bluuebunny/crossref_metadata_2025 huggingface dataset:
Phase I Study of Vaccination with Dendritic Cell Myeloma Fusions.
Authors: Avigan David , Rosenblatt Jacalyn , Vasir Baldev , Wu Zekui , Bissonnette Adam , MacNamara Claire , Uhl Lynne , Lenahan Corrinne , Miller Kenneth , Joyce Robin , Levine James D. , Lowe Karen , Dombagoda Dilani , Richardson Paul , Anderson Kenneth , Munshi Nikhil , Kufe Donald
Published: October 2019

We have developed a cancer vaccine in which patient derived myeloma cells are fused with autologous dendritic cells (DCs). In this way, multiple tumor antigens are presented in the context of the immune stimulating machinery of the DC. We are conducting a phase I trial in which patients with multiple myeloma (MM) undergo serial vaccination with DC/myeloma fusions in conjunction with GM-CSF. Eligibility criteria included ≥20% plasma cells in the bone marrow and measurable paraprotein or light chain. To date, 16 patients have been enrolled and 15 have undergone therapy. Patients underwent leukapheresis collection and DCs were generated from adherent mononuclear cells cultured for 5 days with GM-CSF and IL-4 and matured by exposure to TNFa for 48–72 hours. The mean yield of DCs was 1.23 x108 cells with a mean viability of 88%. Patient derived myeloma cells were isolated from bone marrow aspirates. The mean yield of myeloma cells was 92.5 x 106 cells with a mean viability of 90%. Fusion cells were generated by coculture of DCs with myeloma cells in the presence of 50% polyethylene glycol. Fusion cells were quantified by determining the percentage of cells that co-expressed unique DC and myeloma antigens. Mean fusion efficiency and viability was 40% and 84%, respectively. As a measure of immunologic potency, DC/MM fusions stimulated allogeneic T cell proliferation, with mean stimulation indexes of 48, 36, and 9 for the DC, fusion and myeloma cell populations, respectively. To date, 15 patients have completed vaccination at a dose of 1–5x106 fusion cells. Adverse events potentially related to vaccine included injection site reactions, edema, rash, fever (infection), chills, fatigue, muscle aches, pruritis, and diarrhea. One patient with a history of prior deep venous thrombosis (DVT) developed a DVT and pulmonary embolus of uncertain relation to the vaccine. To date, 6/9 evaluable patients have demonstrated immunologic response, defined by at least 2 fold increase in IFNγ expression by CD4 and/or CD8 T cells in response to ex vivo exposure to autologous tumor lysate. Vaccination was associated with an increase in circulating tumor specific T cells as evidenced by an increase in CD8+ cells binding the MUC1 tetramer. The impact of vaccination on circulating levels of regulatory T cells is being examined. Humoral immune responses are being assessed by SERAX analysis. Three patients have ongoing stable disease at 3.5, 6, and 19 months following their initial vaccination. Six patients demonstrated disease stability with subsequent progression ranging from 9 weeks to 8 months following their first vaccine. Vaccination with DC/MM fusions has been well tolerated, associated with immunologic response, and disease stabilization in a majority of patients with multiple myeloma.
Effects of Celecoxib on Human Acute Promyelocytic Leukemia Cell Line and Its Mechanism.
Authors: Xu Yun , Huang He , Zhao Yanmin , Zeng Fenfang , Zhou Qian
Published: October 2019

Acute promyelocytic leukemia (APL) is a special subtype of acute myelogenous leukemia (AML) which is characterized for a specific translocation between chromosome 15 and 17 [t(15;17)] and the expression of PML/RARα fusion gene. Celecoxib, one of the specific inhibitors of cyclooxygenase-2 (COX-2), has been reported to induce anti-neoplastic activity on many solid human tumor cell lines in recent years. In our study, ATRA resistant APL cell line MR2 cells were used to investigate the effects of celecoxib on hematological malignancy. MR2 cells were treated with celecoxib at different concentration (0, 20, 40, 80, 120 and 160μmol/L). The proliferation of MR2 cells was observed by MTT assay and apoptosis was detected by DNA fragmentation analysis and flow cytometry using Annexin V-FITC/PI staining. Western blot was used to detect the change of caspase-8, -9, -3 and PARP in MR2 cells. The expression of mRNA of fusion gene PML/RARα, COX-2 and survivin, bcl-2/bax, CIAP1 and CIAP2 was assessed by reverse transcription polymerase chain reaction (RT-PCR). Cell cycle analyzed by flow cytometry with PI staining and western blot was used to detect the expression of cell-cycle-regulating proteins. The telomerase activity of MR2 cells was analyzed by PCR-ELISA. The expression of hTERT mRNA and c-myc mRNA was assessed by RT-PCR. MR2 cells viability in presence of celecoxib decreased markedly in a dose- and time- dependent manner. After treated with celecoxib (20-160μmol/L) for 12-48h, the proliferation of MR2 cells were inhibited significantly, in comparison with the control group (P<0.01). 50% growth inhibition (IC50) at 24h and 48h was 80.93μmol/L and 71.72μmol/L, respectively. A DNA ladder pattern of internucleosomal fragmentation was observed. The translocation of phosphatidylserine at the outer surface of the cell plasma membrane could be induced by celecoxib and its level increased following the augmentation of the drug concentration. MR2 cells exposure to 40-160μmol/L celecoxib for 24h caused 9.59%, 24.00%, 36.10% apoptotic cells, which was more than that of the untreated group 2.84% (P<0.01). The expression of survivin mRNA decreased dramatically, while no significant change with PML/RARα and COX-2. Treatment with celecoxib for 24h resulted in the activation of caspase-3 and caspase-9, cleavage of PARP. 40-160μmol/L celecoxib led to cell cycle arrest in G1/S phase, and CyclinD1 and CyclinE decreased, accompanied with up-regulation of P21waf/cip1, P27KIP, P16INK4a. celecoxib could inhibit the telomerase activity of APL cell line, and the inhibition was dose- and time- dependent. The expression of hTERT mRNA and c-myc mRNA were down-regulated by celecoxib in dose- dependent manner. These results indicated that celecoxib could inhibit MR2 cells proliferation by inducing apoptosis, cell cycle arrest and suppression of telomerase activity.
Exclusion from Acute Myeloid Leukemia (AML) Clinical Trials of Patients Unlikely To Respond.
Authors: Tsimberidou Apostolia-Maria , Kantarjian Hagop M. , Faderl Stefan , Freireich Emil J. , Estey Elihu
Published: October 2019

Introduction: Patients with newly diagnosed AML are increasingly being enrolled on industry-sponsored clinical trials. These trials routinely exclude patients with Zubrod performance status (PS) >2, bilirubin >1.9 mg/dL, or creatinine >1.9 mg/dL; such patients are well-known to be relatively unlikely to respond (UTR). Here we quantify the effect of this practice on inclusion of UTR patients in clinical trials.

Patients and Methods: Overall, 2323 adults with newly-diagnosed AML (no acute promyelocytic leukemia) were registered on clinical trials (industry-sponsored and other) from 1991–2006. Considering patients as UTR if they fulfilled any of the criteria noted above, we examined rates of enrollment of UTR patients over time:

Results: Thus, while more patients are being enrolled on clinical trials, fewer UTR patients are being enrolled particularly over the last 5 years (p = .04) with a concomitant decrease in the proportion of such patients enrolled (p<.0001 over the 16 years in question). Findings are qualitatively the same in older patients.

Conclusion: Newer clinical trials may be less applicable to UTR patients than previous trials. There is a need for clinical trials specific to the UTR population, the group most in need of novel therapies.

Rates of enrollement of unlikely to respond patients on clinical trials for newly diangosed AML Year No. of pts. treated on clinical trial (pts. age≥60 yrs) UTR pts. enrolled (pts. age≥60 yrs) % of UTR pts. enrolled (pts. age ≥60 yrs) 1991 101 (50) 17 (13) 17 (26) 1992 100 (48) 20 (14) 20 (29) 1993 96 (46) 18 (9) 19 (20) 1994 109 (41) 23 (13) 21 (32) 1995 102 (48) 20 (10) 20 (21) 1996 104 (63) 14 (10) 13 (16) 1997 145 (81) 30 (20) 21 (25) 1998 124 (63) 14 (8) 11 (13) 1999 164 (86) 26 (16) 16 (19) 2000 169 (96) 25 (19) 15 (20) 2001 187 (98) 30 (19) 16 (19) 2002 162 (91) 17 (11) 10 (12) 2003 195 (111) 4 (2) 2 (2) 2004 184 (109) 17 (11) 9 (10) 2005 185 (117) 15 (7) 8 (6) 2006 196 (109) 17 (12) 9 (11) Total 2323 (1257) 307 (194) 13 (15)
Efficacy of Nilotinib (AMN107) in Patients (Pts) with Newly Diagnosed, Previously Untreated Philadelphia Chromosome (Ph)-Positive Chronic Myelogenous Leukemia in Early Chronic Phase (CML-CP).
Authors: Cortes Jorge , O’Brien Susan , Jabbour Elias , Ferrajoli Alexandra , Borthakur Gautam , Burger Jan , Wierda William , Garcia-Manero Guillermo , Thomas Michelle , Letvak Laurie , Kantarjian Hagop
Published: October 2019

Background: Nilotinib is a novel, oral tyrosine kinase inhibitor with increased selectivity against Bcr-Abl that is approximately 30-fold more potent than imatinib. High response rates have been reported with nilotinib therapy in CML after imatinib failure.

Methods: We evaluated the efficacy of nilotinib as first line therapy in pts with newly diagnosed Ph-positive CML-CP. The primary objective was to estimate the proportion of pts attaining major molecular response (BCR-ABL/ABL ratio ≤0.05% in our lab) at 12 months (mo).

Results: Thirty-two pts have been treated with nilotinib 400 mg orally twice daily for a median of 5 months (mo) (range, 1 to 31 mo). The median age was 47 years (yrs) (range, 24–73 yrs). The Sokal risk at pretreatment was low in 21 (70%) pts, intermediate in 6 (20%), and high in 3 (10%). The rate of complete cytogenetic response [CCyR] (Ph 0%) at 3, 6 and 12 mo compares favorably to those observed in historical controls treated with imatinib 400 mg or 800 mg daily:

The median QPCR with nilotinib at 3, 6, and 12 months were, respectively, 0.52% (range, 0.0–29.5%), 0.03% (range, 0.0–9.13%), and 0.09% (range, 0.0–16.21%). At 3 mo follow-up, major molecular response (MMR; BCR-ABL/ABL ratio ≤0.05%) was observed in 3/22 patients (14%), 7/13 (54%) at 6 mo, and 5/11 (45%) at 12 mo. 12-mo rates of MMR for the historical imatinib groups treated at 400 mg and 800 mg were 24% and 47%, respectively (p=0.02). None of the molecular responses has been lost while on therapy. Grade 3–4 neutropenia was observed in 2 (7%) pts, and thrombocytopenia in 1 (3%). Other grade 3–4 adverse events included elevation of lipase (n=3, 9%), or bilirubin (n=2; 6%), and amylase elevation, back pain, and infection (1 each). Twelve pts had transient treatment interruptions (median 11 days), most frequently due to pain (n=3; musculoskeletal 2, abdominal 1), lipase elevation (n=2). Seven patients had their dose reduced to 400 mg daily, 2 to 200 mg twice daily, and 1 to 200 mg daily due to extramedullary toxicity. Three patients decided to change therapy after 4, 6 and 8 months; 2 switched to imatinib and 1 received SCT.

Conclusion: Nilotinib 400 mg orally twice daily suggest significant efficacy manifested by complete cytogenetic responses in nearly all patients as early as 3 months after the start of therapy with a favorable toxicity profile.

Percent with CCyR (No. evaluable) Months on Therapy Nilotinib Imatinib 400 mg Imatinib 800 mg P value 3mo 95 (22) 37 (49) 62 (202) < 0.0001 6mo 100 (13) 54 (48) 82 (199) <0.0001 12mo 100 (11) 65 (48) 86 (197) 0.0007
Inhibitory Effects of Homoharringtonine on Leukemic Stem Cells and BCR-ABL Induced Chronic Myeloid Leukemia and Acute Lymphoblastic Leukemia in Mice.
Authors: Chen Yaoyu , Hu Yiguo , Michaels Shawnya , Brown Dennis , Li Shaoguang
Published: October 2019

The Abl tyrosine kinase inhibitors (TKIs) imatinib mesylate (IM) and dasatinib, targeting BCR-ABL for the treatment of Philadelphia-positive (Ph+) leukemia including chronic myeloid leukemia (CML) and B-cell acute lymphoblastic leukemia (B-ALL), have produced impressive results in terms of therapeutic outcome and safety for patients. However, clinical resistance to these TKIs likely at the level of leukemic stem cell negates curative results in Ph+ leukemia. At present, an anti-stem cell strategy has not been developed for treating these leukemia patients. Homoharringtonine (HHT) (omacetaxine mepesuccinate - USAN/INN designation) has shown significant clinical activity in CML in combination with IM or alone for patients failing IM. However, little is known about whether HHT has an inhibitory effect on leukemic stem cells. The purpose of this study is to determine whether HHT inhibits BCR-ABL-expressing leukemic stem cells (Lin-c-Kit+Sca-1+) that we identified previously (Hu et al. Proc Natl Acad Sci USA 103(45):16870–16875, 2007) and to evaluate therapeutic effects of HHT on CML and B-ALL in mice. We find that in our in vitro stem cell assay, greater than 90% of leukemic stem cells were killed after being treating with HHT (12.5, 25, and 50 nM) for 6 days, and in contrast, greater than 75% or 92% of leukemic stem cells survived the treatment with dasatinib (100 nM) or imatinib (2 mM). We next treated CML mice with HHT (0.5 mg/kg, i.p., once a day). 4 days after the treatment, FACS analysis detected only 2% GFP+Gr–1+ myeloid leukemia cells in peripheral blood of HHT -treated CML mice and in contrast, 41% GFP+Gr–1+ myeloid leukemia cells in placebo-treated mice. We also treated mice with BCR-ABL induced B-ALL with HHT, and found that only 0.78% GFP+B220+ lymphoid leukemia cells were detected in peripheral blood compared to 34% GFP+B220+ lymphoid leukemia cells in placebo-treated mice. Furthermore, HHT significantly inhibited in vitro proliferation of K562 and B-lymphoid leukemic cells isolated from mice with B-ALL induced by BCR-ABL wild type and BCR-ABL-T315I resistant to both imatinib and dasatinib. In sum, HHT has an inhibitory activity against CML stem cells, and is highly effective in treating CML and B-ALL induced by BCR-ABL in mice.
Molecular Basis Explanation of Imatinib Resistance of Bcr-Abl Due to T315I and P-Loop Mutations from Molecular Dynamics Simulations.
Authors: Lee Tai-Sung , Potts Steven , Kantarjian Hagop , Cortes Jorge , Giles Francis , Albitar Maher
Published: October 2019

Molecular dynamics (MD) simulations on the complex of imatinib with the wild-type, T315I, and other 10 P-loop mutants of the tyrosine kinase Bcr-Abl have been performed to study the imatinib resistance mechanism at the atomic level. MD simulations show that large scale computational simulations could offer insight information that a static structure or simple homology modeling methods cannot provide for studying the Bcr-Abl imatinib resistance problem, especially in the case of conformational changes due to remote mutations. By utilizing the Molecular Mechanics/Poisson-Boltzmann surface area (MM-PBSA) techniques and analyzing the interactions between imatinib and individual residues, imatinib resistance mechanisms not previously thought have been revealed. Non-directly contacted P-loop mutations either unfavorably change the direct electrostatic interactions with imatinib, or cause the conformational changes influencing the contact energies between imatinib and other non-P-loop residues. We demonstrate that imatinib resistance of T315I mainly comes from the breakdown of the interactions between imatinib and E286 and M290, contradictory to previously suggested that the missing hydrogen bonding is the main contribution. We also demonstrate that except for the mutations of the direct contact residues, such as L248 and Y253, the unfavorable electrostatic interaction between P-loop and imatinib is the main reason for resistance for the P-loop mutations. Furthermore, in Y255H, protonation of the histidin is essential for rendering this mutation resistant to Gleevec. Our results demonstrate that MD is a powerful way to verify and predict clinical response or resistance to imatinib and other potential drugs.
Glycoprotein Ibα Forms Catch Bonds with von Willebrand Factor A1 Domain but Not with Mutant A1 Domains Exhibiting Properties of Type 2B von Willebrand Disease.
Authors: Wu Tao , Yago Tadayuki , Yang Jun , Miner Jonathan , Coburn Leslie , Lopez Jose A. , Cruz Miguel A. , McIntire Larry V. , McEver Rodger P. , Zhu Cheng
Published: October 2019

Interactions between glycoprotein Ibα (GPIbα) and von Willebrand factor (VWF) initiate platelet adhesion to injured vascular surfaces, which is enhanced by arterial blood flow. The flow requirement for adhesion is reduced (i.e., gain-of-function) by single-residue substitutions of the VWF-A1 domain, e.g., R543Q that occurs in some patients with type 2B von Willebrand disease (VWD) and R687E that was designed to exhibit type 2B VWD properties. Yet the mechanisms for flow-enhanced adhesion through GPIbα-VWF interactions are not understood. By probing single bonds with atomic force microscopy, we showed that lifetimes of GPIbα/VWF-A1 bonds first increased (catch) and then decreased (slip) with increasing force applied to the bond. Remarkably, the catch bond aspect of the GPIbα/VWF-A1 bonds was eliminated by the R543Q and R687E mutations, which exhibited slip bonds only with prolonged lifetimes at low forces. Flow chamber experiments showed that catch-slip transitional bonds governed flow-enhanced rolling of platelets and GPIbα-coated microspheres on wild-type A1, such that rolling velocities first decreased and then increased with increasing flow. By contrast, slip bonds governed rolling velocities on R543Q and R687E A1 mutants, which increased monotonically with increasing flow. We changed fluid viscosity by adding Ficoll to the medium, tether force by using microspheres of different radii, and platelet deformability by using a chemical fixative. The rolling velocity vs. flow curves aligned with tether force but not with transport parameters and were minimally affected by fixation, which respectively rule out transport-enhanced GPIbα/VWF-A1 bond formation and force-induced enlargement of platelet-surface contact area as the causes for flow-enhanced rolling. Flowing platelets agglutinated with microspheres bearing R543Q and R687E A1 mutants but not with those bearing wild-type A1, suggesting that GPIbα/VWF-A1 catch bonds prevent agglutination of circulating platelets via binding to VWF multimers and that platelet agglutination in patients with type 2B VWD may be explained by the prolonged lifetimes at low forces resulting from elimination of catch bonds in the interactions of GPIbα with VWF-A1 mutants.
Glyoxalase-I Is a Potential Therapeutic Target for Chronic Myelogenous Leukemia Cells That Have Acquired Drug Resistance through Adaptation to Hypoxia.
Authors: Takeuchi Miki , Kuroda Junya , Kimura Shinya , Ashihara Eishi , Umezawa Kazuo , Yasui Eiko , Kawatani Makoto , Osada Hiroyuki , Hodohara Keiko , Fujiyama Yoshihide , Maekawa Taira
Published: October 2019

Growth under hypoxic conditions can lead to resistance to a variety of cell death stimuli in cancer cells. Bone marrow (BM) has less vasculature than other organs, and the rapid growth of leukemic cells in BM frequently outpaces the oxygen supply. Thus, adaptation to hypoxia may be critical to the survival and proliferation of leukemic cells. However, the role of hypoxia in the treatment of leukemia has received little attention. We hypothesized that leukemic cells acquire drug resistance by adapting to hypoxia in the BM, and that identifying the underlying molecular mechanisms for this adaptation would reveal new approaches to the development of new therapeutic targets in drug- resistant leukemia cells. To test this hypothesis, we cultured five chronic myelogenous leukemia (CML)-derived cell lines (BV173, K562, KCL22, KT-1, and MYL) continuously in 1.0% oxygen. Among them, hypoxia resistant (HR) subclones of K562 and KCL22 cells (K562/HR and KCL22/HR) were selected under 1.0% oxygen conditions, and these cells were able to grow for several months and to be suspended in RPMI1640 plus 10% FCS without any additional nutrients, although their growth rate was slower than parental cells in 20% oxygen. Both K562/HR and KCL22/HR cells produced less ATP (78% and 92%, respectively), perhaps due to impairment of the oxygen-dependent energy production pathway, and HR cells showed higher expression and enhanced kinetics of glyoxalase-I (Glo-I), a detoxification kinase of methylglyoxal (a cytotoxic byproduct of glycolysis), compared to parental cells. Glo-I has previously been implicated in drug resistance in cancer, including leukemia. Autophosphorylation of Bcr-Abl was reduced in HR cell lines, while the phosphorylation status of the downstream signalling molecules Akt and Stat5 was unchanged, which suggests that there are alternative, hypoxia-specific activators of these pathways. K562/HR and KCL22/HR cells were less sensitive to chemotherapeutic agents such as the Bcr-Abl tyrosine kinase (TK) inhibitors imatinib, dasatinib, nilotinib, and INNO-406, and the alkylating agents daunorucbicin and busulfan. In contrast, K562/HR and KCL22/HR cells were more sensitive to cell death induced by a Glo-I inhibitor, COTC, compared to parental cells. These results indicated that Glo-I plays an important role in survival under hypoxic conditions, and that Glo- I may be an alternative molecular target in CML cells that have acquired resistance to conventional chemotherapeutics through adaptating to hypoxia.
Combination of Imatinib with Cisplatin and Nutlin-3: Functional and Molecular Effects on Bcr-Abl Positive Cells.
Authors: Skorta Ioanna , van der Kuip Heiko , Henkes Martin , Oren Moshe , Aulitzky Walter E.
Published: October 2019

Imatinib is highly effective in inducing remissions in chronic phase CML. However, complete eradication of the malignant clone by Imatinib monotherapy is rare. This prompted us to explore the efficacy of combination of Imatinib with the DNA damaging agent Cisplatin and with Nutlin-3, a compound which induces p53 accumulation by interfering with its binding to Mdm2. We used a Bcr-Abl positive cell line characterized by the loss of Imatinib sensitivity in the presence of optimal growth factor concentrations. Whereas combined treatment of Imatinib with either Cisplatin or Nutlin-3 in low and nontoxic doses induced ∼15–20% cell death, simultaneous treatment of all three compounds was highly effective (∼50% cell death) even in the presence of optimal growth conditions. Importantly, comparable effects were also seen in CFU assays performed with primary CD34 positive cells from 5 CML patients. To examine the molecular mechanisms of these effects we analyzed p53 dependent cellular response to Cisplatin in our cell line model in the presence or absence of Imatinib and/or Nutlin-3. The active Bcr-Abl kinase caused superinduction of p53 protein after DNA damage. We found both an upregulated p53 accumulation and enhanced p21 and Mdm2 RNA and protein levels upon Cisplatin treatment. This phenomenon could be reversed both by siRNA-mediated inhibition of Bcr-Abl expression and by Imatinib-induced inhibition of Bcr-Abl kinase activity: despite of optimal growth conditions inhibition of Bcr-Abl caused a significant reduction of p53 expression and activation. In the presence of Nutlin-3, p53 expression was significantly upregulated upon Cisplatin treatment both with and without Imatinib. However, Nutlin-3 was not capable to restore the Imatinib-induced blockade of the transcriptional activity of p53 on mdm2. The reduced p53 activation observed in Bcr-Abl positive cells treated with Imatinib was paralleled by a shift from cell cycle arrest to cell death both in the presence and absence of Nutlin selectively in Bcr-Abl positive cells rendering them hypersensitive to Cisplatin. In summary, combined treatment of Imatinib with low concentrations of a DNA damaging agent and a p53 activator is highly effective in vitro. Such combined treatment may prove to be clinically relevant for complete eradication of the malignant clone in CML, provided that conditions are found where it does not affect adversely normal hematopoietic cells in vivo.
Skin Changes on Continued Imatinib Therapy in Patients with Ph+ CML.
Authors: Saikia Tapan , Hazarika Nandini , Dhabhar Boman
Published: October 2019

We reported the skin changes (generalized hypopigmentation) caused by imatinib mesylate (IM) in patients with Ph+ CML or GIST (EHA 2004 & ASH 2004). Subsequent reports from MDACC confirmed such findings. Such changes could be different and more profound among the brown skinned people of India. A cohort of 200 Ph+ CML patients at various phases of the disease receiving IM in a daily dose of 300 mg - 800 mg (children 200 mg daily) were studied over a median period of 14 months (range 5 – 72 months). Skin changes of any kind or severity were noted in 120 (60%) patients. Hypopigmentation alone (usually generalized) was seen in 75 patients (32.5%), combined hypopigmentation and hyperpigmentation in 39 (19.5%), hypopigmentation + extreme thinning of the skin in 3 (1.5%) and the classical skin rash of grade 3/4 in 2 (1%). Skin changes occurred at all dose levels. Characteristically the hyperpigmented patches were localized to the facial area over the forehead and/or in a butterfly pattern across the malar areas and bridge of the nose. IM dose was not modified among patients with pigment changes alone. IM causes dose-independent hypo- or hyperpigmentation of the skin in a significant number of Indian patients with Ph+ CML. This apparently happens due to modified melanin metabolism through the tyrosine pathway. Some of these changes (hyperpigmentation and extreme thinning) are bothersome from cosmetic viewpoint.
Evaluation of Immune Responses to Seasonal Influenza Vaccination in Healthy Volunteers and in Patients after Stem Cell Transplantation.
Authors: Avetisyan Gayane , Aschan Johan , Hassan Moustapha , Ljungman Per
Published: October 2019

Influenza causes significant morbidity and mortality in immunocompromised stem cell transplant (SCT) recipients. Measurement of cellular and humoral immunological responses might increase our understanding of how to estimate a protective response to influenza vaccination. Eighteen healthy subjects and 14 SCT patients tested before and 4 weeks after influenza vaccination were included in the study. Peripheral blood lymphocytes were stimulated with four influenza peptides; three based on sequences from the hemagglutinin and one from the M1 protein. Elispot assays were performed to measure the production of intracellular interferon-gamma (IFN-) and interleukin-13 (IL-13). Pre-labeled MHC class I pentamers were used for the detection of influenza-specific CD8+ T-cells. B-cell Elispot was performed to enumerate influenza-specific antibody-secreting cells (ASCs). Finally, haemagglutination inhibition (HIA) was used to determine the serum titers of neutralizing antibodies. Influenza vaccination elicited strong cell-mediated immune responses in the healthy volunteers (p≤0.003 for all four peptides) and SCT patients (p≤0.008). The percentage of CD8+ influenza-specific cells increased significantly after vaccination of both volunteers (p=0.005) and patients (p≤0.003). The number of influenza-specific ASCs increased as well (in volunteers p=0.009, in patients p=0.01). Only 29% of the SCT patients developed protective antibody levels to the influenza A H1/N1 serotype while no patient had protective antibody levels to H3/N2 or influenza B types. All responses were lower in patients studied less than 6 months after SCT compared to those studied later after SCT. These results indicate that seasonal vaccination against influenza can boost the cellular immune response in SCT patients and healthy controls, while B-cell responses are suboptimal in SCT patients.
Symptomatic Improvement Following Intravesicular Cidofovir for the Management of BK Virus Associated Cystitis.
Authors: Buie Larry W. , Rao Kamakshi V. , Epstein Stacy , Shea Thomas C. , Comeau Terrence , Gabriel Don A. , Irons Robert , Eron Barbara , Serody Jonathan
Published: October 2019

BACKGROUND: Hemorrhagic cystitis (HC) is a complication of hematopoietic stem cell transplantation, occurring in 10 to 70% of transplant recipients. Factors that may contribute to the risk of developing HC include intensive preparative regimens, especially those including busulfan, high-dose cyclophosphamide or ifosfamide, transplantation from matched unrelated donors, infection with adenovirus, and the development of graft-versus-host disease. The reactivation of latent BK virus in the kidney is thought to lead to the development of late-onset HC in hematopoietic stem cell transplant patients. Early reports of cidofovir’s efficacy in BK virus-associated HC used intravenous cidofovir, but treatment was often complicated or compromised by excessive toxicity, especially renal compromise. Based on small single patient reports of the use of intravesicular cidofovir in the management of other viral causes of cystitis, we evaluated the use of intravesicular cidofovir for the management of BK virus associated cystitis.

METHODS: In patients undergoing stem cell transplant, BK viral loads were routinely monitored twice weekly. The presence of cystitis or hematuria in a patient with detectable BK viruria was considered the trigger for initiation of therapy. Patients were treated with cidofovir at a dose of 5 mg/kg. Drug was diluted with 60mL of normal saline and administered via foley catheter, which was clamped for one hour post administration. Weekly repeat dosing was allowed as long as symptoms persisted. In addition to symptom control, BK viral loads were measured before, during, and after treatment.

RESULTS: Four patients were included in this initial evaluation. All were recipients of allogeneic stem cell transplants; three were recipients of matched unrelated donor transplants and one received a transplant from a matched sibling. Patients received between 1 and 4 doses of cidofovir for initial therapy. All 4 patients experienced complete resolution of symptoms. The range for symptom resolution ranged from 2 days to 3 weeks. Symptoms reappeared in only 1 patient after discontinuation of therapy. Monitoring of BK urine viral loads revealed that 3 of the 4 patients had a greater than 50% decrease in urine BK viral load with initial dosing of cidofovir, indicating presence of antiviral activity when the drug is administered by this novel route.

CONCLUSION: Intravesicular cidofovir is safe, easy to administer, and associated with both elimination of symptoms and reduction in viral loads in patients with hemorrhagic cystitis following high dose therapy and allogeneic transplantation.
Adaptation of Coagulation and Fibrinolysis in Allogeneic Stem Cell Transplantation.
Authors: Pinomaki Anne , Volin Liisa , Joutsi-Korhonen Lotta , Virtanen Jussi Oskari , labtech Marja Lemponen , Ruutu Tapani , Lassila Riitta
Published: October 2019

Allogeneic stem cell transplantation (SCT) seems to result in activation of coagulation and fibrinolysis. We characterised the outcome of SCT and graft versus host disease (GvHD) in association with the adaptive mechanisms of coagulation and fibrinolysis. 30 patients given myeloablative conditioning with cyclophosphamide and total body irradiation underwent allogeneic SCT for a hematological malignancy. 19 patients received the transplant from a sibling and 11 from an unrelated donor. GvHD prophylaxis consisted of cyclosporine and methotrexate, and in addition methylprednisolone in case of a sibling donor. Eight patients developed GvHD during the 3-month follow-up. Several coagulation and fibrinolysis activities were serially assessed: antithrombin, protein C (PC), FVIII, prothrombin fragments 1+2 (F1+2), D-dimer, tissue plasminogen activator (tPA) and plasminogen activator inhibitor (PAI-1). During the conditioning therapy as an early sign of thrombin generation, F1+2 increased 3-fold and D-dimer 4-fold, compatible with enhanced fibrin turnover. The activity of tPA reached its maximum already before the engraftment and that of PAI-1 diminished accordingly. FVIII increased steadily from normal to reach maximum (up to 273% ±104%, median ±SD, p<0.001) after engraftment. Interestingly, the natural anticoagulant, PC rose (up to 189% ± 63%) in parallel with FVIII, but showed a 5-fold individual variability. After the engraftment the FVIII, PC and antihrombin levels were highly interrelated. Clinically relevant prognostic association was observed between early low PC activity and the appearance of GvHD: the level of PC lower than 90% during the conditioning therapy was associated and even predicted (p=0.007) acute GvHD (OR=16.7). After the transplantation the level of F1+2 over 2.5 nmol/l associated and predicted the non-relapse mortality (p=0.024). Three patients with the largest early thrombin generation all died during a longer follow-up (6–24 months). Also, elevated PAI-1 activity (>10 U/ml) predicted the non-relapse mortality (p=0.013). In conclusion, early activation of coagulation and fibrinolysis is followed by increased FVIII and PC activities in SCT. Early marked thrombin generation and elevated PAI-1 associated with the non-relapse mortality and low PC activity with the appearance of acute GvHD.
Allogeneic Hematopoietic Stem Cell Transplantation for Myeloma: Does the Melphalan Dose Matter?.
Authors: Mehta Jayesh , Chunduri Sandeep , Dobogai Lisa , Singhal Seema , Rondelli Damiano
Published: October 2019

The dose of melphalan affects the outcome of autotransplantation in myeloma significantly. Regimens not containing melphalan or doses of melphalan lower than 200 mg/m2 have been shown to result in inferior disease-free (DFS) and overall (OS) survival (Mehta J, Singhal S. High-dose chemotherapy and autologous hematopoietic stem cell transplantation in myeloma patients under the age of 65 years. Bone Marrow Transplantation advance online publication, 6 August 2007; doi:10.1038/sj.bmt.1705799). However, whether the melphalan dose affects the outcome of allogeneic HSCT is unknown. The outcome of 30 patients undergoing submyeloablative allogeneic HSCT after 100 mg/m2 (n=24) or 140 mg/m2 (n=6) melphalan was studied to determine the effect of melphalan dose-intensity. The higher melphalan dose was combined with 150 mg/m2 fludarabine. As the figures below show, DFS and OS appeared to be better after the higher dose of melphalan on unadjusted analysis.

Figure Figure

The following variables were studied for their effect on OS: patient age, donor age, chemorefractoriness, HLA mismatch, melphalan dose, performance status, and LDH level. The factors found to be significant in univariate analysis - donor age, performance status, LDH, and chemorefractoriness - were entered into a Cox model. The dose of melphalan was forced into the final model. The table below shows the factors found to significantly affect relapse, DFS and OS in multivariate analysis. No factor was found to affect transplant-related mortality significantly.

Adverse variable RR P Relapse Refractory disease 5.5 0.02 DFS Elevated LDH 3.5 0.017 Donor age >45 3.4 0.024 Refractory disease 3.5 0.044 OS Elevated LDH 8.0 0.002 Performance status 2/3 5.8 0.003 Donor age >45 4.1 0.031

The melphalan dose did not affect relapse (P=0.37), DFS (P=0.16) or OS (P=0.12) significantly. These data suggest that 100 mg/m2 and 140 mg/m2 melphalan doses appear to result in comparable outcomes after allogeneic HSCT in myeloma. This is probably because the immunologic graft-versus-myeloma effect compensates for the reduced anti-myeloma efficacy of the lower melphalan dose. Whether higher melphalan doses, used successfully as pre-allograft conditioning for hematologic malignancies (Singhal S, Powles R, Treleaven J, Horton C, Mehta J. Melphalan alone prior to allogeneic bone marrow transplantation from HLA-identical sibling donors for hematologic malignancies: alloengraftment with potential preservation of fertility. Bone Marrow Transplant1996; 18:1049–1055), confer any benefit for myeloma allografts remains to be seen.
Outcomes of Allogeneic Hematopoietic Cell Transplantation (HCT) after Non-Myeloablative Conditioning in Relapsed Diffuse Large B-Cell Lymphoma (DLBCL).
Authors: Rezvani Andrew R. , Norasetthada L. , Gooley T. , Sorror Mohamed , Forman Stephen J. , Agura Edward , Chauncey T. , Maziarz Richard T. , Maris Michael , Shizuru J. , Bruno B. , Wade James C. , Lange T. , Yeager Andrew , Sandmaier Brenda M. , Storb Rainer F. , Maloney David G.
Published: October 2019

BACKGROUND: We report on outcomes after non-myeloablative allogeneic HCT in patients (pts) with relapsed DLBCL.

PATIENTS: Thirty-two pts with relapsed DLBCL were treated at 11 centers with allogeneic HCT from related (n=21) and unrelated (n=11) donors following non-myeloablative conditioning with 2 Gy total body irradiation, with or without fludarabine. Median age was 51.8 years. Twenty-four pts (75%) had undergone prior high-dose therapy (HDT) with autologous HCT: 18 (56%) had failed HDT, and 6 (19%) underwent planned tandem autologous/allogeneic HCT. The remaining 8 pts (25%) were medically ineligible for HDT. Twenty-three pts (72%) had chemosensitive disease at HCT, while 9 (28%) were chemorefractory. Disease status at HCT was complete response (CR, 14/32, 44%), partial response (PR, 9/32, 28%), or relapsed/refractory (9/32, 28%).

RESULTS: All patients engrafted. Cumulative incidences of acute GVHD grades II-IV, acute GVHD grades III-IV, and chronic GVHD were 53%, 19%, and 47%, respectively. Median follow-up of surviving patients was 45 months. Of pts with measurable disease at HCT (n=18), responses were seen in 7/18 (39%), with 6 CR’s and 1 PR. Nine of these eighteen pts progressed, and 2/18 were not evaluable due to early death from non-relapse causes. Of pts in CR at HCT, 10/14 (71%) remained in CR while 4/14 (29%) progressed. Three-year estimated overall (OS) and progression-free survival (PFS) were 45% and 35%, respectively. Three-year cumulative incidences of relapse and non-relapse mortality were 41% and 25%, respectively (see Fig. 1). In multivariate models, chemosensitive disease and receipt of >4 lines of chemotherapy prior to HCT were associated with better overall survival. Pts with chemosensitive disease at HCT had 3-year OS and PFS of 56% and 43%, respectively, while those with chemorefractory disease had 3-year OS/PFS of 11% (see Fig. 2). Prior failed or tandem autologous HCT did not have a significant effect on outcome.

CONCLUSION: Non-myeloablative allogeneic HCT can produce sustained disease responses in pts with chemosensitive relapsed DLBCL. Patients with relapsed DLBCL who have failed or are ineligible for autologous HCT lack effective therapeutic options. Non-myeloablative allogeneic HCT is a promising treatment for such patients.

Figure Figure Figure Figure
Tacrolimus and Sirolimus without Methotrexate as Acute GVHD Prophylaxis after Matched Related Donor Reduced Intensity Conditioning (RIC) Stem Cell Transplantation (SCT).
Authors: Ho Vincent T. , Kim Haesook , Li Shuli , Cutler Corey , Koreth John , Armand Phillipe , Soiffer Robert J. , Antin Joseph H. , Alyea Edwin P.
Published: October 2019

We have previously shown that the combination of tacrolimus and sirolimus (TAC/SIR) without methotrexate (MTX) is a safe and effective regimen for GVHD prophylaxis after myeloablative SCT. However, TAC/SIR has not been investigated in the RIC setting. We hereby report a prospective phase II trial testing TAC/SIR as GVHD prophylaxis after RIC SCT from matched related donors (MRD). All patients received fludarabine (FLU) 30 mg/m2 IV and intravenous busulfan (BU) 0.8 mg/kg IV daily × 4 days (day -5 thru day -2) as conditioning, followed by transplantation of filgrastim mobilized peripheral blood stem cells. Filgrastim 5 mcg/kg SC QD was started on day +1 until neutrophil engraftment. Tacrolimus and sirolimus were started on day-3, and doses were adjusted to maintain target serum trough levels 5–10 ng/ml and 3–12 ng/ml, respectively. Twenty-six patients have been transplanted, with a median age of 52 years (range 29–64 yrs). Diagnoses include NHL (8), AML (6), HD (5), CLL/SLL (3), CML (2), MDS (1), MM (1). Median CD34+ cells infused was 8.06 × 106 cells/kg (range: 2.96–38.0 × 106 cells/kg). All patients had sustained hematologic engraftment. Only 4 (15%) patients developed neutropenia below ANC 500, and 6 (23%) had platelet counts below 20K. One patient had late graft failure at 7 months post transplant. Grade II-IV acute GVHD incidence was 20%, with grade III-IV incidence of 12%. No hepatic VOD or thrombotic microangiopathy was observed. Day +100 transplant related mortality (TRM) was 0%. The cumulative incidences of TRM and relapse at 1 year were 4% and 38%, respectively. The cumulative incidence of chronic GVHD at 1 year was 74%. A high level (>90%) of donor-derived hematopoiesis was achieved in 68% by 1 month post transplant. Median donor chimerism at between day 20–50 post transplant was 93.5% (range 32%-99%), and 94% (range 22%-100%) at day 85–115. With a median follow-up of 13.5 months among survivors (range 5.5–19 months), progression-free survival (PFS) and overall survival (OS) at 1 year were 58% and 79%, respectively. Compared to a historical cohort of 47 MRD transplant recipients treated with the same FLU/BU conditioning regimen and using TAC/SIR ± mini-MTX (5 mg/m2 IV day +1,3,6) as GVHD prophylaxis, there was no statistical difference between TAC/SIR vs. TAC/SIR/MTX in the incidence of grade II-IV acute GVHD (20% vs 11%, p= 0.48), cumulative incidence of relapse at year (38% vs. 57%, p= 0.25), 1-yr PFS (58% vs. 41%, p= 0.33), or 1-yr OS (79% vs. 73%, p= 0.79). These results demonstrate that omission of mini-MTX is permissible, and that the combination of TAC/SIR alone as GVHD prophylaxis following reduced intensity conditioning with FLU/BU is associated with low rates of acute GVHD, TRM, and high levels of donor chimerism after MRD SCT.
Myelofibrosis and Reduced Intensity Allogeneic Hematopoietic Stem Cell Transplantation (RISCT).
Authors: Popat Uday R. , Rondon Gabriela , Alousi Amin M. , Anderlini Paolo , Andersson Borje S. , de Lima Marcos J. , Giralt Sergio A. , Hosing Chitra M. , Kebriaei Partow , Jones Roy , Nieto Yago , Shpall Elizabeth J. , Verstovesek Srdan , Qazilbash Muzaffar H. , Champlin Richard
Published: October 2019

Background: Reduced intensity allogeneic transplantation was developed to harness graft versus leukemia immune effect to treat older patients and patients with comorbidities who are not eligible for conventional myeloablative transplant. Limited but encouraging data are available on outcome of patients with myelofibrosis undergoing this therapy. We therefore sought to prospectively study the safety and efficacy of reduced intensity HCT in patients with myelofibrosis.

Methods: Patients with intermediate or high risk MF were eligible if they had adequate organ function and at least 9/10 matched related or unrelated donor. Patients were conditioned with Fludarabine 40mg/m2 × 4 (days −5, −4, −3 −2) and Busulfan 130mg/m2 × 2 (day −3,−2). Thymoglobulin 2.5 mg/kg × 3 (day −3,−2 and −1) was additionally given to patients receiving unrelated donor graft. Tacrolimus and mini Methotrexate were used as graft versus host disease prophylaxis. All patients received standard supportive care.

Results: Twelve consecutive patients with myelofibrosis were enrolled between 1/2006 and 7/2007. There were 6 males and 6 female with a median age of 59 (range 40–65). Four patients had primary MF, 3 post PV MF and 5 Post ET MF. Based on Lile criteria, 8 patients had intermediate risk disease and 4 had high risk disease. Eight patients had circulating blasts (1%-8%). Six patients had splenectomy prior to transplant; median spleen size was 15 cms (range 5–28cms) below the costal margin in the remaining 6 patients. Seven patients had mutated Jak 2. Median peripheral blood CD 34 count was 77/μl (range 2–3770/μl). Karyotype was abnormal in 6 patients and diploid in 6. Donors were siblings for 3 patients, matched unrelated for 6, and one antigen or allele mismatched unrelated for 3 patient. All but 1 patient received peripheral blood stem cells. All patients engrafted with a median time to neutrophil engraftment of 13 days (0–27) days) and a median time to platelet engraftment of 14 (0–74) days. Day 100 non relapse mortality was 0%. One patient developed grade 3 acute graft versus host disease (GVHD) and died. One patient developed limited chronic GVHD. All Jak 2 positive patients achieved molecular remission post transplant with negative JaK 2. Three patients relapsed: 2 in blast phase and 1 in chronic phase. With a median follow up of 177 (29–563) days, six month overall survival is 87%.

Conclusion: RISCT induces molecular remission with very low non relapse mortality in older patients with myelofibrosis.
Bax Degradation Activity Is a Novel Poor Prognosis Marker of Chronic Lymphocytic Leukemia.
Authors: Agrawal Samir G. , Liu Feng-Ting , Wiseman Catherine , Shirali Sima , Liu Hongxiang , Lillington Debra , Du Ming Qing , Court Denise Syndercombe , Newland Adrian C. , Gribben John G. , Jia Li
Published: October 2019

B-cell chronic lymphocytic leukemia (CLL) is a highly heterogeneous disease, with prognosis varying from a normal lifespan to death early after diagnosis. A large number of prognostic markers are now available in CLL. Better understanding of the mechanisms underlying the association of these markers with poor outcomes may guide their use in the clinic, as well as identify new therapeutic targets. Bax is an essential pro-apoptotic protein and decreased levels in malignant cells lead to resistance to apoptosis and chemotherapy. Using a Bax degradation activity (BDA) assay, CLL cells were found to show variable Bax instability. However, BDA did not correlate with Bax protein levels, BDA positive and negative cases each showing high and low baseline Bax levels. We looked at a cohort of patients with CLL and compared Bax degradation activity with established prognostic markers: Ig VH mutation status, Zap-70, CD38, cytogenetics, as well as clinical stage and lymphocyte doubling time (LDT). BDA positive cases showed a marked accumulation of poor prognostic markers - unmutated immunoglobulin heavy chain variable genes (VH), ZAP-70 and CD38 positivity, 11q22 and 17p13 deletion, and short lymphocyte doubling time. Patients with BDA positive cells had a shorter median overall survival (OS) (126 months v. not reached, p = 0.011) and shorter time to first treatment (16 v. 156 months, p = 0.029) than BDA negative cases; while patients with dual BDA and ZAP-70 positivity had a median OS of 84 months (p = 0.012). The BDA assay reflects the differential intrinsic ubiquitin/proteasome activity of CLL cells and measures dynamic changes in Bax protein levels, as opposed to single time point measurements. Mechanistically, Bax protein instability may represent a final common pathway for disparate prognostic markers, as well as being itself an indicator of poor prognosis.