bluuebunny/crossref_metadata_2025_split huggingface dataset:
Single Agent Metronomic Cyclophosphamide in Refractory,Non-Hodgkin's Lymphoma.
Authors: Hamm Caroline , Kanjeekal Sindu M. , Mathews John , Alam Yasmin , Yoshida Sam , Elfiki Tarek 
Published: October 2019

Abstract
Abstract 4776
n this retrospective review, we report a case series of eight patients with refractory, heavily pre-treated non-Hodgkin's lymphoma(NHL), treated with low dose cyclophosphamide. Median number of prior treatment regimens was 3. The median disease free survival was 15.5 months with three patients in ongoing CR at the time of reporting with 15 - 36 months of followup. In a comparative group of 6 patients with other malignancies beside NHL (3 chronic lymphocytic leukemia, 2 breast cancer, 1 thymoma, and one small cell carcinoma) the median progression free survival was significantly worse at 3 months. Interestingly the thymoma had a progression free survival of 36 months.
Toxicity was minimal. One patient stopped because of thrombocytopenia. One of the patients with ongoing CR at 36 months stopped after 11 months of treatment. One patient died in CR of anal cancer after 15 months of treatment.
These results demonstrate an easily tolerable and surprisingly effective treatment option in patients with non-Hodgkin's lymphoma who are not candidates for bone marrow transplantation.
The mechanism of action of metronomic chemotherapy has been proposed as anti-angiogenic. We suggest an alternate mechanism of action: that of immunomodulation. We are suggesting this alternate explanation in view of the significant difference in the response rate between those disease that are known to immunogenic and those that may be less so, as well as the well-known immunogenic affect of cyclophosphamide. Further studies are ongoing to investigate this hypothesis.

Disclosures:
No relevant conflicts of interest to declare.
Combination of Enzastaurin and HDAC Inhibitors Synergically Induce Apoptosis in Lymphoma Cells.
Authors: Bodo Juraj , Sedlak Jan , Maciejewski Jaroslaw P. , Hsi Eric D. 
Published: October 2019

Abstract
Abstract 4783

Introduction
Histone deacetylase inhibitors (HDACis) are approved for use in the setting of cutaneous T-cell lymphoma with modest benefit. Enzastaurin is an investigational PKCβ inhibitor that has growth inhibitory and pro-apoptotic effects in both B and T-cell lymphoma. Specifically, enzastaurin-induced inhibition of PKC leads to rapid accumulation of β-catenin that triggers c-Jun dependent induction of p73, followed by apoptosis. We investigated the cytotoxicity and mechanisms of cell death of combination enzastaurin and low concentrations of HDACis in B-cell lymphoma and T-cell lymphoma cell lines and primary lymphoma/leukemia cells.

Experimental design
Apoptosis was measured by flow cytometry and PARP cleavage. Phospho-GSK3β (S9), pS6, phospho-c-jun (S73) and β-catenin were analyzed by Western blot or quantum-dot immunoflourescence as measures of PKCβ inhibition. Cytotoxicity was determined by WST-1 proliferation assay and colony forming cell (CFC) assays.

Results
As expected, enzastaurin induced dephosphorylation of GSK3β and S6RP associated with increased β-catenin expression followed by phosphorylation of c-jun (S73) and PARP cleavage in SU-DHL-6 (diffuse large B-cell lymphoma line) cells. Treatment with low concentrations of suberoylanilide hydroxamic acid (SAHA) showed slight or no changes in studied proteins. Combined enzastaurin/SAHA treatment resulted in strong synergistic apoptosis in two treated germinal center B-cell-like and two activated B-cell-like lymphoma cell lines, two T-cell lymphoma cell lines and four different primary lymphoma/leukemia samples. Similarly, combined enzastaurin/ valproic acid treatment induced synergistic apoptosis in SU-DHL-6 cell line, suggesting the synergy is generalizable to other HDACis. In comparison to the single agent treatment, combined enzastaurin/ SAHA treatment resulted in activation of proapoptotic MAPK, c-jun N-terminal kinase, further increase of phospho c-jun (S73) levels, increased FasL levels, and amplification of PARP cleavage. Quantitative immunofluorescence assay showed a more rapid increase of β-catenin levels with the combination than either agent alone. Furthermore, compared to the low dose SAHA treatment alone, hyperacetylation of histone H3 was detected in samples when enzastaurin was added in combination with low dose SAHA, likely the consequence of displacement of HDAC by β-catenin. In addition, no change in CFC output in normal bone marrow exposed to this combination was observed.

Conclusion
Enzastaurin/ HDACi therapy can synergistically inhibit growth and induce apoptosis in lymphoid malignancy through increased biochemical effects attributed to each agent. These data support further investigation of addition of PKCβ inhibitors to HDACi in order to increase their anti-lymphoma effects.

Disclosures:
No relevant conflicts of interest to declare.
Hepatitis B Virus Reactivation in Malignant Lymphoma with Occult HBV Carrier.
Authors: Fukushima Noriyasu , Mizuta Toshihiko , Urata Chisako , Tanaka Mariko , Yokoo Masako , Ide Masaru , Hisatomi Takashi , Sueoka Eisaburo , Kimura Shinya 
Published: October 2019

Abstract
Abstract 4794

Background
In surface antigen of hepatitis B virus (HBsAg)-positive carrier for anticancer treatment of malignant lymphoma, it is well recognized that reactivation of hepatitis B virus (HBV) occasionally occurs. However, there have been only a few studies of HBV reactivation in serum HBsAg-negative and hepatitis B core antigen (HBcAb)-positive occult HBV carriers. We looked at both retrospective and prospective studies to determine the prevalence, clinical course and risk factor of HBV reactivation during chemotherapy in lymphoma patients.

Patients and methods
Forty-nine of 128 (38.3%) lymphoma patients were HBsAg negative and HBcAb positive, and 25 of these patients were then given liver function tests and HBsAg tests monthly and serum HBV DNA every 3 months.

Results
HBV reactivation was observed in two patients (4.0%) who had received intensive chemotherapy including steroid and rituximab. Immediate administration of entecavir therapy after elevation of HBV DNA level was conducted, and this resulted in reduction of it and improvement of liver function test.

Conclusions
Rituximab plus steroid-containing regimens may increase the risk of HBV reactivation in HBsAgnegative and HBcAb-positive lymphoma patients. More ambitious prospective studies are required to establish clinically useful or cost-effective follow-up methods for control of HBV reactivation in lymphoma patients with occult HBV infection.

Disclosures:
No relevant conflicts of interest to declare.
Population Pharmacokinetic of Native Escherichia Coli Asparaginase.
Authors: Borghorst Stephan , Pieters Rob , Kuehnel Hans Juergen , Boos Joachim , Hempel Georg 
Published: October 2019

Abstract
Abstract 4803

Introduction
Native Escherichia Coli Asparaginase (ASNase) is an integral component in the therapy of acute lymphoblastic leukemia (ALL) and non-Hodgkin's Lymphoma (NHL). There is a great interindividual variability in treatment intensity in patients treated with the same dose of ASNase. Population pharmacokinetics (PopPK) provides the possibility to divide the overall variability of a population in an inter- and intraindividual element and to develop more precise dosing recommendations. Furthermore, pharmacokinetic parameters can be estimated as well as possible covariates that may influence the pharmacokinetics of the drug can be identified.

Patients and Methods
The model building dataset consisted of 16 patients (233 samples) receiving 5000 U/m2 ASNase (Asparaginase Medac®) 8 times according to the DCOG-ALL 10 treatment protocol. Asparaginase activity was measured in a randomized clinical Phase 2 study comparing the pharmacokinetic and pharmacodynamic of a newly developed recombinant ASNase with that of the established ASNase (Asparaginase Medac®)[R. Pieters et al. Blood. 2008 Dec 15. 112(13):4832-8]. The PopPK-model was developed using NONMEM (version VI) with First Order Conditional Estimation (FOCE) method and INTERACTION option.

Results
A linear 2-compartmental model with a combined proportional (0.9%) and additive (48.1U/l) error model described the data adequately. The pharmacokinetic parameters estimated were: Total systemic clearance 0.135 ± 12.8% l/h/70kg, volume of distribution of the central compartment 4.27 ± 13.1% l/70kg, volume of distribution in the peripheral compartment 0.83 ± 80.4% l/70kg and intercompartmental clearance 0.058 l/h/70kg (mean ± interindividual variability). Body weight was identified as the most important covariate. Validity of the model was verified by simulating different dosages of ASNase (2500U/m2 and 10000U/m2) in induction and reinduction of the ALL-BFM treatment protocol. The median and mean ASNase activity was compared with published data [E. Ahlke et al. Br J Haematol. 1997 Mar. 96(4):675-81 and Boos et al. Eur J Cancer. 1996 Aug. 32A(9):1544-50]. Furthermore pharmacokinetic data obtained by a noncompartmental analysis [R. Pieters et al. Blood. 2008 Dec 15.112(13):4832-8] were compared with the pharmacokinetic data estimated by the PopPK model. Both procedures indicated on face validity of the PopPK model.

Conclusion
This PopPK analysis provides the first step in the development of a PopPK model for ASNase. Face validity of the PopPK model could be demonstrated and will be confirmed with an independent dataset.

Disclosures:
Pieters: Medac GmbH: Research Funding. Kuehnel:Medac GmbH: Employment. Boos:Medac GmbH: Honoraria. Hempel:Medac GmbH: Honoraria.
Myeloma and the Microenvironment: Macrophages Are a Protector of Myeloma Cells Apoptosis Induced by Chemotherapy Drugs.
Authors: Yang Jing , Zheng Yuhuan , Cai Zhen , Qian Jianfei , Hong Sungyoul , Li Haiyan , Matthews Jairo A. , Wang Michael , Yi Qing 
Published: October 2019

Abstract
Abstract 4804
Multiple myeloma is a B-cell malignancy characterized by the proliferation of plasma cells in the bone marrow. It is the second most common hematological malignancy and is still largely incurable. One of the major problems is that myeloma cells develop drug resistance upon interaction with bone marrow stromal cells. To better understand the importance of different stromal cell components in the bone marrow microenvironment, we examined the effects of macrophages on myeloma cell survival and myeloma cell response to chemotherapy. We report here that macrophages, in particular tumor-associated macrophages obtained by culturing macrophages with myeloma cell culture supernatants, are a protector of myeloma cells. Macrophages protected both myeloma cell lines and primary myeloma cells isolated from patients from spontaneous and chemotherapy drug-induced apoptosis via attenuating the activation of caspase-dependent apoptotic signaling. The protective effect was dependent on direct contact between macrophages and myeloma cells. Although tumor-associated macrophages secreted large amounts of IL-6, which is the most important survival factor for myeloma cells, our results showed that IL-6 neutralizing antibodies fail to significantly affect the protective effects of tumor-associated macrophages. The reduced numbers of apoptotic tumor cells in the cocultures were not the result of macrophage-uptake of apoptotic cells, because macrophages with or without the ability to phagocytose apoptotic cells provide similar protection to myeloma cells against chemotherapy-induced apoptosis. These findings are clinically relevant, because we examined bone marrow biopsies of patients by immunochemical analysis and found that CD68+ macrophages are heavily infiltrated in the bone marrow (tumor bed) of patients with myeloma but not control patients. Thus, our results indicate that macrophages are an important component of the bone marrow stromal cells and may contribute to myeloma cell survival and resistance to chemotherapeutic treatment in vivo.

Disclosures:
No relevant conflicts of interest to declare.
LACK of IRON Binding by Pixantrone IS ASSOCIATED with Reduced PRODUCTION of Reactive Oxygen SPECIES and Myoctye Cytotoxicity IN VITRO.
Authors: Hacker Miles , McKennon Marc , Singer Jack W. 
Published: October 2019

Abstract
Abstract 4806

Introduction
Pixantrone (PIX), an aza-anthracenedione, which has successfully completed a phase 3 trial (J Clin Oncol 2009; 27:15s, No. 8523) was designed to enhance clinical efficacy while significantly decreasing cardiotoxicity compared to doxorubicin (DOX) and mitoxantrone (MIT). Multidose administration, in animal models of equitoxic doses of PIX, MIT, and DOX, with or without prior therapy with DOX, resulted in minimal evidence for PIX cardiotoxicity compared with the severe histologic lesions seen with these other agents (Cavaletti et al: Investigational New Drugs 2007; 3:187-95). Both DOX and MIT contain a dihydroquinone structural element known to interact with iron. Additionally, DOX contains an aliphatic ketone which, once metabolized to the corresponding secondary alcohol metabolite doxorubicinol, is implicated in release of free iron and the chronic cardiotoxicity observed with DOX. In contrast, PIX has a nitrogen containing heterocycle which replaces the dihydroquinone, forming an aza-anthracenedione structure. PIX also does not contain an aliphatic ketone and cannot form metabolites analogous to doxorubicinol.

Methods
To validate the proposed mechanisms underlying the observed differences in cardiotoxicity, we used established spectrophotometric techniques to quantify iron:drug interactions that are thought to be mechanistic for chronic doxorubicin cardiotoxicity (Menna et al: Cardiovasc Toxicol 2007; 7:80–85).

Results
Adding increasing amounts of iron to drug solution, we observed that DOX and MIT underwent changes in visible range absorbance patterns, characteristic of drug:iron complex formation, confirming the expected 1:3 Fe(II)-drug ratio for both DOX and MIT. In contrast, no spectrophotometric changes were observed with iron added to PIX, clearly demonstrating that PIX does not bind iron. In vitro studies using H2C9 rat myocardial cells indicate that PIX (ID50 >50 μg/ml) is far less toxic than DOX (ID50= 1 μ/ml). Moreover, PIX does not induce significant reactive oxygen species (ROS) production in the H2C9 cells compared to DOX.

Conclusion
These results demonstrate that PIX does not bind iron and that its inability to bind iron and its reduced propensity to generate ROS may be the mechanism for reduced PIX cardiotoxicity in animal models compared to DOX or MIT.

Disclosures:
McKennon: Cell Therapeutics, Inc: Employment. Singer:Cell Therapeutics, Inc: Employment.
Selective Protein Kinase C β Inhibition Induces Apoptosis and Arrests Cell Cycle in AIDS-Related Non-Hodgkin Lymphoma Cell Lines.
Authors: Saba Nakhle S , Safah Hana F , Levy Laura S 
Published: October 2019

Abstract
Abstract 4807
AIDS-related Non-Hodgkin Lymphoma (AIDS-NHL) constitutes an aggressive variety of lymphomas characterized by increased extranodal involvement, relapse rate and resistance to chemotherapy. In the present era of target-specific therapy, PKCβ targeting showed promising results in preclinical studies, as well as phase II clinical trials involving a wide variety of cancers. Studies describing the role of PKCβ in AIDS-NHL are primitive if not lacking, and HIV infection has been considered an exclusion criterion in clinical studies involving PKCβ inhibition in NHL patients. In the present study, we measured the sensitivity of a variety of AIDS-NHL cell lines to PKCβ inhibition. Three cell lines were studied: 2F7 (AIDS-Burkitt's Lymphoma), BCBL-1 (AIDS-Primary Effusion Lymphoma) and UMCL01-101 (AIDS-Diffuse Large B Cell Lymphoma). Cells were tested for PKCβ1 and PKCβ2 expression by immunoblot. Cell viability was measured in the presence of a PKCβ-specific inhibitor at concentrations of 5, 10, 20 and 30 μM for 48 hours in the presence of 10% fetal bovine serum (FBS). The IC50 at 48 hours was determined for each cell line, and cells were cultured in the presence of the correspondent IC50 for 24 and 72 hours. MTS assay was performed to quantify cell viability, and TUNEL assay with propidium iodide staining was used to detect apoptotic induction and effect on cell cycle. Results showed that 2F7 and UMCL01-101 cell lines express PKCβ1 and PKCβ2, while BCBL-1 expresses only PKCβ1 and lacks PKCβ2 expression. 2F7 and BCBL-1 were sensitive to PKCβ inhibitor with IC50 of 14 and 15 μM of inhibitor, respectively. UMCL01-101 showed relative resistance to PKCβ inhibition with an IC50 of 28 μM. Incubation in the presence of the correspondent IC50 induced significant apoptosis in all cell lines starting after 2 hours of treatment; UMCL01-101 cell line was not significantly affected when incubated in the presence of 14 μM. Our results also showed cell cycle inhibition in 2F7 and BCBL-1 starting respectively after 2 and 8 hours of incubation with the correspondent IC50; UMCL01-101 showed no features of cell cycle inhibition when cultured in the presence of high concentration (IC50) or low concentration (14 μM) of the inhibitor. BCBL-1 cell line findings were unexpected in the absence of PKCβ2 expression and implicate PKCβ1 as a regulator in these cells. These results indicate that PKCβ plays an important role in AIDS-related NHL survival, and suggest that PKCβ targeting should be considered in a broader spectrum of NHL. Ongoing studies will detail the mechanism of PKCβ inhibition and uncover the underlying mechanism of resistance in PKCβ expressing UMCL01-101 cells. This mechanism may be considered for exclusion from treatment.

Disclosures:
No relevant conflicts of interest to declare.
L-Gossypol Inhibits NF- κB, Down Regulates Mcl-1 and Induces Apoptosis of Primary Acute Myeloid Leukaemia Cells.
Authors: Pepper Chris J , Osman Hani Y , Hewamana Saman , Walsby Elisabeth J , Burnett Alan K , Knapper Steven 
Published: October 2019

Abstract
Abstract 4813
Standard treatments for acute myeloid leukaemia (AML) result in a median survival of approximately 1 year. There is now a realisation that in order to significantly improve outcomes in this disease more targeted therapies that take account of the specific biology of the tumour cell are required. L-Gossypol is a polyphenolic oil cotton seed extract that has anti-tumour activity against a range of haematological malignancies but has never been evaluated in AML cells. It is known to act as a BH3-mimetic, binding to the BH3 pocket of anti-apoptotic proteins and displacing pro-death partners to induce apoptosis. However, knowledge of the molecular events that underpin its downstream effects is limited. In this study we analysed the in vitro effects of L-Gossypol in 50 primary AML samples in order to determine its efficacy and mode of action. Apoptosis was induced in all the samples tested in a dose- and time-dependent manner as evidenced by increased Annexin V / propidium iodide labelling and the activation of caspase-9 and caspase-3. The median LD50 value (the concentration of drug required to kill 50% of the cells) was 27.5μM ± 18.3μM. There was no association between LD50 and age, sex, presenting white cell count, FLT3 mutation status or karyotype. Mechanistically, L-gossypol decreased the DNA binding activity of the NF-κB subunit, Rel A, in a concentration-dependent manner; this inhibition was evident after only 4 hours and preceded the induction of apoptosis. Furthermore, treatment with L-Gossypol inhibited the transcription of the NF-κB-regulated genes CFLAR, BCL2, BIRC5 and MCL1 in the same timeframe. Finally, studies of Mcl-1 protein expression showed down regulation in a dose-dependent manner but this was only apparent after 8 hours exposure to L-Gossypol. Taken together, our data demonstrate that L-Gossypol works, at least in part, through the inhibition of NF-κB and our data provides a rationale for clinical investigations of this agent in AML patients.

Disclosures:
No relevant conflicts of interest to declare.
A Modified Foamy Viral Envelope Enhances Gene Transfer Efficiency and Reduces Toxicity of Lentiviral FANCA Vectors in Fanca-/- HSCs.
Authors: Sun Zejin , Li Yan , Li Jingling , Cai Shanbao , Zeng Yi , Lindemann Dirk , Pollok Karen E , Hanenberg Helmut , Clapp D. Wade 
Published: October 2019

Abstract
Abstract 696
Fanconi anemia (FA) is a recessive DNA repair disorder characterized by bone marrow failure, genomic instability, and a predisposition to malignancies. Stem cell gene transfer technology is a potentially promising therapy, however, we have previously shown that prolonged ex vivo culture of cells using gamma retroviruses, results in a high incidence of apoptosis and predisposes the surviving reinfused cells to hematological malignancy in a murine model of FA. Here, we developed a lentiviral vector encoding the human FANCA cDNA and tested the ability of this construct pseudotyped with either VSV-G or a modified foamy virus (FV) envelope to correct murine Fanca-/- stem and progenitor cells. An overnight transduction protocol was utilized to minimize genotoxic stress due to extended ex vivo manipulations. Transduction and expression of hFANCA was confirmed by three classical functional and biochemical measures in vitro: improved survival of clonogenic progenitors in the presence of mitomycin C (MMC), correction of MMC-induced G2/M arrest, and by the restoration of Fancd2 mono-ubiquitination. Furthermore, in vivo competitive repopulation experiments demonstrated that the repopulating ability of Fanca-/- stem cells transduced with the lentivirus encoding hFANCA was equivalent to that of wild-type stem cells, and the genetically-corrected reconstituting Fanca-/- cells were resistant to MMC and TNF-αa. Importantly, while a significant toxicity was observed using the VSV-G envelope, the toxicity of the FV envelope to murine c-kit+ cells was limited.
In parallel experiments, human umbilical cord blood CD34+ cord blood cells were transduced with either a VSV-G- or FV envelope-pseudotyped lentivirus encoding the EGFP reporter gene. Transplantation of 4×105 cells into NOD/SCID/gamma-chainnull yielded a peripheral blood human chimerism comparable to the untransduced control cells (∼30%) regardless of the envelope. However, a much higher gene transfer efficiency of CD34+ cells was observed prior to transplantation when the FV envelope was employed (∼60%), as compared to the VSV-G envelope (∼15%). Furthermore, a similar 4 fold increase in transduction efficiency was observed in peripheral blood and in progenitors isolated from the bone marrow of both primary and secondary long term reconstituted mice. Collectively, these data indicate that the lentiviral construct pseudotyped with FV envelope can efficiently correct murine FA HSC/progenitor cells in a short transduction protocol and that the modified foamy envelope offers significantly greater transduction efficiency at comparable titers in long term reconstituting human cells in a xenograft model. This envelope may offer significant advantages compared to VSV-G in moving forward to phase 1 clinical trials.

Disclosures:
No relevant conflicts of interest to declare.