bluuebunny/crossref_metadata_2025 huggingface dataset:
Systemic Relapses In Primary CNS Lymphoma (PCNSL) - Unexpected Clonal Diversity
Authors: Fischer Lars , Hummel Michael , Korfel Agnieszka , Lenze Dido , Jahnke Kristoph , Kiewe Philipp , Weller Michael , Thiel Eckhard
Published: October 2019

Abstract 3125

Background:

PCNSL is a rare form of extranodal diffuse large B-cell lymphoma (DLBCL). Relapses after initial therapy are usually within the CNS. Systemic relapses (SR) have been reported only occasionally. We conducted a systemic analysis of immunocompetent patients with PCNSL und SR using both clinical data and tumor samples at first diagnosis and SR to determine clonal relatedness.

Methods:

Records of patients included in a multicenter therapy trial (n=411) and all PCNSL patients treated at our institution outside the trial after 1993 (n=39) were reviewed for SR. PCR analysis of the rearranged Ig heavy-chain (IgH) and light-chain (IgL) genes was performed according to the BIOMED-2 protocol.

Result:

All patients had received high-dose methotrexate based chemotherapy with our without adjuvant whole brain irradiation as first line therapy. Of 450 patients 26 (5.8%) were identified with SR. Median time from PCNSL diagnosis to SR was 14.5 months (range 2–61). SR was extranodal in 22 (85%) patients: testicular n=3, gastric n=3, mamma n=3, liver n=2, bone/bone marrow n=3, intestinal n=1, [sub-]cutaneous/muscular n=5, pancreatic n=1, pulmonary n=1. Four patients had concomitant CNS relapse. Median overall survival from diagnosis was 38 months, from SR 9.8 months. Causes of death were SR in 13, CNS relapse in 3, toxicity of chemotherapy in 3 and unknown in 1 patients; 6 patients were alive at last follow up. All 26 patients initially had DLBCL. SR was biopsied in 20 patients: 19 had DLBCL, 1 a Burkitt-like DLBCL. DNA of 7 patients was sufficient for PCR analysis. The same clone was found in the CNS and SR in 4 patients, in 3 patients the clones were different. Clonally related SR occurred after 4, 8, 14 and 25 months; the unrelated SR after 30, 35 and 36 moths.PCR products in 2 of 4 clonally related samples were suitable for sequencing. One case exhibited 6 identical somatic mutations in both biopsies. The second case harbored 35 mutations in the CNS sample and 45 in the SR sample. Here 26 mutations were identical in both samples, 3 involved different nucleosides, 16 were only present in SR, and 6 mutations exclusively present in the CNS sample.

Conclusion:

SR is a rare and frequently fatal complication in PCNSL. The localization of SR is predominantly extranodal. A proportion of SR is not clonally related to the primary tumor and then most likely represent a secondary lymphoma. In clonally related SR the presence of somatic mutations unique to the CNS tumor and the SR might be explained by a common ancestor, whose localization however remains unknown.

Disclosures:

No relevant conflicts of interest to declare.
Growth Factor Independent-1 (Gfi1) Is Critically Required for T-Cell Acute Lymphoblastic Leukemia (T-ALL) Tumor Initiation and Maintenance
Authors: Phelan James D. , Khandanpour Cyrus , Horman Shane , Gaudreau Marie-Claude , Zhu Jinfang , Paul William E , Dührsen Ulrich , Grimes H. Leighton , Moroy Tarik
Published: October 2019

Abstract 3156

T cell acute lymphoblastic leukemia (T-ALL) is one of the most common childhood cancers associated with mutations in NOTCH1. The Growth factor independent-1 (Gfi1) transcriptional repressor gene was originally discovered as a common target of Moloney murine leukemia virus (MMLV) proviral insertion in murine T-ALL. In fact, the Gfi1 locus is the most frequently activated gene in MMLV-induced T cell leukemia. Therefore, we investigated whether the most commonly activated gene in MMLV-induced murine T-ALL, Gfi1, could collaborate with the most commonly activated gene in human T-ALL, NOTCH1. Here, we show that GFI1 expression is associated with Notch signaling in human T-ALL (p'0.0003). Functionally, Gfi1 collaborates with Notch-induced murine T-ALL by accelerating an already rapid disease model (p=0.03) without altering the lymphoblastic nature of the disease. Furthermore, inducible deletion of Gfi1 is counter-selected in both Notch-driven retroviral and transgenic mouse models of T-ALL; whereas, constitutive absence of Gfi1 completely prevents transgenic Notch-induced T-ALL (p≤0.04). However, T-ALL tumors can form in Gfi1-/- animals using either ENU-mutagenesis or MMLV-infection, yet tumor formation is delayed (p≤0.02, p≤0.03 respectively). This suggests that Gfi1 deletion does not prevent the formation of the T-ALL initiating cell and that Gfi1 might be absolutely required for Notch-induced T-ALL. Most striking is that Gfi1 is required for T-ALL maintenance in vitro and in vivo. Using three separate Tal1-initiated murine T-ALL cell lines, the overexpression of the Gfi1 dominant-negative, Gfi1N382S, was quickly and completely counter-selected. As Gfi1 has previously been found to regulate pro-apoptotic genes in T cells, we attempted to rescue the above loss of function phenotype by overexpressing the anti-apoptotic factor Bcl2. Notably, counter-selection of Gfi1N382 is not observed or is significantly delayed in all three cell lines. In vivo, inducible deletion of Gfi1 leads to both mutagen- or Notch-induced tumor regression as measured by ultrasound. In fact, levels of Gfi1 expression directly correlate to tumor regression and disease free survival of T-ALL. Finally, targeting Gfi1 enhances the efficacy of radiation therapy and bone marrow transplantation. Deletion of Gfi1 sensitizes T-ALL tumors and T cells to p53-dependent apoptosis after exposure to DNA-damaging agents such as radiation, Etoposide or Daunorubicin by de-repression of the pro-apoptotic Gfi1 target gene Bax. These data extend the role of Gfi1 to human T-ALL and suggest that T-ALL is dependent upon Gfi1 to repress pro-apoptotic genes for tumor survival, ultimately highlighting a new therapeutic target in the fight against lymphoid malignancies.

Disclosures:

No relevant conflicts of interest to declare.
Prevalence of Venous Thromboembolism In Patients with Secondary Polycythemia
Authors: Nadeem Omar , Gui Jiang , Ornstein Deborah
Published: October 2019

Abstract 3164

Background:

Polycythemia vera is associated with an increased risk for venous thromboembolism (VTE). Although phlebotomy is employed as an adjunct to treatment with hydroxyurea and/or aspirin for VTE risk reduction, emerging data suggest that hematocrit is less of a determinant of VTE risk than leukocyte count and JAK2 V617F gene mutation allele burden. The role of secondary polycythemia as a risk factor for VTE is unknown, but phlebotomy for thrombosis risk reduction is frequently practiced. Based on the polycythemia vera model, however, we hypothesize that secondary polycythemia does not increase VTE risk. The purpose of this study was to determine the prevalence of VTE in patients with secondary polycythemia and to investigate the factors associated with VTE in this population.

Methods:

We performed a case control study that included patients admitted to Dartmouth-Hitchcock Medical Center with a diagnosis of chronic obstructive pulmonary disease (COPD) and a hematocrit greater than or equal to 50% from August 2004 to July 2009. The controls were matched for age and sex and carried a diagnosis of chronic obstructive pulmonary disease (COPD) without evidence of secondary polycythemia. Data were collected on body mass index (BMI), VTE history, comorbid conditions, thrombophilia and smoking history. Clinical characteristics of patients with and without secondary polycythemia were analyzed using chi square and t-test to evaluate for significant differences in the two populations.

Results:

Eighty-six patients with secondary polycythemia and 86 controls were included in the study. The mean hematocrit was 53.5% in the case group and 43.7% in the control group (p=<0.005). Among cases, a history of VTE was documented in 17/86 (19.8%), 10 of which (58.8%) were judged to be idiopathic. In the control group, VTE was documented in 12/86 (14%), 4 of which (33.3%) were judged to be idiopathic. There was no statistically significant difference in the number of total (OR=1.52, p=0.42) or idiopathic (OR=2.7; p=0.16) VTE between cases and controls, respectively. There were no statistically significant differences noted in age, sex, body mass index, presence of diabetes mellitus, smoking history or the presence of malignancy in the two groups. Patients with VTE in both groups had higher BMI, however, compared to patients without VTE.

Conclusions:

We did not observe an increased prevalence of VTE in patients with secondary polycythemia compared to age- and sex-matched controls. Our findings suggest that the high prevalence of VTE observed in patients with secondary polycythemia is more likely related to known risk factors such as obesity rather than hyperviscosity due to increased cell mass. The role of phlebotomy for VTE risk reduction in patients with secondary polycythemia is therefore questionable.

Disclosures:

No relevant conflicts of interest to declare.
Is Higher Soluble P-Selectin a Risk Factor for Chronic Venous Insufficiency?
Authors: Bryan Locke J , Callas Peter , Criqui Michael , Cushman Mary
Published: October 2019

Abstract 3166

Background:

Chronic venous insufficiency (CVI) represents a spectrum of disease characterized by impaired venous return. Deep venous thrombosis (DVT) when complicated by post thrombotic syndrome (PTS) is a well established etiology for CVI. P-selectin is a cell adhesion molecule involved in thrombus formation; higher levels are related to DVT risk. Thus, P-selectin may play a role in CVI pathophysiology.

Objective:

We evaluated whether higher levels of soluble P-selectin are a risk factor for CVI.

Methods:

Data were collected from the San Diego Population Study, an established cohort of 2,404 men and women, in whom CVI was evaluated by symptoms, physical examination, and duplex venous ultrasound. A case-control study was conducted within the cohort to analyze the association of P-selectin with risk and severity of CVI. 352 controls without venous abnormalities were matched by age, race, and sex to 352 cases. Cases consisted of four groups with increasing severity of CVI based on clinical and imaging characteristics. Soluble P-selectin was measured in cases and controls using a commercial immunoassay. Multivariable models were used to determine associations of P-selectin with CVI, controlling for potential confounders.

Results:

P-selectin levels did not differ by CVI risk factors including age, sex, race, body mass index (BMI), smoking status, diabetes, prior DVT, venous surgery, and history of cancer (not shown). Overall, P-selectin was not associated with risk of CVI. However, comparing the two most severe case groups to controls, the age, sex, and race adjusted odds ratios (ORs) of CVI for P-selectin in the top two versus bottom tertiles were increased at 1.4 and 2.3, respectively (see table), with the top tertile OR being statistically significant. The association of P-selectin in the top tertile with CVI were slightly weakened with adjustment for BMI, as the case groups with higher P-selectin levels had on average a higher BMI. Successive addition of other risk factors to the model, shown in the table, had no impact on the ORs.

Conclusions:

Higher levels of soluble P-selectin are associated with risk of more severe, but not mild, CVI. This suggests that thrombosis potential or platelet function is important in the pathogenesis of this disease.

Disclosures:

No relevant conflicts of interest to declare.
The Metabolic SYNDROME and the RISK of Venous Thrombosis: RESULTS of AN Individual LEVEL Patient Meta-ANALYSIS
Authors: Dentali Francesco , Ay Cihan , Jang Moon , di Minno Matteo , Pabinger Ingrid , Oh Doyeun , di Minno Giovanni , Squizzato Alessandro , Grandi Anna Maria , Ageno Walter
Published: October 2019

Abstract 3177

Background:

The metabolic syndrome (MS) is a cluster of interrelated risk factors that identify patients at increased risk of cardiovascular events. Recent studies also suggested an association between MS and venous thromboembolism (VTE). However, the role of the individual features of MS and whether MS and its features are more important than obesity alone to predict VTE remain to be established.

Methods:

We performed an individual patient level meta-analysis of case-control studies comparing the prevalence of MS in patients with unprovoked VTE and in controls. MEDLINE, EMBASE databases, and abstract books were searched up to January 2010. Odds ratios (OR) and 95% confidence intervals of pooled results were calculated. The influence of individual variables (age, sex, BMI and MS) on the likelihood of VTE was compared using logistic regression analysis. Multivariate analysis was subsequently performed including the individual components of MS in the place of MS. The impact of increasing number of individual components of MS on the risk of VTE was investigated.

Results:

Four studies were identified and analyzed, for a total of 1332 patients (479 cases and 833 controls). Mean age was 53.3 and 52.7, respectively (p=n.s.), 49.5% cases and 42.4% controls were males (p=0.0003), 38.8% and 30.0% were obese (p=0.0001). MS was significantly associated with VTE (OR 1.97, 1.57–2.47), and the association linearly increased with the number of MS features (p for trend <0.001). At multivariate analysis, MS but not obesity remained associated with VTE (OR 1.92, 1.50–2.46 and 1.14, 0.88–1.47, respectively). All individual features of MS, but HDL cholesterol, were independently associated with VTE.

Conclusions:

The results of this meta-analysis confirm the association between MS and VTE and suggest that MS (and visceral obesity defined by increased waist circumference) could be a more important predictor of VTE than obesity defined by BMI.

Disclosures:

No relevant conflicts of interest to declare.
Differential Pathogenesis of Bernard-Soulier Syndrome-Associated Mutations In the Platelet Glycoprotein Ibβ Ectodomain
Authors: Yang Wenjun , McEwan P.A , Mo Xi , Emsley Jonas , Li Renhao
Published: October 2019

Abstract 3202

Eight missense mutations in the ectodomain of glycoprotein (GP)Ibβ have been identified in patients with Bernard-Soulier syndrome (BSS) that is characterized by the deficiency of functional GPIb-IX complex on the platelet surface, clearly highlighting the importance of GPIbβ ectodomain in assembly of the GPIb-IX complex. To understand the molecular pathogenesis of these mutations, we have characterized their effects on the expression, secretion, folding of the isolated GPIbβ ectodomain as well as its interaction with GPIX ectodomain in the context of full-length complex. Each of the 8 mutations — C5Y, R17C, P29L, N64T, P74R, Y88C, P96S, and A108P — was constructed into genes encoding HA-tagged GPIbβ ectodomain or full-length GPIbβ subunit, and the mutant gene transfected transiently, along with GPIba and GPIX genes if desired, into Chinese hamster ovary (CHO) cells. Flow cytometry and Western blot analysis indicated that while all 8 mutations impeded formation of the disulfide bonds between GPIba and GPIbβ and significantly decreased the surface expression level of GPIb-IX complex comparing to the wild-type, the extent of disruption varies with each mutation. Further characterization in the context of isolated GPIbβ ectodomain revealed that the majority of 8 mutations — C5Y, R17C, P29L, N64T, Y88C, P96S — are detrimental to proper folding of the GPIbβ ectodomain, resulting in secretion defects and/or domain misfolding. In contrast, two mutations, P74R and A108P, preserved structural integrity of the GPIbβ ectodomain since the mutant ectodomains exhibited wild-type-like secretion levels and formed no inter-molecular disulfide bonds. However, neither of the two mutations, in the context of full-length GPIbβ, were able to support surface expression of GPIX in transfected CHO cells as the wild-type, indicating that P74R and A108P disrupt the interaction between GPIbβ and GPIX ectodomains. Thus, our results demonstrated although all 8 BSS mutations in GPIbβ share the same phenotype, they impair expression of the GPIb-IX complex by two different mechanisms — disrupting folding of the GPIbβ ectodomain or disrupting interactions between GPIb-IX subunits. Furthermore, our results suggest that Pro74 and Ala108 may be located in the interfacial region between GPIbβ and GPIX ectodomains, helping to shed light on the structure of GPIb-IX complex.

Disclosures:

No relevant conflicts of interest to declare.
A Reassessment of the Red Blood Cell Folate Assay
Authors: Hoyer James , Oliveira Jennifer , Baumann Nikola , Saenger Amy
Published: October 2019

Abstract 3209

Background:

The red blood cell folate (RCF) assay has historically been recommended as a more reliable indicator of tissue folate stores compared to the serum folate (SF) assay, as it is not affected by recent ingestion of food. However, the RCF assay suffers from inherent problems with imprecision and accuracy, which are not encountered with SF measurements. Furthermore, following the advent of required folic acid supplementation of many foods by the Food and Drug Administration (FDA) in 1992, folate deficiency is increasingly rare. Very few studies have looked at the value of the RCF versus the SF. We undertook a 10 year retrospective analysis of RCF and SF results to determine the clinical utility of RCF beyond that of SF.

Methods:

We retrieved all RCF and SF results from the laboratory information system at Mayo Clinic (Rochester, MN) ordered on inpatients and outpatients between 1999–2009. Data for patients who had concurrent orders for SF and RCF were analyzed and chart reviews were conducted on those patients with normal SF but low RCF. Abnormal values were defined by the National Health and Nutrition Examination Surveys (NHANES)/Center for Disease Control (CDC) criteria for folate deficiency (SF< 3.0 ng/ml, RCF < 140 ng/ml).

Results:

A total of 152,166 SF and 15,708 RCF were performed over the decade of the study. The prevalence of folate deficiency using only SF values was 0.39% and 0.27% using only RCF values. There were 1082 patients in which SFA and RCFA were ordered concurrently (Table 1).

Only 1 patient (0.09%) had both abnormal SF and RCF. Chart reviews of the 4 patients with a normal SF but low RCF were as follows: 1) a 6 year old (yo). male with known folic acid transporter deficiency treated with Leucovorin. 2) a 58 yo male with history of gout, hypertension, psoriasis, and hyperlipidemia with normal hemoglobin (Hb) and MCV. 3) a 65 yo male with chronic diarrhea and suspected alcohol abuse; slight macrocytosis (MCV=100.3 fL) but normal Hb. 4) a 51 yo male with multifactorial gait disorder and alcohol abuse. There was a previous history of vitamin B12 deficiency but B12 levels were normal at this time. The CBC was notable for macrocytosis (MCV=115.1 fL) without anemia. Only in patient 4 did the RCF value result in the institution of folic acid supplementation.

Conclusions:

The RCF provides no additional information beyond that provided by the SF in virtually all situations. Thus SF alone is sufficient for assessment of folate stores. However, there is no evidence to support routine ordering of either SF or RCF, as true folate deficiency in the current era of FDA mandated folic acid supplementation is exceedingly rare.

Disclosures:

No relevant conflicts of interest to declare.
Connectivity Mapping Identifies HDAC Inhibitors as Suitable Candidates for the Treatment of t(4;11)-Positive Acute Lymphoblastic Leukemia In Infants
Authors: Stumpel Dominique JPM , Schneider Pauline , Seslija Lidija , Osaki Hikari , Williams Owen , Pieters Rob , Stam Ronald
Published: October 2019

Abstract 3222

To date, MLL-rearranged infant Acute Lymphoblastic Leukemia (ALL) remains the most aggressive type of childhood leukemia characterized by a high rate of early relapses and a grim prognosis. This type of leukemia arises from chromosomal translocations involving the Mixed Lineage Leukemia (MLL) gene. The genesis of such translocations seems to be initiated in utero. Hence, MLL-rearranged ALL is typically diagnosed in infants at or shortly after birth. The most common MLL translocation among infant ALL patients, occurring in about 50% of the cases, is t(4;11) generating the oncogenic fusion product MLL-AF4.

MLL-rearranged ALL is distinguishable from other ALL subtypes by unique gene-expression profiles. While inappropriate gene activation seems to be largely driven by specific histone modifications, we recently showed that gene inactivation in MLL-rearranged infant ALL can be explained by aberrant promoter hypermethylation. Focusing on genome-wide DNA methylation, we here uncovered that MLL-rearranged infant ALL is characterized by the activation of particular set of (proto-onco)genes as a result of abnormal promoter hypomethylation.

In search for therapeutic agents capable of targeting these potential cancer-promoting genes, we applied connectivity mapping on a gene expression signature based on the genes most significantly hypomethylated in t(4;11)-positive infant ALL as compared with healthy bone marrows. Connectivity map analyses revealed that histone deacetylase (HDAC) inhibitors represent the most suitable compounds to reverse this gene signature. We show that six different HDAC inhibitors effectively induce leukemic cell death in primary t(4;11)-positive infant ALL cells in vitro. These cytotoxic responses were accompanied by down-regulation of MYC, SET, RUNX1 and RAN, and were preceded by degradation of the oncogenic MLL-AF4 fusion product.

Our data shows that DNA methylation profiles allow the identification hypomethylated proto-oncogenes, and underlines the essential role for epigenetic de-regulation in MLL-rearranged ALL. Furthermore, we show for the first time that connectivity mapping can indirectly be applied on DNA methylation patterns, providing a rationale for HDAC inhibition in t(4;11)-positive leukemias. Given the presented potential of HDAC inhibitors to target important proto-oncogenes including the leukemia-driving MLL fusion in vitro, these agents should urgently be tested in in vivo models and subsequent clinical trials.

Disclosures:

No relevant conflicts of interest to declare.
Adolescent and Young Adult Patients Treated with Modified Augmented Berlin-Frankfurt-Muenster Therapy
Authors: Rytting Michael E. , Kantarjian Hagop M. , Thomas Deborah A , Jabbour Elias , Franklin Anna , Ravandi Farhad , Faderl Stefan , Garris Rebecca , Kadia Tapan , Schroeder Kurt , O'Brien Susan
Published: October 2019

Abstract 3250

Pediatric protocols for acute lymphoblastic leukemia (ALL) are currently employed by many groups to treat adolescents and young adults (AYA) with ALL. Adults up to age 55 are presently enrolled in such treatment regimens, and investigators have reported excellent results in these young adult patients (pts). Augmented Berlin-Frankfurt-Muenster therapy has been shown to be effective therapy for adolescents with ALL. To evaluate the efficacy and toxicity of this therapy in young adults, we designed a protocol of modified augmented Berlin-Frankfurt-Muenster (ABFM) therapy for patients with ALL from age 12 to 40. 70 pts have been enrolled on protocol and are evaluable for toxicity. 60 pts with de novo Philadelphia chromosome negative ALL have completed at least 29 days of therapy (induction) on protocol. They have the following characteristics: 52 (87%) pts have pre-B ALL and 8 (13%) have T-ALL. The median age is 20 (mean=22; range=13-39). The median presenting WBC=5.2 (mean=32; range= 0.4–494). 58/60 (97%) pts have achieved a remission. There have been no induction deaths. 49(82%) pts have attained remission by 15 days of induction, while 11 (18%) have been slow responders. 5/60 (8%) pts have required an extension of induction by 2 weeks. At day 29 of therapy, 38 (63%) pts had negative minimal residual disease (MRD) by flow cytometry, 15 (25%) patients were positive for MRD, 7 were suspicious or not available. By day 84 of therapy, 46 (77%) pts were negative for MRD and 9 (15%) were positive. Currently, there have been 10 relapses and 9 deaths. Toxicities in the entire group include severe asparaginase allergy in 15 (21%) pts, thrombus formation in 15 (21%) pts, hyperbilirubinemia grade III-IV in 19 (27%) pts, ALT/AST grade III-IV in 20 (28%) pts, CNS stroke-like symptoms in 6 (8%) pts, and avascular necrosis in 6 (8%) pts. Hepatic toxicity has resolved completely within two weeks in almost all pts as has the CNS toxicity. Grade III-IV hepatic toxicity have been transient. For pts 25 years of age and younger (n=25), the overall survival (OS) and disease free survival (DFS) at 2 years are 91% and 85% respectively. For pts > 25 years of age (n=15), the OS and DFS at 2 years are 55% and 61% respectively. This difference is statistically significant for the two groups; p=0.04 for DFS and p=0.004 for OS. Continued enrollment is anticipated to further evaluate the survival differences between these two patient groups.

Disclosures:

No relevant conflicts of interest to declare.
Leukemia-Specific Ligands Against Childhood Acute Lymphoblastic leukemia
Authors: Satake Noriko , Sarangi Susmita , Chang Astra , McLaughlin Bridget , Zhou Ping , Kenny Elaina , Kraynov Liliya , Arnott Sarah , McGee Jeannine , Nolta Jan A. , Lam Kit
Published: October 2019

Abstract 3269

The one-bead-one-compound (OBOC) combinatorial library method was invented by Kit Lam to identify cancer cell surface targeting ligands (Peng L et al. Nat. Chem. Biol. 2006, Aina OH et al., Mol. Pharm. 2007). Using this method, LLP2A, a ligand against activated a4b1 integrin, was discovered that targets malignant lymphoma with high affinity and specificity (Peng L et al., Mol Cancer Ther. 2008). We tested whether childhood acute lymphoblastic leukemia (ALL) cells expressed activated a4b1 integrin and bound to LLP2A using primary leukemia cells and leukemia cells engrafted in NOD/SCID/IL2Rg null (NSG) mice. Expression of activated a4b1 integrin was observed in three primary leukemia samples (1 T-ALL, 1 precursor B (preB) ALL, and 1 relapsed preB ALL) by the rainbow beads (color-coded polystyrene bead mixture displaying unique ligands against known receptors such as a4b1, a3b1, avb3, a5b1 integrins (Luo J et al., J. Comb. Chem. 2008). To quantify activated a4b1 integrin expression, primary leukemia cells were analyzed by flow cytometry using biotinylated LLP2A/streptavidin-PE. Nine samples (7 preB ALL and 2 relapsed preB ALL) were analyzed. All samples, except for one relapsed preB ALL sample, showed expression of activated a4b1 integrin at different levels. In order to determine the specificity of activated a4b1 integrin expression, hematopoietic stem cells from three normal bone marrow (BM) samples were analyzed and showed very low levels of activated a4b1 integrin expression. Furthermore, we have identified 32 analogues of LLP2A using three fresh preB ALL samples to screen two LLP2A-focused libraries. Peptide microarrays using a polystyrene slide coated with neutravidin and biotinylated ligands will be prepared with these analogues. We will determine the binding affinity and specificity of these analogues to a series of primary ALL cells. Binding specificity to leukemia cells will be determined by excluding ligands which bind to normal hematopoietic progenitor (CD34) cells. The property of the selected ligands will be tested in vivo using NSG mice engrafted with primary ALL cells. Updated results with these new ligands will be presented at the meeting.

Disclosures:

No relevant conflicts of interest to declare.
Development of Impaired Cytochrome C Oxidase Activity In Apheresis Platelets During Storage
Authors: Diab Yaser A. , Thomas Adia , Luban Naomi L.C. , Wong Edward , Wagner Stephen J. , Moroff Gary , Levy Richard J.
Published: October 2019

Abstract 3347

Background:

With current storage requirements, the shelf life of platelet (PLT) products is largely limited by the development of deleterious in vitro changes associated with overall reduction in therapeutic efficacy collectively known as the “Platelet Storage Lesion” (PSL). PSL is characterized by a number of biochemical changes including lactate accumulation, bicarbonate depletion and a fall in pH. We hypothesize that these changes reflect a state of impaired oxidative phosphorylation associated with increased reliance on anaerobic glycolysis that evolves during PLT storage. In this study we evaluated the function and expression of “Cytochrome C Oxidase” (COX), a key mitochondrial enzyme in oxidative phosphorylation, with relation to several in vitro markers of PSL. The studies were performed in apheresis PLT stored for up to 7 days under standard blood bank conditions.

Methods:

Apheresis PLT concentrates were collected in 100% plasma using a Trima Cell Separator (CaridianBCT, Lakewood CO) to provide products with at least 3 × 1011 PLT. All products were collected in Trima® storage bags and were stored in a flatbed reciprocal agitator at 22 ± 2°C for 7 days. Multiple standard in vitro assays were performed on Days 0 (baseline measurement), 2, 4 and 7 of storage including PLT count, mean PLT volume, pH, pO2, pCO2, bicarbonate, lactate & glucose levels, aggregation and ATP release studies (ADP/Collagen), soluble CD40 ligand levels (supernatant) and total intracellular PLT ATP content. In addition, steady state COX kinetics and protein immunoblotting for COX subunits I and IV, were performed using isolated PLT mitochondria from simultaneously collected samples. Data are reported as mean ± 2 SD (n =10 experiments). One-way Analysis of Variance (ANOVA) and post-hoc Tukey's range test were used to compare data obtained at different time points. Differences were considered statistically significant only if the p value was <0.001 taking into the account that performing such large number of tests in the study increased the probability that a significant p value was incorrectly obtained by random chance.

Results:

PLT COX function declined significantly throughout storage (Table). Steady-state levels of COX I and IV remained essentially unchanged. This decrease in COX function paralleled progressive ATP depletion and time-dependent changes that were consistent with the development of the PSL.

Conclusion:

During storage of apheresis PLT for 7 days, COX function decreased progressively in association with ATP depletion indicating acquired impairment in oxidative phosphorylation. These findings suggest that bioenergetic failure is associated with PSL. Further studies are required to determine if mitochondrial dysfunction is a cause of PSL, and if it can be prevented or is amenable to therapeutic intervention.

Disclosures:

No relevant conflicts of interest to declare.
Interferon Gamma (IFNγ) Interferes with the Effects of Tyrosine Kinase Inhibitors (TKI) In CML Cells.
Authors: Held Stefanie AE , Bringmann Anita , Heine Annkristin , Kock Grethe , Körber Ruth-Miriam , Seltmann Nina Manon , Brossart Peter
Published: October 2019

Abstract 3393

Introduction:

Chronic myeloid leukemia (CML) is a myeloproliferative disease which resolves in the constitutive activity of the BCR-ABL protein. Since the introduction of imatinib as first line therapy for CML, the overall survival of patients could be significantly extended. However, mutations of the BCR-ABL protein and environmental effects mediated by adhesion molecules, cytokines and growth factors were shown to induce resistance to TKIs. For the development of immunotherapeutic approaches that combine TKIs with vaccination strategies or donor lymphocyte infusions it is important to understand the mechanisms of possible interactions and inhibition. Therefore, we thought to determine the role of activated T cells and to analyze a possible antiapoptotic effect of released IFNγ on the CML cells.

Methods:

CML cell lines and primary cells from patients with CML were treated with imatinib or nilotinib and incubated with activated (PMA/ionomycin) or resting T cells in transwell experiments. Induction of apoptosis was analyzed by flow cytometry. In addition, IFNγ was used together with TKIs and apoptotic cell death and proliferation was determined. Immunoblotting was performed to analyze the involved pathways.

Result:

Incubation of CML cells in transwell assays with activated but not resting T lymphocytes resulted in significant inhibition of the apoptosis induction by imatinib and nilotinib. To further determine the mechanisms mediating these effects, CML cells were treated with IFNγ in addition to the TKIs. In line with the results from previous experiments, IFNγ reduced the rate of cell death and proliferation inhibition mediated by imatinib and nilotinib. Interestingly, in contrast to IFNα, IFNγ inhibited the TKIs induced downregulation of MHC-class I molecules on CML cells. In line with these results, Western blot analysis of K-562 cells incubated with imatinib or nilotinib showed an increased caspase-3 activation and PARP cleavage as well as reduced down regulation of anti-apoptotic molecules such as xIAP as compared to IFNγ treated cells. Furthermore, we observed a diminished TKI induced downregulation of Jak-2 and STAT-5 phosphorylation as well as increased nuclear expression of RUNX-1 in cells treated with IFNγ.

Conclusion:

Our results demonstrate that IFNγ released by activated T-cells can interfere with the action of TKIs in CML patients and might have important implications in the development of immunotherapeutic approaches.

Disclosures:

No relevant conflicts of interest to declare.
Prognostic Significance of the Lost of a Major Molecular Response In Philadelphia Chromosome-Positive Chronic Myelogenous Leukemia (Ph+CML).
Authors: García-Gutiérrez J. Valentin , Herrera Pilar , Jimenez-Rolando Marta , Tenorio María , Calbacho María , Blanchard M. Jesús , Rey Maria Dolores , Ramos Paloma , Ramos ML , Lopez Javier
Published: October 2019

Abstract 3435

Background:

Albeit of well-known, dramatic improvements, there remain some questions to be solved around Ph+CML in treatment with tyrosine kinase inhibitors (TKI). Among these, the significance of the amount of minimal residual disease (MRD) measured by RT-PCR. For instance, loss of a so-called major molecular response (MMR) is claimed to be a Òsuboptimal responseÓ and following the ELN recommendations, a change in treatment should be considered in these patients.

Aims:

To evaluate the relevance of a loss of MMR in patients with complete cytogenetic response (CCR).

Study Group and Methods:

We have analized 81 patients treated with imatinib for CML in chronic phase with a median follow up of 66 months. 36 patients started imatinib after interferon failure and 45 as front line therapy. Major Molecular Response (MMR; BCR-ABL/ABL ratio<0.1% IS) at any time was achieved by 63 patients.

Results:

22 patients (34%) lost MMR (documented al least twice). The risk of losing MMR was higher in late MMR (>18 months) compared with those cases whose MMR came much earlier (<18 months): 70% vs 18% (p=. 000). We have found no correlation among the lost of MMR and classical prognostic factors (Sokal-Index, mutations at the TK domain or imatinib plasma levels). Of these 22 patients, 7 (32 %) recovered MMR later with no therapy changes, 8 (36%) experienced fluctuations in the BCR-ABL transcript-levels without losing CCR, 4 (19%) did not attain a MMR but remained in stable CRR, and 3 (13%) lost CCR. These regained MMR after being treated on second generation TKI. The results show how the stability of the early MMR is greater than late MMR (table1).

Conclusions:

In our experience, one third of the patients who lost MMR recovered it later on the same treatment. And only 13% went on to treatment failure. Perhaps some similar cases (after first losing MMR) should be closely monitored before a change in treatment. Also of note is, of course regarding only our experience, that the risk of a loss of MMR seems to be maximal in patients who achieve a late MMR.

Disclosures:

No relevant conflicts of interest to declare.
Stem Cell Transplant without Growth Factor In Multiple Myeloma: Engraftment Kinetics, Bacteremia, and Hospitalization.
Authors: Gertz Morie Abraham , Gastineau Dennis , Lacy Martha , Dispenzieri Angela , Hayman Suzanne R. , Kumar Shaji , Dingli David , Leung Nelson , Wolf Robert , Hogan William J , Buadi Francis
Published: October 2019

Abstract 3469

Introduction:

Stem cell transplantation is important in the management of multiple myeloma. In the United States, the standard of care is administration of growth factors to accelerate neutrophil recovery after stem cell transplant. The need for growth factors after transplant has not been investigated recently.

Patients:

We analyzed a cohort of 166 patients at our institution who underwent autologous transplant for multiple myeloma without receiving growth factors after transplant and compared them with 498 patients who received standard filgrastim beginning on posttransplant day 5. TABLE

Results:

A neutrophil count of 500/μL was achieved in a median of 12.5 days in patients receiving growth factor, compared with 13.5 days in those not receiving growth factor (P<.001) Fig 1. Platelet engraftment was identical (median, 14.5 days; P=.12) in both groups, despite a lower median number of CD34+ cells infused in patients who did not receive growth factors. Incidence of nonstaphylococcal bacteremia was identical in both groups. The median hospital stay was 3.5 days shorter in the group not receiving growth factor. Bacteremia impacted platelet but not neutrophil engraftment. Figure 2

Conclusion:

It is feasible and reasonable to perform autologous stem cell transplant for multiple myeloma without administering growth factors. Estimated savings would be $4,600 for each transplanted patient. Growth factor administration is not required for a safe outcome and decreases the number and severity of some of the fluid-related complications after transplant such as acute respiratory distress syndrome, allergic reactions, alveolar hemorrhage, and rarely splenic rupture.

Disclosures:

Gertz: Celgene: Honoraria; Millenium: Honoraria, Membership on an entity's Board of Directors or advisory committees. Lacy:Celgene: Research Funding. Dispenzieri:Celgene: Honoraria, Research Funding; Binding Site: Honoraria. Kumar:Celgene: Consultancy, Research Funding; Millennium: Research Funding; Merck: Consultancy, Research Funding; Novartis: Research Funding; Genzyme: Consultancy, Research Funding; Cephalon: Research Funding.
Clinical Significance of Polymorphisms During Hepatopoietic Stem Cell Transplantation.
Authors: Ahn Jae-Sook , Kim Hee Nam , Kim Nan Young , Li Yu– , Lee Il-Kwon , Kim Yeo-Kyeoung , Bae Soo-Young , Yang Deok-Hwan , Lee Je-Jung , Kim Hyeoung Joon
Published: October 2019

Abstract 3497

Conidtioning plays an important part in hematopoietic stem cell transplantation (HSCT) and is a major cause of early mortality. Most of conditioning regimens include the ionizing radiation and alkylating agent. The tissue damage made by chemotherapy or radiation is the major cause of acute toxicity and the development of graft versus host disease (GvHD). DNA repair and drug metabolism gene polymorphisms influence the transplantation associated mortality and GvHD by causing incomplete DNA repair and interference with metabolism of drugs. To explore whether these polymorphisms influence clinical implication and the adverse outcomes related to chemo-radiotherapy, we analyzed total 12 polymorphisms in DNA repair (ERCC1, ERCC2, XRCC1, XRCC3, XRCC4, XPC, MGMT and OGG1) and drug metabolizing pathway (GST T1, GST P1, CYP1A1 rs1048943 and CYP1A1 rs4646903).

The total 175 patients were assessed from a HLA identical sibling (n=122, 69.7%) and unrelated donors (n=53, 30.3%). Patients have either acute leukemia (n=123, 70.3%), MDS (n=18, 10.3%), CML (n=15, 8.6%) or others (n=19, 10.8%). We evaluated the association between SNPs with adverse effects such as venoocclusive disease (VOD), mucositis, treatment related mortality and acute/chronic GvHD. Multivariate analysis was performed adjusting for clinical factors; recipient age, donor type, diagnosis, disease status, gender mismatch, HLA mismatch, stem cell source, conditioning regimen and GvHD prophylaxis. Three SNPs were associated with clinical significance. XRCC4 (rs1056503) was associated with increased risk of acute gut GvHD (RR:6.84, 95% CI:1.69-28.2, p=0.007), CYP1A1 (rs1048943) with mucositis (RR:3.12, 95% CI:1.22-7.96, p=0.017) and XPC (rs2228001) with VOD (RR:38.23, 95% CI:2.08-702.18, p=0.014).

Our findings show that the acute adverse events after allogeneic HSCT were correlated DNA repair and drug metabolism gene polymorphisms and these findings suggest that XRCC4, CYP1A1 and XPC might provide useful information for the risk stratification of HSCT

Disclosures:

No relevant conflicts of interest to declare.
Reduced Intensity Versus Myeloablative Conditioning in Adults with Acute Myeloid Leukemia Transplanted with Umbilical Cord Blood
Authors: Oran Betul , Wagner John E. , Defor Todd , Weisdorf Daniel J. , Brunstein Claudio
Published: October 2019

Abstract 3531

Background:

Reduced intensity conditioning (RIC) umbilical cord blood (UCB) transplantation is increasingly used in hematopoietic stem cell transplantation (HCT) for older and medically unfit patients. Data on the efficacy of HCT after RIC relative to myeloablative conditioning (MAC) are limited. We compared the outcomes of acute myeloid leukemia (AML) patients > 18 yrs who received UCB grafts after either RIC or MAC.

Methods:

One hundred nineteen adult patients with AML in complete remission (CR) underwent an UCB transplant after RIC (n=74, 62%) or MAC (n=45, 38%) between January 2001 and December 2009. Conditioning was either reduced intensity and consisted of cyclophosphamide 50 mg/kg, fludarabine 200 mg/m2 and total body irradiation (TBI) 200 cGy or myelablative and consisted for cyclophosphamide 120 mg/kg, fludarabine 75mg/m2 and TBI 1200–1320cGy. All patients received cyclosporine (day -3 to day +180) and mycophenolate mofetil (day -3 to day +45) post-HCT immunosuppression and hematopoietic growth factor. Use of RIC was reserved for patients >45 years (n=37, 82%) or pre-existing severe co-morbidities (n=8, 18%). The two groups were similar except RIC had more preceding myelodysplastic syndrome (RIC=28% vs. MAC=4%, p<0.01) and older age (median, RIC=55yrs vs. MAC=33yrs; p<0.01). Remission status (CR1; RIC=65% vs. MAC=56%, p=0.9), CR1 duration ≥ 1 yr for CR2 HCT (RIC=46% vs. MAC=45% p=0.9) and the HCT-comorbidity index were similar (score >2; RIC=50% vs. MAC=40%, p=0.1). Furthermore, most patients received double UCB graft (RIC=85% vs. MAC=89%, p=0.5) with a maximum HLA disparity of 4/6 and 5/6 in 52% and 34% of RIC recipients vs. 62% and 29% of MAC recipients (p=0.5).

Results:

Median follow-up for survivors was 4.2 (RIC) and 3.5 years (MAC). The 3-year leukemia-free survival (LFS) was 31% vs. 55% (p=0.02) and 3-year relapse incidence was 43% vs. 9% (p<0.01) in recipients of RIC and MAC, respectively (Figure). One-year transplant related mortality (TRM) was similar (RIC=16% vs. MAC=24%; p=0.55). In multivariate analysis (Table), RIC recipients and those in CR2 with short CR1 duration had higher risk of relapse and poorer LFS. No factors predicted TRM.

Conclusion:

UCB with RIC extends the use of allogeneic HCT for older and less fit patients with limited TRM. However, the increased relapse risk and poorer LFS in the RIC setting indicates the need for novel strategies to improve leukemia control post-transplantation, especially in CR2 patients with short CR1 duration.

Disclosures:

Wagner: CORD:USE: Membership on an entity's Board of Directors or advisory committees; VidaCord: Membership on an entity's Board of Directors or advisory committees. Weisdorf:Genzyme: Research Funding; Hospira.
Outcomes In Patients with Multiple Myeloma Following Autologous Stem Cell Transplantation Using Intravenous Busulfan and Melphalan: A Matched Comparison to a Double Autologous Transplant Strategy
Authors: Blanes Margarita , De La Rubia Javier , Lahuerta Juan-Jose , Gonzalez José David , Ribas Paz , Solano Carlos , Sanz Miguel Angel
Published: October 2019

Abstract 3560

Background:

We have previously reported a high response rate with a conditioning regimen consisting of intravenous busulfan (ivBu) and melphalan (Mel) in patients with multiple myeloma (MM) undergoing autologous stem cell transplant (ASCT) (Blanes M et al, Leuk Lymphoma 2009). In this case-matched study, we analyze the efficacy and toxicity of ivBuMel as conditioning regimen before ASCT in 51 patients with newly diagnosed MM. Their clinical outcome was compared with that of a control group of 102 pair mates included in the Spanish PETHEMA/GEM2000 study of tandem transplant (control group).

Patients and Methods:

Patients in the control group were transplanted between 2002 and 2005, while patients in the ivBuMel group underwent ASCT between 2005 and 2009. Controls were matched with respect to sex, age, Durie-Salmon and ISS stage at diagnosis, and disease status at transplant. The conditioning regimen comprised iv Bu (3.2 mg/kg in a single daily dose, days -5 to -3) and Mel (140 mg/m2, day -2). Patients in the control group underwent a first ASCT after Mel 200 mg/m2 (Mel 200) in a single dose on day -2 followed in patients failing to achieve a complete response (CR) by a second autologous transplant. The conditioning regimen administered in the second transplant was either Mel 200 or a combination of cyclophosphamide, carmustine, and etoposide.

Results:

There were no differences in the hematopoietic recovery and mild or moderate mucositis was the toxicity most frequently observed in both groups of patients. Two (4%) patients died due to transplant-related complications in the ivBuMel group and 5 (5%) in the control group. The CR/near CR (nCR) rate was 51 % in both groups. After median follow-up of 32 and 40.5 months in the ivBuMel and control group, respectively, median progression free survival (PFS) was 37.6 for patients who received ivBuMel and 26.5 months for those in the control group. At 4 years, overall survival and PFS was 65±10% and 42±9% in the ivBuMel and 62±5% and 32± 5% in the control group, respectively (P = NS). For patients achieving CR/nCR after ASCT, the 4-years PFS rate was 52±14% and 40±8% in ivBuMel and control group, respectively (P = 0.3). Finally, in patients achieving partial response or less after transplant, the 4-years PFS rate was 33±12% and 23±6% in ivBuMel and tandem transplant group, respectively (P = 0.3).

Conclusion:

Results of this case-matched study suggest that the use of ivBuMel conditioning regimen before ASCT in patients with newly diagnosed MM is associated with low transplant-related morbidity and mortality and a high anti myeloma efficacy that compares favourably with a tandem transplant strategy.

Disclosures:

No relevant conflicts of interest to declare.