bluuebunny/crossref_metadata_2025 huggingface dataset:
Bone Marrow Neutralizes Human Regulatory T-Cells to Permit Graft-Versus-Tumor Immunity In Humanized Mice
Authors: Guichelaar Teun , Rozemuller Henk , Emmelot Maarten , Martini Bianka , Storm Gert , Lokhorst Henk M , Martens Anton , Mutis Tuna
Published: October 2019

Abstract 1017

While application of regulatory T-cells (Tregs) has become a promising tool to prevent Graft-versus-Host Disease (GvHD), Tregs pose the risk of suppressing the curative Graft-versus-Tumor (GvT) effect induced by therapeutically transfused donor lymphocytes. Towards clinical application of Tregs, we explored the in vivo impact of human Tregs on GvT and GvHD in a humanized GvT model. In this model we have previously shown that infusion of human PBMC into tumor bearing Rag2-/-gc-/- mice induces a profound GvT effect but also results in lethal xenogeneic(x) GvHD. We observed that co-infusion of human CD4+CD25+ Tregs suppressed x-GvHD, but allowed effective GvT against human tumors localized within the bone marrow, despite efficient Treg-homing to bone marrow. Remarkably, however, Tregs abrogated the GvT effect against the same tumor when located extramedullary, suggesting that permittance of the GvT effect in the bone marrow was due to the inactivation of Tregs in the bone marrow micro-environment. In detailed exploration of this remarkable phenomenon, we discovered that stromal cells derived from bone marrow neutralize the suppressive phenotype and promotes interleukin-17 in human Tregs through secretion of IL-6/IL-1β. In conclusion, this study for the first time provides a novel in vivo mechanism of how human Tregs can control undesired inflammation while permitting GvT against hematological tumors residing in bone marrow. This mechanism involves the bone marrow stroma, which creates a tumor micro-environment that significantly drives conversion of Tregs into non-suppressor T-cells.

Disclosures:

No relevant conflicts of interest to declare.
Evolution of Human BCR-ABL1 lymphoblastic Leukaemia-Initiating Cells.
Authors: Notta Faiyaz , Mullighan Charles , Wang Jean , Poeppl Armando G , Doulatov Sergei , Phillips Letha A , Ma Jing , Minden Mark D. , Downing James R. , Dick John E.
Published: October 2019

Abstract 1023

Many tumour types are composed of genetically diverse cells, however little is known of how diversity evolves or the impact that diversity has on functional properties. Here, using xenografting and DNA copy number alteration (CNA) profiling of human BCR-ABL1 acute lymphoblastic leukaemia, we demonstrate that genetic diversity occurs in functionally defined leukaemia-initiating cells and that many diagnostic patient samples contain multiple genetically distinct subclones. Reconstruction of the subclonal genetic ancestry of several samples by CNA profiling demonstrated a branching multiclonal evolution model of leukaemogenesis, rather than linear succession. For some patient samples, the predominant diagnostic clone repopulated xenografts, while in others it was outcompeted by minor subclones. Reconstitution with the predominant diagnosis clone was associated with more aggressive growth properties in xenografts, deletion of CDKN2A/CDKN2B, and poor patient outcome. Our findings link clonal diversity with function and underscore the importance of developing therapies that eradicate all subclones.

Disclosures:

No relevant conflicts of interest to declare.
Treatment for Primary Acute Myeloid Leukemia: Results of Two Consecutive Trials From the Spanish CETLAM Group Showing Improvement with the Use of G-CSF Priming and Precise Risk-Adapted Therapy.
Authors: Sierra Jorge , Hoyos Montserrat , Nomdedeu Josep F. , Esteve Jordi , Duarte Rafael F. , Ribera Josep-Maria , Tormo Mar , Guardia Ramon , Llorente Andreu , Bueno Javier , Besalduch Juan , Queipo M. Paz , Marti-Tutusaus Josep M. , Pedro Carmen , Torres Pio , Garcia Antoni , Heras Inmaculada , Bargay Joan , Font Llorenç , Gonzalez Jose , Sampol Antonia , Salamero Olga , Escoda Lourdes , Gallardo David , Calabuig Marisa , Oriol Albert , Arnan Montserrat , Pratcorona Marta , Brunet Salut
Published: October 2019

Abstract 1066

Different approaches have been investigated to improve the prognosis of adult patients with primary acute myeloid leukemia. In two consecutive phase II trials our group has explored the use of intermediate-dose cytarabine in induction associated with idarubicin and etoposide, the addition of G-CSF priming to the previous combination, and risk-adapted postremission therapy.

Objective:

To compare the results of two consecutive trials for primary AML and to analyze the factors influencing the outcome.

Patients and methods:

Adult patients between 17 and 60 years of age with de novo AML, diagnosed between 1999 and 2009, were included in the CETLAM AML-99 and AML-03 trials. Induction chemotherapy (CT) included one or two courses of idarubicin 12 mg/m2 IV days 1,3,5, cytarabine 500 mg/m2/12h over 2h IV days 1,3,5,7 and etoposide 100 mg/m2 IV days 1,2,3. This was followed by one consolidation with mitoxantrone 12 mg/m2 IV from day 4 to 6, and cytarabine 500 mg/m2/12h IV from day 1–6. In the AML 03 trial, patients also received G-CSF priming, 150 mg/m2 subcutaneously (SC) from day 0 to the last day of induction and consolidation CT. Postremission therapy consisted of high-dose cytarabine (CBF AML), autologous or allogeneic hematopoietic transplantation depending on cytogenetics, courses to complete remission (CR), and in the AML-03 protocol also based on molecular abnormalities involving FLT3 or MLL genes and/or the persistence of minimal residual disease after consolidation.

Results:

Overall, 788 patients were included, 353 in the AML-99 trial and 435 in the AML-03. Median age of the patients was 46 years (range 17–60). There were no differences between patients included in the two protocols regarding age, gender, leukocyte counts, cytogenetics and proportion of favourable and unfavourable molecular cases in the group with intermediate-risk karyotype. The main results achieved appear in the table.

Multivariate analysis confirmed the favourable impact of AML-03 protocol on outcome. Other significant factors influencing survival were age, leukocyte counts and cytogenetics.

Conclusion:

G-CSF priming improved the CR rate of adult patients with primary AML and favourable or unfavourable cytogenetics. This fact and a more precise risk-adapted therapy taking into account genetic data and MRD studies translated into improved overall survival.

Disclosures:

No relevant conflicts of interest to declare.
Outcome of Saddle Pulmonary Embolism: A Nested Case-Control Study.
Authors: Bose Gauruv , Gandara Esteban , Carrier Marc , Erkens Petra MG , Rodger Marc , Wells Philip
Published: October 2019

Abstract 1102

Introduction:

The management of saddle pulmonary embolism (PE) is controversial. Evidence about outcomes and management strategies is scarce in the literature due to the small prevalence of saddle PE. Historically it has been recommended that this group of patients should be treated aggressively.

Purpose:

To determine the prevalence and outcomes of patients diagnosed with saddle PE.

Methods:

Retrospective cohort study of consecutive patients with saddle PE diagnosed at the Ottawa Hospital between January 2007 and December 2008. Patients were included if a thrombus was present on computed tomographic pulmonary angiography (CTPA) in the main pulmonary arteries spanning the bifurcation of the main pulmonary trunk. These cases were each matched with two non-saddle controls with proximal PE (thromboemboli in the main pulmonary arteries) based on age, sex, systolic blood pressure greater than or less than 90 mmHg, and the presence or absence of cancer. Demographics, prognostic factors, treatment, and outcomes were collected. Patients were followed over a 30 day period following the diagnosis. RESULTS: A total of 32 (5%) of 724 patients with PE had a saddle event. Baseline characteristics are depicted in Table 1. Differences between the saddle case group and non-saddle control group include the presence of right ventricular dilation (59% of saddle cases vs. 22% of controls, p-value: 0.0007) and in the proportion of patients managed as outpatients (7% of saddle cases vs. 33% of controls, p-value: 0.02). At 30 days no differences were found in patients with saddle PE or proximal PE for all cause mortality (6% vs. 10%; OR: 0.64; 95% CI: 0.08–3.2), PE related mortality (0% vs. 6%; OR: 0.52; 95% CI: 0.01–6.1), major bleeding (3% vs. 5%; OR: 0.65; 95% CI: 0.02–6.4), or recurrent venous thromboembolism (6% vs. 10%; OR: 0.64; 95% CI: 0.08–3.2).

Conclusions:

Patients with saddle PE do not have a worse 30-day prognosis than patients with proximal PE matched by age, sex, systolic blood pressure, and presence of cancer.

Disclosures:

No relevant conflicts of interest to declare.
Prothrombin Complex Concentrate (Octaplex®) In Patients Requiring Immediate Reversal of Vitamin K Antagonist Anticoagulation.
Authors: Song Madeline , Warne Connor , Crowther Mark A.
Published: October 2019

Abstract 1115

Rapid reversal of vitamin K antagonists (VKAs) is required when patients experience acute bleeding or are in need of urgent surgery. Traditionally, in Canada, the only therapeutic option for warfarin reversal was fresh frozen plasma (FFP); recently a four factor prothrombin complex concentrate (PCC) has become available. In this retrospective chart review, we examined the use of Octaplex®, a four factor (FII, FVII, FIX, FX), solvent/detergent (S/D) treated and nanofiltered PCC, in a tertiary care centre over 18 months (November 2008 to May 2010), to determine its efficiency and safety in VKA reversal. The PCC was used 85 times in 82 patients. All patients had received warfarin prior to PCC administration. Patients received a mean dose of 1792 (1000 – 3500) IU of PCC for treatment of bleeding (36), urgent surgery (40) and excessively elevated international normalized ratio (INR) (8). One patient received PCC for an unknown indication and is excluded from the results presented here; a further 6 patients did not have INR tests done immediately before and immediately after PCC administration and are excluded from the laboratory results presented here. The mean INR before administration of the PCC was 5.10 (range 1.2 – 25). The mean INR after administration of the PCC was 1.43 (range 0.9 – 3.3). Forty-seven of seventy-nine [59.5%, 95% CI 47.9 to 70.4%] administrations resulted in an INR of 1.3 or less and 60/79 [75.9%, 65.0 to 84.9%] resulted in an INR of 1.5 or less. Of the 40 patient encounters where PCC was administered due to need for urgent surgery, 39 were able to undergo the procedure. Three patients experienced a thromboembolic event; one had a venous thromboembolism due to a clot extension of a previously diagnosed left leg DVT 8 days after PCC administration, another had a non-occlusive renal vein clot 8 days after PCC administration and the last had a cilioretinal artery occlusion 238 days after receiving PCC. No other adverse events potentially related to PCC were observed. We conclude that the prothrombin complex concentrate Octaplex® provides efficacious, safe and rapid reversal of VKA therapy.

Disclosures:

Crowther: Pfizer: Consultancy, Research Funding; Sanofi Aventis: Consultancy, Research Funding; Leo Laboratories: Consultancy, Research Funding; Artisan: Consultancy. Off Label Use: The product described in this abstract is not available in the US. The described use is “on label” in Canada.
Short Polyphosphates Inhibit the Procoagulant Activities of Long-Chain Polyphosphate: A Novel Role for Platelet-Derived Pyrophosphate?.
Authors: Smith Stephanie A. , Morrissey James H.
Published: October 2019

Abstract 1140

Introduction:

Inorganic polyphosphates are negatively charged, linear phosphate polymers that influence hemostasis via accelerating factor V activation, triggering the contact pathway, and enhancing fibrin polymerization. The latter two effects require long-chain polyphosphates for optimal activity (>1000mers for activating the contact pathway and >250mers for enhancing fibrin polymerization). We explored whether short polyphosphates could inhibit the procoagulant effects of long polyphosphate polymers.

Methods:

Polyphosphate was size-fractionated by gel electrophoresis. “Long-chain polyphosphate” (LCP) was a heterogeneous polyphosphate preparation containing polymers ranging from 400 to several thousand phosphates long. Short-chain polyphosphate preparations were narrow fractions of homogeneous size. We also tested additional phosphate-containing molecules including ADP, ATP, monophosphate, pyrophosphate (PPi), and triphosphate. Clotting assays were performed using pooled normal plasma spiked with 0–20 μM LCP and 0–500 μM small (poly)phosphate; clotting was initiated by factor Xa plus CaCl2, or by CaCl2 alone (contact pathway). Fibrin was formed by clotting 2.6 mg/ml fibrinogen with 10 nM thrombin in the presence of 0–500 μM small (poly)phosphate plus 150 μM LCP and CaCl2.

Results:

Small polyphosphates (size range 23–83mer) reduced the ability of LCP to trigger the contact pathway of blood clotting, while monophosphate, PPi, triphosphate, ADP, and ATP all failed to influence the procoagulant activity of LCP. None of the small (poly)phosphates antagonized the ability of LCP to enhance factor V activation. Interestingly, monophosphate, PPi and triphosphate all inhibited the ability of LCP to enhance fibrin clot structure, with PPi being the most potent. Although ADP and ATP contain di- and tri-phosphates, they did not recapitulate the potent inhibitory activity of PPi. On the other hand, PPi had no measurable effect on the turbidity of fibrin clots formed in the absence of LCP.

Conclusions:

Short-chain polyphosphates inhibit the ability of LCP to initiate the contact pathway of coagulation. Platelet dense granules contain abundant PPi which is secreted in response to platelet agonists, although biological roles for platelet-secreted PPi are unclear. We propose that PPi is a novel modulator of fibrin clot structure, acting to regulate the effects of longer-chain polyphosphates on fibrin fibril formation.

Disclosures:

Smith: University of Illinois: Patents & Royalties. Morrissey:University of Illinois: Patents & Royalties.
Potentiation of Bortezomib-Induced Killing of Lymphoma Cells by Inhibition of Autophagy and Prevention of I-κBα Degradation
Authors: Jia Li , Gopinathan Ganga , Sukumar Johanna T , Gribben John G.
Published: October 2019

Abstract 116

Previous studies have shown that germinal center B cell-like (GCB) diffuse large B-cell lymphoma (DLBCL) are resistant to proteasome inhibitors such as bortezomib. The mechanism by which GCB-DLBCL cells escape from proteasome inhibitor-induced apoptosis is unclear. To investigate this further, we examined the proteasomal pathway, expression of anti-apoptotic proteins and autophagy. Using bortezomib or MG-262, we show that DLBCL cells have differential sensitivity to proteasome inhibitor-mediated cell death, even though the effects on proteasome inhibition were similar. DLBCL cells that either over-express anti-apoptotic proteins such as Bcl-2, or lack pro-apoptotic proteins including Bax/Bak, are more resistant to proteasome inhibitor-induced reduction of mitochondrial membrane potential and activation of caspase-3. Treatment with bortezomib induced autophagy in both sensitive and resistant DLBCL cells, as demonstrated by an accumulation and aggregation of the autophagy marker protein LC3-II. Bortezomib induced accumulation of ubiquitinated proteins and a decrease in the adaptor protein p62, indicating activation of autophagic degradation. Fluorescent microscopy revealed that bortezomib induced p62 recruits both ubiquitinated proteins and LC3-II, suggesting that p62 leads ubiquitinated protein to autophagosomes. Treatment with bortezomib thereby promotes I-kBα degradation, demonstrating that the route of I-κBα degradation is not via the ubiquitin-proteasome degradation system. Bortezomib-induced I-kBα degradation was detected in both DLBCL cell lines and primary DLBCL and follicular lymphoma samples. Chloroquine (CQ), an inhibitor of autophagy, significantly increased bortezomib-induced accumulation of p62 and ubiquitinated proteins, including I-κBα, Bax and p53. CQ alone induced a dose-dependent increase in I-kBα protein levels, indicating that I-κBα protein can be degraded by autophagy even in the absence of proteasome inhibition. Importantly, the combination of proteasome and autophagy inhibitors shows great potential to kill apoptosis-resistant lymphoma cells. Proteasome inhibitor with or without CQ induced cell death in DLBCL cells cannot be blocked completely by either caspase inhibitors or knockdown of Bax/Bak proteins, suggesting that cell death occurs via a caspase-independent pathway. In summary, proteasome inhibitors induce autophagy and confer DLBCL cells resistance by eliminating I-κBα and possibly other pro-apoptotic proteins. Addition of autophagy inhibitors to bortezomib has the potential to induce increased killing in patients with resistant lymphoma.

Disclosures:

Gribben: Roche: Consultancy; Celgene: Consultancy; GSK: Honoraria; Napp: Honoraria.
Hematopoietic Growth Factors In Aplastic Anemia Patients Treated with Immunosuppressive Therapy: Systematic Review and Meta-Analysis - 2010 Update.
Authors: Gurion Ronit , Gafter-Gvili Anat , Vidal Liat , Paul Mical , Ben-Bassat Isaac , Yeshurun Moshe , Shpilberg Ofer , Raanani Pia
Published: October 2019

Abstract 1164

Background:

Immunosuppressive therapy (IST) is the treatment for patients with severe aplastic anemia (SAA) not eligible for transplantation. It is controversial whether there is a role for hematopoietic growth factors (HGF) as an adjunct to IST in these patients.

Objectives:

A meta-analysis evaluating the role of HGF in this setting was published by our group in 2009. Since then, results of the largest conducted clinical trial by the Aplastic Anemia Working Party of the EBMT have been reported. We therefore updated our meta-analysis in order to evaluate if in 2010 there is still a role for the addition of HGF to IST in patients with SAA.

Methods:

Systematic review and meta-analysis of randomized controlled trials comparing treatment with IST and HGF to IST alone in patients with SAA. An updated search in The Cochrane Library, MEDLINE, conference proceedings and references was conducted in July 2010. Two reviewers independently assessed the quality of the trials and extracted data. Outcomes assessed were: all-cause mortality, overall hematologic response, infections and clonal evolution (transformation to myelodysplastic syndrome or acute leukemia). Relative risks (RR) with 95% confidence intervals (CIs) were estimated and pooled.

Results:

Our search yielded 7 trials, randomizing 619 patients, including the 205 patients included in the EBMT trial recently published. Trials were conducted between the years 1991 and 2008. The IST regimen for most trials consisted of anti-thymocyte globulin, cyclosporine and steroids. The HGF in 6 trials was G-SCF and in 1 trial GM-CSF and erythropoietin. The addition of HGF to IST, compared with IST alone yielded no difference in all cause mortality at 100 days (RR 1.33, 95% CI 0.56–3.18) and at 5 years (RR 0.91, 95% CI 0.64–1.30, Fig.1). There was no difference in overall hematologic response at 12 months between the two arms (RR 1.16, 95% CI 0.91–1.47). There was no increase in the incidence of clonal evolution in the HGF arm compared to the control (RR 1.45, 95% CI 0.42–5.07). In addition there was no difference in the number of infections between both arms (RR 0.98, 95%CI 0.82–1.17).

Conclusions:

The addition of HGF to IST in SAA does not influence all-cause mortality, long term response, or the incidence of infections. The cumulative data in our updated meta-analysis is consistent with the results of our previous report. Therefore, HGFs should not be recommended routinely as an adjunct to IST for patients with SAA.

Disclosures:

Shpilberg: Roche: Consultancy, Honoraria.
Clinical Grade Isolation of Regulatory T Cells From G-CSF Mobilized PBSC Improves with Initial Depletion of Monocytes.
Authors: Mahmud Dolores , Park Youngmin , Mahmud Nadim , Rondelli Damiano
Published: October 2019

Abstract 1176

We have recently demonstrated that G-CSF mobilized peripheral blood stem cells (PBSC) CD4+CD25+FoxP3+ cells (Tregs) prevent anti-CD34+ hematopoietic stem cells T cell alloreactivity in-vitro and co-transplantation of CD34+ cells and Tregs does not affect human stem cell engraftment in NOD/SCID mice (Mahmud D et al. Biol Blood Marrow Transplant, 2010). Since only a small number of Tregs can be isolated from normal peripheral blood we examined whether PBSC can be a useful source of Tregs for future clinical trials. Five leukapheresis products from healthy donors who received rh G-CSF at 10 ug/kg daily for 5 days were processed using the CliniMACS instrument (Miltenyi Biotec, Auburn, CA). CliniMACS CD34 reagent was initially used to isolate CD34+ cells. To isolate Tregs a 2-step procedure was initially utilized. A cocktail of clinical grade CD14, CD8 and CD19 reagents was mixed with the CD34- cells and depletion of monocytes, cytotoxic T cells and B cells was obtained by using the Depletion 2.1 program. The CD4+ cells were then enriched in Tregs by positive selection of CD25+ cells using a clinical grade CD25 reagent (Miltenyi). Because PBSC contain large amount of myeloid cells, and particularly monocytes, this clinical scale 2-step strategy was compared with a 3-step method that included an initial negative selection of CD14+ monocytes, followed by negative selection of CD8+ and CD19+ cells and a positive selection of CD25+ cells. Prior to isolation, the average proportion of CD4+CD25+ cells in PBSC was 0.77±0.26% in 5 separate PBSC products. After the 2-step process the proportion of CD4+CD25+ cells was 35±33% (n=3) vs 72±1% after the 3-step process. Therefore, utilizing the 3-step approach a better yield of Tregs was observed (10 vs 60%). Intracellular expression of FoxP3 was on average 74% in CD4+CD25+ cells obtained with a 3-step process. Contamination of different cell subsets in the final products enriched in Tregs was largely superior following the 2-step as compared to 3-step isolation method. Contaminating monocytes were, on average, 43 vs 5.7%, and contaminating CD8 and CD19+ cells were 12 vs 1.7% and 0.9 vs 0.3%, respectively. In the two procedures using a 3-step approach the final absolute number of Tregs isolated from products containing on average 30 × 109 mononuclear cells, was 95 and 93 × 106, respectively. These findings obtained using clinically available reagents and device, suggest that depletion of monocytes may improve the purity of Treg cell population isolated from PBSC. PBSC may represent a valuable source of Tregs for future clinical trials.

Disclosures:

No relevant conflicts of interest to declare.
RHPS4, An Anti-Telomere Agent, Specifically Targets the Subgroup of AML Cells with Loss or Gain of Parts of Chromosomes.
Authors: Pallis Monica , Clarke Simone , Grundy Martin , Richardson Jaineeta , Ojo Dotun , Anderson Susan , Russell Nigel H. , Seedhouse Claire
Published: October 2019

Abstract 1195

Cytogenetic abnormalities vary greatly in AML and those with gains and losses of chromosome parts, terminal deletions and whole chromosome losses could theoretically derive from telomere dysfunction. Cells with these abnormalities are characterised by telomere shortening and abnormal function characterised by activation of both the negative telomere regulator TRF1 and also of hTERT. The G quadruplex ligand RHPS4 (Pharminox Ltd, Biocity, Nottingham, UK) binds telomeres and induces DNA damage at higher doses in the short term and critical telomere shortening in longer term growth assays. We have previously reported cytostatic and cytotoxic effects for this agent in AML cell lines, whilst others have found that normal haematopoietic progenitor cells are not targeted. We have now examined effects of RHPS4 on the in vitro growth and survival of primary AML cells, comprising 9 samples with normal and 8 with abnormal karyotypes (5 complex, one -7, one 5q- and one with additional material on chromosomes 2 and 9). Cells were incubated with RHPS4 in standard 14 day colony assays and for 7 and 21 days in a novel suspension culture medium supplemented with supernatant from cultured normal human osteoblasts. In the 7 day cultures, no effects were observed on cell survival. In the longer term growth assays, a submicromolar IC50 for RHPS4 was observed in 7/8 samples with aberrant cytogenetics in contrast with only 1/9 with normal cytogenetics (P=0.003, Fisher's exact test). Furthermore, RHPS4 significantly sensitised samples with abnormal cytogenetics to daunorubicin reducing the median IC50 from 27nM to 10nM (n=8, p<0.05).

A qPCR method was used to measure telomere length of untreated DNA from 15 of the samples; the qPCR assay had previously been performed in a separate patient cohort alongside a Southern technique and comparable values were attained for the two different methods. There were no significant differences in telomere lengths between the normal and abnormal cytogenetic groups suggestive that telomere length is not a determinant of RHPS4 sensitivity in these assays. Samples with adverse cytogenetics have previously been associated with an up-regulation of basal DNA damage markers and may therefore be sensitive to compounds that can exacerbate levels of damage. We therefore studied the levels of p-ATM by immunocytochemistry in RHPS4-treated cells and found a significant increase compared to untreated cells - hence RHPS4 may be acting via a DNA damage mechanism.

We conclude that telomere targeting has exciting potential as therapy for a subset of AML patients including those with adverse cytogenetics who have a poor response to conventional chemotherapeutics.

Disclosures:

No relevant conflicts of interest to declare.
Jak2 Regulates Bcr-Abl In CD34+ Cells From Imatinib Mesylate-Resistant CML Patients.
Authors: Samanta Ajoy K. , Perazzona Bastianella , Chakraborty Sandip N. , Sun Xiaoping , Modi Hardik , Bhatia Ravi , Arlinghaus Ralph B.
Published: October 2019

Abstract 1220

Despite the success of imatinib mesylate (IM), complete molecular remission is difficult to achieve in IM-treated CML patients. After continuing treatment with IM, a population of leukemic stem/progenitor cells remains, which are insensitive to IM. Our previous studies have shown that Jak2 regulates Bcr-Abl signaling and maintains levels of Bcr-Abl in CML cell lines, fresh cells from blast crisis patients, and mouse hematopoietic cells transformed by wild-type and imatinib-resistant mutants of BCR-ABL (Samanta et al. ASH 2009 meeting, submitted). We hypothesize that Jak2 plays similar role in Bcr-Abl+ CD34+ stem/progenitor cells, and that inhibition of Jak2 will induce apoptosis by inhibiting critical signaling pathways. In our current studies we have examined Bcr-Abl CD34+ progenitor cells from CML blast crisis patients. These cells are resistant to IM but quite sensitive to cell death induction by the selective Jak2 inhibitor TG101209 at 5–10 μM concentrations for 48 h. Similar results were obtained with cells from IM-resistant accelerated phase and IM-resistant chronic phase patients. Western blotting of CD34+ cells from an IM-resistant blast crisis CML patient established that a four h treatment with TG101209 at 10 μM severely reduced levels of Bcr-Abl protein and levels of phosphotyrosine 177 within Bcr-Abl.

To begin to assess whether Jak2 effects on Bcr-Abl signaling also occur in early stage Bcr-Abl induced disease, we assayed CD34+ cord blood cells transformed by Bcr-Abl in short-term culture (Chu S et al. Cancer Res 2007; 67: 7045–7053). The results showed that a six h treatment with TG101209 (10 μM) reduced levels of: Jak2 protein; activated Jak2 (lower levels of pTyr 1007/8); Tyr 177 within Bcr-Abl; and the Bcr-Abl protein itself. These results support our earlier findings that Jak2 regulates Bcr-Abl signaling in CML cell lines (Samanta AK, et al. Cancer Res 66 (13):6468-72, 7/2006; Samanta AK, et al., Oncogene 28 (14):1669-81, 4/2009; Tao W., et al., Oncogene 27(22):3194-200, 5/2008). Furthermore, these results suggest that Jak2 plays a critical role in progenitor CML cells from early stage and late stage patients with CML disease.

Disclosures:

Bhatia: Novartis: Consultancy.
Survival and Response Outcomes to Different Treatment Schedules in CML Patients Starting Therapy with Imatinib. Results from the CML Spanish Registry (RELMC)
Authors: Casado Luis Felipe , Massague Isabel , Giraldo Pilar , Perez-Encinas Manuel , De Paz Raquel , Martínez-López Joaquín , Bautista Guiomar , Osorio Santiago , Requena Maria José , Palomera Luis , Peñarrubia Maria Jesús , Calle Carmen , Maestro Begoña , Steegmann Juan Luis
Published: October 2019

Abstract 1237

Background:

The RELMC is a multicentric, 17-hospitals-based cancer registry whose aim is to describe the treatments received by patients with CML, their outcomes, and the variables that influence treatment choices.

Aim:

To study the response and survival outcomes, in newly diagnosed CML patients treated with Imatinib (Im) as first line treatment.

Patients and methods:

249 newly diagnosed CML patients have been included. They are distributed in the following subgroups according to treatments received Im400. 166 patients received only Im400.

Result:

A summary of response and outcome is included in Table 1.

Complete cytogenetic response with regards to the best response, the CCyR rate was lower in patients with Im400-HDIm-2GTKI (60%) and Im400-2GTKI (62%). The rates were 84% in Im400, 83% in Im400-HDIm and 85% in HDIm; P Chi2 8,381(a) p=0,079. The CCyR cumulative incidence was also lower in patients with Im400-HDIm-2GTKI and Im400-2GTKI in comparison to the other groups, although second line response was faster in patients who changed to 2GTKI after Im400. The frequency of CCyR as best response in the Hasford high risk patients was low in all groups (66%,50%,50%,50&55%).

Major molecular response MMR as best response was lower in patients with Im400-HDIm-2GTKI (50%) and Im400-2GTKI (47%). The rates were 83%, 81% and 77% in the Im400, Im400-HDIm and HDIm groups respectively; P Chi2 19,4(a)p=0,001. The MMR cumulative incidence was higher in the HDIm group, lower in those treated with Im400-HDIm-2GTKI, and intermediate and similar in the other three groups. MMR as best response in the Hasford high risk patients was also low in all groups (60%, 75%, 50%, 50% & 33%).

Complete molecular response regarding best response, the CMR rate was lower in patients with Im400-HDIm-2GTKI (37,5%), Im400-2GTKI (31,6%) and Im400-HDIm (34%). In the other groups, the rate was 48% (Im400), and 72% (HDIm); P Chi2 17,4(a) p=0,002. The CMR cumulative incidence was higher in the HDIm group, and nil in those treated with Im400-HDIm-2GTKI.

Salvage therapy after suboptimal response (SR) or Failure (F). Two-thirds of patients with SR or F were able to obtain an optimal response and avoid transformation with a timely therapy change. All but one of the options (Im400-HDIm-2GTKI group) were similarly effective.

Survival:

6 patients progressed (2,4%) (4 AP, 2 BC), and died; 6 patients changed to allo BMT and were censored; 6 patients died of non-CML related causes.

Conclusion:

Disclosures:

Palomera: Janssen Cilag: Honoraria. Steegmann:Bristol-Myers Squibb: Honoraria, Participated in advisory boards, Research Funding; Novartis: Participated in advisory boards, Research Funding.
Treosulfan, Etoposide and Cyclophosphamide as Non TBI-Conditioning Regimen Followed by Allogeneic SCT for Acute Lymphoblastic Leukemia
Authors: Kröger Nicolaus , Stelljes Matthias , Bornhäuser Martin , Bethge Wolfgang A. , Arnold Renate , Martin Hans , Schmid Christoph , Kolbe Karin , Goekbuget Nicola , Hoelzer Dieter , Zander Axel R. , Beelen Dietrich
Published: October 2019

Abstract 1327

Total body irradiation (TBI) is an essential part of the conditioning regimen for acute lymphablastic leukaemia (ALL) patients who undergo allogeneic stem cell transplantation. Due to high toxicity and mortality induced by TBI, we investigate in a multicenter prospective phase II study the toxicity and efficacy of a non TBI-based regimen consisting of treosulfan (3 × 12 g/m2), etoposide (30 mg/kg), and cyclophosphamide (120 mg/kg) in patients with ALL who underwent allogeneic stem cell transplantation. Major inclusion criteria were complete remission, indication for allogeneic stem cell transplantation according GMALL, non eligibility for TBI or patient's wish to avoid TBI. Graft-versus host prophylaxis consisted of cyclosporine A and short course methotrexat and in case of unrelated donors also of anti-lymphocyte globulin (ATG). This preliminary analysis is based on 36 patients (female: n=17; male: n=19) with a median age of 42 years (range: 18–60y). Remission-status at transplantation was either 1. CR (n=27), 2.CR (n=7) or >2.CR (n=2). Donors were HLA-identical sibling (n=6), matched unrelated (n=21) or mismatched unrelated (n=9). 39% of the patients were Philadelphia chromosome positive. Primary graft failure was observed in 1 pts. The median time to achieve leukocyte (> 1×109/l)) and platelet (>20 × 109/l) engraftment was 22 (range, 11–47) and 24 days (range, 11–95), respectively. The toxicity was moderate including VOD (6%) and liver toxicity grade III (8%) and IV (3%). Acute graft versus host disease grade II-IV and grade III/IV was noted in 22% and 11%, respectively. Chronic graft versus host disease at 1 year was seen in 27%, which was extensive in 7% of the patients. After a median follow up of 12 months, the cumulative incidence of non-relapse mortality (NRM) at 1 year was only 9% (90% CI: 1–17%) and for relapse 37% (90% CI: 21–52%). The estimated 1-year disease free survival was 56% (90% CI:40-71%) and significantly better for patients transplanted in 1.CR vs 2. or higher CR (68% vs 15%, p=0.05). The estimated 1-year overall survival was 81% (90% CI: 69–92%). This preliminary results of a treosulfan, non TBI based conditioning regimen followed by allogeneic stem cell transplantation shows favourable toxicity profile with low non-relapse mortality. Longer follow up is needed to determine long-term freedom from disease.

Disclosures:

Kröger: MEDAC: Research Funding. Off Label Use: Treosulfan is not approved as conditioning regimen for stem cell transplantation.
Higher Non-Relapse Mortality with BuCyVP Compared to BuCy In Allogeneic Hematopoietic Stem Cell Transplant
Authors: Duong Hien , Bolwell Brian J. , Rybicki Lisa , Kalaycio Matt , Andresen Steven , Sobecks Ronald , Dean Robert , Sweetenham John William , Tench Shawnda , Copelan Edward A.
Published: October 2019

Abstract 1340

We analyzed results of allogeneic hematopoietic stem cell transplantation from 1/2000 to 1/2009 in 520 adult patients, receiving either bone marrow or peripheral stem cells, in order to identify factors associated with non-relapse mortality (NRM). Median age at transplant was 47 years (range 18–70). Median follow up was 43 months (range 5–122 months). Two hundred eighty four (55%) patients were male. Diagnoses included: acute myeloid leukemia 210 (40%), acute lymphoblastic leukemia 68 (13%), myelodysplastic syndrome 66 (13%), non-Hodgkin or Hodgkin lymphoma 64 (12%), chronic myelogenous leukemia 45 (9%), or other (13%). High scores (≤3) on hematopoietic cell transplant-specific comorbidity index were identified in 162 (31%) patients. For preparative regimens, 187 (36%) received BuCy (busulfan 1mg/kg oral given every 6 hours × 16 doses days -8 to -4, followed by cyclophosphamide 60mg/kg × 2 days -3 to -2), 91 (18%) received BuCyVP (busulfan 1mg/kg oral every 6 hours × 16 doses days -9 to -5, followed by etoposide 60mg/kg days -5 to -4, then cyclophosphamide 60mg/kg days -3 to -2), 225 (43%) received TBI-based therapy, and 17 (3%) received other therapy. Two hundred ninety five (57%) underwent related transplant. A majority (67%) received stem cells derived from bone marrow.

In univariable analysis, male recipient (p=0.07), high comoborbidity score (p=0.004), more prior chemothepary regimens (p=0.037), BuCyVP (p<0.001) or TBI-based preparative regimen (relative to BuCy, p=0.045) and unrelated donor (p=.001) were associated with high NRM. In multivariate analysis, male recipient (p=0.008), high comorbidity score (p=0.002), and BuCyVP compared to BuCy (p<0.001) remained independent risk factors for NRM. There were no significant differences in the proportions of identified cause of death between regimens. Figure 1 illustrates the sustained influence of preparative therapy on NRM:

Factors associated with poor overall survival (OS) and relapse-free survival (RFS) on multivariate analysis included male recipient (p=0.001, p=0.003, respectively), older age at transplant (p=0.048 for RFS), high comorbidity score (p<0.001, p<0.001), more prior chemotherapy regimens (per 1 regimen increase, p=0.22, p=0.02 respectively), and BuCyVP compared to BuCy (p=0.007, p=0.021).

Long-term follow-up of a large cohort of patients indicates that compared to BuCy, BuCyVP is associated with significantly higher NRM and lower OS and RFS.

Disclosures:

Sweetenham: Pfizer: Research Funding.
Prognostic Index for Predicting Overall Survival and Event-Free Survival In Total Therapy 3 Patients
Authors: Hoering Antje , Mitchell Alan , Sexton Rachael , Crowley John , Barlogie Bart
Published: October 2019

Abstract 1342

Background:

Today's front-line treatment of Multiple Myeloma is frequently based on the MIRT experience with Total Therapy 3 (TT3). We developed prognostic models and a web tool based on those models to predict survival and progression free survival.

Patients and methods:

Data from 303 patients enrolled on UARK 2003-33 and 177 patients treated on UARK 2006-66 was examined. Multivariate Cox regression analysis was used to determine the best prognostic model for overall survival (OS) and event-free survival (EFS) both in the presence and absence of baseline GEP-based risk information.

Results:

Variable selection resulted in consistent models; both OS and EFS models contained similar elements when the selection process terminated. When GEP-defined 70-gene risk information was available, variable selection for OS resulted in a model containing GEP-risk, β2M, cytogenetic abnormalities and LDH. All variables were chosen to be dichotomous. When GEP-defined 70-gene risk information was not available, we obtained two competing models for both OS and EFS. The variables significant for those models were β2M, cytogenetic abnormalities and LDH. In addition to those variables, albumin entered the competing model. Cumulative R squared, measuring the amount of statistical variability accounted for in a statistical model, was comparable in both. We therefore chose the model that is complementary to the model derived when GEP- based risk information was available. All statistical models used for predicting survival and event-free survival have a cumulative R squared of at least 30. The parameter estimates determined in those models were used to evaluate the predicted OS and EFS for a patient with specific baseline features. Two examples for a predicted OS curve with available GEP data are shown below. Figure 1 shows predicted overall survival with 95% confidence interval for a patient with low risk GEP, no cytogenetic abnormalities, β2M ≤ 5.5 mg/L and LDH < 190 U/L. Figure 2 displays the predicted overall survival with 95% confidence interval for a patient with high-risk GEP, cytogenetic abnormalities, β2M > 5.5 mg/L and LDH ≥ 190 U/L.

Conclusion:

A web tool was developed with radio-buttons that enable selection of variables based on observed baseline features of a specific patient. The tool then evaluates and displays the predicted OS and EFS curves with 95% confidence intervals based on the parameter estimates of the underlying multivariate model described above.

Disclosures:

No relevant conflicts of interest to declare.
High Dose Melphalan with Autologous Stem Cell Transplantation Overcomes Poor Prognosis of Primary Amyloidosis
Authors: Parmar Simrit , Khan Mubeen , Rondon Gabriela , Shah Nina , Bashir Qaiser , Tung Shawndeep , Shah Jatin J , Weber Donna , Orlowski Robert Z. , Giralt Sergio , Champlin Richard E. , Qazilbash Muzaffar H.
Published: October 2019

Abstract 1350

Background:

Systemic Primary AL Amyloidosis is a rare but potentially fatal disease resulting from tissue deposits of amyloid fibrils derived from monoclonal immunoglobulin light chains. High-dose melphalan followed by autologous hematopoietic stem cell transplant (auto HCT) is associated with hematologic and organ responses and improved survival.

Methods:

In this retrospective analysis we identified 46 patients with primary AL amyloidosis who received auto HCT between 01/1998 to 05/2010 at MDACC. Organ responses were determined using Amyloidosis Consensus Criteria.

Results:

The median age at auto HSCT was 56 years (34-74) where 61% were males and 35% were older than 60 years of age. 61% had lambda light chain restriction and only 4% had cytogenetic abnormalities. Disease characteristics are summarized in Table 1. The median time from diagnosis to auto HCT was 6.6 months (2.2-29.4 months). 22 pts (47.8%) had one organ, 19 pts (41.3%) had 2 organ and 4 pts (8.7%) had 3 organ involvement. 11 pts (23.9%) had heart and 35 pts (76.1%) had kidney involvement. The median follow up from the time of diagnosis was 22.4 months and from time of auto HCT was 16.7 months. High dose Melphalan dose was 200mg/m2 in 24 pts (52%) and 140mg/m2 in 22 (47.8%). There were 4 early deaths and 4 pts whose follow up was less than 3 months and their response was not assessed. Out of the 38 evaluable patients, the post-transplant organ responses were as follows ≥PR 25(66%), ≥stable disease 35(92%) (Table2). The hematologic responses were: CR=5 (13%), ≥VGPR=10(26%), ≥PR=26 (68%), ≥SD=37(97%). One patient had progressive disease. There was a correlation between organ response and hematologic response (chi square;p<10-3). The day-100 treatment related mortality (TRM) was 8.7% and 1-yr TRM was 13%. The median progression-free (PFS) and overall survival (OS) from auto HCT was 73.8 months and not reached (from transplant). The median PFS and OS from diagnosis were 93 months and 59.8 months respectively. In multivariate analysis, heart involvement (p=0.01), female sex (p=0.011), age ≥60 years (p=0.002), bone marrow plasma cells≥10% (p=0.043) and Beta-2 microglobulin>3.5mg/l (p=0.02) were associated with poor OS. Improved OS correlated with organ response (52.6 vs 11.4 months; p=0.01) and hematologic response (52.6 vs.6.1months; p=0.002). Hemoglobin <10 g/dl (p=0.047), bone marrow plasma cells≥10% (p=0.043) and age≥60 years (p=0.075) were associated with shorter PFS. Hematologc response (p=0.48) and organ response (p=0.12) were not associated with improved PFS.

Conclusion:

In this analysis the outcome of patients with primary systemic AL amyloidosis was durable with auto HCT with acceptable mortality risk and improved survival.

Disclosures:

No relevant conflicts of interest to declare.
TP53 Positively Regulates Cell Death through TRAILR2 (DR5) but Negatively through TRAILR1 (DR4) In Myeloma Cells
Authors: Surget Sylvanie , Gomez-Bougie Patricia , Chiron David , Humphreys Robin , Moreau Philippe , Le Gouill Steven , Harousseau Jean-Luc , Bataille Regis , Amiot Martine , Pellat-Deceunynck Catherine
Published: October 2019

Abstract 136

Mapatumumab and lexatumumab are human antibodies that bind and activate death receptors TRAILR1/TNFSF10A/DR4 and TRAILR2/TNFSF10B/DR5, respectively. Treatment of primary myeloma cells and myeloma cell lines with these mAbs induced cell death. Mapatumumab induced cell death more effectively than lexatumumab in a panel of 30 human myeloma cell lines (HMCLs). Interestingly, sensitivity to mapatumumab and lexatumumab was mutually exclusive and related to TP53 status (p=0.006). Indeed, wildtype TP53 HMCLs (n=9) were sensitive to lexatumumab (mean of death 40%) but resistant to mapatumumab (mean of death 7%). In contrast, abnormal (n=21) TP53 HMCLs were resistant to lexatumumab (mean of death 7%) but sensitive to mapatumumab (mean of death 44%). Of note, killing by lexatumumab was correlated to TRAILR2 expression while no correlation was found for TRAILR1 expression and lapatumumab killing. Transcriptomic analysis of 30 HMCLs revealed that HMCLs with abnormal TP53 status underexpressed 4 well-known p53 target genes, MDM2, CDKN1A, Bax and TRAILR2 (p<0.01). To activate p53 pathway in myeloma cells, we used melphalan at low doses. Melphalan treatment of wildtype HMCLs, but not of TP53 abnormal HMCLs, increased TRAILR2 expression and cell death mediated by lexatumumab. In contrast, melphalan did not alter TRAILR1 expression or mapatumumab-induced killing, suggesting that TRAILR2 but not TRAILR1 is a p53 target in myeloma cells. Silencing TP53 significantly increased mapatumumab apoptosis (>50% p<0.01). In good agreement with Bax underexpression in TP53 abnormal HMCLs, extensive silencing of key molecules of both extrinsic (caspase 8, caspase 3) and intrinsic pathways of apoptosis (caspase 9, Bid, Bim, Bax) showed that mapatumumab killing was dependent on the extrinsic pathway of apoptosis only: only silencing of caspase 8 or caspase 3 inhibited mapatumumab killing. Altogether, these data show that killing through TRAIL receptors is differentially regulated by p53 in myeloma cells, positively for TRAILR2 but negatively for TRAILR1. Interestingly, myeloma cells with an abnormal p53 that are more resistant to all drugs are more sensitive than wt ones to killing through TRAILR1 making this pathway very attractive for p53 deficient myeloma cells.

Disclosures:

No relevant conflicts of interest to declare.