bluuebunny/crossref_metadata_2025 huggingface dataset:
Purified Gr1+CD11b+ Cells Induce Neovascularization in An Ischemic Hind Limb Mouse Model.
Authors: Kim Jeong A , March Keith , Chae Heedon , Johnstone Brian , Cook Todd , Merfeld-Clauss Stephanie , Broxmeyer Hal E.
Published: April 2020

Neovascularization in ischemic tissues is enhanced by circulating endothelial precursor cells, some of which have been identified as bone marrow (BM)-derived (BMEPC). However the identity of BM-derived populations that participate in the process of neovascularization is not clear; and indeed several subsets have been proposed as important to this process. Gr1+CD11b+ cells have been recently described as myeloidderived suppressor cells that may be involved in tumor angiogenesis. However, there has been little investigation of the role of Gr1+CD11b+ cells in non-tumor angiogenesis. To study the relationship of Gr1+CD11b+ cells to neovascularization in an ischemic hindlimb model of C57BL/6 mice, we tested the following hypotheses:

The number of tissue-resident Gr1+CD11b+ cells increases in BM and ischemic muscle after the ischemic injury; prospectively isolated Gr1+CD11b+ cells can manifest characteristics of endothelial differentiation; and Gr1+CD11b+ cells modulate neovascularization in ischemic hind-limbs of C57BL/6 mice upon direct injection into ischemic muscle.

Results demonstrated that the number of Gr1+CD11b+ cells markedly increased in the ischemic muscle at 4 days post-surgery (n=5 each; 1.08±0.65x105/g tissue in non-surgically treated mice vs 13.43± 7.52x105/g tissue in the ischemic muscle, p<0.05); then returned to basal level at day 10. The number of Gr1+CD11b+ cells from BM in the ischemic group at day 4 trended towards an increase when compared with cells from BM of non-surgically treated C57BL/6 mice (n=5 each; 1.05±0.31x107/2 femur+ 2 tibia in ischemic group vs 0.70±0.32x107/2 femur+ 2 tibia in non-surgically treated C57BL/6 mice). Following culture under endothelial cell growth conditions, purified Gr1+CD11b+ cells from BM exhibited cobblestone and spindle-shaped morphology. FACS analysis demonstrated that cobblestone-shaped cells exhibited development of a CD144+ phenotype, a marker for mature endothelium; while spindle-shaped cells expressed desmin. Following direct injection of purified Gr1+CD11b+ cells from BM or medium control, laser Doppler imaging revealed an increase in blood flow recovery by 14 days post-femoral artery dissection in the cell injected group compared to the control group (n=7 each, 41.14±1.28% in Gr1+CD11b+ cell injected group vs 35.83±2.57% in control group). The mobilization of Gr1+CD11b+ to regions of muscle ischemia, taken together with the novel finding of endothelial differentiation from this BM-derived population, and enhanced flow recovery following local provision of exogenous Gr1+CD11b+ cells, support the hypothesis that this cell population plays an important role in the systemic and local response to ischemic injury.
Mir-128b Is a Potent Glucocorticoid Sensitizer in MLL-AF4 Acute Lymphocytic Leukemia Cells and Exerts Synergistic Effects with Mir- 221.
Authors: Kotani Ai , Ha Daon , Rao Prakash , Hsieh James , Schotte Diana , den Boer Monique L. , Armstrong Scott , Lodish Harvey F.
Published: April 2020

MLL-AF4 Acute Lymphocytic Leukemia (ALL) has a poor prognosis. We show that re-expression of miR-128b and miR-221, downregulated in MLL rearranged ALL, synergistically sensitizes MLL-AF4 ALL cells to glucocorticoids. Target genes downregulated by miR-128b include MLL, AF4, and both MLL-AF4 and AF4-MLL fusion genes; miR-221 downregulates CDKN1B. Finally we identified a novel A to G point mutation in the miR-128b gene in some MLL-AF4 ALL cells, which significantly reduces the processing of precursor miR-128b miRNA to the mature miRNA. These results demonstrate that downregulation of miR-128b and miR-221 induces glucocorticoid resistance, restoration of their levels is a promising therapeutic in MLL-AF4 ALL, and the A to G point mutation might pathophysiologically contribute to MLL-AF4 ALL.
Remarkable Activity of Bortezomib Combined with Chemotherapy in a Phase I Study of Relapsed Childhood Acute Lymphoblastic Leukemia (ALL). A Report from the Therapeutic Advances in Childhood Leukemia (TACL) Consortium.
Authors: Messinger Yoav H. , Gaynon Paul S. , Raetz Elizabeth , Hutchinson Raymond , DuBois Steven , Bender Julia Glade , Sposto Richard , van der Giessen Jeannette , Eckroth Elena , Bostrom Bruce C.
Published: April 2020

The outcome of relapsed ALL remains very poor, and many re-induction regimens are highly toxic. We report here the Phase I component of a Phase I/II study of bortezomib added to 4-drug induction with vincristine, dexamethasone, pegylated asparaginase, and doxorubicin in children with relapsed ALL. A total of 10 patients were enrolled, 5 in 1st marrow relapse (2 early and 3 late relapses) and 5 in 2nd or subsequent relapse. Four patients were enrolled at dose level 1 (1 mg/m2) to obtain 3 evaluable patients. One patient was inevaluable for toxicity because of an error in dexamethasone doses and was removed after 8 days. No dose-limiting toxicity (DLT) was observed. Six patients were enrolled at dose level 2 (1.3 mg/m2), the standard single agent dose. One patient had a DLT (hypophosphatemia and rhabdomyolysis) after1 dose of bortezomib, and that patient died from diffuse zygomyces infection by day 17. Five additional patients were enrolled with no further DLT’s. As planned, no further dose escalation was pursued. The regimen had predictable toxicity related to the chemotherapy drugs. Only 2 patients had mild peripheral neuropathy (grades 1 and 2). Seven of the10 patients (70%) achieved a CR, and 1/10 had bone marrow CR with persistent central nervous system (CNS) leukemia despite weekly intrathecal (IT) methotrexate, for a total of 80% bone marrow CR rate. In conclusion: the combination of bortezomib with 4 standard drugs is highly active with acceptable toxicity in heavily pretreated relapsed pediatric ALL patients. We are expanding the MTD cohort for a phase II estimate, and intend to report updated results at the ASH meeting.

Day 1 2 4 8 11 14 15 18 22 29 - 35 Bortezomib (1 or 1.3 mg/m2) B B B B Evaluate Vincristine (1.5 mg/m2) V V V V Doxorubicin (60 mg/m2) Dox Dexamethasone (10 mg/m2) x 14 days —— —— —— —— —— —> PEG-Asparaginase (2500 U/ m2) PEG PEG PEG PEG IT Ara-C ITA CNS Negative: IT Methotrexate ITM CNS Positive: IT Triples ITT ITT ITT
The Reduction of Leukemic Blasts In Bone Marrow Aspirate on Day 6 of Remission Induction Treatment Is Predictive for Complete Remission Rate and Survival in Adult Acute Myeloid Leukemia: The Results of Multicenter, Prospective Polish Adult Leukemia Group Study.
Authors: Holowiecki Jerzy , Grosicki Sebastian , Kyrcz-Krzemien Slawomira , Kuliczkowski Kazimierz , Kielbinski Marek , Skotnicki Aleksander B. , Piatkowska-Jakubas Beata , Hellmann Andrzej , Wierzbowska Agnieszka , Seweryn Marek , Stella-Holowiecka Beata , Sadus-Wojciechowska Maria
Published: April 2020

The goal of this study was to pr ove the significance of an early evaluation of leukemic blast reduction based on cytological examination of bone marrow aspirates obtained on day 6 of remission induction treatment.

Patients: 152 adult AML patients aged 18–60 (median 47) registered to the PALG prospective trial evaluating the efficacy of 3 remission induction protocols:

DAF: daunorubicine (DNR) 60 mg/m2/d iv, d 1–3; cytarabine (AraC) 200 mg/m2/d ci, d 1–7, and fludarabine 25 mg/m2 2h inf. iv d 1–5) DAC (like DAF but cladribin is used at 5 mg/m2 instead of fludarabine) and the standard DA 3+7 regimen.

Patients achieving CR received two courses of subsequent intensive consolidation:

HAM (HD AraC, mitoxantrone) HD AraC. In case of partial remission (PR) after the first induction course the same regimen was repeated.

Patients with no remission (NR) or with PR after 2 induction courses were withdrawn from the study. Bone marrow aspirates were performed before the treatment and on the 6-th day of the first remission induction course, and the MGG stained marrow smears were evaluated at each centre by two experienced hematologists.

Results: Based on the proportion of blasts in bone marrow specimen on day 6 patients were arranged into 2 groups:

“responders” with <5 % of blasts (n=59) and “non responders” with >5% of blastic cells in bone marrow (n=31).

The complete remission rate (CR) in the “responders” group equaling 86% (82/95) was significantly higher in comparison to that found in the “non responders” group 33%(19/57), p=<0,00001. The probability of overall survival (OS) was also higher in the group of “responders” if compared to “non responders” one; 63% versus 33% respectively, p=0,0009. In multivariate analysis including bone marrow examination, cytogenetic risk group, age, leukocyte count at diagnosis and the induction arm, only the persistence of leukemic blasts >5% in bone marrow specimens on 6-th day of induction was associated with higher risk of not achieving CR (HR = 51,1; p=0,00002), and with shorter OS (HR=2,9; p=0,004).

Conclusion: This prospective, multicenter study demonstrates that a simple and commonly accessible evaluation of the leukemic blasts reduction in bone marrow smear on the 6-th day of remission induction treatment is a reliable and independent predictor for achieving CR and for overall survival. This offers a decision point for an early modification of the treatment strategy.
Stem Cell Transplantation in Elderly Patients with Acute Lymphoblastic Leukemia (ALL) in First Complete Remission (CR1).
Authors: Arnold Renate , Beelen Dietrich , Bornhaeuser Martin , Bunjes Donald , Finke Juergen , Kroeger Nikolaus , Stadler Michael , Stelljes Matthias , Goekbuget Nicola , Hoelzer Dieter
Published: April 2020

In the German Multicenter ALL studies (GMALL) patients aged >55 years with high risk (B-lineage ALL with WBC at diagnosis >30000, late CR, t (4; 11), complex aberrant karyotype or prae-T or mature T-ALL) or very high risk (Ph+/BCR-ABL+) ALL are increasingly candidates for allogeneic stem cell transplantation (allogeneic SCT with a HLA identical sibling donor, MRD or a matched unrelated donor, MUD) or autologous SCT. Here, we report on 31 elderly patients transplanted within the GMALL studies 06/99 and 07/03. Median age of the patients was 61 years (56–65). 22 patients belonged to the very high risk group (VHR), 8 patients to the high risk group (HR) and 1 patient from the standard risk group (SR) was transplanted because of detection of minimal residual disease.17/31 patients were transplanted from a matched unrelated donor, 9/31 patients from a HLA identical sibling donor and 5/31 patients underwent autologous SCT. Conditioning regimens for MRD SCT were myeloablative (MAC) in 6 patients (TBI 12 Gy and chemotherapy n=2, radioimmunotherapy + chemotherapy n=2, chemotherapy only n=2) and 3 patients received reduced intensity conditioning (RIC).Conditioning regimens for MUD SCT changed over time with an increasing number of RIC in the study 07/03. In total, 7/17 patients received MAC (TBI 12 Gy and chemotherapy n=5, chemotherapy only n=2) and 10/17 patients received RIC. Conditioning regimens in autologous SCT were myeloablative (MAC) in 5/5 patients.

Results: After allogeneic MRD SCT 4/9 patients (44%) are alive in CCR (d+ 24, d+ 611, d+ 1721, d+ 2321), 3/9 patients died due to leukemia, 2/9 due to transplant related mortality (TRM). After allogeneic MUD SCT 8/17 patients (46%) are alive in CCR (from d+ 165 to d+ 2176). 1 further patient is alive after re- SCT for treatment of relapse. 7/17 patients died due to TRM and 1 patient died due to relapse. After autologous SCT 2/5 patients are alive in CCR (d+ 1703, d+ 1731), 3/5 died due to relapse. Risk factors for TRM: In allo SCT and MAC 8/13 patients died due to TRM in contrast to 1/13 patients with RIC. In auto SCT none of the patients died due to TRM. Risk factors for relapse: In allogeneic MRD SCT 3/9 patients died due to relapse and 2/17 patients relapsed after MUD SCT. Due to the small number of patients, no difference between MAC and RIC could be found. In autologous SCT 3/5 patients died due to relapse.

In conclusion: The study shows that allo MRD but also MUD SCT is very effective in a selected population of elderly ALL patients. Since the survival of elderly patients with chemotherapy only is about 25%, more patients should be encouraged to have a MRD or MUD SCT.
Sym001, the First Recombinant Polyclonal Rhesus-D Specific Antibody Product, Was Safe and Well-Tolerated in a Placebo-Controlled Randomized Phase I Trial.
Authors: Hjelmström Peter , Hallén Bengt , Lindenstroem Ewa , Gyaw Shwe
Published: April 2020

Sym001 is the first compound in a new class of biopharmaceuticals in clinical development for the treatment of Immune Thrombocytopenic Purpura (ITP), and the prevention of hemolytic disease of the newborn by anti-D prophylaxis (ADP). Sym001 is a recombinant polyclonal antibody product consisting of 25 different anti-Rhesus D specific (RhD) antibodies. The primary objective of this clinical study was to assess the safety of Sym001 following a single intravenous (iv) dose in RhD+ and RhD− healthy male volunteers. Secondary objectives were to assess the potential immunogenicity and pharmacokinetic (PK) profile of Sym001. Fifty-nine RhD+ subjects and 18 RhD− subjects were randomized to receive up to 12.5 μg/kg or 75 μg/kg, respectively, in seven dose cohorts as shown below:

Number of Subjects Cohort Dose (μg/kg) RhD+ Sym001 RhD+ Placebo RhD− Sym001 RhD− Placebo 1 0.25 5 2 0 0 2 1.0 5 2 0 0 3 4.0 7 2 7 2 4 12.5 7 2 7 2 5 25 7 2 0 0 6 50 7 2 0 0 7 75 7 2 0 0

Treatment with Sym001 was well tolerated and there were no serious adverse events (SAEs), no AEs of severe intensity and no AEs that resulted in discontinuation of the study in either the RhD+ or RhD− population. The frequencies of treatment-emergent AEs (TEAE), i.e. AEs occurring after drug administration, were similar among Sym001 and placebo treated subjects and there was no dose-dependent pattern of TEAEs. No significant decrease in mean hemoglobin level compared to baseline was observed at any dose level during the study, and no decrease in hemoglobin > 2 g/dL in any subject was seen. No other laboratory findings were suggestive of clinically significant hemolysis. In addition, no clinically significant changes in vital signs or electrocardiograms (ECGs) were observed, and there were no significant changes in clinical chemistry variables that did translate into SAEs or were assessed as being related to study medication. No immunogenicity to Sym001 was detected and analysis of PK is in progress. In conclusion, Sym001 was found to be safe and well-tolerated in healthy volunteers in doses up to 75 μg/ kg. Sym001 is now being evaluated in a red blood cell (RBC) challenge trial for the ADP indication and in a Phase II trial for treatment of patients with ITP.
Alemtuzumab-CHOP for Aggressive T Cell Lymphoma. A Phase II HOVON 69 Trial.
Authors: Kluin-Nelemans J.C. , Van Marwijk Kooij Marinus , Lugtenburg P.J. , Wijermans Pierre W , Van Putten Willem JL , van Imhoff Gustaaf W
Published: April 2020

The prognosis of patients with aggressive mature T cell lymphoma is very poor. We wondered whether the addition of the anti-CD52 monoclonal antibody alemtuzumab to 2-weekly CHOP chemotherapy as 1st line treatment would result in acceptable toxicity, improved response, and outcome. In a multicenter phase II design, 20 patients were to receive 8 cycles of CHOP14 (d 1: cyclophosphamide 750 mg/m2, doxorubicin 50 mg/m2, vincristine 1.4 mg/m2, 2 mg max, and day 1–5 prednisone 100 mg; repeat at day 15). Alemtuzumab 30 mg subcutaneously was added at day 1, 5 and 10 of each cycle (total of 24 administrations). Valaciclovir, cotrimoxazol and fluconazol were given as prophylaxis for herpes viral, PCP and fungal infections, respectively; G-CSF was given to enhance neutrophil recovery. Blood products were irradiated. Monitoring for CMV (re)activation by PCR or pp65 antigenemia was mandatory. Between Nov 2005 and Oct 2007, 20 patients with newly diagnosed T-NHL were included by 10 different centers (55% male; median age 50, range 20 to 65 years; 19 stage III/IV; 11 IPI high/high-intermediate). T-NHL histologies were: peripheral T cell lymphoma NOS (n=10), angioimmunoblastic lymphoma (n=6), subcutaneous panniculitis-like lymphoma (n=3), enteropathy-associated T cell lymphoma (n=1). Patients received a median of 8 CHOP cycles, 85% received 6 or more. Response to treatment was: 12 CR, 6 PR (ORR 85%) and 2 NR. At a median follow-up of 18 months, 11 patients are still alive, 9 patients have relapsed; the median FFS is 20 months (range 2–25) and median OS is 23 months (5–29). Toxicity was considerable, with CMV reactivation in 7/20 patients; one patient developed CMV disease; hospital admissions because of (mostly neutropenic) fever occurred in 8 patients. However, none of these patients died from these adverse events. One patient died in CR, 5 months after treatment due to sepsis complicating an extensive varicella zoster infection. Three patients (two with peripheral T cell lymphoma, one with angioimmunoblastic lymphoma without initial EBV activity) developed an EBV-related lymphoproliferative disorder. These patients ultimately died of relapse T-NHL.

In conclusion, an intensive alemtuzumab-CHOP14 regimen is feasible and effective in aggressive T-NHL as far as response and early outcome are concerned. Toxicity, especially the high percentage of herpes viral reactivation including EBV-related lymphoproliferation requires careful monitoring.
Chemical Genomic Screen Identifies Ionophores as Modulators of Notch1 in T-ALL
Authors: Roti Giovanni , Ross Kenneth N. , Mecucci Cristina , Ferrando Adolfo A. , Aster Jon C. , Stegmaier Kimberly
Published: April 2020

Although in recent years intensive chemotherapy regimens have improved outcome for patients with T-cell acute lymphoblastic leukemia (T-ALL), side effects and relapse-rates remain high. An alternative therapeutic approach would be to target constitutively active mutated Notch1. Notch1, a trans-membrane receptor that acts as a transcription factor following proteolytic activation by the γ-secretase complex, harbors gain of function mutations in over 50% of T-ALL. Early clinical trials attempting to modulate Notch1 with γ-secretase inhibitors (GSI) were closed secondary to gastrointestinal toxicity, suggesting that alternative approaches to inhibiting Notch1 signaling are needed. Transcription factor complexes are difficult to target pharmacologically with conventional high-throughput screening approaches. We therefore sought to target Notch1 with a new gene expressionbased approach to small molecule library screening. Signatures for the Notch1 on versus off states were defined using microarray expression profiling of 7 different Notch1 mutant T-ALL cell lines treated with vehicle (Notch1 on) versus a GSI (Notch1 off). We adapted this 32-gene signature to our gene expression-based high-throughput screening (GE-HTS) assay, which uses ligation-mediated amplification (LMA) and a Luminex bead-based detection system. As proof of principle, we confirmed that an shRNA against Notch1 induced the Notch1 off signature in DND41 cells (a T-ALL cell line bearing a mutated Notch1 allele). In order to facilitate future clinical translation, we screened a collection of 4,500 bioactive compounds highly enriched for FDA-approved drugs in DND41 cells. Twenty-eight compounds that induced the Notch1 off signature were identified as top hits. These compounds were tested in a secondary screen in a 2-fold dilution series in two GSI sensitive cell lines, DND41 and KOPTK1, and one GSI resistant cell line, PF382. Ion channel modulators emerged as the top class of compounds confirmed to induce the Notch1 off signature across all cell lines. We next focused on two representative compounds, the ionophores salinomycin and ionomycin. Similar to GSI, both of these molecules induced cell cycle arrest and markedly decreased levels of the activated intracellular portion of Notch (ICN). Both salinomycin and ionomycin, as well as numerous other ionophores that scored as inhibitors in the screen, are known to increase cytosolic calcium. These studies are consistent with a role for calcium in the activation of oncogenic forms of Notch1 and suggest a potential role for ion channel modulators in the treatment of T-ALL.
Comparing Standard IPI with Revised-IPI in Patients with Diffuse Large B-Cell Lymphoma: Which Has a More Differential Potential for Predicting the Outcomes after R-CHOP Chemotherapy.
Authors: Yang Deok-Hwan , Ahn Jae Sook , Kim Yeo-Kyeoung , Lee Je-Jung , Choi Young Jin , Shin Ho Jin , Chung Joo-Seop , Moon Joon Ho , Chae Yee Soo , Kim Jong Gwang , Sohn Sang Kyun , Kim Hyeoung-Joon
Published: April 2020

The utility of International Prognostic index (IPI) in the era of immunochemotherapy could not be determined by the available randomized trials. Recently, the study of British Columbia group suggested that a revised IPI (R-IPI) which redistributed the IPI factors into 3 distinct prognostic groups provided a more clinically useful prediction of outcome for patients with diffuse large B-cell lymphoma (DLBCL). We investigated the clinical outcomes of R-CHOP chemotherapy in patients with DLBCL for determining which has a more differential potential of predicting the prognosis among R-IPI or standard IPI.

Patients and Methods: We analyzed a total of 348 patients with newly diagnosed DLBCL from National University Hospitals of Southern Korea (NUHSK) between September 2002 and July 2008. R-CHOP conducted standard doses of chemotherapy and rituximab (375 mg/m2) administered a 21-day interval. The limited-stage patients were treated with 3rd or 4th chemotherapy followed by involved field radiation therapy (IFRT) and the advanced-stage patients were treated with six to eight cycles of chemotherapy.

Results: The median age was 61 years (range, 18–85) with 51.6% of patients aged above 60. After a median follow-up of 25 months (range, 2-66.8), 280 patients (80.5%) were alive and 76 patients (21.8%) had relapsed or progressed. 16 patients (4.6%) underwent autologous stem cell transplantation (ASCT). The 4-year probability of overall survival (OS) and progression-free survival (PFS) were 73.7 ± 3.4 % and 69.1 ± 3.6 %, respectively (Fig. 1A). The distribution of patients and 4-year OS rates according to IPI factors were followings; 16.7% patients had zero risk factor with 95.4%, 33.9% patients had 1 factor with 90.0%, 19.3% patients had 2 factors with 69.9%, 19.0% patients had 3 factors with 55.7%, 10.3% patients had 4 factors with 28.4% and 0.9% patients had 5 factors without reached 4-year OS rates (Fig. 1B). Both standard IPI and revised IPI showed a significant potential as prognostic variable in OS and PFS. However, the standard IPI distinguished clinical outcomes between patients with 3 risk factors and patients with 4–5 risk factors. In addition, there was no survival difference between patients with zero risk factor and patients with 1 risk factor (Fig. 2).

Conclusion: The standard IPI had more differentiation potency in predicting prognosis than R-IPI in patients with DLBCL treated with R-CHOP chemotherapy.

Fig. 1 Overall outcomes (A) and outcomes according to the number of IPI factors (B) Fig. 1. Overall outcomes (A) and outcomes according to the number of IPI factors (B) Fig. 2 Comparison of OS and PFS according to the standard IPI and revised IPI Fig. 2. Comparison of OS and PFS according to the standard IPI and revised IPI
First-Line Treatment with Rituximab Combined with Intravenous or Oral Fludarabine for Patients with Extranodal Mucosa Associated Lymphoid Tissue (MALT) Lymphoma.
Authors: Salar Antonio , Domingo-Domènech Eva , Estany Cristina , Canales Miguel , Servitge Octavio , Gallardo Fernando , Fraile Guadalupe , Montalbán Carlos
Published: April 2020

Backgroud: Synergistic antitumor effect with combination of fludarabine and rituximab has been demostrated on MALT cells. Addition of rituximab to other drugs has improved outcomes in several types of NHL without a significant addition of toxicity. Our aim was to evaluate the safety and efficacy of rituximab combined with fludarabine (RF) in first-line therapy for extranodal MALT lymphoma.

Patients and methods: Adult patients with untreated extranodal MALT lymphoma who were included and received rituximab 375 mg/m intravenously (IV) on day 1 and fludarabine 25 mg/m (IV) given on days 1–5 (days 1–3 in > 60 years), every 4 weeks; after the first cycle, oral fludarabine was permitted. After 3 cycles, a work-up was done. Patients in complete remission (CR) received an additional cycle and, if partial remission (PR), a total of 6 cycles.

Results: 22 patients were included. Characteristics: median age: 60 years (32–83); 45% male; PS 0–1 (100%); site of lymphoma origin: gastric (61%) and extragastric (39%); stage: I (45%), II1 (23%), II2 (5%) and IV (27%). A total of 101 cycles of RF were administered and 21 pts were evaluable for response. After the third cycle, 13 pts (62%) achieved CR and 8 pts (38%) PR. At the end of therapy, 19 pts (90%) achieved CR and 2 pts (10%) PR. Univariate analysis identified primary extragastric disease as an adverse factor to reach CR after 3 cycles of RF (HR 23.3 (95% CI, 2.0–273.3)). The median follow-up time was 23 months (95% CI, 18–27 months). Progression free survival (PFS) at 24 months was 88 % (95% CI, 80–100%). PFS at 24 months in gastric and extragastric MALT lymphoma were 100% and 79%, respectively. Tolerance to oral fludarabine was excellent. Mild neutropenia was the most common toxicity, usually presenting after the third cycle and 2 pts had prolonged mild thrombocytopenia. No grade 3–4 infections were observed.

Conclusions: Immunochemotherapy with RF, either with intravenous or oral fludarabine, achieves a high CR rate in both gastric and extragastric MALT lymphoma, although the firsts responded faster. With only four cycles of RF, two thirds of patients achieves CR. RF is associated with a good safety profile being mild granulocytopenia and thrombocytopenia the main adverse events. The long-term benefit of this therapy will require prolonged follow-up.
Depletion of the Shwachman-Diamond Syndrome Gene Product, SBDS, Leads to Growth Inhibition and Increased Expression of OPG and VEGFA.
Authors: Nihrane Abdallah , Sezgin Gulay , Dsilva Sehba , Ellis Steven , Liu Johnson M.
Published: April 2020

Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by bone marrow failure and leukemia predisposition, pancreatic exocrine dysfunction, and skeletal abnormalities, manifesting as skeletal dysplasia and osteoporosis. Mutations in SBDS have been shown to cause SDS, but the function of the SBDS gene product is unclear. Accelerated angiogenesis has recently been described in bone marrow cells from SDS patients. To clarify the unknown function of SBDS, we performed experiments analyzing the cellular effects of depleting SBDS by constitutive or inducible RNA interference. The growth of HeLa and NIH3T3 cells constitutively depleted of SBDS was markedly hindered when compared to cells stably transfected with siRNA against an irrelevant control gene. However, the growth abnormality was related to a significantly increased propensity to apoptosis in NIH3T3 more than in HeLa cells. We believe these differences are related to inactivation of p53 in HPV-18 positive HeLa cells, suggesting that depletion of SBDS may partially sensitize cells to p53-mediated apoptosis. As in constitutive knockdown, HeLa cells induced by doxycycline to express siRNA against SBDS exhibited growth inhibition, which was associated with modestly increased levels of apoptosis, suggesting a partial contribution of this process in inducible knockdown cells. By microarray analysis of knockdown cells, we found marked differences in expression of genes in multiple pathways, and we chose to examine a selected subset more closely using quantitative PCR arrays. In constitutive and inducible SBDS-depleted HeLa cell clones, we found 3- to 6-fold elevated mRNA levels of osteoprotegerin (OPG or TNFRSF11B) and vascular endothelial growth factor-A (VEGF-A). We confirmed significant overexpression of both secreted proteins by ELISA from supernatants of SBDS-depleted HeLa cells. Osteoprotegerin and VEGF-A are known to have diverse effects on osteoclast differentiation, angiogenesis, and monocyte/macrophage migration, all processes that may be aberrant in SDS, and we propose that overexpression of these factors may contribute to its pathology.
Inhibition of BCR Signaling Using the SYK Inhibitor R406 Antagonizes Migratory and Pro-Survival Responses in CLL Cells.
Authors: Quiroga Maite P. , Kurtova Antonina , Wierda William , Keating Michael , Burger Jan A.
Published: April 2020

Disrupting B cell receptor- (BCR-) induced signaling by inhibiting the spleen tyrosine kinase (SYK) represents a novel therapeutic approach in CLL. R406 is an ATP-competitive SYK inhibitor, and its prodrug, R788 (Fostamatinib disodium), displays clinical activity in patients with B cell lymphoma and CLL (Friedberg J. et al. Ann Oncol. 2008 Jun; 19 Suppl 4). In normal lymphocytes BCR engagement and SYK activation modulate cell adhesion and migration, suggesting that this pathway affects chemokine receptor and adhesion molecule function/signaling. The effects of SYK on CLL cell migration, activation and survival are largely unknown. Therefore, we examined the effects of BCR stimulation and its inhibition with R406 on CLL cell chemotaxis, expression of adhesion- and costimulatory- molecules, and survival. Activation of BCR with polyclonal goat F(ab’)2 fragments to human IgM significantly increased chemotaxis towards CXCL12 to 135.4% ±5.2% of controls and towards CXCL13 to 170.5% ± 31.5% of controls (Mean ± SEM, n=15). Engagement of BCR also significantly increased expression of CD40, CD44, CD54, and CD62L and decreased the expression of CXCR4. Pre-treatment with R406, which was kindly provided by Rigel Pharmaceuticals, Inc., abrogated the increased chemotaxis, adhesion- and co-stimulatory molecule expression, and pro-survival signals seen in response to BCR engagement. As displayed in Fig. 1, CLL cell viability at 24 hours was significantly increased to 60±5.2% with anti-IgM, compared to controls (41.8±6.7%) or CLL cells treated with anti-IgM plus R406 5 μM (39±6.6%, mean± SEM, n=9). Moreover, we found that R406 induced apoptosis in CLL cells in suspension culture and co-cultures with nurselike cells (NLC) and marrow stromal cells (MSC). When compared to CLL cells cultured with NLC (100%), the mean relative viability of CLL cells at 48h was reduced to 71.2±6.4% with 5 μM R406 and to 75.4±5.6% with 2.5 μM R406, whereas the same CLL samples cultured without NLC displayed a mean relative viability of 87±2.9% in controls, 58.9±6.8% with 5 μM R406, and 59.9±7.3%% with 2.5 μM R406 (mean±SEM, n=10). Similar levels of apoptosis were observed when CLL cells were incubated with R406 in MSC co-cultures.

Collectively, this study demonstrates that BCR engagement increases CLL cell migration, adhesion- and co-stimulatory-molecule expression, and survival. R406 effectively antagonizes these responses and induces CLL cells apoptosis, providing an explanation for the clinical activity of R406 in CLL patients.

Figure 1 Figure 1.
Dual Action of CD160 on CLL Cells: Giving Life to Prepare for Death!.
Authors: Agrawal Sami r G , Liu Feng-Ting , Giustiniani Jerome , Jia Li , Gribben John G. , Bensussan Armand
Published: April 2020

CD160 is a cell surface antigen expressed on normal human NK cells, as well as a subset of T cells. We have previously reported that almost all cases of chronic lymphocytic leukaemic (CLL) express CD160 and the activation of CD160 by monoclonal anti-CD160 antibodies (mAb) induces cellular activation (proliferation and cytokine production). In this study, using fresh, peripheral blood mononuclear cells from patients with lymphocyte counts of > 40 x 109/L, we found that the activation of CD160 protected CLL cells from spontaneous cell death (n = 18). The rates of spontaneous cell death of CLL cells ranged from 5 to 64% (average 33 ± 18%) after culture for 18 hours. However, the spontaneous death rate was reduced to 20 ± 11% (p=0.012) when the CLL cells were incubated with anti-CD160 MAb. The protective effect on cell survival was more marked in the cases showing the greatest spontaneous cell death: in those cases with > 30% cell death (ie, greater than the average for the whole population), cell death was reduced from 48±9.9% to 28±8.2% (p=0.0001). In the cases which were resistant to spontaneous cell death (< 30%), CD160 reduced cell death from 16 ± 6.6% to 11 ± 5.9% (p = 0.15). The PI3- kinase/AKT pathway is a well known survival pathway and CD160 activity in NK cells has been shown to involve PI3-kinase. CLL cells were incubated with anti-CD160 mAb and found to show AKT phosphorylation, I-kB degradation, Bcl-2 upregulation, but also Bax upregulation. Interestingly, the PI3-kinase inhibitor, wortmannin, completely blocked AKT phosphorylation, but had no effect on CD160-mediated cell survival and did not prevent activation of NF-kB nor upregulation of Bcl-2 or Bax. In view of the upregulated Bax expression, we hypothesised that despite CD160-induced increases in Bcl-2 and the improved survival, CLL cells may have simultaneously become more sensitive to death signals. CLL cells (from 9 cases) were treated with chlorambucil (20mg/ml) with or without the anti-CD160 antibody overnight and apoptosis was determined with Annexin-V/propidium iodide by flow cytometry. Chlorambucil alone induced 63%±16 apoptosis with 34%±23 spontaneous cell death. Co-treatment with chlorambucil and anti-CD160 mAb increased apoptosis to 72±14% with spontaneous rate at 23%±13.

In summary, the activation of CD160 protected CLL cells from spontaneous cell death in vitro via a PI3-kinase/AKT independent pathway. However, this improved survival was also associated with increased Bax and sensitisation to chemotherapeutic agents. The mechanism by which the activation of CD160 plays dual roles in the destiny of CLL cells remains unclear, but this represents a potential novel therapeutic approach. Data will be presented showing the activity of a chimeric, humanised anti-CD160 mAb on CLL killing.
Stem Cell Transplant (SCT) for Patients (pts) with Chronic Myeloid Leukemia (CML) Resistant to Tyrosine Kinase Inhibitors (TKI) with BCR-ABL Kinase Domain (KD) Mutation T315I.
Authors: Velev Nikolai , Cortes Jorge , Champlin Richard , Kantarjian Hagop M. , Rondon Gabriela , Giralt Sergio , Borthakur Gautam , De Lima Marcos
Published: April 2020

Background: Resistance to TKI therapy is associated with development of KD mutations in approximately 50–60% of pts. Although many imatinib-resistant mutations respond well to second generation TKI, T315I is insensitive to all currently available TKI (imatinib, dasatinib, nilotinib) in vitro and in the clinic. SCT is frequently recommended for these pts but there is no available data about the efficacy of SCT in such pts.

Aims: To investigate the efficacy and safety of SCT for patients with TKI-resistant CML with a T315I mutation.

Methods: We reviewed the outcome of all pts with T315I that have received a SCT at MD Anderson Cancer Center.

Results: Seven pts received 8 transplants. Their median age was 44 years (yrs) (range, 26 to 64 yrs). The median time from diagnosis to SCT was 42 months (mo) (range, 9–160 mo). All pts had become resistant to with imatinib; 5 received dasatinib and 1 nilotinib after imatinib failure, and 6 pts received other additional therapy prior to SCT. At the time of SCT 2 pts were in chronic phase (CP), both in partial cytogenetic response; 2 in accelerated (AP) with active disease; and 3 in second or greater CP from lymphoid blast phase (BP) (1 in minor cytogenetic response, 2 major molecular response, 1 complete molecular response –CMR-). Six transplants were from matched unrelated donors and 2 from cord blood. Best response after SCT was CCyR in 3 (2 AP, 1 BP), CMR in 4 (2 CP, 2 BP), and 1 unknown (died early). After a median follow-up of 11 months from SCT, 4 pts are alive; the 2 transplanted in CP are alive after 11 and 42 months after SCT and in CMR; 1 pt transplanted in AP has a sustained CCyR 20 mo after SCT with persistent T315I representing 94% of transcripts by pyrosequencing; and 1 in BP has a CMR sustained 6 mo after a second SCT (relapsed 5 months after first SCT) 3 pts have died: 1 AP and 2 BP, all with relapse.

Conclusion: SCT appears to be an effective strategy for pts with CML with T315I, although longer follow up is needed. Results are significantly better when pts are transplanted in CP. Thus, SCT should be considered in pts with resistance to TKI once T315I is identified, ideally in CP.
Inhibition of BCR/ABL-T315I by Dismantling the Hydrophobic Spine.
Authors: Azam Mohammad , Seeliger Markus , Powers John T , Gray Nathanael , Kuriyan John , Daley George Q.
Published: April 2020

Mutation in the ABL kinase domain is the principal mechanism of imatinib resistance (IMR) in patients with chronic myelogenous leukemia (CML). The second generation BCR/ABL inhibitors, Nilotinib and Dasatinib, are effective in inhibiting essentially all IMR variants, but not the gatekeeper mutant T315I. Substitution of a bulky hydrophobic residue for the gatekeeper threonine not only causes steric blockade to the inhibitor but also stabilizes the active kinase conformation through a network of hydrophobic connections dubbed the hydrophobic-spine. In this study we describe the molecular mechanisms employed by the gatekeeper mutation to stabilize the active conformation, and demonstrate that these structural components can be targeted by the small molecule inhibitor compound #14, which efficiently inhibits native and gatekeeper mutant forms of the BCR/ABL kinase. Structural modeling and mutagenesis of residues constituting the spine suggests that compound #14 inhibits the kinase by disrupting the hydrophobic spine. Screening for drug resistance in vitro selected for clones having compound mutations involving both the P-loop and gatekeeper residues. Our studies provide structural guidance for the design of inhibitors against the gatekeeper mutant, and suggest that combination therapy may be required to prevent the emergence of compound resistance mutations.
Pre-Transplant Bone Status Evaluation in Patients Undergoing Allogeneic Stem Cell Transplantation.
Authors: Yakoub-Agha Ibrahim , Wibaux Cecile , Magro Leonardo , Juilliard Audrey , Legroux-Gerot Isabelle , Coiteux Valerie , Jouet Jean-Pierre , Cortet Bernard
Published: April 2020

In view of the high observed frequency of bone events following allogeneic haematopoietic stem cell transplantation (allo-SCT), the aim of this prospective study was to evaluate pre-transplant bone status in allo-SCT patients. In the month before transplantation, bone-loss risk factors were documented for 27 patients. We measured the levels of bone remodelling markers (BRMs: plasma osteocalcin and bone & total alkaline phosphatases for bone formation; CTX and telopeptides for bone resorption), together with plasma creatinine, intact PTH, TSH, testosterone, LH, FSH, 25 OH vitamin D, serum calcium & phosphorus and calciuria. In addition, bone mineral density (BMD) at the lumbar spine, femoral neck and hips was measured using double-energy X-ray absorptiometry (DEXA). Spine X-ray measurements were also made. Bisphosphonate, oral calcium and vitamin D or testosterone were administered according to the results of the evaluation.Between June 2006 and March 2007, 13 males and 14 females underwent allo-CST for haematological malignancies. The median age at transplantation was 44 years (range: 22–60). Eighteen had received prior corticosteroid therapy, 10 were smokers and 2 had a history of alcohol abuse. All had received prior chemotherapy, including 2 patients having already undergone autologous SCT. Ten of the 14 women were post-menopausal but none was on hormone replacement therapy. The median BMI was 24 kg/m2 (19–35). The daily calcium intake was low, with a median value of 950 mg/day (467–1852). While serum calcium, phosphorus and creatinine levels were within the corresponding normal ranges for all patients, 15 individuals displayed vitamin D deficiency and 6 had calciuria < 100 mg/day. Two patients suffered from hyperparathyroidism. In terms of BRMs, the patients respectively displayed a normal profile (n=9), high bone resorption activity (n=12) or high bone formation/resorption activity (n=5). One patient had hyperthyroidism and another presented hypotestosteronaemia. Bone density results were normal in 16 patients and abnormal in 11 (41%), including 8 with osteopaenia and 3 with osteoporosis. Vertebral fractures were observed in 4 patients. Overall, 18 patients (67%) were considered as having a pathological bone status and required treatment with bisphosphonate alone (n=5), vitamin D supplementation alone (n=11) or both (n=2).This study revealed that a large proportion of allo-CST patients have pre-existing abnormal bone status and thus demonstrates the importance of pre-transplant bone status evaluation in allo-CST candidates. The implementation of appropriate bone-related treatments may reduce the frequency of post-transplant bone events.
TNF-a Polymorphism Modulates the Outcome of Multiple Myeloma Patients Treated with Bortezomib
Authors: Buda Gabriele , Martino Alessandro , Maggini Valentina , Orciuolo Enrico , Galimberti Sara , Gentile Massimo , Brunori Marino , Morabito Fortunato , Rossi Anna Maria , Petrini Mario
Published: April 2020

TNF-a is a potent mediator of inflammation and bone resorption that seems to be involved in the malignant transformation of plasma cells, since mononuclear cells, obtained from Multiple Myeloma (MM) patients and exposed in vitro to TNF-a and interleukin-4, produced monoclonal plasma cells. In addition, TNF-a can stimulate plasma cell proliferation, by triggering interleukin-6 secretion, and shows proangiogenic properties in vitro (1). In addition, TNF-a determination was reported to be a good parameter for estimating tumour mass and for monitoring therapy outcome during treatment with different protocols (2). In particular, TNF-a is able to activate NF-kB, which is the main target of bortezomib (3). Variation in TNF-a levels can be related to gene expression, which is regulated at transcriptional level, as well as to genetic polymorphism. Single nucleotide polymorphisms (SNPs) have been identified at position -308 and -238 in the gene promoter. In particular, a G to A substitution at position -308 is associated with higher levels of TNF-a (1). The aim of this study was to investigate the effect of this TNF-a polymorphism on the outcome of refractory and relapsed MM patients (pts), receiving Bortezomib containing regimen as second line therapy. From September 2005 to April 2008 we selected 90 MM pts, who received at least one cycle of chemotherapy before treatment with Bortezomib and at least one cycle of high dose chemotherapy with peripheral blood stem cell transplantation in 25 pts. TNF a polymorphism at position -308 was determined on genomic DNA extracted from blood samples. The genotype frequencies obtained (71/90 G/G; 19/90 G/A) were in agreement with Hardy–Weinberg equilibrium. Patients were categorized as responders (Complete + Partial response =R) or non responders (Stable + Progression disease=NR) to treatment. The overall response in our patients was about 74% (67/90). In MM pts carrying the rarest A allele the overall response was reduced to 58% (11/19 pts), whilst GG carriers showed a better response rate of 79% (56/71 pts) (OR = 0.37). Our data indicate that the SNP at position -308 of the TNF-a gene promoter may be associated with different outcome in patients affected by MM and treated with bortezomib containing regimen. Similar conclusion was reached in a previous study where TNF-a gene promoter could predict the outcome after thalidomide therapy (1). To investigate whether TNF-a SNP is an independent predictor for response to MM therapy, larger studies are necessary, however if these preliminary results will be confirmed, they may represent a rationale for targeting TNF-a in novel therapeutic approaches to MM (4).
15 Year Follow-up of ABMT for NHL: Surprisingly Favorable Outcome for Younger Adults.
Authors: Duong Hien , Rybicki Lisa , Andresen Steven , Copelan Edward , Dean Robert , Kalaycio Matt , Pohlman Brad , Sobecks Ronald , Sweetenham John , Chan Josephine , Sungren Shawnda , Bolwell Brian
Published: April 2020

Despite the fact that autologous bone marrow/stem cell transplantation (ABMT) has been successfully used to treat patients with relapsed Non Hodgkin’s Lymphoma (NHL) for over 25 years, few series report follow-up beyond 10 years. We have followed a cohort of 110 NHL patients who underwent ABMT from 9/1988 through 12/1993, all treated with a uniform preparative regimen of CBV (Cyclophosphamide, BCNU, Etoposide (VP-16)). At the present time, 38 (35%) are alive and 72 patients have expired. Among the 38 living patients, the median follow-up is 15 years. NHL histology for all transplanted patients was based on the Working Formulation, and 21% had low grade, 54% intermediate grade, 25% high grade (the majority of the high grade group had diffuse immunoblastic large cell lymphoma). Median age at transplant was 47 years; 85% were sensitive to salvage chemotherapy; 31% had elevated LDH at the time of diagnosis; age adjusted IPI at ABMT was 0 (7%), 1 (56%), 2 (29%), 3 (7%). Multivariable analysis analyzing prognostic factors for survival predictably revealed that sensitivity to chemotherapy, and LDH status at the time of transplant, had significant impact on survival. Additionally, age at transplant was also highly significant (p=0.002). This is shown graphically below:

Figure Figure

The incidence of low grade histologies was similar for all three groups. The younger group had a lower incidence of relapse mortality, as shown below:

Figure Figure

It is commonly believed that age is of limited importance with respect to survival after ABMT. However, this data suggests that age may play a very important prognostic role. The causes are not well defined, but the continued relapse and mortality risk may warrant surveillance indefinitely for patients undergoing ABMT for NHL.