bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison between Progenitor Cells from Mobilized Peripheral Blood (PB) in Healthy Donors and Non-Hodgkin’s Lymphoma (NHL) Patients.
Authors: Villaron Eva M. , Almeida Julia , Lopez-Holgado Natalia , Sanchez-Guijo Fermin M. , Alberca Mercedes , Blanco Belen , Sanchez-Abarca Luis I. , Lopez Olga , Perez-Simon Jose A. , Miguel Jesus F. San , del Cañizo Consuelo
Published: October 2019

INTRODUCTION: Peripheral blood stem cell (PBSC) mobilization is impaired in patients receiving chemotherapy but, as far as we know there is no data about the impact of chemotherapy on different PB progenitor cell subpopulations.

AIM: to ascertain whether or not immature or committed progenitor cell are affected by chemotherapy prior PBSC mobilization in NHL patients.

MATERIAL AND METHODS: a total of 27 PB samples from NHL patients and 36 PB samples from healthy donors were studied. Immunophenotypic analysis of CD34+ cell subpopulations was performed using the following four colour combinations of monoclonal antibodies (FITC/PE/PC5/APC): CD90/CD133/CD38/CD34 and CD71/CD13/CD45/CD34. In order to study committed progenitor cells “in vitro”, standard colony-forming assays were used and, in order to investigate the behaviour of the uncommitted progenitors Delta Assays of plastic adherent progenitor cells (PΔ) were performed.

RESULTS: The comparison between NHL patients and healthy donors is shown in Table 1. The relationship between data obtained by flow cytometry and cultures was statistically significant (p<0.05, r>0.568) for all the progenitors analysed.

Table 1: Results of Immunophenotypic and Functional Assays

LNH patients Healthy donors p Data expressed as median (range). 1. Percentage among CD34+ cells. 2. Number of CFU/10 5 planted cells. 3. Number of CFU/10 6 planted cells % CD34 0.16(0.04–3.65) 0.57(0.11–1.81) 0.013 Immunophenotypic Data Erithroid 1 0.05(0.01–0.60) 0.14(0.02–0.42) 0.098 Myelo–monocytic 1 0.11(0.02–2.41) 0.37(0.07–1.18) 0.014 Immature 1 0.01(0.00–0.63) 0.05(0.01–0.19) 0.014 CFU-GM 2 70(4–440) 90(0–904) 0.327 Clonogenic and Delta Assays data BFU-E 2 62(6–172) 85(0–240) 0.046 CFU-Mix 2 18(0–124) 42(0–140) 0.018 CFU Δ3 356(0–3509) 953(90–8320) 0.033

CONCLUSIONS: We can conclude that in NHL, mobilized committed and above all immature progenitors are impaired when compared with healthy subjects, both analysed by immunological and functional assays. Only granulomonocytic progenitors analysed by a functional approach seemed to be preserved.
Amifostine Can Reduce Oral Mucositis after High-Dose Melphalan Conditioning in Patients with Multiple Myeloma: A Retrospective Study.
Authors: Italiano Antoine , Chamorey Emmanuel , Foa Cyril , Peyrade Frédéric , Otto Josiane , Thyss Antoine
Published: October 2019

Autologous peripheral blood stem cell (PBSC)- supported high-dose melphalan (HDM) is now considered standard therapy in the treatment of multiple myeloma, at least for patients under 65 years. Oral mucositis is a frequent non-hematological complication which causes severe pain, interferes with patient nutrition and can lead to systemic infection. Amifostine (WR-2721; Ethyol), a phosphoaminothiol, is a prodrug that protects a broad range of normal tissues from the cytotoxic damage induced by anticancer agents. We retrospectively compared two groups of patients with stage II/III previously untreated multiple myeloma who received between April 96 and May 2004 an induction chemotherapy with 3 or 4 cycles of VAD (vincristine, adriamycin, dexamethasone) followed by HDM (200 mg/m²) and autologous PBSC transplantation. These two groups either received (group A, n = 10 ) or did not receive (group B, n= 32) amifostine (740 mg/m²) before HDM. The occurrence of grade 3/4 oral mucositis was significantly decreased in group A in comparison to group B (10% versus 53%, p =0.023) with no difference for the time to mucosal recovery. Supportive care differed between the two groups: only 2 patients (20%) needed opioid treatment in group A versus 22 patients (69%) in group B ( p=0.005) and 1 patient (10%) required parenteral nutrition in group A compared to 16 patients (50%) in group B (p= 0.015). The occurrence of severe infectious complications did not differ between the two groups (0% versus 12,5%, p= 0.56). Amifostine did not affect haematological recovery, the median time to granulocyte recovery to > 500/μl was similar in the two groups (8,4 days versus 9,9 days, p=0.22). Moreover, there is no statistically significant difference between the amifostine and control group for the disease response. The tolerability of amifostine was excellent and no adverse effects were reported. This study suggest that amifostine can reduce mucosal damage associated with high dose melphalan-based therapy, reducing, as a consequence, the necessity of nutrition and analgesic support, without compromising therapeutic benefit. Obviously, these interesting results have to be confirmed by larger randomised trials.
Quantitative Analysis of the Correlation of Human Polyomavirus and Hematuria in Patients after Stem Cell Transplantation.
Authors: Lu Yuhong , Radonic Aleksandar , Thulke Stefanie , Nitsche Andreas , Siegert Wolfgang
Published: October 2019

Objective: quantitative evaluation of viruria (DNA-uria) and hematuria in patients with SCT and analyze both relationship clinically.

Methods and Materials: two PCR assays for detecting viruses in clinical specimens are established: quantifying virus load by real-time PCR and distinguishing BKV and JCV by seminested PCR. Intensity of hematuria, that is the dominant sign of hemorrhagic cystitis, was assessed by urinestick and classified into 6 grades: negative (−), trace or 1–10 (+), 11–25 (++), 26–80 (+++), 81–200 (++++) erythrocytes in 1 milliliter urine, and macrohematuria (+++++). 531 urine specimens were collected from 41 patients with SCT. Data were analyzed through SPSS software package.

Results: Prevalence rate of polyoma viruria and hematuria following SCT were 80% and 73% in this study cohort and 45% and 20 % in urine specimens. No viral genomes were detected in DNA extracted from blood cells and plasma. BK, BK integrating with JC, JC and non-identified virus-variants, respectively, were shed in 15, 11, 3, 4 patients in urine and were detected in 99, 21, 32, 11 urine samples. Data of 25 patients with specimens collected before SCT showed that only 5 cases sheded polyoma virus in urine, comparing with 21 cases with polyoma viruria post-SCT, demonstrates significant difference (P<0.0001). Risk factors of polyoma viruria were not found. The median of maximal virus load was higher in male than female (2.1x109 vs. 9.1x106, P=0.031, Mann Whithney), and in patients with hemorrhagic cystitis than without (2.5 x1010 vs. 4.7x107, P=0.043). There were no documents of hematuria pre-SCT. Recipients with stem cells originating from bone marrow underwent more frequently hematuria than from peripheral blood cells. This is likely due to more patients with standard conditioning therapy in BMT group (90% vs. 57%, P=0.032). The application of anti-thymocyte glolubin impacted likely on incidence of hematuria (88% vs. 47%, P=0.008). This is identical with the results of the study reported by Rabenau HF. The significant statistic relationship between polyoma viruria and hematuria is not available by defining entities as positive and negative. Indeed BK-viruria prevalence in 35% urine specimens containing erythrocytes >= ++ and in 17% urine specimens with negative and only trace red blood cells, while hematuria with amount of erythrocytes >=++ presented in 21% BK-viruria, nevertheless in 9% urine specimens with negative or non-amplifying virus-variants, 7% JC-viruria and 6% BK integrating with JC-viruria (P=0.016). Time conformability was observed in onset of most viruria and hematuria within 2nd-3rd weeks post SCT.

Conclusions: the course of SCT exerts explicitly impact on the reactivation of polyomavirus in urinary tract and BK-virus attribute likely to the onset of severer hematuria. Co-factors and pathogenesis are needed to continue exploring.
Safety and Tolerability of Anti-Bacterial Prophylaxis during the Post-Engraftment Phase after Myeloablative Hematopoietic Stem Cell Transplantation.
Authors: Hayes-Lattin Brandon , Bubalo Joseph , Dunn Adam , Subbiah Nan , Allen Bryon , Balleisen Suzanne , Curtin Peter T. , Jacoby Carol , Leis Jose F. , Mauro Michael J. , Simic Aleksandra , Schubach Susan , Maziarz Richard T.
Published: October 2019

Background: Bacterial infections, including infections with Gram-negative organisms, frequently complicate the post-engraftment phase after myeloablative allogeneic hematopoietic stem cell transplantation. In this pilot study, we investigated the safety and tolerability of administering antibacterial prophylaxis from the time of engraftment (absolute neutrophil count of >1500/mm3 for 2 days) through day +100. Moxifloxacin was chosen for its broad spectrum of bacterial coverage and for its ability to target both bacterial topoisomerase IV and DNA gyrase, limiting the development of antimicrobial resistance.

Methods: Twenty-three patients with engraftment after allogeneic transplantation at OHSU received moxifloxacin 400 mg po daily upon discontinuation of any antibiotics used for the treatment of pre-enrollment bacterial infection. Rates of bacterial infection, bacteremia, and colonization were recorded. Comparisons were made with a cohort of 60 consecutive patients undergoing myeloablative transplantation at OHSU without similar antibacterial prophylaxis from 2001–2002.

Results: The median duration of moxifloxacin prophylaxis was 77 days (range 7–87). Five patients had dose interruptions due to nausea associated with graft-versus-host disease and two patients elected not to restart study medication. Only 3 patients had dose interruptions during empiric antibiotic therapy for possible bacterial infection. Overall, moxifloxacin was well tolerated in this patient population, with 5 episodes of grade 3/4 toxicity felt possibly related to study drug (2 orthostatic hypotension in one patient, 2 nausea, and 1 hypertension). Five patients had episodes of transaminitis which were unlikely to be due to study medication, and which responded to discontinuation of antifungal therapy (4) or to treatment of liver GVHD (1). No cardiac arrhythmias or tendonopathy were observed. Compared to historic controls, a marked decrease in post-engraftment phase bacterial infections was observed. The control cohort, 16 patients experienced 17 episodes of Gram-negative and 22 patients had 25 episodes of symptomatic Gram-positive infections during the post-engraftment phase without prophylaxis. In contrast with moxifloxacin prophylaxis, 2 patients experienced 3 Gram-negative infections (χ2, p=0.079), 1 patient had an anaerobic infection, and 7 patients had 9 symptomatic Gram-positive infections.

Conclusions: Prophylactic moxifloxacin dosed throughout the post-engraftment phase has been well tolerated, safe, and associated with a fewer bacterial infections compared to historic controls. The efficacy of this prophylaxis strategy will best be determined in a planned randomized, placebo-controlled trial.
Cidofovir for Adenovirus Infection after Allogenic Hematopoietic Stem Cell Transplantation: A Single-Center Experience of 16 Childrens.
Authors: Ouachée-Chardin Marie , Leruez-Ville Marianne , Rouzioux Christine , Blanche Stéphane
Published: October 2019

Adenovirus is an important cause of morbidity and mortality in pediatric hematopoietic stem cell transplantation (HSCT). We performed a retrospective analysis of 16 patients treated with cidofovir as first-line therapy for adenovirus, from one center. Transplants were performed for primary immunodeficiencies (n=9), other inherited diseases (n=5), acute leukemia (n=1) and Langerhans cell histiocytosis (n=1). Six patients received HSCT from matched sibling donor, 6 from unrelated donor and 4 from a family haploidentical donor. Bone marrow was T-cell depleted in 5 transplants. Nine patients received ATG or Campath as GVHD prophylaxis. Eight patients had a acute GVHD of grade II to IV and 4 received Campath as GVH treatment. Four patients had chronic GVHD. Adenovirus subgenus were A for 10 patients, C for 5 patients and one patient had a coinfection with A and C adenovirus. Symptoms were fever (n=11), diarrhae (n=14), pneumonitis (n=4), hepatitis (n=6) and hematuria (n=1).Eight patient had a viral co-infection. Real-time quantitative PCR was used for detection and monitoring of adenovirus. The median time of first detection was day+7 after HSCT. Four patients had a first detection before HSCT and for 3 of them cidofovir was started before HSCT. All the patients had adenovirus detected from blood with a median time of day+19 afterHSCT. The median number of positive sites before HSCT was 3. Cidofovir was started at a median of 28 days after HSCT. Dosage per infusion was 5mg/kg for 8 patients and 3mg/kg for 4 patients. Four patients received 3 or 5mg/kg.A median number of four infusion was given during a median of 32 days. Median follow-up was 13 months.

Of the 16 children, 14 were successfully treated with cidofovir and two failed therapy and died from disseminated adenovirus disease. One patient died from GVHD. In total, 6 patients had no toxicity, 5 developed a reversible tubulopathy. Of note, 3 children improved under treatment suggestinfg a possible nephritis induced by adenovirus. The discontinuation of the drug was not required in any patient. We conclude that cidofovir may be efficient for adenovirus infection after HSCT in pediatric patients but prospective controlled trials are warranted.
The Disease Status at Transplant Is the Major Factor Association with the Long Term Survival of HLA Haploidentical Peripheral Blood Stem Cells Transplantation.
Authors: Wu Binyi , Wu Lanxiao , Guo Kunyuan , Song Chaoyang , Yan Dingan , Xiao Lulu
Published: October 2019

Objective: To evaluate the factors effecting the long term survival of refractory leukemia patients who received the therapy of HLA haploidentical peripheral blood stem cells transplantation.

Methods: To analysis the factors effecting long term survival of refractory leukemia patients who underwent HLA haploidentical peripheral blood stem cells transplantation. The HLA mismatched locus between the donors and patients, the disease status of patients at transplant, the grafted mononucleaer cells number, and the occurrence of GVHD, the age of patients and other factors were considered and analyzed by data reduction SSP11. From July, 1998 to May, 2004, 30 Patients with refractory leukemia patients including 13 cases of acute non-lymphocytes leukemia, 10 cases of acute lymphocytes leukemia, 6 cases chronic myeloid leukemia and 1 cases of non-Hodgkin lymphoma underwent HLA haploidentical peripheral blood stem cells transplantations. The median age was 25 years old (3– 52 years old). Twelve patients received stem cells from parents, four from daughter and son, and the other from sibling donors. Twelve patients received three HLA locus mismatched stem cells, thirteen patients received two HLA mismatched donors stem cells, and five patients received one HLA mismatched donors stem cells. The conditioning regime consisted of fludara (25mg/m2 X5d), busufan (4mg/kg X4d) and cyclophosphamide ( 60mg/kg X2d). Median dose of rabbit anti-human lymphocyte globulin (5mg/kg X5d) was added in the in some patients. A mean of 6.0 x 108 /kg (3–9 x108 /kg) mono-nucleated cells was grafted. The mean CD34+ cells number was 5.5 x106 /Kg (3–6.5 x106/kg).

Results: Twenty-nine patients were successfully grafted and one failed to graft. The mean time of white cell count more than 1x109/L was 13 days (10–18 days) and 12 days (9–16) respectively. Severe acute graft versus host disease occurred in six patients, and four died. Seven patients suffered from intensive chronic graft versus host disease. Nine patients relapsed and died. The mean relapse time was 10 months (3 months to 24 months). Four patients died from intensive chronic graft versus host disease. Fourteen patients are still disease free survival with high karnofsky performs. The relapse of leukemia was the main cause of death. Five the patients less than 20 year’s old age are still disease free survival with high karnofsky performs scores.

Conclusion: HLA haploidentical peripheral blood stem cells transplantation may be an effect therapy for refractory leukemia. Although graft versus leukemia effect may be strong in HLA haploidentical blood stem cells transplantation, leukemia relapse is still the main cause to death. We suggest that for these patients with leukemia who can not find full matched donor perform related HLA mismatched peripheral blood stem cells transplantation as earlier as to get the better long term outcome.
Increased Risk of Non-Relapse Mortality within One Year after Autologous Stem Cell Transplant in Older Adults with Lymphoma.
Authors: Kalaycio Matt , Pohlman Brad , Sobecks Ronald , Andresen Steven , Summers Kristie , Bolwell Brian J. , Rybicki Lisa , Kuckowski Elizabeth
Published: October 2019

The risk of early mortality after high-dose chemotherapy followed by autologous stem cell transplant (ASCT) is generally less than 5%. However, this mortality rate is often determined by deaths that occur while hospitalized or within 60–100 days of ASCT. We queried our long-term follow-up database to determine non-relapse mortality (NRM) for patients with lymphoma within 1 year of ASCT. From 1/1/99 to 3/30/04 we treated 317 evaluable lymphoma patients with busulfan 14mg/kg, VP-16 60 mg/kg, and cyclophosphamide 120 mg/kg followed by ASCT. Of these 317 patients, 21 (6.6%) had NRM within 1 year of ASCT. There were no significant differences between the 296 patients who did not die of NRM and the 21 who did with regard to diagnosis of non-Hodgkin versus Hodgkin lymphoma, exposure to radiation therapy, exposure to rituximab, stage, and disease status at the time of ASCT. However, the median age of the 21 patients who died of NRM was 56 years compared to 49 years in the 296 who did not (p = 0.003). Of the 21 patients who died of NRM, 13 (62%) died of pulmonary complications. Adult respiratory distress syndrome (ARDS) was listed as the cause of death (COD) in 5 patients, but nearly all patients died at other institutions limiting our ability to confirm COD. Pneumonia was the COD in 5 patients and pulmonary toxicity was the COD in 2 patients. We routinely screen patients with pulmonary function tests including DLCO before and approximately 42 days after ASCT. There was no significant decrease in the median pre-ASCT and the post-ASCT DLCO in either the whole cohort of patients dying of NRM or the 13 patients dying of pulmonary complications. The 8 patients with NRM, but not pulmonary complications, died of various causes including cardiac tamponade, sepsis/multi-system organ failure, cirrhosis, renal failure, and secondary malignancy. The median time from ASCT to NRM was 146 days (range 45 to 287 days). Thus, lymphoma patients remain at risk for NRM for several months after ASCT. Older patients are at particular risk. The most common COD is pulmonary failure that cannot be predicted by screening pulmonary function tests. The cause of the pulmonary failure is uncertain. Busulfan may cause pulmonary toxicity, but the dose in this series of patients is relatively low and we have not seen similar pulmonary complications in 95 patients (median age 52 years) with myeloma treated with the combination of busulfan 16 mg/kg and cyclophosphamide 120mg/kg.
The Application of Sequential and Quantitative Analysis of Donor Chimerism in Adoptive Immunotherapy Following Allogeneic Stem Cell Transplantation.
Authors: Tang Xiaowen , Wu De Pei , Sun Aining , Zhu Ziling , Chang Huirong , Ruan Changgeng
Published: October 2019

Objective In order to investigate the value of sequential and quantitative analysis of chimerism to predict relapse or graft rejection after allogeneic stem cell transplantation, to determine the optional time point of adoptive immunotherapy and to estimate the efficacy of adoptive immunotherapy.

Methods 70 patients who received HLA compatible allo-HSCT were evaluated. Peripheral blood and bone marrow were collected before and after transplantation in different period. Serial and quantitative analysis of donor chimerism (DC) both prior to and following adoptive immunotherapy were performed by multiplex PCR amplification of STR markers (STR-PCR) and capillary electrophoresis with fluorescence detection.

Results 13 of 70 patients relapsed at median time of 6months post transplant (4/13 patients developed hematologic relapse and 9/13 patients appeared cytogenetic or molecular relapse respectively). 2 of 70 patients experienced graft rejection. Before the time of relapse or graft rejection, STR-PCR indicated the decreasing DC in all 15 patients, at levels ranging from 24.8%~86.2%. The declining value of donor chimerism (DC< 90%) was detected in four patients 26 days before relapse or graft rejection diagnosed clinically. Adoptive immunotherapy was used to treat 15 patients(9 CML, 5 AML, 1 MDS). A response to immunotherapy was achieved in 6 patients(6 CML) following withdrawal of CsA and in a further 4 patients(2 CML, 1 M2, 1 MDS) after additional donor lymphocyte infusion(DLI). In these patients, the value of DC increased with convertion to a predominant donor profile (>90%) or converted to stable full donor chimerism(FDC)(6 patients) or stable mixed chimerism(MC)(4 patients) shortly after immunotherapy. All these 10 patients who responsed to immunotherapy developed acute or chronic GVHD. While in the patients without response, the level of DC decreased persistently or declined after transient increasing. Three patients without response received second DLI but remain failed to therapy.

Conclution The results demonstrate that the decreasing value of DC can identify the patients who have high risk of relapse or graft failure and can be used to guide adoptive immunotherapy implementation at early stage. The sequential and quantitative monitoring of DC has been shown to be a valuable tool to determine the optional time point of immunotherapy and to early evaluate the efficacy of treatment. Furthermore, it can present a rational basis for treatment intensification in patients who did not respond to first-line DLI treatment
Reduced Intensity Conditioning Regimen for Allografting Following Cytoreductive Autografting in Relapsed/Resistant Hodgkin’s Lymphoma.
Authors: Carella Michele , Beltrami Germana , Corsetti Maria T. , Greco Mario M. , Scalzulli Potito , Cascavilla Nicola , Musto Pellegrino , La Sala Antonio , Dell’Olio Matteo , Carella A.M.
Published: October 2019

Background: We postulated that it is possibile to safely extend the benefit of allografting to relapsed or resistant Hodgkin’s Lymphoma (HL) by combining cytoreduction with high-dose therapy/autologous stem cell transplant (HDT/ASCT) and graft versus HL effect mediated through transplanted donor immune cells with nonmyeloablative allografting (RICT).

Methods: Twenty-two patients, 32 years of age (range, 16–39) with heavily pretreated disease were given HDT/ASCT. At a median of 92 days after HDT/ASCT the patients received fludarabine 30 mg/m2/day x 3 days and cyclophosphamide 300 mg/m2/day x 3 days and non-T depleted donor peripheral blood stem cells from HLA-identical siblings. Postgrafting immunosuppression consisted of methotrexate and cyclosporine. Cyclosporine was withdrawn early in patients with mixed chimerism or disease progression. Donor lymphocyte infusions were given to 12 patients with stable mixed chimerism and/or progressive disease.

Results: Seventeen patients (77%) achieved full chimerism and 5 patients mixed chimerism. Twelve patients (55%) achieved CR after tandem transplant. The first signs of responsiveness in CR patients began at day ≥60 and the maximal response was achieved on day ≥80. The remitters patients achieved full chimerism and developed acute/chronic GVHD before regression of the disease; moreover, 5 patients achieved CR after DLI. At last follow-up (July 31, 2004), 9 out of 12 CR patients are alive at a median of 1844 days (range, 1092 to 2045). Subsequently, 5 patients relapsed and are now alive on therapy and four patients maintain CR after a median follow-up of 1874 days (range, 1092–2045). Acute GVHD grade ≥ II occurred in 9 patients (40%); chronic GVHD developed in 5 patients (23%), four of them with extensive chronic GVHD requiring further immunosuppressive therapy. Two patients died of GVHD/infections, 1 patient died of extensive chronic GVHD and 7 patients of progressive disease.

Conclusions: This tandem auto-allotransplant protocol demonstrated to be highly effective also in advanced resistant Hodgkin’s Lymphoma patients. The exploitation of graft-versus-lymphoma effect is a promising finding
Fludarabine and Busulfan (FluBuP Regimen) as Myeloablative Conditioning Regimen for Allogeneic PBSCT in 71 Leukemic Patients (A Unicenteric Study).
Authors: Iravani Masoud , Tavakoli Maryam , Shamshiri Ahmad Reza , Evazi Mohammad Reza , Mousavi Asadollah , Bahar Babak , Ghavamzadeh Ardeshir
Published: October 2019

Following 3 other centers in USA, Canada and Germany, we are evaluating fludarabine (40 mg/m2 on days −6 to −2) and busulfan (4 mg/kg/day on days −5 to −2) as a new conditioning regimen for allogeneic peripheral blood stem cell transplantation in leukemic patients with matched related donors. Seventy one patients were enrolled, 18 with high and 53 with standard risk (18 ALL, 35 AML, 16 CML and 2 MDS; F=29 M=42). The median patient age was 23.7 years(range, 2.4– 46.7). Cyclosporine was used as a prophylactic agent for GVHD (3mg/kg IV till +4, 10 mg/kg oral from day +5). The median follow-up was 269 days (range, 50–459 days). 91.5% and 15.5% developed mucositis and hepatic toxicity respectively which resolved with conservative therapy. There was no cardiac toxicity (except one patient with mild pericardial effusion and another with tachycardia). The median of highest serum creatinin level during hospitalization were 1.6 mg/dl (range, 0.8–3.7; 24.3% with Cr>2) and serum cyclosporine level, at the same time, was 246 ng/ml (range, 9–814). 7% experienced hemorrhagic cystitis (infection was ruled out) and 36.6% experienced moderate to severe headache. 38% and 14.1% of the patients showed grade 1, 2 and grade 3 acute GVHD respectively. Grade 4 acute GVHD was found in one patient. 50% and 6% showed limited and extensive chronic GVHD. 27% of patients became CMV+ (min +17, max +69). The median time for neutrophil and platelet recovery were 10 (min 0, max +26) and 12 (min 0, max +30) days. In day +38, 86.7% of the patients had 90% or more, mononuclear chimerism (with STR-PCR technique; median, 97%; range, 25–100). 5 ALL and 8 AML patients relapsed (18.3% of all patients) and 6 (8.45%) died after relapse. Nonrelapse mortality was 13% (9 patients; acute GVHD grade IV=1, CMV infection and GVHD=2, CMV infection=2, pneumonia=2, infection=2). With a median follow up of 9 months (range, 1.6–15.3 months), the probability of overall survival and disease free survival were 79.68% and 81.26% respectively. It could be beneficial to use fludarabine versus cyclophosphamide in standard conditioning regimen for leukemic patients because of reduced toxicity, low incidence of acute GVHD and facilitated donor engraftment.
Purified T Depleted Peripheral Blood and Bone Marrow CD34 Transplantation from Haploidentical Mother to Child with Thalassemia.
Authors: Sodani Pietro , Andreani Marco , Polchi Paola , Gaziev Javid , Centis Filippo , Giardini Claudio , Nesci Sonia , Cicconi Silvia , Spitaleri Luca , Cassiani Flaminia , Lucarelli Guido
Published: October 2019

Approximately 60% of thalassemic patients can not apply to “gene terapy today” which the insertion of one allogenic HLA identical stem cell into the empty bone marrow as the vector of the normal gene for beta globin chain synthesis. We studied the use of the haploidentical mother as the donor of hematopoietic stem cells assuming that the immuno-tollerance estabilished during the pregnancy will help to bypass the HLA disparity and allow the hemopoietic allogeneic reconstitution in the thalassemic recipient of the transplant.We have employed a new preparative regimen for the transplant in nine thalassemic children aged 3 to 8 years ( median age 5 years ) using T cell depleted peripheral blood stem cell (PBSCTs) plus bone marrow (BM) stem cells.. All patients received hydroxyurea (OHU) 60 mg/kg and azathioprine 3 mg/kg from day −59 untill day−11, fludarabine (FLU) 30 mg/m 2 from day −17 to day −11, busulphan (BU) 14 mg/kg starting on day −10, and cyclophosphamide(CY) 200mg/kg, Thiotepa 10 mg/kg and ATG Sangstat 2.5 mg/kg, followed by a CD34 + t cell depleted (CliniMacs sistem), granulocyte colony stimulating factor (G-csf) mobilized PBSC from their HLA haploidentical mother. The purity of CD34+ cells after MACS sorting was 98–99%, the average number of transplanted CD34+ cells was 15, 4 x 10 6/kg and the average number of infused T lymphocytes from BM was 1,8 x 10 5/Kg.The patients received cyclosporin after transplant for graft versus host disease( GVHD) prophilaxis. Four patients rejected the transplant and are alive with thalassemia: one patient received a different dose of CD3 without cyclosporine after transplant, two patients received a lower dose of CD34+, in the fourth patient the donor has been the haploidentical father instead than the mother. One of the nine patients, after the failure of the transplant from the mother, received a second transplant using purified CD34+ cells from the father, using the same preparative regimen and achieved a complete hematopoietic reconstitution. Six patients are alive disease free with a median follow up of 19 months (range 7–30). None of the six patients showed AGVHR. This preliminary study suggest that the transplantation of megadose of haploidentical CD34+ cell from the mother is a realistic therapeutic option for those thalassemic patients whithout genotipically or phenotipically HLA identical donor.
Stem Cell Transplantation from Mismatched Related Donors for Paediatric Haematological Disorders : The Hospital for Sick Children, Toronto, Experience.
Authors: Nicolin Gary , Das Prabodh , McDougall Elizabeth , Gassas Adam , Sung Lillian , Lau Wendy , Saunders E. Fred , Doyle John , Calderwood Stan
Published: October 2019

Between June 1994 and December 2003, 31 patients with haematological disorders presenting at the Hospital for Sick Children, Toronto, Canada, received a stem cell transplant (SCT) from a mismatched related donor. Diagnoses included ALL (15), AML (10), CML (1), JMML (1), SAA (2), de novo MDS (1) and Fanconi anaemia (1). Patients were aged between 0.9 to 17.8 years (mean 8.5 years). There were 20 males and 11 females. HLA typing included A, B and DRB1 on all patients (Class 1 alleles by low resolution molecular technique and Class 2 by high resolution technique). 22 patients had 1 major mismatch and 9 had 2–3 major mismatches. The source of stem cells was bone marrow in 15 patients (mean total nucleated cell dose 4.44 x 108/kg, range 1.89–6.63), peripheral blood (PBSC) in 15 patients (mean CD34 dose 11.80 x 106/kg, range 3.19–42.18) and one patient received both BM and PBSC. Of those who received PBSC, 9 had CD34 selection, 3 had CD34 selection and T-cell depletion, and 3 received an unmodified product. Conditioning regimens included cyclophosphamide and total body irradiation (TBI) (15), etoposide and TBI (6), busulfan and cyclophosphamide (6), etoposide and cyclophosphamide (1), fludarabine, cyclophosphamide and TBI (1), fludarabine and cyclophosphamide (1) and fludarabine, melphalan and thiotepa (1). Graft versus host disease (GvHD) prophylaxis included combinations of cyclosporine A, methotrexate, anti-thymocyte globulin and methylprednisolone. There was one primary graft failure and 3 secondary graft failures. Mean time to neutrophil engraftment was 17 days (range 9–36). The incidence of acute and chronic GvHD was 48% and 51% respectively. Of the 31 patients, 15 are alive (12 in complete remission (one of these with a secondary brain tumour) and 3 with relapsed disease) and 16 are dead (3 of relapsed progressive disease, 12 from transplant related mortality (TRM - 9 of these were in CR) and in one the cause is not known). Three year actuarial OS and EFS for the entire group were 50±9% and 45±9%, OS and EFS for those matched at 5/6 loci were 56±11% and 44±11% and OS and EFS for 3/6 and 4/6 matched were 38±16% and 36±16%. The mean follow-up time for survivors is 5.6 years (range 11.6 months to 9.7 years). In this series, there was no significant difference in survival when comparing BM vs. PBSC harvests; in one vs. 2–3 mismatch donors; and in the presence vs. absence of acute or chronic GvHD. These results confirm the feasibility and efficacy of using mismatched related donors for paediatric haematological disorders. Steps should be taken to decrease the TRM to achieve a better survival rate.
Comparison of Conditioning Regimens with or without Total Body Irradiation in Bone Marrow Transplantation from an Alternative Donor.
Authors: Yabe Hiromasa , Kato Shunichi , Yabe Miharu , Kudo Kazuko , Yagasaki Hiroshi , Horibe Keizo , Yamamoto Masuji , Ohara Akira , Kuroki Fumiko , Tsukimoto Ichiro , Kojima Seiji
Published: October 2019

An optimum conditioning regimen is indispensable in improving outcomes for patients with acquired aplastic anemia (AA) who receive bone marrow transplantation (BMT) from an alternative donor. Although high-dose total body irradiation (TBI) is effective in enhancing engraftment, intensive conditioning regimens are associated with increased regimen-related toxicity, which results in a decreased success rate. Currently, two different approaches are primarily used; a conditioning regimen including low-dose TBI and a fludarabin-based regimen without TBI. We attempt to clarify the efficacy and regimen-related toxicity of these two approaches. Ten patients (7 males, 3 females; median age 12y; range 9–17y) underwent an alternative donor transplant following a conditioning regimen with low-dose TBI (TBI group); patients received cyclophosphamide (50 mg/kg/d on days −5 to −2), antithymocyte globulin (Thymoglobulin, 2.5 mg/kg/d on days −5 to −2) and TBI (2.5 Gy x 2 on day −1). The donor was unrelated in 9 patients (3 were serologically HLA A or B antigen mismatched; 3 were serologically HLA DR antigen mismatched, 1 was both serologically and genotypically DR antigen mismatched; 1 was genotypically DRB1 mismatched; 1 was a genotypically complete match) and was a family member in 1 patient (serologically HLA A-mismatched father). Twelve patients (6 males, 6 females; median age, 11 y; range 2–20 y) received a conditioning regimen of fludarabine (25 mg/kg/d on days −5 to −2), cyclophosphamide (750 mg/m 2 /d on days −5 to −2), thymoglobulin (1.5 mg/kg/d on days −5 to −2) and thoracoabdominal irradiation (TAI, 3Gy x 1 on day −1) (TAI group). The donor was unrelated in 11 patients (8 were genotypically complete matches, and 3 were genotypically DRB1 antigen mismatched) and a family member in 1 patient (3 antigen-mismatched mother). The source of stem cells was unmanipulated bone marrow. Graft versus host disease (GVHD) prophylaxis consisted of tacrolimus (0.02 mg/kg from day −1) and short-term methotrexate. Rejection was seen in one patient in each group. The doses of infused nucleated marrow cells were less than 2.0 x 10 8 /kg in both patients, and both patients then received a second transplant and achieved full donor chimerism. Acute GVHD grade II-IV developed in 2/10 evaluable patients in the TBI group and in 3/12 patients in the TAI group. Chronic GVHD developed in 5/9 evaluable patients in TBI group and 3/10 evaluable patients in TAI group. Median follow up was 9 months (4–27 months) in the TBI group and 21 months (3–33 months) in the TAI group. Both conditioning regimens were well tolerated and effective even in HLA-mismatched unrelated donor transplantation. Long-term follow up is warranted in order to compare the late sequelae in both regimens.
Allogeneic Stem Cell Transplantation from Unrelated Donors after Reduced Intensity Conditioning in Patients with MDS/sAML. A Report from the Chronic Leukemia Working Party (CLWP) of the EBMT.
Authors: Kroeger Nicolaus , Brand Ronald , Martino Rodrigo , Guardiola Philippe , van Biezen Anja , Zander Axel , Niederwieser Dietger , De Witte Theo
Published: October 2019

We analysed the results of 67 patients with MDS/sAML who were transplanted with allogeneic stem cell transplantation from unrelated donors after a reduced intenisity conditioning and reported to the EBMT. The median age was 52 years (range 17–70 years) and stem cell source was bone marrow (n = 30) or peripheral blood progenitor cells (n = 33).. The graft was HLA matched in 57 patients while 8 patients received SCT from HLA-mismatched donor. The MDS classification was as follows: RA/RARS: n=8, RAEB/CMML: n = 14, RAEB-t/sAML: n = 22. The conditioning regimen consisted of fludarabine/busulfan (n=15), fludarabine/melphalan (n=6), fludarabine and TBI (n=8) or fludarabine and others (n=36)At time of transplantation only 12 (18%) were in first complete remission. The Kaplan-Meier estimates of the probability of 2 years overall and disease free survival were 33 % (95% CI: 21–45 %) and 24 % (95% CI: 12–36 %), respectively. The probability of relapse at two years was 58 % (95% CI: 40–76 %) and of one year treatment-related mortality 37 % (95% CI %: 23–51 %).

In an univariate analysis assessing source of stem cells, age, disease type, T-cell depletion, and HLA-matching no factor was significant for OS, EFS, TRM and Relapse. Allogeneic stem cell transplantation after a reduced intensified conditioning followed by unrelated SCT seems to be a feasible approach in those patients who were no candidates for a standard conditioning but is associated with a considerable number of relapses.
Addition of Purine Analogues to Induction/Consolidation Regimen Does Not Impair Peripheral Blood Stem Cell Mobilization and Bone Marrow Harvest for Autotransplantation in Acute Myeloid Leukemia.
Authors: Holowiecki Jerzy , Grosicki Sebastian , Hellmann Andrzej , Skotnicki Aleksander , Jedrzejczak Wieslaw W. , Dmoszynska Anna , Marianska Bozena , Robak Tadeusz , Giebel Sebastian , Wojnar Jerzy , Krawczyk-Kulis Malgorzata , Wojciechowska Maria
Published: October 2019

In the previous study by the Polish Adult Leukemia Group (PALG 1999 Study) we demonstrated that addition of cladribine to standard daunorubicine + cytarabine (DA-7) induction potentiates antileukemic activity of the regimen in acute myeloid leukemia (AML). However, there is a concern that the treatment with purine analogues may affect a successful collection of hematopoietic cells for transplantation (autoHCT).

The goal of this study was to compare the efficacy of peripheral blood and bone marrow hematopoietic CD34+ cell harvest in patients who both in induction and consolidation were treated with or without purine analogues and who were intended for autoHCT.

Sixty-seven AML patients, aged 41 (17–58) years, were included in this study; 33 patients received cladribine-containing regimen (DAC-7), one patient - fludarabine-containing regimen (DAF-7), 33 patients received standard treatment (DA-7, HAM, HD cytarabine). In the DAC-7 treated patients cladribine was also given as a adjunct to second HD cytarabine consolidation. AutoHCT using bone marrow (autoBMT) or peripheral blood (autoBCT) as a source of hematopoietic cells was performed in first complete remission after completion of consolidation therapy. An additional course of AraC 2x2g/m2 on days 1, 3, 5 + G-CSF 10 mg/kg since day 7 was used as mobilization in case of autoBCT.

The number of collected CD34+cells/kg was similar for patients pre-traeted with purine analogues and those not receiving purine analogues: 2.55 (0,79-9,25) x106/kg vs 2.5 (1,41–23,5) x106/kg (p=NS) for peripheral blood and 1.62 (0,32–3,01) x106/kg vs 1.55 (0,5–2,45) x106/kg (p=NS) for bone marrow, respectively. In 90% and 95% of patients for both subgroups sufficient number of hematopoietic cells for transplantation could not be collected (p=NS). The proportion of unsuccessful bone marrow harvest was significantly lower for patients pre-treated purine analogues with purine analogues compared to those not receiving purine analogues (90% vs 56%, p=0,04), wheras not difference was found with respect to peripheral blood cell collection (95% vs 62%, p=NS). All patients who received autoHCT engrafted. The time to neutrophil and platelet recovery was similar for both study subgroups.

We conclude that treatment with purine analogues in course of induction/consolidation therapy does not impair the hematopoietic cell harvest and does not decrease a chance to perform autoBCT or autoBMT in AML patients.
Mel-140 and TBI vs Mel-200 Prior to Autologous Peripheral Blood Stem Cell Transplantation for Multiple Myeloma - a Single Institution Experience.
Authors: Janakiraman Nalini , Chamarthy UshaSree , Acharya Radhika , Donthireddy Vijaya , Kamboj Ginny , Oja-Tebbe Nancy , Divine George
Published: October 2019

Between 1991–98, twenty-nine patients with myeloma received Mel-140 and TBI (960 cGy) followed by Cytoxan mobilized autologous PBSCT. Between 1998 and April, 2004, 41 patients received Mel-200 alone followed by autologous PBSCT. The two groups were comparable regarding age, gender, stage, number of prior regimens and ß2 microglobulin at diagnosis. However, the Mel-200 group had greater proportion of patients with IgG myeloma (78% vs 38%) and shorter time from diagnosis to transplant (301 days vs 581 days). Results: Neutrophil engraftment was similar in the two groups. However, platelet engraftment, transfusion requirements, length of stay and number of days on antibiotics were worse in the Mel-TBI group. The complete and very good partial responses were high in both groups Mel TBI 58.6% vs Mel-200 56% (p=0.81). Median overall survival (OS) was 3.5 years and 3.8 years for Mel TBI and Mel-200 groups, respectively (p=0.75). Event-free survival (EFS) was 1.9 years and 2.3 years, respectively (p=0.88). Survival after relapse post-PBSCT was short, median 1.1 and 1.8 years, respectively. We present here the results of two groups of myeloma patients treated consecutively in a single institution. The Mel-200 patients had a higher proportion of IgG myeloma and were transplanted earlier in the phase of their disease. As expected, regimen-related toxicities were higher in the Mel-TBI group. The response rates, OS, and EFS were similar. These results are in keeping with published reports of prospective randomized studies. Mel-200 is now the standard conditioning regimen for autologous transplant for myeloma.
Generation of Novel, High Titre, Multi-Cistronic Retroviral Vectors Which Simultaneously Express Individual Proteins in Different Subcellular Locations of Hematopoietic Cells.
Authors: Milsom Michael D. , Gavin Catherine , Baldwin Rebecca , Margison Geoff , Fairbairn Leslie J.
Published: October 2019

As gene therapy strategies become ever more sophisticated, there is a need for vectors which efficiently deliver and express more than one gene product to target cells e.g. to enable in vivo selection; to correct multiple genetic loci; to provide a suicide mechanism for safety purposes. Employing alternate codon usage, we have synthesised a novel version of the FMDV 2A self-processing moiety which encodes the same primary amino-acid sequence as the wild type protein (and which possesses equivalent cleavage activity to the wild type protein). By including this synthetic FMDV 2A sequence in retroviral vectors which also contain the wild type sequence, we are able to facilitate the efficient co-translational separation of 3 different proteins encoded by a single transcript. This vector configuration also permits the inclusion of an IRES sequence and thus expression of a fourth protein. Using this technology we have created two tetracistronic retroviral vectors which express eGFP; O6-methylguanine DNA-methyltransferase and truncated human nerve growth factor receptor courtesy of 2A mediated cleavage, and a mitochondrially targeted dsRED via an IRES. Stable amphotropic producer lines containing these vectors generated supernatant which contained a high titre of retroviral particles (>106 IU/ml) which was sufficient for the transduction of K562 human erythroleukemic cells. Analysis of transduced cells confirmed that all four proteins were expressed individually and segragated to their correct subcellular locations. Furthermore, limiting dilution assay revealed that proteins expressed courtesy of the 2A sequences were still generated at high levels, even when cells were infected with a low MOI of virus. We propose that this technology will allow the creation of more sophisticated vectors for use in gene therapy of hematopoietic and other cells.
VEGF Specific siRNAs Act Synergistically with Bortezomib and Steroids in Inhibiting VEGF Gene Expression, Proliferation and Induction of Apoptosis in Multiple Myeloma Cells.
Authors: Koldehoff Michael , Steckel Nina K. , Trenschel Rudolf , Beelen Dietrich W. , Elmaagacli Ahmet H.
Published: October 2019

Multiple myeloma (MM) is a clonal B-cell malignancy characterized by the accumulation of malignant plasma cells within the bone marrow (BM). Vascular endothelial growth factor (VEGF), a glycoprotein produced by normal and neoplastic cells is an important regulator of physiological and pathological angiogenesis. MM cells secrete VEGF, which promotes production of cytokines in bone marrow stromal cells, as well as migration and proliferation of the tumor cells. Inhibition of VEGF activity or disabling the function of its receptors has been shown to inhibit both tumor growth and spread of metastases in a variety of animal tumor models. RNA interference (RNAi) is rapidly being established as a post-transcriptional gene silencing method and holds promise to specifically inhibit gene expression in mammals. Another novel class of antitumor agents is based on the inhibition of the ubiquitin-proteosomal system which represents the major nonlysosomal pathway through which intracellular proteins are degraded in eukaryotic cells. Bortezomib, a reversible proteosome inhibitor, shows remarkable anticancer activity in various malignant cell types, including MM cells that are resistant to conventional therapies. We studied the effect of transfection with small interfering RNA (siRNA) targeting VEGF in MM cells in terms of proliferation, induction of apoptosis, and cell differentiation. Further, we evaluated if the effects of post-transcriptional gene silencing by VEGF specific siRNA can be augmented by bortezomib and/or steroids in the cell line OPM-2. A mean reduction of VEGF gene expression to 38% as determined by real-time PCR was observed with 0.8 ug VEGF siRNA in OPM-2 cells compared to controls (controls were set up to 100%). Simultaneous administration of bortezomib and siRNA was able to reduce VEGF gene expression down to 23% compared to VEGF siRNA alone demonstrating a synergistic effect of combined bortezomib and siRNA treatment. We found a 2.5-fold increase in induced apoptosis in OPM-2 cells subsequent to VEGF siRNA administration but we saw no additional stimulation of apoptosis after combination of VEGF siRNA with bortezomib and/or steroids. Proliferation in OPM-2 cells was strongly inhibited (about 91%) following combination treatment as opposed to only 62% after administration of VEGF siRNA alone. The transfection of VEGF siRNA in OPM-2 cells had no influence on the expression levels of differentiation markers such as CD38, CD138, CD19, CD34, CD45, and CD7AAD. Our findings suggest that synergistic effects of VEGF siRNA with bortezomib and dexamethason may offer new therapeutic options in the treatment of MM.