic activities; CYP3A5*1 and *3 genotypes represent expressor and non-expressor, respectively. In the present study, we investigated whether the tacrolimus concentration per dose is associated with CYP3A5 gene polymorphisms.

Methods: 51 Japanese SCT patients (21 AML, 12 ALL, 9 CML, 3 NHL, 1 HL, 2 ATL, 1 MM, 2 AA), receiving tacrolimus were studied. DNA was extracted from peripheral blood samples. CYP3A5 genotype was analyzed by PCR-restriction fragment length polymorphism (RFLP) using specific primers and a restriction enzyme, SspI. Blood tacrolimus concentrations were measured twice a week by a commercially available EIA kit.

Results: The frequency of the CYP3A5 *1/*1, *1/*3 and *3/*3 genotype was 7.8, 31.4, and 60.8% of 51 patients, respectively, and the genotype distribution was not different significantly from that of 121 Japanese healthy volunteers (2.5, 38.8, and 58.8%, respectively). The frequency of CYP3A5 genotypes of white Americans is reported to be 2.2, 12.0 and 86.0%, respectively, indicating the ethnic difference of distribution. The dose-adjusted concentrations of tacrolimus 5 to 7 days following SCT were significantly higher in CYP3A5 *3/*3 patients than in CYP3A5 *1/*1 and *1/*3 patients (691.7±298.0 ng/ml per mg/kg vs. 451.8±153.8 ng/ml per mg/kg, P=0.0017). The dose needed to achieve the target concentration (10 to 15 ng/ml) between 40 to 60 days following SCT, when the blood concentrations of tacrolimus tended to be stable in most patients, is significantly lower in CYP3A5 *3/*3 patients than in CYP3A5 *1/*1 and *1/*3 patients (0.008±0.005 mg/kg vs.0.016±0.006 mg/kg, P<0.001). Above grade two adverse events related to tacrolimus were observed only in CYP3A5 *3/*3 patients (3 patients in grade 4 and 1 patient in grade2), suggesting the contribution of a higher concentration to renal toxicity by tacrolimus.

Conclusion: The tacrolimus concentration per dose in SCT patients is influenced by the CYP3A5 gene polymorphism. This is the first report showing the relationship between CYP3A5 gene polymorphism and the tacrolimus concentration in SCT patients to the best of our knowledge. Information of CYP3A5 genotype may be helpful to determine the initial tacrolimus dose adequate for each patient.
Evaluation of Pre-Transplantation T-Cell Receptor Excision Circles (TREC) in Reduced-Intensity Stem Cell Transplantation (RIST) Recipients.
Authors: Morita-Hoshi Yuriko , Heike Yuji , Yamagata Shizuka , Tajima Kinuko , Taniguchi Shuichi , Tobinai Kensei , Takaue Yoichi
Published: October 2019

〈Background and Methods〉: Previous studies have shown that the recipient’s thymic function before hematopoietic stem cell transplantation (HSCT) is critical for subsequent immune reconstitution and that the value of pre-transplantation T-cell receptor excision circles (TREC) is related to the incidence of graft-versus-host disease (GVHD), infection and survival. However, these observations have been mainly based on younger patients, who were eligible for myeloablative HSCT. In contrast, little is known about thymic function in elderly patients and its influence on the outcome of reduced-intensity stem cell transplantation (RIST). In this study, TREC values were measured in a total of 180 HSCT recipients who received myeloablative HSCT (n=60: mean age, 34 y) or RIST (n=120, 51 y). Quantification of thymic signal joint TREC was performed by real-time quantitative PCR and TREC values were calculated as TREC copies per 10 ng DNA obtained from peripheral blood mononuclear cells.

〈Results〉: The mean and median pre-transplantation TREC values were 3.9 and 1.3, respectively, and these showed a significant inverse correlation with patient age (p=0.0002). The mean TREC value was 3.0 in males and 5.1 in females (p=0.22), and was significantly lower in 19 patients who had received previous radiation therapy involving the mediastinum (0.96, p=0.003). The TREC value was 3.7 in 99 patients who had a disease duration of longer than 365 days, and 4.2 in 81 patients with a shorter duration (p=0.61). The TREC value was 5.5 for solid tumor (n=10), 5.0 for CML (n=10), 4.8 for AML (n=52), 3.7 for MDS (n=23), 4.0 for ALL (n=23), 2.8 for lymphoma (n=60) and 0.4 for myelofibrosis (n=2), with no difference between those in complete remission and others (both 3.9). The clinical outcome in relation to the TREC value was analyzed in 98 patients who received RIST from an HLA-identical donor. A comparison of 51 patients with lower pre-transplant TREC values (<1.3) and 47 patients with higher values (>1.3) disclosed that the incidence of acute GVHD was 64% vs 57% (p=0.52), that of chronic GVHD was 69% vs 77% (p=0.56), CMV reactivation was 72% vs 60% (p=0.20) and progression-free survival at day 100 was 72% vs 100% (p=0.0001).

〈Conclusion〉: Pre-transplant TREC values showed an inverse correlation with age, and were significantly lower among patients who received radiotherapy involving the thymus. Although these differences were not statistically significant, patients with lymphoid malignancy, patients with longer disease duration and male patients tended to have lower TREC values, while the disease status did not influence the TREC value. This study provides the baseline reference data regarding the thymic function in patients undergoing RIST, although further study is required to evaluate the significance of TREC on the clinical outcome of RIST.