ALBERT

Description: HM1.24, an immunologic target for multiple myeloma (MM) cells, has not been effectively targeted with therapeutic monoclonal antibodies (mAbs). In this study, we investigated in vitro and in vivo anti-MM activities of XmAb5592, a humanized anti-HM1.24 mAb with Fc-domain engineered to significantly enhance FcγR binding and associated immune effector functions. XmAb5592 increased antibody-dependent cellular cytotoxicity (ADCC) several fold relative to the anti-HM1.24 IgG1 analog against both MM cell lines and primary patient myeloma cells. XmAb5592 also augmented antibody dependent cellular phagocytosis (ADCP) by macrophages. Natural killer (NK) cells became more activated by XmAb5592 than the IgG1 analog, evidenced by increased cell surface expression of granzyme B–dependent CD107a and MM cell lysis, even in the presence of bone marrow stromal cells. XmAb5592 potently inhibited tumor growth in mice bearing human MM xenografts via FcγR-dependent mechanisms, and was significantly more effective than the IgG1 analog. Lenalidomide synergistically enhanced in vitro ADCC against MM cells and in vivo tumor inhibition induced by XmAb5592. A single dose of 20 mg/kg XmAb5592 effectively depleted both blood and bone marrow plasma cells in cynomolgus monkeys. These results support clinical development of XmAb5592, both as a monotherapy and in combination with lenalidomide, to improve patient outcome of MM.

Description: Recurrence of multiple myeloma (MM) after therapy suggests the presence of tumor-initiating subpopulations. In our study, we performed flow cytometry–based Hoechst 33342 staining to evaluate the existence of a MM population with stem-like features known as side population (SP) cells. SP cells exhibit substantial heterogeneity in MM cell lines and primary MM cells; express CD138 antigen in MM cell lines; display higher mRNA expression and functional activity of ABCG2 transporter; and have a higher proliferation index compared with non-SP cells. We observed evidence for clonogenic potential of SP cells, as well as the ability of SP cells to regenerate original population. Moreover, SP cells revealed higher tumorigenicity compared with non-SP cells. Importantly, lenalidomide decreased the percentage and clonogenicity of SP cells, and also induced phosphorylation changes in Akt, GSK-3α/β, MEK1, c-Jun, p53, and p70S6K in SP cells. Adherence to bone marrow stromal cells (BMSCs) increased the percentage, viability, and proliferation potential of SP cells. Lenalidomide and thalidomide abrogated this stimulatory effect of BMSCs and significantly decreased the percentage of SP cells. Our studies demonstrate a novel mechanism of action for lenalidomide, namely targeting SP fraction, providing the framework for new therapeutic strategies targeting subpopulations of MM cells including presumptive stem cells.

Description: Background Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis and progression of cancer, and blood level of VEGF has been known to predict of outcome in several types of cancer. However, the impact of blood VEGF levels on prognosis of diffuse large B-cell lymphoma (DLBCL) has not been fully elucidated. Here we investigated the prognostic implication of circulating VEGF levels in patients with DLBCL. Patients and Methods The study involved 127 DLBCL patients treated by rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) at National Cancer Center, Korea. Both serum and plasma of the patients were obtained at diagnosis. VEGF were measured using ELISA kit (R&D systems) according to manufacturer’s guidance. We investigated the correlation between clinical parameters and blood VEGF levels. Survival rates and hazard ratios (HRs) in terms of risk for overall survival were determined using Kaplan-Meier method and Cox proportional hazard regression analysis. Results The median patient age was 56 years, and 75 (59%) patients were men. Clinical characteristics and international prognostic index (IPI), including age, performance, lactate dehydrogenase (LDH), Ann Arbor stage and extranodal involvement were evaluated. The mean (±SD) value of serum and plasma VEGFs were 713 (±599.8) and 107 (±164.4) pg/mL, and they were correlated as r=0.61 (p

Description: Background The optimal peripheral blood stem cell (PBSC) collection is a key step for successful outcome in hematopoietic stem cell transplantation (HSCT). Many indicators including preharvest white blood cell (WBC), mononuclear cell (MNC), and CD34 positive cell counts have been used for deciding the adequate time for collection of PBSCs, but each indicator has limitations. Here we investigated hematopoietic progenitor cell (HPC) count as an indicator for PBSC collection. Methods: Data from 851 autologous PBSC collections from 233 patients at the National Cancer Center, Korea, were analyzed. The correlations between harvested CD34 cell counts with preharvest WBC, MNC, CD34 cell counts, and HPC were analyzed, as were correlations by disease and mobilizing agent. Also how the outcome for engraftment can be predicted based on HPC count was studied. Results: The median age of patients was 41 years (range 0.1-72 years). The most frequent diseases were multiple myeloma (n=64) and non-Hodgkin lymphoma (n=56). The correlation coefficient between collected CD34 cells and preharvest CD34 count was (r=0.669, p

Description: Abstract 1428 Immune thrombocytopenic purpura (ITP) occurs in a few patients with malignancies such as breast, gastric, lung, cervical, ovarian cancers, and hematologic malignancies. The existence of ITP in these malignancies often has been regarded as a poor prognostic factor. However, recent researchers suggest that platelets have significant roles in cancer metastasis by promoting angiogenesis, maintaining vascular integrity and protecting tumor cells from NK cells. Therefore, low platelet counts might influence on the outcome the associated malignancies. This study was performed to get the clinical information whether malignancies accompanying ITP have poor outcome or not. A total of 18 patients with malignancies diagnosed as ITP at the National Cancer Center in Korea between 2000 and 2010 were included. The diagnosis of ITP was confirmed when their platelet counts were less than 140,000/mm3 after exclusion of other causes of thrombocytopenia. No one had an evidence of disseminated intravascular coagulation at the time of ITP diagnosis and 14 patients had been examined for bone marrow biopsy to rule out other diseases than ITP. Of the 18 patients included in this study, 7 patients were men and 11 patients were women with median age 59 years (range, 40 to 79 years). The most common cancer was the gastric cancer (n=5). Colorectal cancers (n=4), breast cancers (n=3), lung, pancreas, thyroid, ovarian, endometrial cancer and acute myeloid leukemia were examined. ITP was diagnosed concomitantly in 3 cases and after detection of malignancies in 15 cases with a median interval 8.5 months (range, 1 to 75 months). All of gastric cancer patients had Helicobacter pylori (H. pylori) infection detected by H. pylori IgG or stomach biopsy methods. The other 3 out of 13 patients (23%) showed H. pylori infection by the same methods. Of the 16 patients checked for anti-platelet antibody, only 2 (13%) patients revealed positive results. No one showed anti-HCV antibody. Six patients (30%) died of disease progression. Two patients (10%) are alive with residual disease and 10 patients (60%) are alive in disease-free statuses. Except 2 patients, no one had distant metastatic lesions at the initial diagnosis of cancer. At the time of malignancy diagnosis, there were 2 patients with distant metastasis and 5 patients at the time of last follow-up. Two gastric cancer patients who are alive in advanced stages (stage III, 43 months; stage IV 5.7 months) have survived similar or more than expected in the report (Park JM et al, Gastrointest Cancer Res 2009). A colorectal cancer patient with prostate cancer has survived 67 months. Another patient who died of stage IV colon cancer survived 19 months which was comparable to the report of the same stage colorectal cancer with median survival of 18 months in Korea (Park YJ et al, World J Surg 1999). ITP was stable in 8 patients without treatment. Nine patients received corticosteroid or immunoglobulin therapy. Anti-D immunoglobulin was administered to 2 patients who had been refractory to corticosteroid and immunoglobulin. Splenectomy was performed 3 patients who were not responsive to medical treatment and showed clinical improvement. There were only 2 patients who had shown platelet count increase after treatment of malignancies. ITP associated with malignancies were stable and responsive to therapy in this study. Although there was a limitation of interpretation due to the heterogeneity of this study group, clinical outcomes of the cancer patients with ITP did not show poor prognosis. Especially, rare patients showed distant metastasis during the disease course. Further experimental and clinical studies are needed to investigate the relationship between platelet and tumor progression including metastasis. Disclosures: No relevant conflicts of interest to declare.

Description: BACKGROUND. X-ray repair cross-complementing group I (XRCC1) is a DNA repair gene. Polymorphisms in DNA repair genes may alter protein function and therefore the efficacy of DNA damaging chemotherapy. We retrospectively evaluated the association of three polymorphisms in XRCC1 codon 194 (Arg to Trp), 280 (Arg to His), and 399 (Arg to Gln), with grade 3 or 4 toxicities of primary rituximab plus cyclophosphamide/doxorubicin/vincristine/prednisone (R-CHOP) therapy in 145 patients with diffuse large B cell lymphoma (DLBCL). METHODS. A total of 145 patients who received R-CHOP chemotherapy as a frontline regimen DLBCL were included in this retrospective study from 3 hospitals in Korea. The genotypes of XRCC1 at the Arg194Trp, Arg280His, and Arg399Gln were determined by direct sequencing methods. The clinical characteristics, treatment outcomes and grade 3 and 4 toxicities of the patients were compared using Chi-square, Fisher exact, Mann-Whitney U tests, or logistic regression according to the XRCC1 polymorphisms. RESULTS. Carrying at least one variant XRCC1 Arg194Trp alleles (194Arg/Trp or 194Trp/Trp) and XRCC1 Arg399Gln variant alleles (399Arg/Gln or 399Gln/Gln) was associated with a significantly increased risk of grade 3 or 4 gastrointestinal toxicity (Arg194Trp alleles; odds ratio, 3.489; 95% confidence interval, 0.126–1.568; P=0.001, Arg399Gln alleles; odds ratio, 2.577; 95% confidence interval, 0.144–1.096; P=0.011, respectively). The carriers of XRCC1 Arg280His variant alleles (280Arg/His or 280His/His) was associated with grade 3 or 4 neutropenia (odds ratio, 2.047; 95% confidence interval, 0.008–0.450; P=0.043). No differences in the patient characteristics, disease characteristics, response, and survival in patients with DLBCL who received frontline R-CHOP chemotherapy were observed according to three XRCC1 polymorphims. CONCLUSIONS. This study demonstrates that the patients carrying at least one variant XRCC1 Arg194Trp or XRCC1 Arg399Gln alleles have a 2.5-to 3.5-fold increased risk of grade 3 or 4 gastrointestinal toxicity and the patients carrying at least one variant XRCC1 Arg280His allele have a 2.0-fold increased risk of grade 3 or 4 neutropenia when treated with frontline R-CHOP chemotherapy and this has implications for optimizing treatment with such agents.

Description: Neutrophil-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) have been lightened as a prognostic factor in many types of solid tumor. However, there was a limited report for diffuse large B cell lymphoma (DLBCL). Thus, we investigated the impact of NLR and PLR on prognosis of patients with DLBCL. This study involved 234 DLBCL patients treated by rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisolone (R-CHOP) at National Cancer Center, Korea. The median patient age was 57 years, and 135 patients (58%) were men. Clinical characteristics as international prognostic index (IPI) including age, performance, LDH, stage and extra-nodal involvement, were evaluated. NLR and PLR at diagnosis were calculated by CBC data from automated hematologic analyzer XE2100 (Sysmex, Kobe, Japan) or differential count which was done by manually using stained peripheral blood slide when CBC data showed flags in auto-analyzer. Hazard ratios (HRs) were determined in terms of risk for overall survival using Cox proportional hazard regression analysis. The mean percentage of neutrophils and lymphocytes was 60 (range, 5-95) and 12 (2-79), and the number of platelets was 257 × 10^9/L (28-839). The mean and standard deviation of NLR and PLR was 3.62 (±4.77) and 14.7 (±20.16), respectively. IPI and stage were predictors for prognosis (p

Description: Abstract 994 Background: Angiogenesis and angiogenic factors are increased in most lymphomas and it has been associated with adverse outcome or more aggressive behavior of malignant lymphomas in previous studies. There is substantial inherited genetic variability within VEGF and one of its receptors, VEGF receptor 2 (VEGFR2), including multiple single nucleotide polymorphisms (SNPs). The level of VEGF expression has been found to vary depending on the presence of a genetic polymorphism. Moreover, VEGFR2 gene polymorphisms affect the binding efficacy of VEGF to VEGFR2. We therefore assessed the association between VEGFA and VEGFR2 polymorphisms and survival outcomes in patients with diffuse large B cell lymphoma (DLBCL). Patients and Methods: This study included 494 patients with de novo DLBCL treated at 5 hospitals throughout Korea from August 2001 through August 2009. Patients were included if they were (1) ethnic Koreans; (2) had blood samples taken at diagnosis; (3) had been treated with R-CHOP (rituximab, cyclophosphamide, adriamycin, vincristine, prednisolone) with curative intent; and (4) were available for follow-up at the treating institution. Genes and polymorphisms known to modulate angiogenesis were selected. Criteria included: (1) SNPs involved in the VEGF pathway; (2) potentially functional SNPs predicting alterations in protein function; (3) SNPs relevant to outcomes in other settings; (4) and SNPs with a minor allele frequency 〉5% in the study population. We selected a total of five genotypes, three in the VEGFA gene (rs699947, rs833061, and rs3025039) and two in the VEGFR2 gene (rs1870377 and rs2305948). Results: There was a trend towards greater proportions of patients 〉 60 years (P=0.078) patients with bulky disease (P=0.072) and patients who did not achieved complete response (P=0.068) among the VEGF2R rs1870377 TT type than the TA+AA genotype patients. Of the five polymorphisms, VEGF2R rs1870377T〉A was significantly associated with both OS and PFS; in the dominant model, the TT genotype had worse OS (P=0.002) and PFS (P=0.004) than the AA+TA genotype. Among patients with low IPI scores (0 to 2), those with the VEGFR2 rs1870377 AA+TA genotype had significantly better OS (P=0.035); a similar difference for this genotype was observed among patients with high IPI scores (3 to 5) (P=0.043). Patients in the moderate (P=0.031) and high (P=0.043) risk subgroups, according to revised IPI scores, with the VEGFR2 rs1870377 AA+TA genotype also had better outcomes than those with the TT genotype. Multivariate analysis showed that the rs1870377 genotype was an independent prognostic factor for OS (HR for TT vs AA+AT, 1.71; 95% CI, 1.21 to 2.43; P=0.002) and PFS (HR for TT vs AA+AT, 1.57; 1.13 to 2.17; P=0.007). Age 〉 60 years (P=0.0001 for OS; P=0.0001 for PFS), LDH level 〉 normal (P=0.005 for OS; P=0.003 for PFS), extranodal disease 〉 1 (P=0.013 for OS; P=0.017 for PFS) and presence of B symptom (P=0.0001 for OS; P=0.0001 for PFS) were also independent prognostic factors for the survival of patients with DLBCL. Conclusion: The VEGFR2 rs1870377 polymorphism may affect survival in patients with DLBCL. These findings suggest that increased angiogenic activity may be related to tumor progression in patients with DLBCL. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 4898 Introduction The technique of fluorescence immunophenotyping and interphase cytogenetics as a tool for the investigation of neoplasms has recently been introduced to detect molecular cytogenetic abnormalities in plasma cell myeloma of bone marrow (BM) aspirate. However, in case of sub-optimal BM aspirate or the focal distribution of myeloma in the BM, the plasma cells are significantly lower in the BM aspirate than those of biopsy section. Therefore, we have developed a sensitive fluorescence in situ hybridization (FISH) technique which is combined with immunochemistry and is applicable to BM biopsy section for molecular cytogenetic study of plasma cell neoplasms. Patients and Methods Conventional cytogenetic analysis and FISH results of BM samples of 35 multiple myeloma (MM) patients at the time of diagnosis have been evaluated. The probe for IgH rearrangement has been used for hybridization with myeloma cells coupled with CD138 immunostain at BM biopsy section. Results Nineteen patients (54.3%) had abnormal FISH IgH results in biopsy section, whereas seven (20%) cases had abnormal findings in BM aspirate. FISH IgH analysis at biopsy section revealed various signal patterns and proportions (range 6-87%) of cells with atypical signals out of CD138 positive cells. Among five cases with

Description: Bruton tyrosine kinase (Btk) has a well-defined role in B-cell development, whereas its expression in osteoclasts (OCs) further suggests a role in osteoclastogenesis. Here we investigated effects of PCI-32765, an oral and selective Btk inhibitor, on osteoclastogenesis as well as on multiple myeloma (MM) growth within the BM microenvironment. PCI-32765 blocked RANKL/M-CSF–induced phosphorylation of Btk and downstream PLC-γ2 in OCs, resulting in diminished TRAP5b (ED50 = 17nM) and bone resorption activity. PCI-32765 also inhibited secretion of multiple cytokines and chemokines from OC and BM stromal cell cultures from both normal donors (ED50 = 0.5nM) and MM patients. It decreased SDF-1–induced migration of MM cells, and down-regulated MIP1-α/CCL3 in MM cells. It also blocked MM cell growth and survival triggered by IL-6 or coculture with BM stromal cells or OCs in vitro. Importantly, PCI-32765 treatment significantly inhibits in vivo MM cell growth (P 〈 .03) and MM cell–induced osteolysis of implanted human bone chips in SCID mice. Moreover, PCI-32765 prevents in vitro colony formation by stem-like cells from MM patients. Together, these results delineate functional sequelae of Btk activation mediating osteolysis and growth of MM cells, supporting evaluation of PCI-32765 as a novel therapeutic in MM.