Citations

Abstract Abstract 601 A paucity of validated kinase targets in human multiple myeloma (MM) has delayed clinical deployment of kinase inhibitors in treatment strategies. We therefore conducted a kinome-wide small interfering RNA (siRNA) lethality study in MM tumor lines bearing common t(4;14), t(14;16) and t(11;14) translocations to identify critically vulnerable kinases in MM tumor cells without regard to preconceived mechanistic notions. Primary screening was performed in duplicate using an 1800-oligo siRNA library in a single-siRNA-per-well format. siRNA were transfected at low concentration (13nM) to minimize off-target effects using conditions that resulted in transfection of >95% cells and <5% background cytotoxicity. After 96 hours, viability was measured by ATP-dependent luminescence. Fifteen kinases were consistently vulnerable in MM cells, including AKT1, AK3L1, AURKA, AURKB, CDC2L1, CDK5R2, FES, FLT4, GAK, GRK6, HK1, PKN1, PLK1, SMG1, and TNK2. While several kinases (PLK1, HK1) were equally vulnerable in epithelial cells, others and particularly the G-protein coupled receptor kinase, GRK6, appeared selectively vulnerable in MM. GRK6 inhibition is selectively lethal to KMS11, OPM1, H929, KMS18 and OCI-MY5 myeloma cells and has minimal effect on 293, MCF7, SF767, A375 or A549 epithelial cells. Persistent expression of FLAG-GRK6 via cDNA rescued KMS11 cells from the lethal effect of a 3'UTR-targeted GRK6 siRNA, but not from control siRNA, validating identification of GRK6 as an essential myeloma survival kinase. Furthermore, concordant results were obtained using four different exon-based GRK6 siRNA, all of which induced GRK6 silencing and similar inhibition of KMS11 proliferation and viability. Significantly, GRK6 is ubiquitously expressed in lymphoid tissues and myeloma, but by comparison appears absent or only weakly expressed in most primary human somatic tissues. From co-immunoprecipitation experiments we demonstrate that GRK6 is highly expressed in myeloma cells via direct association with the HSP90 chaperone. Inhibition of HSP90 with geldanamycin blocks GRK6 protein expression. Importantly, direct GRK6 silencing causes rapid and selective suppression of STAT3 phosphorylation that is associated with sustained reductions in total MCL1 protein levels and MCL1 phosphorylation (within 24 hours), providing a potent mechanism for the cytotoxicity of GRK6 inhibition in MM tumor cells. GF109203X is an inhibitor of both protein kinase C and of GRK6 that causes near total inhibition of these kinases in vitro at distinct concentrations of 0.1μM and 1-10μM respectively. Notably, GF109203X was substantially cytotoxic to 10/14 myeloma tumor lines at concentrations most consistent with GRK6 inhibition (5-20μM), and was selectively more cytotoxic to myeloma tumor cells than to non-myeloma cell lines (P=0.01), highlighting the potential of GRK6 as a pharmaceutical target for selective therapeutic intervention in myeloma. As mice that lack GRK6 are healthy, inhibition of GRK6 represents a uniquely targeted novel therapeutic strategy in human multiple myeloma. Disclosures: Perkins: MMRC: Employment. Reeder:Celgene: Research Funding; Millennium: Research Funding. Fonseca:Otsuka: Consultancy; BMS: Consultancy; Amgen: Consultancy; Medtronic: Consultancy; Genzyme: Consultancy.

Abstract The objective of this study was to analyze molecular changes that occur at the protein level in WM and IgM-MGUS in order to identify proteins dysregulated at disease progression and design new therapies. We employed antibody protein microarrays (BD Clonetech, CA) to measure changes in the patterns of protein expression between WM, IgM-MGUS and normal lymphoplasmacytic cells as control. The antibody array is a novel technique that assays protein differences directly by hybridizing fluorescently labeled protein mixtures from cell extracts onto a glass slide spotted with 512 different monoclonal antibodies specific for human proteins. CD 19+ and CD138+ purified lymphoplasmacytic cells were obtained from cryopreserved bone marrow samples of 5 newly diagnosed patients with WM and 5 patients with IgM-MGUS. Control plasma cells were obtained from 3 pooled CD19+ and CD138+ purified normal donor bone marrow cells. To assess differential expression, the mean of the ratios of Cy5/Cy3 for each sample were analyzed using the Clontech software to calculate an internally normalized ratio. The normalized data were analyzed by the Genespring software and changes of protein expression ≥2 fold in 60% of the samples as compared to normal control were identified in both WM and MGUS groups. There were 20 proteins upregulated in WM and 21 proteins in MGUS as compared to normal control cells These included proteins from the small GTPases such as the Ras, Rho and Ran families (Rab 4, MONA, RCC1, and CDC42GAP), Kinases/signaling proteins (ROK alpha, p62 dok, JAK, 14-3-3e), cyclin dependent kinase (CDK2), fatty acid synthase (HDAC3), NFkB pathway (IKKalpha, IKBe), and histone deacetylase. The WM and IgM MGUS differed in only 3 proteins using the 2-fold threshold. The WM samples showed overexpression of the heat shock protein Hsp90, the Ras family protein CDC25C, and the chemotaxis protein p43/EMAPII as compared to the MGUS samples. This is the first proteomic study of WM and MGUS patients. It identifies novel proteins dysregulated in WM that may serve in future clinical trials as targets for therapy such as Hsp90, NFkB, HDAC and fatty acid synthase. In addition, it identifies 3 proteins that identify disease progression from MGUS to WM, which could be useful for monitoring MGUS patients and designing targeted therapies to prevent progression to WM.

Abstract Background: CP-751,871, a fully human IgG2 subtype monoclonal antibody, is a potent and specific inhibitor of the insulin- like growth factor type I receptor (IGF-IR). The IGF-IR regulates the growth, survival, adhesion and invasiveness of multiple myeloma cells. High IGF-IR expression is observed in poor-prognostic subtypes of multiple myeloma and its inhibition has been long proposed as a potential therapeutic approach for treatment of this disease. Methods: A phase 1 dose escalation study was conducted to define the safety and tolerability, and to characterize the pharmacokinetic and pharmacodynamic (granulocyte surface IGF-IR expression and serum IGF-I levels) properties of CP-751,871 in patients with multiple myeloma. Patient’s eligibility included previously treated multiple myeloma in relapse or refractory phase including those less than complete remission to autologous stem cell transplant or tandem transplant. Results: Following informed consent and screening, 47 patients were enrolled into 11 dose-escalation cohorts of CP-751,871 at doses from 0.025 to 20 mg/Kg. Median patient age was 60 years. Median number of previous regimens was 4 (range 1–8). CP-751,871 was given as an IV infusion on Day 1 of 4-week cycles. Patients with a suboptimal response to CP-751,871 alone were eligible to receive CP-751,871 in combination with either oral dexamethasone and/or rapamycin at the discretion of the investigator. Twenty-seven patients received CP-751, 871 in combination with dexamethasone, while four patients received rapamycin in combination with either CP-751,871 or CP-751,871 and dexamethasone. Median number of treatment cycles was 3 (range 1–16). Ten patients were dosed at the highest cohort of 20 mg/Kg. No CP-751,871 related dose limiting toxicities were identified. Grade 3 toxicities were all observed at the 20 mg/Kg cohort (1 hyperglycemia, 1 anemia, 1 AST increase, 1 accidental fall, 1 muscle weakness). Plasma CP-751,871 exposure increased with dose, and the pharmacokinetic characteristics were consistent with target-mediated disposition. Granulocyte IGF-IR expression was maximally down-regulated for the entire duration of the dosing period at doses ≥1.5 mg/kg, indicating a saturation of circulating targets. CP-751,871 also led to a dose-dependent increase in circulating IGF-I concentrations. Tumor response was assessed according to Blade criteria. Two remissions and 4 partial remissions were reported in patients treated with different doses of CP-751,871 in combination with dexamethasone. Interestingly, the 2 patients with remission were previously found to be refractory to dexamethasone treatment. Conclusions: These data indicate that CP-751,871 is well tolerated either as a single agent or in combination with dexamethasone. Furthermore, CP-751,981 in combination with dexamethasone, may constitute a novel and effective therapeutic approach for patients with multiple myeloma.

Abstract Abstract 1824 Introduction. Chromosomal abnormalities are prevalent in multiple myeloma (MM) and have been useful in delineating disease subtypes and prognosis groups. Globally, a dichotomous developmental pathway based on ploidy status seems to exist. The accumulation of chromosomal abnormalities in MM often gives rise to complex copy number profiles which are difficult to be captured due to the limitations of karyotyping in a relatively non-proliferative tumor. It is therefore unclear if genomic complexity as a reflection of chromosomal instability (CIN) is of biological and clinical relevance in MM. In this study, we introduce a novel measure of CIN, chromosome instability genome event count (CINGEC), validated its prognostic relevance in myeloma, and subsequently derived a gene expression (GEP) signature, CINGECS, investigated the gene contents of it to elucidate biological mechanisms related to CIN, and assessed its association with survival in the context of other gene expression based prognostic factors. Method. The rationale for CINGEC is that the more unstable is a genome, the more genome events does it harbor irrespective of the size of altered segments. Hence, we first obtain a copy number level sequence from a segmentation result of copy number profile signals from aCGH or SNP chip. We developed an algorithm to estimate the minimum number of genome events to account for the observed complex genome profiles. CINGEC is the sum of genome event counts from all autosomal chromosomes. The GEP signature, CINGECS, is derived from a public dataset that has both aCGH and GEP (GSE26849 and GSE26760, n=246) by performing differential gene expression analysis (SAM, p<0.001, q<0.001, fold-change 2) using GEP data between samples grouped into the top and bottom quartiles based on their CINGEC obtained from the analysis of aCGH data. The biological mechanisms distinctive between high and low CINGEC samples are assessed with biological pathways such as Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO). Results. We first show that CINGEC is associated with MM patient survival by analyzing a previously published Mayo Clinic aCGH dataset (n=64, Chng WJ et al. Leukemia 2010; 24: 833–842). Furthermore, CINGEC is a stronger prognostic factor than another aCGH-based measure of CIN derived from breast cancer, the Genome Instability Index (Chin et al. Genome Biol 2007;8; R215). The CINGECS comprise of 160 differentially expressed genes with 144 up-regulated and 16 down-regulated. KEGG pathway and GO analysis of this set of genes showed enrichment by those involved in (1) cell cycle checkpoints and progression such as cell cycle, cell division, spindle organization, mitosis, (2) DNA damage responses such as response to DNA damage stimulus, DNA repair, nucleotide-excision repair, DNA gap filling, and (3) generic cancer related processes such as DNA replication, cell proliferation. Finally, CINGECS is shown to be significantly associated with poorer overall as well as progression free survival in both newly diagnosed (UAMS TT2 dataset, n=351, GSE 2658) and relapsed patients (Millenium Apex Dataset, n=188, GSE9782). Furthermore, it is found to be a significant prognostic signature independent of diverse GEP signatures known to be associated with survival for MM patients, including the Proliferation Index, the Centrosome Index, the UAMS 70-gene High-Risk Signature, the IFM High-Risk Signature, the IL-6 signature derived from cell lines, as well as GEP-based signatures of genomic instability derived from other cancers such as the CIN70 signature (Carter et al. Nat Genet 2006; 38:1043–1048) and CINSARC67 (Chibon et al. Nat Med 2010; 16: 781–787). Conclusions. We conclude that CINGEC can account for CIN with high resolution copy number profile data. Also, its corresponding GEP signature, CINGECS, potentially encompasses changes reflective of both cause and consequences of the CIN phenotype and is an independent prognostic signature in MM. Disclosures: Fonseca: Consulting:Genzyme, Medtronic, BMS, Amgen, Otsuka, Celgene, Intellikine, Lilly Research Support: Cylene, Onyz, Celgene: Consultancy, Research Funding.

Abstract Abstract 995 PCNSL is an aggressive primary brain tumor characterized by a perivascular accumulation of malignant lymphoid cells. Most PCNSLs (90%) are diffuse large B-cell lymphoma (DLBCL); the remaining 10% are poorly characterized low-grade, Burkitt, and T-cell lymphomas. Since most patients are biopsed, genomic analyses are challenging. To determine the pattern of genetic alterations in PCNSL, frozen samples and formalin fixed embedded paraffin sections from 17 EBV and HIV negative and immunocompetent patients were studied by array-based comparative genomic hybridization (aCGH) using Sureprint G3 (1 million probes) array (Agilent). B-cell differentiation status was characterized by immunostains for CD10, MUM-1, and BCL-6. All cases were characterized by complex genomic aberrations with a median of 21 copy-number abnormalities (CNA) per patient (range 10–49). Overall, 22 minimal deleted regions (MDR) and 14 minimal amplified regions (MAR) were found in more than 20% of patients. Focal deletion affecting CDKN2A (9p21) was the most common CNA, found in 14 of 17 cases (82%); biallelic in six cases. Losses of 6q were observed in 71% of cases. Deletions of 6q23.3 (TNFAIP3) and 6q21 (PRDM1) were found in 59% (10/17) and 47% (8/17) of cases, respectively. Other common CNA were deletions of 6p21 (9/17; 53%), 3p21.1 (5/17; 29%), 3q26.32 (5/17; 29%), 8q12.1 (5/17; 29%), 10p14-p15.3 (5/17; 29%), 12q24.31 (5/17; 29%) and gains of 12q21-q24 (9/17; 53%), 7q21-q31 (6/17; 35%), 19q13 (6/17; 35%), 3q27.3 (5/17; 29%) and 11q24.1-q25 (5/17; 29%). Interestingly, several CNA were unique to PCNSL and were not identified in related entities as the typical DLBCL. Besides in CDKN2A, homozygous deletions were recurrently found in TMEM30A and TOX, the latter a regulator of T-cell development. Another 64 genes, including B2M, CD58, ETV6, LAPTM, MHC class II genes, PRDM1, TNFRSF10A and TNFRSF10B were also homozygously deleted. CD58, which encodes for a member of the immunoglobulin family and regulates the adhesion and activation of T lymphocytes, was also recurrently affected by focal monoallelic losses from 15 nucleotides to 1–2 exons, affecting the Ig-like C2-type domain as was confirmed by DNA resequencing. Focal heterozygous deletions affect TBL1XR1, a negative regulator of the NF-kB and Wnt pathways, and the putative tumor suppressor BCL7A in 29% of cases each. Pathway analysis done including the most commonly affected genes (Ingenuity Pathway Analysis) highlights the importance of networks associated with apoptosis and lymphocyte differentiation and proliferation, especially of T lymphocytes. In summary, this study showed evidence for a highly complex genome and identified target genes of potential relevance in the pathogenesis of PCNSL. The genomic profile described here is unique to PCNSL, thus helping to genetically differentiate this entity from the typical DLBCL and other related lymphomas. Disclosures: Fonseca: Genzyme: Consultancy; Medtronic: Consultancy; BMS: Consultancy; AMGEN: Consultancy; Otsuka: Consultancy; Celgene: Consultancy, Research Funding; Intellikine: Consultancy; Cylene: Research Funding; Onyx: Research Funding; FISH probes prognostication in myeloma: Patents & Royalties.

Abstract Abstract 863 Background: Patients with MM who have progressed after multiple novel agents have limited treatment options. Pomalidomide (CC4047) is the newest immunomodulatory (IMiD) agent. Pom/dex using a dose of 2 mg/day has demonstrated response rates (≥PR) of 63% in relapsed MM (Lacy, JCO 2009, 27:5008-5014) and 32% in a lenalidomide-refractory cohort (Lacy, Leukemia, in press). The maximum tolerated dose has been determined to be 4 mg/day for 21 of 28 days (Richardson, ASH, 2009), We opened two sequential phase II trials using the Pom/dex regimen at differing doses to study the efficacy of this regimen in patients who have failed both lenalidomide and bortezomib. Methods: Patients refractory to both lenalidomide and bortezomib therapy; defined as relapsing on or within 60 days of stopping each regimen, were enrolled. Pomalidomide was given orally 2 mg daily (Cohort A) or 4mg daily (Cohort B) on days 1–28 of a 28-day cycle with oral dexamethasone given 40 mg daily on days 1, 8, 15 and 22. Response was assessed by the International Myeloma Working Group Uniform Response criteria. All patients received aspirin 325 mg daily for DVT prophylaxis. Results: 35 patients with relapsed and resistant/refractory to both lenalidomide and bortezomib were enrolled in each cohort. The median age was 62 years (range, 39–77) in Cohort A and 61 (range, 45–77) years in Cohort B. The median time from diagnosis to enrollment was 57 months for Cohort A (range 12–249) and 72 months(range, 13–183) for Cohort B. 15 patients had high risk molecular markers in Cohort A and 16 in Cohort B. The median number of prior regimens was 6 in both groups. The median (range) duration on treatment was 5(1-13) and 2(0-6) cycles in cohorts A and B respectively. Toxicity at least possibly attributed to drug consisted primarily of myelosuppression: grade 3/4 neutropenia (37% Cohort A vs. 55% Cohort B); grade 3/4 thrombocytopenia (11% Cohort A vs. 13% Cohort B); and grade 3/4 anemia (9% Cohort A vs. 16% Cohort B). Grade 3/4 non-hematologic toxicities occurred in 23% Cohort A vs. 13% Cohort B. Grade 1 or 2 fatigue was the most common non-hematologic toxicity seen in 43% Cohort A vs. 52% Cohort B. Grade 1 or 2 neuropathy occurred in 17% Cohort A vs. 16% Cohort B. Other non-hematologic toxicities occurring in <5% included pneumonitis, hyperglycemia, renal failure, thrombosis. One patient in cohort B had grade 4 hepatitis. Confirmed responses in Cohort A consisted of VGPR 14%, PR 11%, and MR 24% (ORR 49%, 95% CI: 31–66), and responses in Cohort B consisted of VGPR 9%, PR 20%, and MR 12% (ORR 40%, 95% CI: 23–58). The median follow-up on alive patients was 7.5 months, and 3 months in Cohorts A and B, respectively. The median PFS in cohorts A and B are respectively 6.4 months (95% CI: 4.7-NR) and 3.3 months (95% CI: 2.3-NR). Conclusions: Pom/dex is remarkably active and well tolerated in this heavily pre-treated population of dual bortezomib/lenalidomide-refractory MM patients. The majority of patients have not progressed and objective responses (MR or better) are seen in 40–49%. This study confirms therapeutic benefit for Pom/dex in patients relapsing after other novel therapies. These studies do not show an advantage for the 4 mg/day on days 1–28 of each 28 day cycle did not show an advantage over the 2 mg/day on days 1–28 of each 28 day cycle. Disclosures: Lacy: Celgene: Research Funding. Gertz:Celgene: Honoraria; Millennium: Honoraria, Membership on an entity's Board of Directors or advisory committees. 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. Fonseca:Genzyme: Consultancy; Medtronic: Consultancy; BMS: Consultancy; AMGEN: Consultancy; Otsuka: Consultancy; Celegene: Consultancy, Research Funding; Intellikine: Consultancy; Cylene: Research Funding; Onyx: Research Funding; FISH probes prognostication in myeloma: Patents & Royalties. Bergsagel:Celgene: Consultancy; Centocor: Consultancy; Genentech: Consultancy; Amgen: Consultancy; Novartis: Consultancy. Stewart:Celgene: Honoraria.

Abstract Abstract 3963 Background: Pomalidomide at doses of 2 or 4 mg/d has demonstrated excellent activity in patients with relapsed multiple myeloma (MM). Between November 2007 and November 2010, we opened 5 sequential phase 2 trials using the pomalidomide at differing doses with weekly dexamethasone (Pom/dex) regimen to study the efficacy of this regimen. Methods: The five cohorts consisted of: Cohort 1 (N=60): relapsed MM with 1–3 prior regimens, 2 mg dose; Cohort 2 (N=34): lenalidomide refractory, 2 mg dose; Cohort 3 (N=35): bortezomib/lenalidomide refractory, 2 mg dose; Cohort 4 (N=35): bortezomib/lenalidomide refractory, 4 mg dose; and Cohort 5 (N=60) lenalidomide refractory, 1–3 prior regimens, 4 mg dose. Pomalidomide was given orally 2 mg daily or 4mg daily on days 1–28 of a 28-day cycle with oral dexamethasone given 40 mg daily on days 1, 8, 15 and 22. Response was assessed by the International Myeloma Working Group Uniform Response criteria. All patients received aspirin 325 mg daily for DVT prophylaxis or full dose anticoagulation. Results: A total of 225 patients were enrolled across all 5 cohorts. One patient was ineligible and excluded from analysis. The median age was 63 years (32.0–88.0). The median time since diagnosis was 53 months. Forty percent had high-risk molecular markers. Eighty-nine percent had received previous IMIDs including thalidomide (53%) and lenalidomide (81%). Sixty-two percent had previous bortezomib and 73% had prior transplant. The median follow-up is 12.6 months, but varies from 9.4 months for the most recent cohort to 30 months for the first cohort. Sixty-nine percent are alive and 30% remain progression free. Toxicities ≥ grade 3 are shown in table 1 and patient outcomes are shown in Table 2. Conclusions: Pom/dex is remarkably active and well tolerated even in heavily pretreated patients. Responses are durable. Response rates and toxicity are similar between the 2 mg and 4 mg doses. Disclosures: Lacy: Celgene: Research Funding. Fonseca:Consulting:Genzyme, Medtronic, BMS, Amgen, Otsuka, Celgene, Intellikine, Lilly Research Support: Cylene, Onyz, Celgene: Consultancy, Research Funding.

Abstract Primary central nervous system lymphoma (PCNSL) is an aggressive and incurable variant of non-Hodgkin lymphoma (NHL) that is confined to the central nervous system. Most PCNSL (90%) are part of the immune-privileged site-associated diffuse large B-cell lymphomas (DLBCL). Unlike nodal DLBCL, only a limited number of genetic studies have been performed in PCNSL, partly due to lack of available tissue specimens. Because of the fragmented knowledge of the genomic basis, it is still a matter of debate whether they differ from systemic DLBCL with respect to their molecular features and pathogenesis and also if there is a CNS specific signature. We performed a comprehensive genomic study in a cohort of 22 immunocompetent (HIV- and EBV-) PCNSL. DNA was extracted from FFPE tissues (N=15) and frozen tissue (N=7). Samples were analyzed using a combination of aCGH (N=18), exome sequencing (N=10), mate-pair genome sequencing (N=2) and targeted sequencing (N=8). We found a complex karyotype with a median of 21 copy-number abnormalities (range 10–47), 6 structural abnormalities, 6 frameshift indels and 67 nonsynonymous mutations. By integrating mutation and copy number data we found a group of genes recurrently mutated and/or deleted in PNCSL. Remarkable findings were the high prevalence of MYD88 activating mutations (L265P/M232T/V217F), found in 69% of cases and the biallelic loss of CDKN2A (60%). A subset of recurrent abnormalities was exclusively found in PCNSL, and not being previously identified in systemic DLBCL. Thus, 11% of PCNSL have biallelic inactivation of TOX (a regulator of T-cell development) and PRKCD (protein kinase C delta), while another 17% of cases show focal monoallelic deletions/mutations in these genes. Finally, recurrent mutations have been identified in ATM, which have not been found in nodal DLBCL. Several other genes affected have been previously identified in nodal DLBCL, such as biallelic loss of TNFAIP3 (16%), PRDM1 (16%), GNA13, TMEM30A, B2M and CD58 (11% each), activating mutations of CD79B (28%) and CARD11 (19%) and translocations of BCL6 (22%). Components of the NF-kB pathways were altered in >90% of PNCSL. Pathway analysis also showed an enrichment of networks associated with immune response, proliferation, regulation of apoptosis and lymphocyte differentiation and activation. Finally, we searched for associations between genetic alterations and clinical outcome. We showed that deletions of 6q21 (PRDM1) and 6q23 (TNFAIP3) were both associated with shorter overall survival (p=0.007 and p=0.03, respectively). In summary, we report a genomic background in PCNSL similar to post-GC DLBCL but reinforcing the existence of a subset of abnormalities specific to PCNSL, suggesting their potential relevance in the disease pathogenesis. Additionally, the results obtained from FFPE samples are encouraging and larger archival tissue collections can now be analyzed in order to complement the still fragmented knowledge we have of the genetic basis of the disease. Disclosures: Stewart: Onyx: Consultancy, Research Funding; Millenium: Honoraria, Research Funding; Celgene: Honoraria; BMS: Honoraria. Fonseca:Medtronic: Consultancy; Otsuka: Consultancy; Celgene: Consultancy; Genzyme: Consultancy; BMS: Consultancy; Lilly: Consultancy; Onyx: Consultancy, Research Funding; Binding Site: Consultancy; Millennium: Consultancy; AMGEN: Consultancy; Cylene: Research Funding; Prognostication of MM based on genetic categorization of the disease: Patents & Royalties.

Abstract Abstract 987 Background: IMiD® compounds are effective in immunoglobulin light-chain amyloidosis (AL), a systemic plasma cell disorder related to multiple myeloma (MM). Pomalidomide is a new IMiD® compound. Methods: Persons with symptomatic, biopsy-proved, previously-treated AL were eligible for treatment with pomalidomide if they had measurable hematologic disease, ECOG PS≥2, adequate hematologic reserve and creatinine ≤2.5 mg/dL. Pomalidomide, 2 mg/d, and dexamethasone, 40 mg/w, were given orally along with low-dose aspirin. Dose-escalation to 4 mg/d was allowed for non-responders. Exclusion-criteria were uncontrolled infection, cancer, NYHA classification >II, serum troponin-T >0.1 ng/mL, ≥grade-3 neuropathy, thrombosis or other AL therapy within 2 weeks of enrollment. 31 subjects enrolled, but only 29 are evaluble for toxicity and response due to recent enrollment. Results: Median age was 66 y (range, 52 82 y); 19 were male. Median time from diagnosis to study-entry was 36 mo (range, 0.9–104.1 mo). 24 had cardiac involvement, 10, renal involvement and 8, nerve involvement. Cardiac biomarker staging was: I, N=3, II, 21, and III, N=7. Subjects were extensively pretreated: 28 received an alkylator (including an autotransplant in 13), 15, prior lenalidomide or thalidomide and 12, prior bortezomib. 193 cycles of pomalidomide were given to 29 subjects. 11 subjects had protocol-specified dose-increases (no response) and 16, dose-reductions (AEs). 20 subjects had dexamethasone dose-reductions. ≥Grade-3 AE's ≥possibly attributed to therapy occurred in 21 subjects. 7 of these subjects had only hematologic AE's. Neutropenia was the most common severe AE (N=10); severe thrombocytopenia occurred in 2 subjects. Other grade-3/-4 AEs included infection/fever (N=3); fatigue (N=4), and pleural effusion, ascites/edema, and edema each in one patient. 1 subject each had dyspnea, increased creatinine, thrombosis, muscle weakness, atrial tachycardia, and diarrhea. There were 2 episodes of syncope attributed to AL rather than therapy. A subject died 5 d after beginning therapy presumably from cardiac AL. As of 7/27/010, median time on-study is 8.4 mo (range, 0.2–16.2 mo). 18 subjects discontinued therapy for: progression/death (N=9); consent withdrawal (N=4), AE (N=3) and physician choice (N=2). Overall hematologic response rate (N=29) is 38% (95% CI, 20–58%) There were 8 PRs and 3 VGPRs including 4 PRs and 2 VGPRs among 14 subjects failing prior lenalidomide and/or thalidomide. 3 subjects had confirmed organ responses (2 heart and 1 kidney) and 4 unconfirmed organ responses. 1 y survival and PFS are 77% (95% CI, 60–98) and 56% (95% CI, 36–85). Conclusion: These data suggest that pomalidomide could provide a safe and effective therapy in previously-treated persons with AL including those failing lenalidomide and/or thalidomide. Disclosures: Dispenzieri: Celgene: Honoraria, Research Funding; Binding Site: Honoraria. Off Label Use: Non-FDA approved indication. Gertz:Celgene: Honoraria. Kumar:Celgene: Consultancy, Research Funding; Millennium: Research Funding; Merck: Consultancy, Research Funding; Novartis: Research Funding; Genzyme: Consultancy, Research Funding; Cephalon: Research Funding. Fonseca:Amgen: Consultancy; Bristol-Myers Squibb: Consultancy; Celgene: Consultancy, Research Funding; Genzyme: Consultancy; Onyx: Research Funding; Otsuka: Consultancy; Medtronic: Consultancy. Bergsagel:Celgene: Consultancy; Centocor: Consultancy; Genentech: Consultancy; Amgen: Consultancy; Novartis: Consultancy. Stewart:Millennium: Consultancy; Celgene: Honoraria. Lacy:Celgene: Research Funding.

Abstract Abstract 1570 B-cell non-Hodgkin lymphoma encompasses a heterogeneous group of B-lymphocyte-derived malignancies that are characterized by chromosomal translocations involving the immunoglobulin (IG) gene loci and specific histological subtypes of disease are associated with a different spectrum of IG translocations. Marginal zone derived B cell lymphomas encompass three distinct entities: extranodal marginal zone B lymphoma (MZL) of mucosa associated lymphoid tissue (MALT), nodal MZL (NMZL), and splenic MZL (SMZL). MALT lymphoma is genetically unique and four mutually exclusive chromosomal translocations have been identified in this disease thus far. However, the known translocations are only present in a subset of cases suggesting that additional uncharacterized translocations or other genetic events may exist that contribute to disease development. In previous studies we characterized the novel t(X;14)(p11.4;q32) translocation in a patient with MALT lymphoma and found that GPR34, an orphan G-protein coupled receptor (GPCR), was highly expressed due to its juxtaposition to the IGHSA2 switch region. We then measured GPR34 mRNA expression in tissue biopsies from a panel of MALT, NMZL, SMZL, lymphoplasmacytic (LPL), diffuse large B cell (DLBCL), follicular (FL), and mantle cell (MCL) lymphomas. Expression of GPR34 was detected in all tissues examined, but was significantly increased in MALT (37 fold), LPL (23 fold), NMZL (18 fold), and SMZL (21 fold) compared to normal CD19+ B cells. GPR34 mRNA expression was also analyzed by gene expression profiling (GEP) of a panel of lymphomas or control lymphoid tissue and we confirmed elevated expression of GPR34 in the t(X;14) case and detected elevated GPR34 mRNA expression in MALT, LPL, NMZL, SMZL, and ABC DLBCL. When we grouped specimens by high and low expression of GPR34, a clear molecular subtype of SMZL could be identified and suggests that GPR34 high expressing tumors have a unique GEP. To confirm the qPCR and GEP analysis of GPR34 expression, we analyzed surface expression of GPR34 by flow cytometry of normal and malignant tissues and found that expression of GPR34 was detected on all MALT and SMZL B cells with an average DMFI of 2.08 (DMFI range 1.26–3.18, n=12) and on the JeKo-1 lymphoma B cell line (DMFI= 4.75). Analysis of CD19+ cells from normal controls (n=3) revealed a low level of GPR34 on (average DMFI =1.12). Little is known about the function of GPR34 and to further characterize the impact of GPR34 overexpression on cell signaling, OCI-LY19 lymphoma B cells and HeLa cells were transduced with GPR34 or a vector control. To determine if MAPK activity, a pathway regulated by GPCRs, was upregulated, the phosphorylation status of ERK1/2 in GPR34 or control cells was analyzed and we found that ERK1/2 was constitutively phosphorylated at significantly higher levels in GRP34 expressing cells. To further explore which signaling pathways were affected by GPR34 overexpression, GPR34 or vector control cells were transfected with an AP-1, CRE, NF-kB, E2F, SRE, NFAT, or ISRE- luciferase reporter plasmid and GPR34 expressing cells had significantly increased luciferase activity driven by AP-1 (5.35-fold), CRE (4.7-fold), NF-kB (2.6-fold), and E2F (2.1) when compared to vector control cells. To determine the biologic impact of GRP34 overexpression, the proliferation rates of vector control and GPR34 cells were compared and we found that proliferation of GPR34 WT cells was significantly higher than that seen in the control. We next tested the effect of the MEK inhibitor on proliferation and saw a dose dependent decrease in proliferation of GPR34 expressing cells. We then tested the ability of GPR34 cells to generate colony formation in a soft-agar colony formation assay. After 3 weeks of culture, the GPR34 cells had formed significantly more colonies than the vector control suggesting that overexpression of GPR34 can result in increased anchorage-independent cell growth. Taken together, our results identify a novel translocation in MALT lymphoma, emphasize a novel role for GPR34 in regulation of gene expression and tumor cell growth, and suggest that MEK inhibitors may be useful in a subset of translocation–harboring or GRP34 high-expressing tumors. Disclosures: No relevant conflicts of interest to declare.