bluuebunny/crossref_metadata_2025 huggingface dataset:
The Incidence and Impact of Thrombocytopenia in Myelodysplastic Syndrome (MDS).
Authors: Kantarjian Hagop M. , Giles Francis , List Alan F. , Lyons Roger , Price Sherry , Deuson Robert , Leveque Joseph
Published: October 2019

Background: MDS is a heterogeneous stem cell malignancy characterized by ineffective hematopoiesis and increased risk of transformation to acute myeloid leukemia. Thrombocytopenia and platelet dysfunction are recognized to contribute to hemorrhagic complications, however, reliable data regarding the frequency and consequences of thrombocytopenia in MDS are lacking.

Objective: To assess thrombocytopenia in MDS based on literature and chart review.

Methods: MDS literature (January 1980 to November 2005) was identified using MEDLINE, EMBASE, and Cochrane databases, society websites, and hand searches of bibliographies. In addition, a retrospective review of all patients referred to the MD Anderson Cancer Center (MDACC) with MDS since 1980 was performed.

Results: A total of 85 references were identified describing clinical consequences of thrombocytopenia, n=16; efficacy and safety of MDS therapies, n=60; guidelines, n=9. Because of variable definitions of thrombocytopenia, a platelet count <100x109/L was applied as the threshold point in the literature analysis. The prevalence of thrombocytopenia in MDS ranged from 40% to 65%. Baseline thrombocytopenia rates were reported in 19/60 (32%) references; the median frequency of thrombocytopenia prior to MDS therapy was 65% (range, 23% to 93%). The MDACC database review identified 1605/2410 (67%) patients with thrombocytopenia (<100x109/L) at referral. Severe thrombocytopenia (<20x109/L) was observed in 425 (18%) patients. Using the International Prognostic Scoring System (IPSS), 399/2410 (17%) patients had intermediate-2 or high risk disease; 83% and 25% of these patients had platelet counts of <100x109/L and <20x109/L, respectively. Corresponding event frequencies decreased to 21% and 3% in patients with low or intermediate-1 risk MDS (264/2410, 11%). The reported incidence of MDS treatment-related thrombocytopenia was >50% in studies involving lenalidomide, azacitidine, or a combination of idarubicin, cytarabine and topotecan. Treatment-related thrombocytopenia was also observed in studies involving tipifarnib, linomide, decitabine, all-trans retinoic acid, and sirolimus. The reported incidence of hemorrhagic complications ranged from 3% to 53%, and the frequency of hemorrhagic deaths ranged from 14% to 24%. At MDACC, 460 patients had a coded cause of death: hemorrhage as a cause of death, 20%; hemorrhage as the only cause of death, 10%.

Conclusions: Thrombocytopenia is common in MDS with a higher prevalence in higher risk IPSS categories. Many approved and investigational MDS therapies cause or exacerbate pre-existing thrombocytopenia. The incidence of severe bleeding in MDS was higher than reported in current guidelines or reviews.
The Novel, Irreversible Proteasome Inhibitor PR-171 Demonstrates Potent Anti-Tumor Activity in Pre-Clinical Models of Multiple Myeloma, and Overcomes Bortezomib Resistance.
Authors: Kuhn Deborah J. , Chen Qing , Voorhees Peter M. , Strader John S. , Shenk Kevin D. , Sun Congcong M. , Demo Susan D. , Bennett Mark K. , van Leeuwen Fred W. , Orlowski Robert Z.
Published: October 2019

Introduction: The ubiquitin-proteasome pathway has been validated as a therapeutic target with the approval of the small molecule proteasome inhibitor, bortezomib (VELCADE®), in multiple myeloma and non-Hodgkin lymphoma. However, the overall response rate of patients with multiple myeloma in phase III clinical trials was 43%, underscoring the need for a next generation of inhibitors with the potential for greater efficacy.

Methods: PR-171 is a novel, tetrapeptide epoxomicin-related inhibitor that binds the proteasome irreversibly, and our objectives were to evaluate its activity and mechanism of action in pre-clinical models of multiple myeloma.

Results: PR-171 potently bound and inhibited the chymotrypsin-like subunit of the proteasome in vitro, in cellulo, and in vivo at low concentrations. At higher concentrations, however, unlike bortezomib, which targeted the chymotrypsin-like and peptidyl-glutamyl peptide hydrolyzing activities in vivo, PR-171 also displayed significant inhibition of the trypsin-like and the peptidyl-glutamyl peptide hydrolyzing activities. PR-171-induced proteasome inhibition was associated with accumulation of polyubiquitinated substrates and pro-apoptotic Bax. Brief pulse PR-171 exposure, which simulates the in vivo pharmacokinetics of bortezomib, led to PR-171-mediated inhibition of cellular proliferation linked to induction of caspase-3-dependent apoptosis through both intrinsic (caspase-9) and extrinsic (caspase-8-dependent) pathways. Pretreatment with caspase-3, -8, and -9 inhibitors rescued the anti-proliferative effect of PR-171. Furthermore, pulse PR-171 treatment activated c-Jun-N-terminal kinase, a key-signaling molecule in proteasome inhibitor-induced apoptosis, and cleavage of poly-ADP-ribose polymerase, while abrogation of c-Jun-N-terminal kinase signaling with a dominant-negative c-Jun inhibited PR-171-induced effects. PR-171 displayed enhanced anti-proliferative activity compared to bortezomib in multiple myeloma cell lines and freshly isolated patient-derived CD138+ plasma cells, associated with enhanced phosphorylation of c-Jun-N-terminal kinase and capase-3, -8, and -9 activation. Lastly, PR-171 was a potent inhibitor of proliferation in a multiple myeloma cell line model resistant to bortezomib and in isolates from two patients, one with primary and the other with acquired bortezomib-resistance.

Conclusions: These data indicate that PR-171 has enhanced activity against preclinical models of multiple myeloma, perhaps owing to its irreversible binding and subunit specificity, and provide a rationale for its translation into the clinic.
Candida tropicalis Arthritis of the Knee in a Patient with Acute Monocytic Leukemia.
Authors: He Jingsong , Ye Xiujin , Zheng Weiyan , Wu Wenjun , Lin Maofang , Huang He
Published: October 2019

The patients with malignant hematopathy are the high-risk group of invasive fungal infections (IFI). Candidemia and IFI caused by Candidas in patients with a hematological malignancy are common but Candida arthritis is rare. Here, we reported a case of Candida tropicalis arthritis of the knee that occurred with acute monocytic leukemia during the recovery period of post chemotherapy myelosuppression and agranulocytosis.

Case report: A 45-year-old Chinese woman was diagnosed with acute monocytic leukemia with normal caryotype in november 2004. Complete remission was achieved upon completion of 2 courses of induction chemotherapy. Treatment was administered via a Hickman catheter in the left elbow. Two days after phase V consolidation chemotherapy, the patient experienced high fever accompanied by the development of arthritis in the right knee. The diagnosis of Candida tropicalis arthritis of the knee was confirmed by the appearance of Candida tropicalis isolated from the synovial fluid, but no leukemic cells and acid-fast bacilli were found. According to the susceptibility test in vitro, itraconazole and amphotericin B injection were used sequentially for therapy for 4–5 weeks, which effectively inhibited bacterial growth. However, the arthritis relapsed after 4–6 weeks of drug withdrawal. The arthritis was fully resolved after 8 weeks of therapy with fluconazol injection at a dose of 400mg/d and douching articular cavity with amphotericin B once a week. And then sequential therapy with oral fluconazole was commenced. There was no any adverse effect occurrence during the course of treatment.

Discussion: Although Candida arthritis in patient with a hematological malignancy is rare, it still occurred in patient with hypoimmunity. Early diagnosis is difficult due to no distinctive clinical manifestation and hysteresis of pathogenic organism detection. It was found that the knee was the sole joint affected according to the review of fourteen other reports of Candida arthritis in patients with a hematological malignancy but the reason remains unclear. Fungal arthritis must be taken into consideration when dealing with patients with immune deficiency accompanied by arthritis, especially gonarthritis. We emphasize that the most important factors for the successful treatment of fungal arthritis are identified diagnosis and adequate dosage through out the course of treatment.
Preclinical Research on Idiotype Vaccine Against B Cell Lymphoma.
Authors: Wang Fuxu , Zhao Bing , Pan Ling , Zhang Xuejun , Luo Jianmin , Dong Zuoren
Published: October 2019

The idiotype (Id) of immunoglobulin expressed by B cell lymphoma can serve as the only widely accepted tumor associated antigen. But the Id vaccines have failed to elicit anti-tumor immunity for its weak immunogenic. Monocyte chemoattractant protein-3 (MCP3) can recruit various subsets of immune cells, such as DCs, which would uptake and properly process and present Id, activating both arms of the immune system, humoral and cellular. So the Id-MCP3 fusion proteins are potential vaccines for immunotherapy of B cell lymphoma. In this study, two vaccine candidates were constructed by fusing allogeneous MCP3 to the amino-(MCP3-scFv) or carboxyl-(scFv-MCP3) terminus of the A20 (BABL/c murine B-lymphocyte) Id scFv with a flexible polypeptide spacer encoding NDAQAPKS to prevent dissociation and keep their respective natural construction and function. And VH and VL domains were linked with a current linker encoding (Gly4Ser)3. Firstly, the cDNAs of Ig VH and Ig VL were amplified by RT-PCR from A20 mRNA, and then assembled into scFv by recombinant PCR method. Secondly the fusion genes of scFv/MCP3 were formed using the same method. After sequencing, MCP3/scFv fusion genes were cloned into pET-39b vector. Lastly MCP3/scFv fusion proteins were expressed in E.coli BL21. And the fusion protein is about 62 kD. We found that, under the same condition, MCP3-scFv fusion protein was expressed successfully and accounted for 40% of the total protein of the bacteria but not scFv-MCP3. Our result indicated that fusing MCP3 to carboxyl-terminus of scFv protein may have cytotoxicity to the host cells or maybe not stable inside the host cells. Next we will determine the activity of the fusion protein MCP3-scFv with cell-chemotatic-experiment in vitro and bearing-tumor mice experiment in vivo. Once the results would suggest that there may be an anti-tumor effect, we can make individual vaccines to lead to a better survival.
Increased Fatty Acid Synthase Is a Therapeutic Target in Multiple Myeloma.
Authors: Wang Weiqin , Zhao Xiaoying , Wang Haiyan , Xu Gengbo , Zhang Xiaohong
Published: October 2019

Multiple myeloma is a malignancy arising from postgerminal nature B cells. The estimated incidence of multiple myeloma accounts for 10% of hematological malignancies and 1% of all malignancies. Despite therapeutic advance, the disease is essentially incurable. Fatty acid synthase (FAS), the principle enzyme in the endogenous synthsis of fatty acid, is highly expressed in a wide variety of human malignancies and their presursor lesions, including carcinoma of the colon, prostate, ovary, endometrium and breast. One therapeutic target that has not previously been considered for multiple myeloma treatment is the pathway of the endogenous synthesis of fatty acid. For this purpose, we first evaluated a series of human multiple myeloma cases, bone marrow samples for FAS expression by immunohistochemistry, then established FAS mRNA and protein expression in multiple myeloma cell lines and human multiple myeloma plasma cells by RT-PCR and immunoblot analysis. Our results showed that 19 of the 27(70.37%) MM cases were scored as FAS positive by immunohistochemistry, whereas 8 of the 27 (29.64%) were scored negative. FAS mRNA was highly expressed in U266, RPMI8226 cells and 22 of the 27 MM patients, but wasn’t detected in healthy donor PBMNCs by RT-PCR. Immunoblot analysis found that FAS protein was detected in 16 of the 27 patients(59.25%) and two above myeloma cell lines but not detected in healthy donor PBMNCs. Encouraged by these findings that human multiple myeloma bone marrow samples and cell lines overexpress FAS, we treated human multiple myeloma cell lines (U266, RPMI8226) in vitro with cerulenin, a native inhibitor of FAS. Cells proliferation was measured by methyl thiazolyl tetrazolium(MTT) analysis and apoptosis was evaluated by flow cytometry. Treat with 5ug/ml—640ug/ml of cerelenin for 24h, the proliferation of U266 cells was obviously inhibited with dosage related effect. The inhibition rate of human skin fibroblast cells are all lower than 30%. When U266 cells are treated for 12h with 20ug/ml of cerulenin, the early stage apoptosis rate revealed by Annxin V/PI using FCM are 56,9%. Our findings suggest that FAS may be an effective target for pharmacological therapy in human multiple myeloma.
In Vivo Radiaprotective Effects of Growth Hormone.
Authors: Chen Benny J. , Deoliveira Divino , Chao Nelson J.
Published: October 2019

Ionizing irradiation can cause bone marrow failure and death. Very few effective therapeutic agents are currently available for this condition. It was previously published that human recombinant growth hormone promotes hematopoietic and immune reconstitution after stem cell transplantation. In this study, we investigated the effects of growth hormone on radiation-induced injury. In a sublethally irradiated model (BALB/c mice, 5 Gy), the recovery of platelets in peripheral blood was significantly accelerated after treatment with human recombinant growth hormone at a dose of 20 μg, i.p., once a day, for 35 days starting within one hour after irradiation. Similar trends were also observed in total white blood cells and all lymphocyte subsets tested (T, B, NK cells). These data suggest that growth hormone can promote hematopoietic and immune recovery after irradiation. We then tested whether growth hormone can rescue animals from lethal irradiation. BALB/c mice were irradiated with 7.5 Gy and treated with growth hormone using the same regimen as that used in the sublethal model. As demonstrated in the figure, 5 out of 10 mice in the growth hormone treated group survived more than 80 days after irradiation, whereas 9 out of 10 mice died by day 25 in the saline control group. Growth hormone was still effective when tested using higher dose of radiation (8.5 Gy). These findings demonstrate that human recombinant growth hormone has significant radioprotective effects even when given after total body irradiation.

Figure Figure
Mechanism for the Immuno-Suppresion Activity of Pentostatin: Selective Apoptosis of Naive T Cell Subsets.
Authors: Smith Melody M. , Powers William , Giver Cindy R. , Waller Ned , Flowers Christopher
Published: October 2019

Background: Nucleoside analogs have potent anti-tumor activity, especially against lymphoid malignancies, with immuno-suppression being a significant toxicity. We have previously described the immunosuppressive effects of fludarabine on murine T-cells, and found selective cytotoxicity against naïve subsets that reduced their allo-reactivity in a GvHD model of murine BMT (Giver et al 2003 BBMT 9:616). In this study we tested the cytotoxicity of fludarabine and pentostatin against human and murine T-cell subsets to determine their immune-suppressive activity, mechanism of action, and their potential utility as agents to decrease the allo-reactivity of allogeneic donor lymphocyte infusions.

Methods: Peripheral blood mononuclear cell samples from normal donors were incubated ex vivo ex vivo treatment with Flu at doses of 5, 50, and 250 mcg/ml or pentostatin at doses of 1, 10, and 100 mcg/ml at 0, 4, 24, and 48-hour time points following 1 or 24 hour drug exposure. All samples that obtained dCF also received 25 mcg/ml of deoxyadenosine (dAdo), which enables dCF to mediate cytotoxicity against lymphocytes. Control samples for fludarabine and pentostatin treatment were untreated or treated with dAdo only, respectively. The total number of viable cells and the fractions of the CD4 and CD8 memory and naive T cell subsets that survived nucleoside exposure were determined by multi-parameter flow cytometry following staining with antibodies to CD3, CD4, CD8, CD45RA, CD6L, Annexin, and 7-AAD.

Results: The decrease in the viability of the human naïve CD4 T cells, whether treated with fludarabine or pentostatin, was rapid and most notably observed after the 24-hour time point. On the other hand, the human memory CD4 T cells displayed a more gradual decline in percent viability in both treatment with fludarabine and pentostatin. Furthermore, the potency of pentostatin was exhibited in that it achieved a comparable effect to fludarabine while at substantially lower doses. Data obtained from the murine model also demonstrated a relative a relative sensitivity of the CD4 naïve T cells as oppesed to the CD4 memory to purine analog exposure.

Conclusions: Future work will determine the optimal dosage and period of incubation for pentostatin and fludarabine that can utilized to induce immunosuppresion ex vivo in donor lymphocytes.

SELECTIVE DEPLETION OF NAÏVE CD4+ T-CELLS FOLLOWING IN VITRO EXPOSURE TO PURINE ANALOGS SELECTIVE DEPLETION OF NAÏVE CD4+ T-CELLS FOLLOWING IN VITRO EXPOSURE TO PURINE ANALOGS
Pre-Transplant Biomarkers Predict GVHD and Treatment-Related Mortality in Patients Treated with Extracorporeal Photopheresis Prior to Allogeneic Bone Marrow Transplantation.
Authors: O’Brien Peter J. , Bolwell Brian , Abhyankar Sunil , Shaughnessy Paul , van Besien Koen , Williamson Paul , Campbell Kim A.
Published: October 2019

Accurate prediction of the onset and severity of GvHD and treatment-related mortality (TRM) can lead to improved outcomes through optimized donor selection and GvHD prophylaxis. Current assessments are typically made after bone marrow transplantation (BMT), and often require inherently unreliable qualitative measures. The persistence of host antigen presenting cells (APC), particularly dendritic cells (DC) has been implicated in the onset of GvHD, and a recent clinical trial sought to confirm previous reports that extracorporeal photopheresis (ECP) can prevent GvHD after allogeneic BMT. We examined blood from a subset of 32 ECP-treated patients to identify peripheral blood cell surface markers that might predict GvHD and TRM. Samples were drawn immediately prior to ECP (at baseline) and immediately after ECP, but prior to myeloablative conditioning, then assayed by flow cytometry. Cytometry data were grouped and modeled to assess their predictive accuracy alone or in combination with clinical laboratory values. Logistic regression showed that markers for specific DC subsets present prior to myeloablation were the best predictors of outcomes. The likelihood of grade II-IV GvHD (aGvHD) increases when baseline lin− HLA-DR+ CD11c+ myeloid cells make up a smaller proportion of circulating mononuclear cells with a predictive accuracy, reflected in the area under the receiver-operator curve (ROC) of 0.83. The best predictive model for TRM was a lower absolute abundance of circulating lin− HLA-DR+ CD123+ plasmacytoid DC at baseline (ROC=0.86). No additional predictive power arose with respect to aGvHD or TRM after including laboratory values. However, models that combined certain clinical laboratory results and demographic factors also predicted these clinical outcomes, and offer a possible alternative to complex cytometric assays. The best such model included baseline measurements of BUN/Creatinine ratio, serum albumin, and the match/relatedness of the graft donor and recipient (ROC=0.82, n=59) for TRM, while baseline neutrophil counts were most predictive of aGvHD (ROC=0.69, n=60). In summary, we have identified biomarkers including host DC subsets, readily measured clinical tests, and demographic factors that, at least in patients receiving ECP, are present before conditioning and can predict outcomes. Although this study does not address the impact of ECP on antigen presenting cells directly, the DC profiles described here highlight the potential role of ECP in modulating DC, and thereby decreasing the risks associated with BMT. These results encourage larger studies with non-ECP control populations to directly address this issue.
Multiple Myeloma Patients Treated with Allogeneic Stem Cell Transplantation Show an Increased Frequency of Bone Marrow-Residing CD4+Foxp3+ T Regulatory Cells.
Authors: Atanackovic Djordje , Cao Yanran , Faltz Christiane , Bartels Katrin , Wolschke Christine , Brummendorf Tim H. , Zander Axel R. , Bokemeyer Carsten , Kroeger Nicolaus
Published: October 2019

BACKGROUND: Immunosuppressive CD4+Foxp3+ T regulatory cells (Treg) play a vital role in immune regulation. Thus, Treg contribute to the prevention of autoimmune disease and graft-versus-host reactions following allogeneic stem cell transplantation (alloSCT) but also to the inhibition of effective anti-tumor T cell responses. It has previously been suggested that the frequency of Treg is increased in the peripheral blood of patients with multiple myeloma (MM). However, little is known about the presence of Treg in the bone marrow and it is unclear whether allogeneic stem cell transplantation might deplete Treg from this immune compartment.

METHODS: In the present study, we analyzed percentages of CD4+Foxp3+ Treg as well as Treg expression of CD45RA and CCR7 in the bone marrow (BM) and in the peripheral blood of MM patients who had received alloSCT (N=42), in newly diagnosed MM patients (N=18), and in healthy controls (N=15) using flow cytometry. In addition, we performed inhibition assays in order to test the functional relevance of peripheral and BM-residing Treg.

RESULTS: While newly diagnosed MM patients and healthy controls showed no significant difference in the proportions of CD4+Foxp3+ Treg in the bone marrow, percentages of BM-residing CD4+Foxp3+ T regulatory cells were markedly higher (p<0.001 and p<0.01) in patients post alloSCT (3.3±0.3%) than in normal BM (1.0±0.3%) or in BM of untreated MM patients (1.8±0.4%). In both groups of patients (p<0.05) as well as in the healthy controls (p<0.001) percentages of Treg were higher in the peripheral blood than in the bone marrow. While there were no differences regarding the percentages of peripheral Treg between the remaining groups, patients post alloSCT had higher percentages of peripheral Treg than newly diagnosed patients (5.6±0.8 vs. 3.2±0.7%, p<0.05). More than 90% of these donor-derived peripheral and BM-residing Treg expressed a memory T cell phenotype, being negative for CD45RA and CCR7. Importantly, peripheral as well as BM-residing Treg of patients post alloSCT were capable of inhibiting the proliferation of autologous non-Treg CD4+ T cells.

CONCLUSION: Our study demonstrates for the first time an increased frequency of immunosuppressive Treg in the bone marrow of MM patients. Remarkably, in our patients these memory-type Treg were all donor-derived and led to an efficient replenishment of Treg in the periphery. These Treg might be necessary for the prevention of graft-versus-host disease in the transplanted MM patients, however, they might also contribute to the failure of an effective graft-versus-myeloma effect in the majority of the patients.
Maternal Haplotype at Time of Banking Is an Effective Strategy to Guarantee the Identification and Traceability of Cord Blood Units.
Authors: Bergamaschi Paola , Perotti Cesare , Viarengo Gianluca , Fante Claudia Del , Parisi Cristina , Marchesi Andrea , Bellotti Laura , Galiazzo Valentina , Pasi Annamaria , Martinetti Miryam , Salvaneschi Laura
Published: October 2019

Stem cell transplantation from unrelated cord blood (CB) donors is nowadays a standard practice for treatment of both malignant and non-malignant haematological disorders. The degree of HLA disparities, the cell dose and the prompt availability of the CB unit strongly influence the donor choice. According to FACT Netcord standards, HLA A, B and DRB1 shall be determined on a pre-cryopreservation sample from each CB. Once a CB unit is identified for potential use, a sample is tested to verify HLA type and confirmation of maternal haplotype is provided. The recipient’s HLA typing and its matching to the donor are confirmed as well prior to CB release for transplant. Quality of the stem cell product represents a basic assumption for the final success of transplant and an essential requirement for CB Banks (CBB). Each CBB shall establish a program of quality assurance that includes identification and traceability. The protection of confidentiality of mother and infant donors and the maintenance of linkage of a particular CB to its mother/neonate pair is of the highest priority. For this purpose each CB is assigned a unique numeric identifier that accompanies the unit in all steps of management and by which it is related to its maternal/infant data and reference samples. At the Pavia CBB, all CB donations are routinely typed for both HLA class I and class II by molecular techniques (low resolution for A and B loci and high resolution for DRB1). The mother haplotype is also assessed by molecular methods at time of banking. Therefore each CB/mother pair is checked before inclusion in the inventory. We retrospectively evaluate the impact of such a strategy on ensuring both identification and linkage of our CB repository. We review the data referring to the CBs stored in our Bank, all typed by LR-DNA for class I loci and by LR/HR-DNA for class II loci. Our inventory consists of 1987 units available for donor selection. In 2 pairs (0.1%) the mother’s typing was found fully disparate as respect to the supposed corresponding CB/neonate. The identification cannot be confirmed leading to discard of the unit. We also review the data pertinent to the 45 units released for transplant by our CBB: in all cases the cord’s confirmatory typing was coherent to the previous typing thus confirming the identity of the product ready for shipment. In our hands, the practice of confirming maternal haplotype at time of banking provides the final evidence of correct assignment and identification before inclusion in the inventory, ensuring the highest quality of the product supplied. This strategy also optimizes the time for the search requests management contributing to prompt availability of the graft.
Large Scale Expansion of Activated T Cells: Defining Procedures and Cryopreservation Methods for Clinical Use as Adoptive Cellular Therapy Post-Transplant.
Authors: Wu Jia-Yan , Webber Susan M. , Hill John M. , Ernstoff Marc , Szczepiorkowski Zbigniew , Meehan Kenneth R.
Published: October 2019

We previously demonstrated a laboratory model of ex vivo expansion of mobilized peripheral blood mononuclear cells (PBMNC) that acquire the CD8+CD56+ NKT cell phenotype and aggressively destroy targets through the NKG2D receptor (Cytotherapy.2006;8:141–8). We developed a large scale culture method for clinical use that allows growth, expansion and cryopreservation of these expanded cells for infusion post-transplant. PBMNC were collected by large volume leukapheresis from healthy donors (n=3). Unselected MNC cells (2.5 × 106/ml) were cultured in Life Cell Bags (Nexell Therapeutics) with serum-free AIM V medium (Invitrogen), IL-2 (Chiron, 1000 IU/ml)and OKT3 (Ortho Biotech, 500 ng/ml) at 37° C and 5% CO2. Six bags were arranged from each collection. Fresh cytokines and medium were added on Days 3 and 5, and the cells were harvested on Day 7. The viability of the ex vivo expanded cell populations was 95% +/− 1.8%. Total MNC cell numbers expanded from 2.5 × 106/ml to 6.4 × 106/ml (2.5 +/− 0.5% fold increase). Phenotypic analyses showed CD3 cells increased from 55% (Day 0) to 94% (Day 7; +/−1.42%); the CD4 cells increased from 30% to 58% (+/− 1.48%); the CD8 cells increased from 11% to 48% (+/−3.98%); the CD56 cells increased from 17.2% to 47.8% (+/− 6.2). Cytotoxicity of Day 7 expanded cells was markedly increased at 74% (+/−7.1%) (Targets: K562 cells; E:T ratio 100:1) compared with 33% (+/− 5.4%) at baseline. We also tested if the ex vivo expanded cells could be cryopreserved for future clinical use. After 7 days in liquid nitrogen, cells were promptly thawed, washed and analyzed. The viability was 91% (+/− 2.1%) and the cytotoxicity against K562 cells was only slightly decreased (65%, +/− 5.4%), when compared to non-cryopreserved expanded cells (p = 0.18). The phenotype was not affected by cryopreservation and thawing. This large scale culture method results in growth and expansion of aggressive effector cells that are cytotoxic against tumor cells. The majority of the expanded cells are CD3+ T cells (equally distributed between CD4+ and CD8+ T cells). NK cells (CD56+) also readily expanded. After cryopreservation of the ex vivo expanded cells and a short-term storage, viability and phenotype of the cells did not change. There is minimal decrease in cytotoxicity that is not statistically significant. This large scale expansion model will be used in clinical trials at our institution to expand cells for adoptive cellular therapy following autologous PBSC transplantation in patients with a hematologic malignancy.
Clinical Efficacy and Feasibility of an Intensive Preparative Regimen with Busulfan, Cyclophosphamide, and Total Body Irradiation Followed by Allogeneic Stem Cell Transplantation for Patients with Myeloid Malignancies.
Authors: Yano Shingo , Asai Osamu , Dobashi Nobuaki , Osawa Hiroshi , Takei Yutaka , Yahagi Yuichi , Takahara Shinobu , Ogasawara Yoji , Otsubo Hiroko , Yamaguchi Yuko , Minami Jiro , Maruyama Dai , Kobayashi Tatsunosuke , Hoshi Yasutaka , Usui Noriko
Published: October 2019

Purpose: High-intensity conditioning regimens before allogeneic stem cell transplantation (all-SCT) could reduce relapse but may increase nonrelapse mortality. We have investigated the efficacy and feasibility of an intensive preparative regimen of busulfan (BU), cyclophosphamide (CY), and total body irradiation (TBI) for patients with myeloid malignancies.

Patients and methods: From November 1990 to February 2005, 80 patients (acute myeloid leukemia (AML) 33, chronic myeloid leukemia (CML) 29, myelodysplastic syndrome (MDS) 18) treated with BU (8 mg/kg), CY (120 mg/kg), and TBI (10 Gy) followed by related (n=46) or unrelated (n=34) allo-SCT. Thirty-eight consecutive patients with AML in 1st remission, CML in chronic phase, and MDS (refractory anemia) defined as a standard risk group, and 42 patients with advanced AML, CML, and MDS (more than 5% marrow blasts) defined as a poor risk group.

RESULTS: With a median follow-up was 6.9 years, the 7-year actuarial overall survival (OS) rate was 58% (standard risk 70% vs poor risk 47%, p=0.0076), and the probability of 7-year relapse rate was 18% (standard risk 17% vs poor risk 18%, p=0.4777). Nonrelapse mortality was 29% (standard risk 26%, poor risk 31%). Multivariate analysis indicated that age older than 40 years (p=0.018) and received allo-SCT before 1998 (p=0.006) had independent predictive value for nonrelapse mortality.

CONCLUSION: The BU- CY- TBI regimen resulted in decreased relapse, especially in poor risk patients. The results of this study suggested that the BU- CY- TBI is an effective and feasible preparative regimen for allo-SCT in patients with myeloid malignancies.
Analysis of Donor Age and Absolute Lymphocyte Count Recovery as Prognostic Indicators of Survival after Allogeneic Blood or Marrow Transplantation.
Authors: Senfield Jeffrey J. , Hahn Theresa , Padmanabhan Swami , McCarthy Philip L. , Battiwalla Minoo
Published: October 2019

Absolute lymphocyte count (ALC) recovery ≥500 cells/mL at day 15 or 30 post-autologous (autoBMT) transplantation has been reported in many studies as an independent prognostic indicator of overall survival (OS) and progression free survival (PFS) in non-Hodgkin lymphoma (NHL), Hodgkin Disease (HD), multiple myeloma (MM) and breast cancer patients. ALC recovery has also been reported in the allogeneic blood and marrow transplant (alloBMT) setting as an important factor in predicting risk of relapse of acute lymphocytic leukemia (ALL) and acute myelogenous leukemia (AML). It has been shown in previous reports that donor age is critically important for OS after alloBMT. In addition, increased donor age correlates with a reduction of common lymphoid progenitor (CLP) differentiation potential which led us to hypothesize that donor age might predict ALC recovery time in related and unrelated alloBMT recipients. We reviewed 76 related and 36 unrelated HLA-matched consecutive patients with hematological malignancies who underwent first alloBMT at Roswell Park Cancer Institute between 4/20/2000 and 3/17/2006. The patient population included 53 males and 59 females with a median (range) age of 44 (4–68) years. Conditioning regimens were FluMel (n=33), CyTBI (n=32), BuCy (n=19), FluCy (n=13), VCT (n=11), and other (n=4). The stem cell source was bone marrow (n=34), peripheral blood (n=77) or both (n=1). Hematologic engraftment was defined as absolute neutrophil count (ANC) reaching 500 cells/ml or more for 3 consecutive days. ALC at day 30 and time to ALC ≥ 500 cells/mL were analyzed. Our results show that peripheral blood (p=0.04) and a higher CD34+ dose (p=0.05) were significantly associated with day 30 ALC ≥ 500. Time to ALC recovery ≥500 cells/mL was not significantly related to donor age for related alloBMT (p=0.340). However, in unrelated alloBMT patients, there was a trend toward increasing donor age associated with a longer time to ALC recovery ≥ 500 cells/mL (p=0.087). ANC recovery correlates with ALC recovery in related (R=0.237, p=0.02) and unrelated (R=0.514, p<0.001) alloBMT recipients. Recipient and donor age were strongly correlated in related alloBMT patients (R=0.89, p<0.001) and weakly correlated in unrelated alloBMT patients (R=0.236, p=0.09). ALC recovery ≥ 500 at day 30 post alloBMT was not a significant predictor of OS in either the related or unrelated subgroups. We will further examine donor-recipient age disparities in unrelated alloBMT patients and present multivariate analyses of predictors for time to ALC recovery.
Reduced Intensity Conditioning Hematopoietic Stem Cell Transplantation (HSCT) with Mesenchymal Stem Cells (MSC) Infusion for Treatment of Metachromatic Leukodystrophy (MLD): A Case Report.
Authors: Meuleman Nathalie , Rocha C. , Bennani Jalil , Tondreau Tatiana , Delforge Alain , Lewalle Philippe , Martiat Philippe , Lagneaux Laurence , Bron Dominique
Published: October 2019

Background: Patients (pts) with MLD have neurological and musculo-skeletal defects with a limited survival. HSCT has been reported as an effective treatment to stabilize or improve defects associated with this disease. Koç and colleagues reported that donor allogeneic MSC infusion is safe and may be associated with reversal of the disease pathophysiology in some tissues (Bone Marrow transplant, 2002). We decide thus to perform a non myeloablative familial HSCT in adult pt with a symptomatic MLD in order to evaluate the safety and the benefit allogeneic HSCT with MSC infusion in a patient with MLD. An informed consent was obtained from the pt.

Patient: a 23 years old women who presented an adult form of MLD for three years was admitted in our department. The most important symptoms associated with the disease were dizziness, proximal weakness of the lower limbs, difficulty to walk, disorder of the memory and urinary incontinence. The reduced intensity conditioning was preferred to decrease the morbidity and mortality of the procedure. MSC were isolated and amplified from the BM-mononuclear cells of the HSC donor. MSC expansion was made in a commercial serum-free medium (UltraCulture, Cambrex, Walkersville, MD) supplemented with a serum substitute (Ultroser, Pall Biosepra, Cergy-Saint-Christophe, France) as previously reported by our group (Eur J Haematol, 2006). Ex vivo expanded allogeneic MSC were intravenously infused at the dose of 1×106 MSC/kg of recipient body weight. The conditioning regimen of the HSCT consisted in Fludarabine and ATG combination and 5× 106 CD34 cells were infused concomitantly with 1.106 MSC cells. We did not observed any immediate or delayed side effects after the MSC infusion. The patient did not presented any complications after the HSCT. At day 8 of the transplantation she had an normal hematological recovery. The platelet nadir was 72.000/mm3 and she did not need any transfusion. With regards of her neurological status, since 22 months the patient had no new deterioration and we observed a stabilisation of the clinical manifestations.

Conclusions: This case report suggests the feasibility and the potential efficacy of reduced intensity conditioning allogeneic HSCT with MSC infusion for patient with MLD. A larger trial is required to confirm this observation.
Impact of Chimerism and Other Transplant Variables on Allogeneic Transplant Outcome after Reduced Intensity Conditioning (RIC) Regimens.
Authors: Corbella Veronica Ortiz , Lê Quoc-Hung , Nicolini Franck E. , Thiébaut Anne , Thomas Xavier , Chelgoum Youcef , Ducastelle Sophie , Mollet Isabelle , Dubois Valérie , Michallet Mauricette
Published: October 2019

RIC regimens followed by allogeneic hematopoietic stem cell transplantation (HSCT) were evaluated in patients with hematologic malignancies who were not candidates for conventional transplantation because of age or medical co-morbidities. In this kind of transplantation, the analysis of chimerism kinetics remains fundamental. The main aims of this study were to evaluate and compare the impact of pre- and post-transplant variables and chimerism on transplantation outcome. Chimerism status was evaluated in total blood (TB) and in purified CD3+ cells using quantitative PCR (STR/SNP) for all. We analysed donor type and kinetics chimerism of 52 patients (32 M - 20 F) who had undergone allogeneic HSCT after RIC for hematological malignancies. Median age was 40 years (range 25–67 years). Diagnosis were multiple myeloma: 16, acute myeloid leukemia: 13, myelodysplasia: 6, chronic lymphocytic leukemia: 5, Non Hodgkin’s lymphoma: 5, acute lymphoblastic leukemia: 1, chronic myeloid leukemia: 2 and primitive myelofibrosis: 4. Conditioning regimens were Fludarabine + Busulfan in 33 patients, Fludarabine + 2 Gys total body irradiation 15, Cyclophasphamide 3 (2 alone/one with Busulfan) and one other chemotherapy. Stem cell source was PB in all except one who received cord blood. Fourty-eight were identical sibling and 4 unrelated donor transplantations. At transplant, 17 patients were in CR, 21 PR and 14 in evolutive diseases. All patients except one engrafted. Twenty-five developed aGVHD ≥ Grade II (16 Grade III-IV). At the last follow-up 22 patients died (9 disease progression and 13 of transplant related mortality, 30 were alive (18 developed chronic graft vs Host disease (cGVHD) 12 extensive/6 limited cGHVD). Among 52 patients, only 49 had a long-term chimerism documentation of CD3 subpopulation (43 TB and CD3+ cells): 15 (14 TB) were full donor chimerism (FDC) throughout the follow-up, and 34 remained mixed chimerism (MC). Among these 34 (29 TB) MC patients at the latest follow-up, 7 (5 TB) remained in MC, 2 (5 TB) converted to recipient profile and 25 (19 TB) converted to donor profile. Univariate analysis demonstrated the close correlation between chimerism status evaluated on PB CD3+ cells only at any time post-transplant and the onset of aGVHD (p = 0.0391) but not cGVHD. Multivariate analysis according to linear regression method did not find any impact of the following variables on chimerism kinetics after RIC transplant: disease status before transplant, age, sex, type of RIC regimen, number of days of ATG, aGVHD (p ≥ 0.11). In conclusion, this study underlines the tight correlation that exists between chimerism status and kinetics on CD3+ PB subpopulations after RIC transplant and acute GVHD development that impacts on transplant outcome.
Novel Tyrosine Kinase Inhibitor Therapy (NTKI) Prior to Allogeneic Stem Cell Transplantation (ASCT) in Patients with Chronic Myeloid Leukemia (CML): No Evidence for Increased Transplant-Related Toxicity.
Authors: Jabbour Elias , Cortes Jorge , Kantarjian Hagop , Giralt Sergio , Giles Francis , Andersson Borje S. , Champlin Richard , de Lima Marcos
Published: October 2019

Introduction: Patients undergoing ASCT for CML are increasingly likely to have received a NTKI after failing imatinib mesylate. It is unknown whether the use of these NTKIs prior to ASCT increases transplant-related toxicity.

Methods: We retrospectively analyzed the outcome of 12 patients with CML (1 in chronic phase, 6 in accelerated phase, and 5 in blastic phase) who received dasatinib (n=2), nilotinib (n=7), or both (n=3) prior to ASCT.

Results: Median age was 41 years (range, 18–59 years). The median time on treatment was 134 days (range, 30–285 days), and the median time from the end of NTKI therapy to ASCT was 34 days (range, 7–130 days). Preparative regimen was ablative in 8 patients and non-ablative in 4. Stem cell source was a matched related donor in 7 patients, a matched unrelated donor in 3, a haplo-identical donor in 1, and unrelated cord blood in 1. All patients engrafted within 13 days. There was no significant early transplant-related toxicity: gastro-intestinal toxicities of grade 3 were encountered in 2 patients. One patient had secondary graft failure 6 months after the first ASCT that required a second ASCT. Acute graft-versus-host disease (GVHD) of grade 2 was observed in 7 patients (58%) and chronic GVHD in 6 (50%). Nine patients achieved a molecular response: four complete and five major (Q-PCR< 0.05%). Three patients had disease progression by day 30 post ASCT. Two patients have relapsed after 6 and 18 months. After a median follow-up of 10 months (range, 3–22 months), 7 patients are alive in molecular response and 5 patients died, 4 of disease progression and one of extensive chronic GVHD.

Conclusion: Previous treatment with NTKI did not increase transplant-related toxicity in this preliminary experience. Further follow-up and larger number of patients will be necessary to confirm these observations.
Defibrotide for the Treatment of Hepatic Veno-Occlusive Disease in Children.
Authors: Uygun Vedat , Tezcan Gulsun , Hazar Volkan , Yesilipek Akif
Published: October 2019

Veno-occlusive disease (VOD) of the liver is a major complication of allogeneic or autologous stem cell transplantation (HSCT), particularly after busulfan-based conditioning regimens. Clinically, VOD is characterized by fluid retention, weight gain, hyperbilirubinemia and painful hepatomegaly. Efficacy and safety of defibrotide for the treatment of VOD, especially in pediatric patients, has not been adequately confirmed or the results are still preliminary. This retrospective report describes experience with defibrotide in children with hepatic veno-occlusive disease (HVOD) following HSCT in a single institution.

Materials and Method: Children who had undergone HSCT between April 2005–June 2006 and who received defibrotide for the treatment of HVOD during their admission were identified. Demographic data and clinical information of these patients were abstracted from their health records.

Results: Eight children (median age: 4.5 years; range: 1–14 years) who underwent HSCT during the study period received defibrotide for the treatment of HVOD; 5 were boys and 3 were girls. Three patients underwent HSCT for hematologic malignancies, 2 for neuroblastoma, 1 for hemoglobinopathy, 1 for severe combined immunodeficiency (SCID) and 1 for fanconi aplastic anemia. Conditioning regimens included busulfan (16 mg/kg) in four, melphelan (140 mg/m2) in two and total body irradiation in 1 patient as being risk factors for HVOD. Two patients with neuroblastoma had undergone autologous HSCT, three patients had been transplanted from matched sibling donor and the other three were either unrelated donor or unrelated cord blood transplantation. HVOD was diagnosed on transplant day +4 to +19 (median: +12 ). The median initial defibrotide dose was 31.5 mg/kg/day (25–46 mg/kg/day). The median duration of defibrotide therapy was 27 days (1–46 days). Defibrotide was discontinued due to clinical improvement (5), gastrointestinal hemorrhage (2), death (1). Two patients in whom defibrotide was discontinued because of gastrointestinal hemorrhage died. The primary cause of death in these two children was: progressive HVOD and multiorgan failue following sepsis.

As a result of this experience and under the light of literature, we believed that defibrotide is an effective treatment for children with HVOD and should be instituted in each children without delay in the event of at least one of the criteria for HVOD is established. But it is necessary to carry out further studies in larger group of patients with longer follow-up to validate the efficacy and safety of defibrotide.
The Impact of Alemtuzumab as a Component of Conditioning Regimens on Transplantation Outcomes in a Setting of CMV-Seropositive Recipient and Donor.
Authors: Ahn Byung Min , Sohn Sang Kyun , Kim Jong Kwang , Chae Yee Soo , Cho Yoon Young , Moon Joon Ho , Yang Deok Hwan , Lee Je-Jung , Kim Yeo-Kyeoung , Kim Hyeoung-Joon
Published: October 2019

Background: Graft-versus-host disease (GVHD) is one of the main issues for allogenic stem cell transplantation (SCT). Previous studies demostrated that in vivo administration of alemtuzumab (an anti-CD52 antibody) was effective to decrease the incidence of GVHD after allogenic SCT. However, due to delayed immune reconstruction, graft failure or infection is concerned. We investigated the impact of in vivo administration of alemtuzumab on the post-transplantation outcomes.

Patients and methods: Thirty-five patients entered a pilot study employing allogenic SCT with alemtuzumab-containing conditioning regimen in two centers (KNUH, CNUH). We evaluated the clinical outcomes, complications and their relationship with dose of alemtuzumab.

Results: The median dose of alemtuzumab was 50mg (range, 20–75) which was not adjusted on body weight and surface. All recipients (9 BM, 26 PB) and donors were CMV-seropositive before transplantion. The median survival was 14.03 months and 2 year survival rate was 47.0%. The probability of nonrelapse mortality at 90 days and 1 year was 11.4% and 22.8%, respectively. The cumulative incidence of grade II-IV acute GVHD at day 90 was 37.1%. The overall incidence of chronic GVHD was 20.8% and the incidence of extensive chronic GVHD was only 8.3%. The cumulative incidence of CMV antigenemia was 70.2%, which has been well controlled with preemptive ganciclovir treatment, and two patients with CMV antigenemia developed into CMV disease. The febrile episode was 37.1% but the duration was relatively short (median, 3 days). There were no differenecies in engraft (P = 0.573), acute and chronic GVHD (P = 0.359, P = 0.335, respectively), relapse (P = 0.849), mortality (P = 0.229), CMV antigenemia (P = 0.227) or overall survival (P = 0.51) between two groups (treated with ≥50mg of alemtuzumab vs. <50mg).

Conclusion: Our study has shown that alemtuzumab is effective for GVHD prevention but correlated with high incidence of CMV antigenemia regardless of its dose. As CMV antigenemia was easily controllable with preemptive ganciclovir, the conditioning regimen containing alemtuzumab seems hopeful especially in a setting of CMV-seropositive recipient and donor with a high risk of GVHD.