bluuebunny/crossref_metadata_2025 huggingface dataset:
Therapeutic Compliance and Efficacy of Dose-Dense R-CHOP-14 Immunochemotherapy Plus Pegfilgrastim for First-Line Treatment of Low Risk Diffuse Large B-Cell Lymphoma Patients: Preliminary Results of An Open-Label Clinical Trial in Spain
Authors: Gonzalez-Barca Eva , Canales Miguel , Salar Antonio , Ferrer Secundino , Bargay Joan , Grande S. , Gardella Santiago , Vidal María Jesús , Oriol Albert , López A. , García J. , Bello Jl , Tomás Fj , Briones Javier , Caballero Dolores
Published: April 2020

Diffuse large B-cell lymphoma (DLBCL) is the most prevalent form of non-Hodgkin lymphoma. Three-weekly R-CHOP has become the standard treatment for DLBCL. Dose-dense immunochemotherapy could improve the efficacy of treatment; however these regimens are worse tolerated due to mielotoxicity. We are conducting a prospective, open-label, single arm clinical trial of treatment with dose-dense R-CHOP supported by pegfilgrastim in patients with DLBCL older than 65 with IPI 0–5 or younger than 65 with IPI 0–2. The aim of this sub-analysis was to evaluate the efficacy and treatment compliance in patients with low risk DLBCL (IPI 0–2). Patients received R-CHOP administered every 14 days followed by pegfilgrastim (6 mg per cycle) on day 2. Patients with tumor masses > 10 cm were allowed to receive radiotherapy with 30 Gy, and those at risk of central nervous system infiltration receive prophylactic lyposomal cytarabine. Fifty-eight patients with IPI 0–2 were included in this analysis, median age was 58 years old (range 18–82), 22 (38%) patients were older than 65 yr, and 32 (53%) were male. Characteristics of the patients at diagnosis were as follows: stage III–IV: 29 (50%), bulky disease: 13 (22%), extra nodal involvement: 34 (59%), ³2 extranodal sites: 6 (10%), B symptoms: 10 (17%), ECOG 0–1: 55 (95%), elevated LDH 21 (36%), elevated b2microglobulin: 15 (26%), IPI 0–1 28 (48%). Fifty-four patients completed 6 cycles of treatment and 4 patients were dropped out, 1 due to progression and 3 due to exitus: 1 respiratory insufficiency, 1 bilateral pneumonia and 1 septic shock. Overall, 335 chemotherapy cycles were administered and 23 (6.9%) were delayed due to: neutropenia (3), febrile neutropenia (4), hepatotoxicity (1), fever or infection without neutropenia (11) and other adverse events (4). Doses of myelotoxic drugs were reduced in 8 (2.4%) cycles (3 patients). Most cycles (91.9%) were administered as scheduled. Serious adverse events were reported in 23 (39.7%) patients. Among them, febrile neutropenia episodes were observed in 10 patients (17.3%). Out of 54 evaluated patients for efficacy after 130-days follow-up, 45 (83%) achieved complete remission, 5 (9%) partial remission, 3 (5.5%) stable disease and 1 (1.8%) progressed. In an intention-to-treat analysis among the 58 patients, 46 (79.3%) patients achieved CR, 6 (10.3%) PR and 2 (3.4%) were not evaluable. High remission rates suggest that dose-dense immunochemotherapy with pegfilgrastim support is an efficacious treatment for DLBCL in low risk patients. The regimen is well tolerated, treatment compliance is high and most of cycles were administered as scheduled.
Pegfilgrastim Use and Risk of Bleomycin Induced Pulmonary Toxicity in Hodgkin Lymphoma
Authors: Janakiram Murali , O’Brien Timothy E.
Published: April 2020

Background Filgrastim may increase the risk of bleomycin induced pulmonary toxicity (BPT) in Hodgkin lymphoma patients, particularly those >age 40 [Martin WG, J Clin Onc, 2005; 23(30): 7614]. It is not clear if pegylated filgrastim, with a longer 1/2 life and delayed onset of action, also increases this risk.

Methods: 38 cases of Hodgkin lymphoma treated with a bleomycin containing regimen were included in this analysis. 29 of these pts (Group A) also received pegfilgrastim and 9 pts (Group B) did not receive any growth factor support. All of the patients who received pegfilgrastim had it administered the day after chemotherapy and in all cases it was given to maintain dose intensity and prevent treatment delays. BPT was defined by the presence of pulmonary symptoms, interstitial infiltrates on chest x ray or computed tomography and the absence of infection.

Results: Grp A: 60% male, 40% female, mean age 39 (19 to 73), stage-I 2/29, II -14/29, III -5/29, IV-8/29; subtype- NS 20, MC 5, LP 3, LD 1. 27/29 received full dose ABVD (which included 10 units/m2 of bleomycin q 2weeks); one received ABD and one received full dose ABVD then had bleomycin held after 3 cycles. Mean number of cycles= 6 (4–8), median duration of treatment =160 days (71 to 231 days). Average ANC during treatment: 6590/mm3 (790 to 17273). Total # of bleomycin treatments: 274. Total # pegfilgrastim treatments: 203. Only 1/29 pts (age 44) developed BPT. There were no serious infections. Response to chemo: 22/29 CR, 5/29 PR, data not available in 2/29. At a median follow up of 18 months (range 4 to 48) 2/29 had relapsed. Grp B: 9 patients; mean age 39 (22 to 51), stage- II: 5/9; III: 4/9; subtype- NS 7, LP 2. All patients received full dose ABVD; ave # cycles= 6 (range 4–8), median duration of treatment= 145 days (30–220 days); average ANC during treatment: 2721/mm3 (2101 to 3535/mm3). Total # bleomycin treatments= 80. 1/9 pts (age 42) developed BPT. Response to chemo: 7/9 CR; 1 PR, 1 w/o response data. At a median follow up of 39 months (range 4–71), 2/9 relapsed.

Conclusions: In this small series, pegfilgrastim use with ABVD chemotherapy did not lead to a high risk of BPT in Hodgkin lymphoma, with only 1/29 patients (which included 203 treatments with both pegfilgrastim and bleomycin) developing BPT, as compared with 1/9 patients treated with ABVD and no pegfilgrastim. Both of the patients who developed BPT were >age 40. Larger studies will need to be performed to confirm our findings.
Result of Paediatric Non Hodgkin’s Lymphoma with Intensified Short Duration Chemotherapy : A Result from Eastern India
Authors: Mukhopadhyay Ashis , Adde Melissa , Magrath Ian , Mukhopadhyay Soma
Published: April 2020

Background: The Non-Hodgkin’s Lymphomas (NHL) in childhood are usually high grade and diffuse histology. They require intensified short duration chemotherapy in contrast to adult NHL. The aim of our study was to observe the result of aggressive short duration chemotherapy in paediatric NHL.

Materials & Methods: We included consecutive 160 paediatric NHL patients in paediatric haemato-oncology department of Netaji Subhash Chandra Bose Cancer Research Institute during period from June 1996 to December 2007. The inclusion criteria were patients less than 25 yrs of age with a diagnosis of NHL and Patients are clinically staged according to the St. Jude’s (Murphy’s) classification. Patients with > 25% blasts in the bone marrow were treated as leukemia and excluded from the study. Each patient received 3 cycles A and 3 cycles B of MCP842 protocol of INCTR. Response was assessed at the completion of 2cycles of chemotherapy (1 each of A and B).

Result: A total of 160 previously untreated patients were entered in this study. The age range was 1 to 25 yrs (median 13.5). Fourty Eight (48) patients had Lymphoblastic Lymphoma (LL), 64 patients (40%) had Burkitt Lymphoma, 40 (25%) diffuse Large B Cell Lymphoma (DLCL) and 8 (5%) had Anaplastic Large Cell. The abdomen was the most common site (56%) of involvement followed by the mediastinum (15.62%). One hundred and Thirty four (134)(83.75%) patients achieved complete response after 2 cycles of therapy, 14 patients (8.75%) achieved partial response and 6 (3.75%) had no response, 6 (3.75%) were not evaluable. With median follow up of 4 & 1/2 yaers (range 7 months – 10 years) a total of 38 (23.75%)patients (19 LL, 13 Burkitt Lymphoma, 5 DLCL and 1 ALCL) have died. The causes of death were progressive disease in 26 (16.25%), infection in 6 (3.75%), tumour lysis in 4 (2.5%), hepatitis in 1 (0.63%), and unknown 1 (0.63%). One hundred and twenty two (122) patients (76.25%) are alive and in complete remission. The patients tolerated chemotherapy well. Grade IV febrile Neutropenia was seen in 34 patients (21.25%).

Conclusion: Result of MCP 842 was promising and we will continue the protocol in future.
Rituximab + IEV/MINE Second Line Approach in Relapsed/Refractory Non Hodgkin Lymphoma (NHL): Efficacy and Safety
Authors: Bongarzoni Velia , Anaclerico Barbara , Borza Paola Anticoli , Cedrone Michele , Chierichini Anna , Fenu Susanna , Ronci Benedetto , Vittori Mariangela , Cantonetti Maria , Petti Maria Concetta , Annino Luciana
Published: April 2020

Background. Relapsed/refractory high grade NHL is a very aggressive pathology characterised by a poor prognosis. High dose therapy followed by peripheral blood stem cell (PBSC) rescue is to date the gold standard therapy. The results of high dose therapy are influenced by remission status after II line therapy as patients who undergo transplantation in complete response have better progression free survival with respect to those in partial response. While Rituximab+chemotherapy has become the approach of choice in 1st line-therapy, this type of schedule has not yet been largely applied in 2nd line therapy. In the aim of testing the efficacy of 2nd line-combined immuno-chemotherapy we designed a protocol including Rituximab + IEV/MINE.

Methods. The planned treatment consists of 3 cycles of IEV or MINE (>65 y patients), plus Rituximab (375 mg/sqm) given on day +1 and on day +14 every cycle. G-CSF was administered from day 7 to ANC recovery and was continued until the CD 34+ collection after the third cycle. From April 2002 to July 2008 30 consecutive patients – 19 males and 11 females, median age 57 y (range 21–73 y) with refractory (12) and relapsed (18) Diffuse Large B Cell Lymphoma (DLBCL) entered the study. According to IPI score 20/30 patients were intermediatehigh risk; 21/30 had extranodal disease.

Results. 27 patients completed treatment and are evaluable; 3 pts are not evaluable because of lethal infection after Ist cicle (2pts) and lost to follow up (1pt). 21/27 (78%) were responders: 12 (44%) achieved CR, 9 (33%) PR, while 6 progressive disease (PD) were withdrawn. Hematological toxicity including WHO grade III or IV neutropenia, anemia and thrombocytopenia was recorded in 19, 8 and 14 pts, respectively. Target CD 34 harvest was reached in 19/27 (70%) pts with CD 34+ median value 7,03 × 106/Kg. Among the responders 4 pts failed CD 34 harvest. 17/27 (63%) patients underwent transplantion, 11 autologous stem cell transplantation (ASCT) and 6 (>60 y) reduced intensity allogeneic-SCT. Of transplanted patients 4 (2 in CR post Mini Allo-SCT, 2 in relapse post ASCT) died 6, 2,10 and 10 months after transplantation, respectively. The remaining 13 (4 mini-alloSCT and 9 ASCT) are alive in CR which is lasting for 23 months median time (11–73). As of July 2008 15/27 (55%) patients are alive: 14 CR and 1 PR. Median follow up post R-IEV/MINE is 28 months (range 12–77), PFS was 38% and median OS is 32 months (range 7–90).

Conclusions. Our experience, if limited to a small series of patients, shows that the addition of Rituximab to second line chemotherapy:

increases response rate even in adverse IPI score cases, allows to obtain a good CD 34 harvest, represents an effective in vivo purging agent.
Antiproliferative Effects of Lenalidomide in Combination with Chemotherapeutic Agents in Lymphoma Cell Lines
Authors: Gandhi Anita K , Kang Jian , Capone Lori , Zhang Ling-Hua , Bartlett Blake , Pietronegro Dennis , Ervin-Haynes Annette , Schafer Peter H
Published: April 2020

Introduction: Lenalidomide (Revlimid®) is approved for the treatment of patients with myelodysplastic syndromes with a del(5q), and in combination with dexamethasone for previously treated multiple myeloma. It is currently being evaluated in lymphomas, as monotherapy or combination therapy. This study aims to identify beneficial combinations of lenalidomide with established or potential chemotherapeutic agents for lymphoma therapy.

Methods: Lenalidomide-sensitive lymphoma cell lines were selected for in vitro tumor cell proliferation studies. These included mantle cell lymphoma (MCL) lines Rec-1 and Jeko- 1, diffuse large-B-cell lymphoma (DLBCL) cell line Farage, the follicular lymphoma (FL) cell line DOHH2, and the chromosome 5-deleted Burkitt’s lymphoma cell line Namalwa CSN.70. Lenalidomide was tested with other therapeutic agents on cells incubated 72 hours and assayed for proliferation using 3H-thymidine incorporation. Twenty-one different therapeutic agents were tested in combination with lenalidomide, at concentrations ranging from 1 pM to 100 μM, depending upon the sensitivity of the cells to the particular agent.

Results: In MCL cells, lenalidomide was at least partially additive with mitoxantrone, rituximab, temsirolimus, bortezomib and azacytidine, and up to partially additive with doxorubicin. Results were similar in Rec-1 and Jeko-1 MCL cells. Lenalidomide was fully additive in the DLBCL cells with gemcitabine, melphalan, dexamethasone, carboplatin, and oxaliplatin, partially additive with cisplatin, mitoxantrone, vincristine, azacytidine, doxorubicin, carmustine, rituximab, etoposide and prednisolone, and non-additive with cytarabine. Synergy with lenalidomide was observed in the FL cells with rituximab, while partially additive with bendamustine and fludarabine, and non-additive with vincristine. In the Burkitt’s lymphoma cells, lenalidomide was synergistic with dexamethasone, rapamycin, and UCN-01. It was partially additive with prednisone and etoposide, and non-additive with other agents tested.

Conclusion: Among the tested lymphoma cell lines, lenalidomide combined synergistically with rituximab, dexamethasone, rapamycin, and UCN-01, and additively with mitoxantrone, temsirolimus, bortezomib, azacitidine, gemcitabine, melphalan, dexamethasone, carboplatin, and oxaliplatin. Thus, these results demonstrate a beneficial antineoplastic pharmacological interaction between lenalidomide and several therapeutic agents including monoclonal antibodies, corticosteroids, mTOR inhibitors, kinase inhibitors, proteasome inhibitors, DNA methyltransferase inhibitors, nucleoside analogs, alkylating agents, and topoisiomerase inhibitors in the treatment of lymphoma.
Pharmacodynamic Effects of Administration of Maytansine Conjugated Anti-CD22 Monoclonal Antibodies to Cynomolgus Monkeys
Authors: Fuh Franklin , Fuji Reina , Poon Kirsten A , Li Dongwei , David Clarissa , Nguyen Quyen , Howell Kathy , Beyer Joseph , Fielder Paul , Manning William , Berry Karen , Ramakrishnan Vanitha , Polson Andrew G. , Ebens Allen , Prabhu Saileta , Williams Marna
Published: April 2020

CD22 is a B cell-specific glycoprotein expressed on the cell surface of all mature B cells. A candidate therapeutic anti-CD22 antibody, 10F4v3, was conjugated to the anti-mitotic agent maytansine (10F4v3-DM1). DM1 disrupts cellular mitosis through inhibition of tubulin polymerization when internalized into cells. The anti-CD22 DM1 conjugate was shown to have significant potency in preclinical efficacy models of NHL. In order to further characterize this antibody-drug conjugate in preclinical studies, we first evaluated the binding characteristics of the 10F4v3 to peripheral blood B cells from various geographical sources of cynomolgus monkeys. 10F4v3 bound to peripheral blood B cells from all cynomolgus monkeys of Indonesian and Mauritian origins, but displayed only limited binding to cynomolgus monkeys of Chinese and Cambodian origins.

Therefore, further pre-clinical evaluation of 10F4v3-DM1 was conducted in Indonesian cynomolgus monkeys to examine the safety, pharmacokinetic, and pharmacodynamic effects in monkeys dosed at 10, 20, and 30 mg/kg (2000, 4000, and 6000 mg/m2 DM1). Pharmacodynamic assessments of peripheral blood and lymphoid tissues included examination of B cells, B cell subsets, CD4+ T cells, CD8+ T cells, and CD3−CD20− (NK) cells. B cell subsets included CD20+, CD20+CD21+, CD20+CD21−, CD20+CD21+CD27+, CD20+CD21+CD27−, and CD20+CD21high lymphocytes which are phenotypically similar to human B cells, mature B cells, germinal center B cells, memory B cells, naïve B cells, and marginal zone B cells, respectively.

B cells and B cell subsets were substantially depleted in peripheral blood at all doses, with no apparent dose-dependent effects. In lymphoid tissue, B cells were also depleted, with substantial depletion of CD20+CD21− and CD20+CD21high B cell subsets in spleen and bone marrow.

Based on the nonclinical data, 10F4v3-DM1 exhibits an encouraging pharmacodynamic profile that supports clinical development for the potential treatment of non-Hodgkin’s lymphoma.
Treatment of Posttransplant Lymphoproliferative Disorder Presenting as Protracted Infectious Mononucleosis with Rituximab
Authors: Chiang Alan Kwok Shing
Published: April 2020

Two young children who received living related-donor orthotopic liver transplantation for congenital biliary atresia developed persistent and progressive symptoms of fever, hepatosplenomegaly, upper airway obstruction, swollen eyes and face, malaise and cervical lymphadenopathy consistent with infectious mononucleosis within one year post-transplantation. Significant anemia, elevated liver transaminases and hypergammaglobulinaemia were observed. Serological assays did not detect any antibodies to Epstein-Barr virus (EBV) pre-transplantation indicating seronegative status. Both patients showed recent seroconversion to EBV (viral capsid antigen IgM and IgG positive but EBV nuclear antigen negative) at the peak of the clinical symptoms. The diagnosis of primary EBV-driven posttransplant lymphoproliferative disorder (PTLD) was established. Lowering or withdrawal of tacrolimus for a period of 2–3 weeks did not lead to any discernible clinical improvement. Both children were treated with four weekly doses of rituximab of 375 mg/m2/dose. They developed infusion reaction of fever, vomiting and unstable blood pressure within half hour of first dose of rituximab requiring stoppage of the drug administration. Subsequent doses were tolerated much better without adverse events. Clinical improvement was gradually observed over the next few weeks with resolution of fever, upper airway obstruction, decreasing liver transaminases and improvement of general condition. Complete resolution of infectious mononucleosis and normalization of hematological and biochemical parameters was achieved over a period of 6 months. No recurrence of PTLD was observed up to one year after treatment with rituximab.
Prognostic Significance of hOCT1 and ABCG2 Expression in Chronic Myeloid Leukemia Patients Treated with Imatinib
Authors: Kim Yeo-Kyeoung , Kim Hee-Nam , Yu Ri , Lee Il-Kwon , Ahn Jae-Sook , Yang Deok-Hwan , Lee Je-Jung , Kim Hyeoung-Joon
Published: April 2020

Background: Imatinib mesylate is a substrate for both the drug influx transporter human Organic Cation Transporter 1 (hOCT1; SLCA22) and the efflux transporter ATP-binding cassette transporters (ABCG2; BCRP). The correlation between the expression of these two transporters of imatinib and the clinical outcome of imatinib in chronic myeloid leukemia (CML) patients remain clarified.

Methods: hOCT-1 and ABCG2 mRNA levels were determined by performing real-time polymerase chain reaction assays on 83 BM samples obtained at initial diagnosis from the patients with chronic phase CML treated with imatinib (started at 400mg/day).

Results: Of patients with higher hOCT-1 mRNA expression than median value (≥ 34.53 a.u.) at diagnosis, 59.5% achieved complete cytogenetic response (CCyR) at 6 months, whereas of patients with no more than a median hOCT-1 expression, 33.3% could achieve CCyR at 6 months (p=0.035). Furthermore, patients with low hOCT-1 showed higher rate of suboptimal response [failed to achieve complete hematologic response (CHR) by 3 months, major cytogenetic response (MCyR) by 6 months, CCyR by 12 months, and major molecular response (MMR) by 18 months] than those with high hOCT-1 expression (55.9% vs. 33.3%. p=0.048). There was no statistical significance in progression-free survival (PFS) or overall survival (OS) according to the hOCT-1 expression levels. Patients with low ABCG2 expression (< 13.53 a.u.) showed higher MMR rate at 18 months (57.1% vs. 35.5 %, p=0.09) and longer PFS (3 yr PFS, 87.2% vs. 64.1%, p=0.07) than those with high ABCG2 expression but there was no statistical significance. The time to achieving MMR was significantly shorter in patients with low ABCG2 expression than in those with high ABCG2 expression (6.9 ± 2.1 ms vs. 12.2 ± 4.2 ms, p=0.09). The patients with high hOCT-1 expression showed shorter time to achieving MMR, but there was no statistical significance (8.1 ± 1.9 vs. 10.5 ± 1.7, p=0.50).

Conclusions: This study demonstrates that pretreatment assay of hOCT-1 and ABCG2 expression may help us to identify the treatment outcome in the CML patients treated with imatinib.
Modification of IGF1R Expression on Malignant Lymphoid Cells by a Proteasome Inhibitor
Authors: Terui Yasuhito , Mishima Yuji , Mishima Yuko , Hatake Kiyohiko
Published: April 2020

Purpose: The type 1 IGF receptor (IGF1R) is a transmembrane tyrosine kinase, which is overexpressed by tumors, and mediates proliferation and apoptosis protection for tumor cells. IGF signaling affects hypoxia signaling, protease secretion, tumor cell motility, and adhesion. ProIGF1R is cleaved into alpha and beta chains by processing, and their heterodyne moves onto plasma membrane. Folding after transfer to Golgi apparatus modifies the beta chain, and it is also known to degradate by ubiquitin-proteasome system. Bortezomib is a proteasome inhibitor for multiple myeloma, and expression of chaperon proteins such as IGF1R could be increased by bortezomib. Here, we examined and analyzed expression of IGF1R on malignant lymphoid cells, and discuss possibility of IGF1R as a target in the treatment of lymphoid malignancies.

Methods: IGF1R expression of malignant T cell lymphoid cell line Jurkat, myeloma cell line IM-9, and bortezomibresistant IM-9 was examined by flow cytometric and western blot analyses. The cell lines were cultured with IGF1 and IGF2 with or without bortezomib, and the growth and cell death were observed.

Results: The IGF1R expression on Jurkat cells was weak by flow cytometry, and western blot analysis revealed that IGF1R beta expression was quite low in spite of higher expression of ProIGF1R and IGF1R alpha. The IGF1R expression on bortezomib-resistant cells was stronger than that on parent IM-9 cells by flow cytometric and western blot analyses. While IGF1 and IGF2 did not give an advantage for the growth of lymphoid cells significantly without bortezomib, there was an advantage for survival of them under exposure to bortezomib.

Conclusion: Difference of IGF1R expression was observed among several malignant lymphoid cells. Cell surface modification by drugs such as bortezomib will make lymphoid cells increased IGF1R expression, and IGF1R inhibition will be a promising way for the treatment of malignant lymphoid cells.
Daunorubicin Loaded Magnetic Nanoparticles of Fe3O4 Greatly Enhance the Responses to Chemotherapy and Induce Apoptosis of Multidrug-Resistant K562 Cells in a Human-Nude Mice Xenograft Leukemia Model
Authors: Chen Bao-An , Lai Bin-bin , Cheng Jian , Gao Feng , Xu Wen-lin , Shen Hui-lin , Ding Jia-hua , Gao Chong , Sun Yun-yu , Wang Jun , Zhao Gang , Song Hui-hui , Bao Wen , Sun Xinchen , Cheng Hong-yan , Deng Yu-xia , Li Guo-hong , Liu Li-jie , Chen Wen-ji , Shao Ze-ye , Xia Guo-hua , Wang Jue-qiong , Yuan Peng , Pei Xiao-ping , Chen Ning-na , Li Qin-ning , Wang Xue-mei
Published: April 2020

Object: To the effect of Fe3O4-magnetic nanoparticle loaded with DNR on multidrugresistant K562 cells in vivo.

Methods: K562-n and its MDR counterpart K562-n/VCR cell were inoculated subcutaneously into both sides of the back of nude mice (5×106 cells/each) to establish a human leukemia xenograft model. The mice were randomly divided into group A receiving normal saline every other day for 20days, group B receiving DNR every other day for 20days, group C receiving Fe3O4-magnetic nanoparticle every other day for 20days, group D receiving Fe3O4-magnetic nanoparticle loaded with DNR every other day for 20days, and group E receiving Fe3O4-magnetic nanoparticle containing DNR every other day for 20days with a magnetic field built on the surface of the tumor tissue. The tumor volume was measured on the day 1, 5, 9, 13, 17 and 21d after the first treatment. Tumor tissues were isolated for examination of the expression of mdr-1, bcl-2, bax and caspase-3 by reverse transcription polymerase chain reaction and Western blotting.

Results: For K562-n/VCR tumor, the tumor volume was markedly lower in groups D and E than in groups A, B and C (group D or E vs group A, B or C, P < 0.05). The transcription of mdr-1 and Bcl-2 gene was significantly lower in groups D and E than in groups A, B and C (group D or E vs group A, B or C, P < 0.05). So did the protein expression of Bcl-2. However, there were no differences among these groups about the protein expression of P-gp. The protein and mRNA expressions of Bax and Caspase-3 in groups D and E were increased significantly compared with groups A, B and C (group D or E vs group A, B or C, P < 0.05). The tumor volume of K562-n was markedly lower in groups C, D and E than in groups A and B (group C, D or E vs group A or B, P < 0.05).

Conclusion: In conclusion, DNR loaded Fe3O4-magnetic nanoparticles can suppress the growth and induce apoptosis further on the MDR K562-n/VCR tumor in vivo compared to DNR alone but not on the K562-n tumor. The external magnetic field failed to improve the antitumor effect.
Pharmacogenetic Outcomes of Pediatric Acute Lymphoblastic Leukemia in Korea
Authors: Kang Hyoung Jin , Kim Hyery , Seo Young Jin , Jang Mi Kyung , Oh Yongtaek , Han Byoung-Don , Park Kyung Duk , Kim Chul Woo , Shin Hee Young , Ahn Hyo Seop
Published: April 2020

Treatment result of pediatric acute lymphoblastic leukemia (ALL) has been markedly improved, but treatment related toxicities and relapse are still remaining problems. Genetic polymorphism is an important factor in the effectiveness and toxicity of anti-leukemic drugs and pharmacogenetics are beginning to emerge as useful research filed to solve those problems. In our experience in the treatment of ALL, many Korean patients could not tolerate full dosages of Western protocols. To make basis for individualized therapy with pharmacogenetics, we analyzed major genes implicated in the treatment of ALL. Fourteen genes of total 103 patients with ALL were analyzed with TotalPlex gene amplification methods (Mol Cell Probes.2008. 22: 193). Among the drug related genes (percentage of mutant type) including CYP3A4*1B (0%), CYP3A5*3 (0%), GSTP1 (22.3%), GSTM1 (20.4%), GSTT1 (16.5%), MDR1 exon 21 (76.3%), MDR1 exon 26 (61.2%), MTHFR (64.1%), MTHFR 1298 (29.1%), NR3C1 1088 (0%), RFC 80 (79.6%), TPMT combined genotype (7%), VDR intron 8 (10.7%), VDR FokI (67%), incidence of mutant was higher in GSTM1 deletion, lower in MTHFR 1298, TYMS enhancer repeat, and VDR intron 8 comparing with the data of Western whites (Rocha J.C. et al. Blood 2005). As we had modified the dose of anti-leukemic agents depending on the toxicity during the treatment, we analyzed the relationship between the dose percent of actually administered dose and the distribution of each mutant to find out polymorphisms affecting toxicities of chemotherapeutic drugs. The mean dose percent of mercaptopurine was lower in patients with variant TPMT then those with wild type (33.2% vs. 53.5%, P=0.04), but there was no polymorphism that influenced the dose percent of methotrexate, daunorubicin, doxorubicin, and L-asparaginase. There was no significant difference in the incidence of genotypes between risk groups and individual mutant did not affect long term survival and relapse in Korean patients with ALL. In conclusion, we found some difference in the incidence of mutant genotypes related to the pharmacogenetics of ALL between Korean and Western whites, but there was no individual genetic polymorphism that affect on the treatment outcome. We expect more extensive researches about pharmacogenetics of Korean to establish the basis for individualized therapy with the consideration of ethnical difference.
Mechanism Research of Reversal of Multidrug Resistance by the Application of 5-Bromotetrandrine and Magnetic Nanoparticle of Fe3O4 Combined with Daunorubicin in a Human-Nude Mice Xenograft Model
Authors: Chen Bao-An , Wu Ya-nan , Cheng Jian , Gao Feng , Xu Wen-lin , Shen Hui-lin , Ding Jia-hua , Gao Chong , Sun Yun-Yu , Wang Jun , Zhao Gang , Song Hui-hui , Bao Wen , Sun Xin-chen , Cheng Hong-yan , Deng Yu-xia , Li Guo-hong , Liu Li-jie , Chen Wen-ji , Wang Jue-qiong , Yuan Peng , Pei Xiao-ping , Chen Ning-na , Wang Xue-mei
Published: April 2020

Objective: To establish the xenograft leukemia model with stable multiple drug resistance in nude mice; to investigate the reversal effect of 5-Bromotetrandrine and Magnetic nanoparticle of Fe3O4 combined with DNR in vivo and to search for the possible reversal mechanisms.

Methods: K562 and K562/A02 cells were respectively inoculated subcutaneously into back of athymic nude mice (1×107 cells/each) to establish the xenograft models. The tumor formation was evaluated by animal ultrasonic inspection. Tumors-bearing nude mice were assigned randomly to five groups which were treated with NS (A group); DNR 1mg/kg (B group); nanoparticle of Fe3O4 combined with DNR 0.63mg/kg(C group): 5-BrTet 2.5mg/kg combined with DNR(D group); 5-Bromotetrandrine 2.5mg/kg and Magnetic nanoparticle of Fe3O4 combined with DNR 0.63mg/kg(E group) respectively. The incidence of tumor formation, growth characteristics, weight and volume of tumor were observed. The histopathologic examination of tumors and organs were detected. For resistant tumors, the protein levels of P-glycoprotein (P-gp) were detected by Western blot.

Results: The tumor incidence was 100% in the nude mice inoculated with either K562 or K562/A02 cells. In 6 to 9 days,the tumors reached a volume of more than 1 00 mm3. In vivo, MTT assay showed K562/A02 tumor maintained the drug resistance. For K562 cells xenograft tumors, there were no apparent differences in tumor suppression effect between the B AC AD AE group. For K562/A02 cells xenograft tumors, 5-BrTet and Magnetic nanoparticle of Fe3O4 combined with DNR significantly suppressed growth of tumor: the inhibition rate was 62.76% while DNR alone be used, the inhibition rate was 3.68%. Pathologic examination of resistant tumors showed the tumors necrosis obviously in E group. Application of 5-BrTet and Magnetic nanoparticle of Fe3O4 inhibited the overexpression of P-gp.

Conclusion: The xenograft leukemia nude mice model was maintain the multiple drug resistance. 5-Bromotetrandrine and Magnetic nanoparticle of Fe3O4 combined with DNR had a significant tumor-suppressing effect on MDR leukemia cells xenograft model.
Effect of Tetrandrine and Toremifene on the Reversion of Multidrug Resistance of K562/A02 Cell Line
Authors: Chen Bao-An , Zhao Qiu-xia , Cheng Jian , Gao Feng , Xu Wen-lin , Shen Hui-lin , Ding Jia-hua , Gao Chong , Sun Yun-Yu , Wang Jun , Zhao Gang , Song Hui-hui , Bao Wen , Sun Xin-chen , Cheng Hong-yan , Deng Yu-xia , Li Guo-hong , Liu Li-jie , Chen Wen-ji , Wang Jue-qiong , Yuan Peng , Pei Xiao-ping , Chen Ning-na , Wang Xue-mei
Published: April 2020

Objective: To study the reversal effect of Tetrandrine(Tet) and the estrogen-receptor inhibitor,toremifene(Tor),on multidrug resistance cell line K562/A02 and to investigate the reversal mechanism of this combination.

Methods: ADM accumulation and the apoptosis percentage of K562 and K562/A02 cells were analyzed by fluorospectrophotometry, respectively. The protein levels of P-glycoprotein (P-gp) were detected by fluorospectrophotometry. The mRNA levels of mdr1 and Survivin were determined by RT-PCR.

Results: The IC50 of ADM for K562/A02 and K562 cells were 57.43±4.55mg/L and 1.16±0.05mg/L respectively. Pretreating K562/A02 cells with toremifene(2.5μmol/L) or Tet(1μmol/L) for 72 hours partially restored the sensitivity of K562/A02 cells to ADM (IC50 were 20.74±1.62mg/L and 14.12±1.20mg/L respectively) but had no effect on K562 cells; IC50 of combined tetrandrine and toremifene was 9.14±1.03mg/L;K562/A02 showed apoptotic characteristics after treated with tetrandrine, toremifene (alone or combination);tetrandrine and toremifene (alone or combination) elevated the intracellular ADM accumulation in K562/A02; P-gp, mdr1 and Survivin mRNA were down regulated.

Conclusion:

Tetrandrine, toremifene (alone or combination) showed significant MDR reversal activity in vitro The reversal activity may be related to the inhibition of P-gp overexpression and down regulation the expression of mdr1 and Survivin mRNA to increase the intracellular accumulation of anticancer drugs, which lead to more K562/A02 cells apoptosis; Multidrug resistance (MDR) can be partially reversed by Tet or Tor of which the combination shows a great synergistic reversal effect.
Variation of p21, P-Gp Expression in K562/A02 Cell Line with Tetrandrine
Authors: Chen Bao-An , Li Jing , Cheng Jian , Gao Feng , Xu Wen-lin , Shen Hui-lin , Ding Jia-hua , Gao Chong , Sun Yun-Yu , Wang Jun , Zhao Gang , Song Hui-hui , Bao Wen , Sun Xin-chen , Cheng Hong-yan , Deng Yu-xia , Li Guo-hong , Liu Li-jie , Chen Wen-ji , Wang Jue-qiong , Yuan Peng , Pei Xiao-ping , Chen Ning-na , Wang Xue-mei , Zhu Min-sheng
Published: April 2020

Object: To study the variation of p21, P-gp expression in reversion of multidrug resistance of K562/A02 leukemic line with different concentration of tetrandrine(TET), to provide new theoretic evidence for the clinical application of TET.

Methods: The IC50 (the concentration causing 50% inhibition of cell growth) of DNR was analyzed by MTT method; The Expressions of p21 were assayed by westernblot; The Expressions of P-gp were assayed by FCM; The expressions of mdr1 were assayed by RT-PCR; The variation of p21 was accentuation with the accrescence concentration of TET(P <0.05).

Results: The variation of p21 protein in K562/A02 cells was accentuation with the accrescence concentration of TET(p<0.05), Mdr1 mRNA was lowly displayed in K562 cells and highly displayed in K562/A02 cells(p<0.01), the variation of mdr1 was attenuation with the accrescence concentration of TET(p<0.05); P-gp was lowly displayed in K562 cells and highly displayed in K562/A02 cells(p<0.01), the variation of P-gp was attenuation with the accrescence concentration of TET(p<0.05).

Conclusion: TET may reverse multidrug resistance of K562/A02 cells by the regulation of p21,P-gp and mdr1.
Factors Involved in Response to Thalidomide in Myelodysplastic Syndrome: Impace of HLA and Prognostic Factors
Authors: Bakanay Sule Mine , Dalva Klara , Topcuoglu Pervin , Civriz-Bozdag Sinem , Arat Mutlu , Beksac Meral , Gurman Gunhan
Published: April 2020

Myelodysplastic Syndrome (MDS) is a clonal disease of hematopoiesis characterized by dysplasia in one or more lineages. Since the syndrome is clinically heterogenous, treatment options are complex and hard to define. Thalidomide (T) is the first agent tested in MDS for its immunomodulatory and anti-angiogenic effects. Twenty to 59 % response rates, which is mainly hematological improvement, have been reported. However, because of excessive toxicity and low response rates, thalidomide has not been widely used as an effective treatment option for MDS. Factors invoved in response to T are not well delineated. Better response to immunosupression (IS) in HLA-DRB1*15(+) patients with aplastic anemia and some MDS subgroups have been reported. In this study, the medical records of MDS patients, who received T at least for ≥ 3 months were retrospectively analyzed. Responses could be evaluated in 47 patients with a median age of 59 years (range, 15–84) and M/F: 29/18. According to WHO classification, there were 7 refractory anemia (RA), 9 refractory anemia with ring sideroblasts (RARS), 5 refractory cytopenia with multilineage dysplasia (RCMD), 2 RCMD-RA, 13 RCMD-RS, 6 Hypoplastic MDS, 1 RAEB, 4 MDS/MF. International Prognostic Scoring (IPSS) risk groups consisted of 42 low/intermediate-1 and 5 intermediate-2/high risk patients. Clonal cytogenetic abnormalities were found in 16 patients (5 trisomy 8, 3 monosomy 7, 2 delY, 2 increased chromosomal breaks, 1 del5q, 1 monosomy 15 and 1 del21q). Thalidomide was administered at a dose range of 100–400mg/day and for a range of 3–48 months. Twenty-one patients had hematological response (10 major and 11 minor). Seven patients among responders had received additional erythropoietin (EPO) combined with T. There was no statistically significant difference between responders and non-responders in terms of age (median 53 vs 60.5 years), sex (M/F: 12/9 vs 17/26) and IPSS risk groups (Low risk/high risk: 19/2 vs 23/3). Response rates were 44% in RA; 50% in RCMD-RA; 56% in RARS; 55% in RCMD-RS; 20% in RCMD; 20% in hypoplastic MDS; 75% in MDS/MF. and none in RAEB patient. The EPO levels did not differ significantly between the responders and non-responders (EPO miu/ml <500 in 93% vs 83%). However, compared with non-responders, majority of the responders had ferritin levels <1000 mcg/L (83% vs 50%, p<0.05). A total of 21 patients had their HLA type screened. HLADRB1*15 was detected in 44% of responders (4/9) and 16% of non-responders (2/12) (p<0.05). Interestingly, among 5 patients who had trisomy 8, 4 responded to T. Fourty-five percent of mainly low risk-MDS patients responded to T. Patients with MDS/MF and RARS/RCMD-RS and patients with less iron overload responded better. HLADRB1*15 predominance and the presence of trisomy 8 in T responders should be underlined.
Prognostic Relevance of Retrospective WHO Re-Classification in 650 MDS Patients Diagnosed by FAB Criteria
Authors: Breccia Massimo , Latagliata Roberto , Cannella Laura , Carmosino Ida , Stefanizzi Caterina , Santopietro Michelina , Alimena Giuliana
Published: April 2020

We retrospectively re-classified according to WHO criteria our series of consecutive 650 MDS cases, in order to validate the prognostic role of this classification in a unicentric series of patients. Based on FAB criteria, 196 patients had been diagnosed as RA, 41 as RARS, 242 as RAEB, 74 as RAEB-t and 83 as CMML. The new WHO classification could be applied to 479 of the 650 patients; of 196 FAB-RA only 76 were classified as “pure” RA, with only anemia and erythroid dysplasia, whereas 87 patients were re-evaluated as RCMD, for the presence of peripheral cytopenias and dysplasia in ≥ 10% of 2 cell lines. Twenty-nine patients had cytopenias but with only unilineage dysplasia in granulocytes or megakaryocytes, and were re-interpreted as MDS, unclassified (MDS-U). Of 41 FAB-RARS, 32 maintained their diagnosis, whereas 9 patients were classified as RCMD-RS, based on criteria listed above. One-hundred-one patients were identified as RAEB-1, for the presence of 5–9% blasts in bone marrow, and < 5% peripheral blasts, whereas 141 patients were classified as RAEB-2 for the evidence of 10–19% and of 5–19% blasts in bone marrow and peripheral blood respectively. We tested the WHO classification in univariate analysis with respect to several clinical features at presentation. Significant correlations were found as to hemorrhagic symptoms in RCMD and RAEB-2 patients (p=0.014), acute transformation, with higher frequency in RCMD and RAEB-2 categories (p=0.002), transfusion dependence, with higher requirement in RAEB-2 (107/141 RAEB-2 vs 59/101 RAEB-1) and in RCMD (63/87 patients) categories; also the application of different scoring prognostic systems, such as the Bournemouth and Spanish, appeared to possess stastical significance (p=0.001 and p=0.002, respectively). No differences were found as to sex, age, infection occurrence and cytogenetic abnormalities among various subgroups. Kaplan-Meier survival test showed high significance, with median overall survival ranging from 55.8 months for pure RA to 22.6 months for RAEB-2 category (p=0.00001). In conclusion, the retrospective application of WHO classification to our series of MDS patients clearly identified prognostic correlations in various disease subtypes. Categorization of MDS entities according to WHO criteria may distinguish parameters of prognostic importance in large groups of patients with morphological similar features.
ABCG2 Expression and Methylated Regulation in Human Multiple Myeloma Cell Lines
Authors: Qu Shiqiang , Fu Chengcheng , Yao Li , Chen Guanghua , Cen Jiannong , Qi Xiaofei , Depei Wu
Published: April 2020

Objective: The expression and regulation of multidrug resistance protein ABCG2 in multiple myeloma (MM) is unclear, expression of ABCG2 mRNA and protein, promoter methylation regulation of ABCG2 in MM cell lines will be investigated.

Methods After human MM cell lines 8226 AU266 and XG-7 were incubated in the demethylation agent 5’-AzaDC, the change of ABCG2 mRNA and protein expression were evaluated separately by using the Taqman probe-based real-time quantitative polymerase chain reaction(RQ-RT-PCR) and flow cytometry. The percentage of methylated alleles was assayed by SYBR green based methylation-specific quantitative PCR (MSQ-PCR). The melting curve was then analyzed to judge the specificity of the PCR product.

Results The three MM cell lines have different expression level of ABCG2 mRNA(Table 1) and protein but all much higher than normal PBMCs and in different promoter methylation status. The highest level of ABCG2 mRNA and protein were detected in the 8266 cell lines and the promotor region is in unmethylation status. Expression in the U266 cell lines was moderate and the methylated percentage is 61.55%±1.98%. Expression in the XG-7 is the least, the methylated percentage is 78.40%±2.46%. ABCG2 expression increased after exposure to demethylation agent 5’-AzaDC to 2 fold in U266 and 3.5 fold in XG-7 cell lines. After MSQ-PCR, the melting curve was analyzed. The Tm value of methylated product and the unmethylated product were 77°C and 75.5°C separately.

Conclution: ABCG2 mRNA and protein expression in 3 MM cell lines were much higher than that in normal peripheral blood mononuclear cells. Demethylation of the promotor increased ABCG2 mRNA and protein expression. So expression of ABCG2 is partially regulated by promoter methylation in myeloma cell lines. The cell line with the more ABCG2 expression may have more potential to effluence the chemotherapy agents. SYBR green I based fluorescence quantitative PCR can calculate the percentage of the methylated alleles in genome.
Increased Thrombin Generation in Severe Hemophiliacs with Mild Clinical Phenotype
Authors: Santagostino Elena , Mancuso Maria Elisa , Tripodi Armando , Chantarangkul Veena , Pasta Gianluigi , Siboni Simona Maria , Mannucci Pier M.
Published: April 2020

Introduction: Some severe hemophiliacs (FVIII/FIX<1%) exhibit a mild bleeding tendency, but the basis for this clinical heterogeneity is poorly understood. This study investigated the relationship between the values of endogenous thrombin potential (ETP) and clinical phenotype in severe hemophiliacs. The impact of FVIII/FIX gene mutations and thrombophilic polymorphisms was also evaluated. Methods: severe hemophiliacs older than 18 years without inhibitor history and treated on demand were eligible. Mild bleeders (MB) and severe bleeders (SB) were defined as follows: spontaneous bleeding episodes per year ≤2 (MB) or 25 (SB) and concentrate consumption <500 (MB) or >2000 (SB) IU/Kg/year. Patients who did not fit these criteria were considered as intermediate bleeders (IB). FVIII was measured by chromogenic assay and ETP was measured in platelet-rich plasma after addition of tissue factor. Results: 22MB, 22SB and 28IB were enrolled. MB had lower clinical and radiological scores when compared with both IB and SB (p<0.005). MB showed an older age at first bleed compared to SB (p < 0.005) and p for trend among the 3 groups was also significant (p < 0.05). The prevalence of severe FVIII/FIX gene defects (null mutations) was lower and ETP values were higher in MB compared with both IB and SB (p<0.05; table 1). Conclusions: our results indicate an extremely low prevalence of null mutations in severe hemophiliacs with mild bleeding diathesis. The measurement of thrombin generation in platelet-rich plasma may allow to identify this subgroup of patients, not otherwise distinguishable by conventional functional assays.

SB (#22) IB (#28) MB (#22) p Age (yr) 38 (21–76) 38 (23–62) 32 (22–73) NS Age 1st bleed (yr) 1 (0–4) 2 (0–6) 3 (1–10) < 0.005 Bleeding episodes/yr 36 (25–60) 10 (3–20) 0 (0–2) < 0.0005 Factor use (IU/Kg/yr) 2207 (2040–8696) 1068 (207–2400) 60 (25–487) < 0.0005 Clinical score 18 (10–35) 10 (0–34) 3 (0–17) < 0.005 Pettersson score 44 (14–62) 28 (0–48) 17 (3–40) < 0.0005 Null mutations (%) 59 70 6 < 0.005 PTG20210A (%) 0 7 5 NS FV Leiden (%) 5 7 0 NS Median ETP (nM) 414 478 850 < 0.05
Clinical and Biological Features of Biclonal Gammopathies. Review of 203 Cases
Authors: Decaux Olivier , Leroy Helene , Ianotto Jean-Christophe , Ruelland Annie , Guenet Lucienne , Sebillot Martine , Jego Patrick , Grosbois Bernard
Published: April 2020

Introduction: Biclonal gammopathies represent about 5% of clonal gammopathies. We describe the clinical and laboratory features of biclonal gammopathies identified in a French university hospital.

Methods: Patients were selected by immunofixation registry of Biochemistry laboratory.

Results: From 1987 to 2008, 203 biclonal gammapathies were identified. Patients were 113 men and 90 women. Median age was 72.0 years (35–95). Seventy eight patients (38.3%) had IgG and IgM components, 64 (31,9%) had two IgG, 24 (11,8%) had IgG and IgA, 23 (11,3%) had two IgM, 8 (3,9%) had IgM and IgA, 5 (2,4%) had two IgA, one (0.5%) had Ig G and IgD (0,5%). Of the 406 light chains, 260 (63,8%) were kappa, 146 (36.2%) were lambda. Eighty nine patients (44.1%) had two kappa light chains, 82 (40,2%) had both kappa and lambda and 32 (15,7%) had two lambda chains. Median gammaglobulin concentration was 13.3 g/L (3–59.9). The most frequent diagnosis was biclonal gammopathy of undeterminated significance (BGUS) in 123 patients (60.6%). Others patients could be divided into 3 groups. Forty seven patients (23.1%) had lymphoproliferative diseases, including Waldenström’s macroglobulinemia (21 cases), non Hodgkin lymphoma (20), chronic lymphocytic leukaemia (6). Eighteen (8.87%) had multiple myeloma. For the last 15 patients (7.4%), biclonal gammopathy was associated with a non lymphoid hemopathy or with systemic disease. Biclonal gammopathy was identified in 12 patients already known to have a monoclonal gammopathy (7 monoclonal gammopathy of undetermined significance-MGUS, 3 myeloma and 2 Waldenström’s macroglobulinemia). In 3 cases, the finding of the second monoclonal component was concurrent to the diagnosis of a MGUS malignant transformation to myeloma (2 cases) or Waldenström’s macroglobulinemia (1 case). Median follow-up was 23 months (12 to 252 months) for the 123 patients with BGUS. In 4 cases (3.2%) a malignant transformation was observed. Three patients developed a multiple myeloma (time to transformation was 2 years for one and 4 years for the 2 others) and one patient Waldenström’s macroglobulinemia (6 years).

Conclusions: As for monoclonal gammopathies, BGUS represent the most frequent diagnosis. However, in contrast to monoclonal gammopathies, biclonal gammopathies are more frequently associated with lymphoproliferative diseases than with multiple myeloma. The apparition of a second monoclonal component during follow up of MGUS could be associated with malignant transformation and should lead to new evaluation. The risk of transformation of BGUS seems similar to MGUS but further studies are necessary to compare the evolution of MGUS and BGUS.