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The purpose of this study was to quantitate the number and labeling index of monoclonal plasma cells in the blood of patients with newly diagnosed multiple myeloma (MM) to learn if these values were independent prognostic factors for survival. Patients were candidates for this study if they had untreated myeloma requiring therapy, were evaluated at our institution between 1984 and 1993, and had a sample of blood analyzed with a sensitive immunofluorescence technique for monoclonal plasma cells and the blood B-cell labelling index (BLI). The % blood monoclonal plasma cells (%BPC) and the BLI were analyzed along with stage, marrow plasma cell LI, % marrow plasma cells, calcium, creatinine, albumin, beta-2-microglobulin, and C-reactive protein as univariate and multivariate factors for survival. Eighty percent of the 254 patients accrued to this study had monoclonal BPC detected. The median % BPC was 6% and 57% (144 of 254) of patients had a high number (> or = 4%). Patients with > or = 4% BPC had a median survival of 2.4 years vs 4.4 years for those with < 4% BPC (P < .001). The BLI was also prognostic (P = .008). In a multivariate analysis, the % BPC, age, albumin, stage, marrow plasma cell LI, and the BLI were independent factors for survival. The %BPC and the marrow plasma cell LI best separated the group into low, intermediate, and high risk myeloma with median survivals of 52, 35, and 26 months, respectively. Patients with high %BPC were less likely to have lytic bone disease from their MM (P = .002). The %BPC and the BLI are independent prognostic factors for survival and are useful in identifying patients as low, intermediate, and high risk. Clonal cells in the blood should be quantified in future clinical trials for myeloma.

Abstract Beta 2-microglobulin (beta 2M) has been proposed as a prognostic factor in multiple myeloma (MM), but beta 2M levels are reported to correlate with other prognostic indicators such as stage and creatinine level. This study addressed the independent prognostic values of these and other variables, including plasma cell labeling indices (LI), in patients with newly diagnosed MM. beta 2M levels were measured with an enzyme-linked immunosorbent assay. LI were determined with a [3H]thymidine autoradiography method. By multivariate analysis and Kaplan-Meier survival analysis, the uncorrected beta 2M level remained the most significant prognostic factor after adjustment for age. Stage and creatinine level were closely related to beta 2M level and were no longer predictive of outcome after adjustment for age and beta 2M. Plasma cell LI varied independently of beta 2M level and remained predictive. A subset of patients with plasma-blastic myeloma had poor survival since beta 2M level and plasma cell LI were high. By using beta 2M level and LI, three risk groups were defined: low (beta 2M less than 4 micrograms/mL and LI less than 0.4%, median survival 48 months); intermediate (beta 2M less than 4 micrograms/mL and LI greater than or equal to 0.4%, median survival 29 months); and high (beta 2M greater than or equal to 4 micrograms/mL, median survival 12 months). Such grouping may better identify MM patients who might benefit from new treatment regimens.

Abstract Abstract 4238 Background Duplication of the long arm of chromosome 1 (1q) is a recurrent finding in hematologic neoplasms. It is most commonly observed as an unbalanced whole arm translocation with chromosome 7, resulting in concurrent deletion of 7q (ISCN as der(1;7)(q10q10)). It is less frequently observed as an unbalanced translocation with a chromosome other than chromosome 7 or as a duplication of only a segment of 1q. The der(1;7) has been described in myelodysplastic syndrome (MDS), and acute myeloid leukemia (AML), and is reportedly less commonly associated with myeloproliferative neoplasms (MPN). The effect of this rearrangement on prognosis is unclear, although it is classified by the International Scoring System (IPSS) as a karyotypic variant of del(7q)/-7 and is thus assigned poor risk. We have initiated a thorough review of all cases of duplication of the long arm of chromosome 1 seen at our institution. Methods After approval from the Institutional Review Board, all cases seen at Mayo Clinic over a 20 year period (1989-2009) were searched for a duplication of the long arm of chromosome 1. The sole exclusion criterion was a complex karyotype (3 or more independent chromosome abnormalities in the clone). Cytogenetic and hematopathology review was done and relevant clinical data were abstracted from the Mayo Clinic record. Survival of MDS patients and MPN patients was determined from time of cytogenetic diagnosis of der(1;7) until death or last contact with the patient, using the Kaplan-Meier method. Results Among 23,375 unique patients at Mayo Clinic with a cytogenetic study for hematologic neoplasm, 229 (0.98%) patients carried a duplication of 1q. Within this group, 99 (0.42%) had duplication of part of 1q, 123 (0.53%) carried a whole arm duplication of 1q, and 7 patients carried an isochromosome 1q (resulting in 4 copies of 1q). The most common karyotype among those with a whole arm duplication of 1q was der(1;7) (n=84), followed by similar whole arm translocations involving chromosomes 9 and 19 (n=7 each) or chromosomes 12, 13, 14, 15, 16, 17, 18, 20, 21, or 22 (n=1 to 6 each). We focused on 65 der(1;7) patients who had adequate hematopathological material that could be reviewed and sub-classified according to WHO criteria. MDS was identified in 30 patients and MPN in 20. MDS (therapy related, RARS, RAEB-1 or 2) was seen in 9 patients (14%), MDS/ refractory cytopenia with multilineage dysplasia (RCMD) in 19 (29%). MDS/Myeloproliferative neoplasm (MPN) in 2 (3%), MPN not otherwise classified in 5 (8%), and MPN / primary myelofibrosis (including 1 post-polycythemic and one post-thromobocythemic) (PMF) in 15 (23%). The bone marrow was apparently normal in 5 patients (8%), CLL was noted in 3 (5%), and 1 each exhibited features of systemic mast cell disease or myeloma. Acute myeloid leukemia (AML) was identified 5 patients (8%). The M:F ratio of the der(1;7) patients was 2.8:1. The median age at occurrence of the der(1;7) was 63 years (range: 13-87 years). All 5 AML patients have died (median survival 9 months). Of the MDS and MPN patients, the median survival was 2.2 and 2.8 years, respectively, from the time of cytogenetic diagnosis. Overall survival was not significantly different between the MDS and MPN group, and they exhibited 30% survival after 5 years (see Figure). Conclusions Duplication of 1q was observed in nearly 1% of the Mayo Clinic patients who had a bone marrow cytogenetic study for a hematologic neoplasm. Among those with a der(1;7), AML was uncommon but MDS and MPN were similar in frequency. Median survival of 2.2 to 2.8 years for the MDS and MPN group justifies assignment to a more favorable group than the current IPSS assignment. Disclosures: No relevant conflicts of interest to declare.

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: Conventional AlloTx for MM is associated with TRM exceeding 20%. Non-myeloablative AlloTx (NST) utilizes the known graft-vs-myeloma (GVM) effect in an attempt to eradicate minimal residual disease. Preliminary studies of NST in MM demonstrated significant CR and low 100 day post-NST TRM (Badros et al Blood97:2574, 2001). However, even though response rates were high, the vast majority of pts with significant MM disease at the time of transplant, relapsed within the first year. To provide maximal tumor cytoreduction to allow for a subsequent GVM effect, the ECOG designed a Phase II trial of AHSCT for maximal tumor cytoreduction followed by a matched sibling NST to allow for a GVM effect. Methods: The study was designed as a two-stage process based upon early transplant mortality in the first 100 days. Eligibility included pts treated as part of induction therapy or salvage therapy. Chemotherapy responsive disease was not required. Only matched sibling donors were allowed. Pts must have had adequate PS and physiologic organ function. After stem cell mobilization, pts received melphalan 200 mg/m2 with AHSCT. Pts underwent NST 100–182 d post-AHSCT following fludarabine 30 mg/m2 daily × 5 and cyclophosphamide 1 g/m2 for 2 days. GVHD prophylaxis included cyclosporine and corticosteroids. For pts with acute GVHD, Zenapax 1 mg/kg for 5 doses q 3 d was recommended. GM-CSF was incorporated post-AHSCT and NST. The primary endpoints were day 100 TRM, early graft failure, extent of acute GVHD, toxicities, DFS and OS. The study was suspended for 6 mo after the first 19 pts to evaluate early TRM-no early TRM was observed and the study was re-opened. Results: From 5/3/01 to 1/14/05, 32 pts from 10 institutions were enrolled on the study. 23 patients, median age 51 (range 32–64) completed both transplants (28% drop-out rate): 7 (30%) CR (2 in CCR), 11 (48%) PR and 2 (9%) NR and 3 (13%) NE as the best overall response. Of the 32 pts who completed AHSCT, there were 12 grade 4 toxicities pre-NST; 11/23 pts had grade 4 toxicities post-NST. Acute grade III/IV GVHD was observed in 4 (17%); chronic GVHD was observed in 9 pts (39%): 5 limited; 4 extensive resulting in 3 CR (33%), 1 CCR, and 3 PR (33%). 1 pt (4%) had graft-failure. Two pts (8.7%; 95% CI 1.6%, 24.9%) had TRM prior to day + 100 post-NST. With a median follow up of 3.5 years from registration, the median PFS was 4.4 years and the 2-year survival rate was 78% (95% CI: 61% ∼ 95%) for the 23 pts who completed both transplants. Conclusions: AHSCT followed by NST is feasible in a Cooperative Group setting with a low early TRM. The PFS and OS were superior to historical reports for single and tandem AHSCT. AHSCT-NST tandem transplants resulted in a 78% overall response, including 30% CR (2/7 CCR) - similar to other reports for AHSCT-NST. 3/9 pts with chronic GVHD achieved CR demonstrating the GVM effect. However, a plateau in PFS or OS was not observed indicating that the GVM effect is not sufficient to produce long- term disease control and is unlikely curative using this treatment schema. Future studies should include novel therapies to induce better responses pre-transplant and consideration for post-NST maintenance to further eradicate MRD while allowing for GVM.

We studied the prognostic significance of plasmablastic (PB) multiple myeloma (MM) in Eastern Cooperative Oncology Group Phase III trial E9486. Two reviewers independently reviewed 453 cases. They agreed on 37 PB (8.2%) cases and 416 non-PB cases, achieving an 85% concordance (P < .0001). These PB cases had significantly lower hemoglobin and serum albumin levels, higher calcium and β 2-microglobuin levels, and higher percentage BM plasma cells (PC) by immunofluorescence. They had higher bone marrow PC labeling indices, higher serum soluble interleukin-6 receptor (sIL-6R) levels, and a higher probability of ras mutations. Three treatment regimens were used: vincristine, bis-chloro-ethyl nitrosourea (BCNU) melphalan, cyclophosphamide, and prednisone (VBMCP) alone; VBMCP with added cyclophosphamide (HiCy); or recombinant interferon α 2 (rIFNα2). Although the numbers are low, patients with PB had a significantly lower response rate versus non-PB MM when treated with VBMCP (treated, 47.1% v nontreated, 66.5% [P = .015]). Patients with nonresponding PB had a significantly higher progression rate than non-PB cases (30.6%v 11.8% [P < .0001]), especially with VBMCP alone (35.3% v 15.8% [P = .002]), and with added HiCy (37.5% v 9.8% [P < .0001]), but not with added rIFNα2. Event-free and overall survival of PB MM was shorter (median years, 1.1 v 2.7 and 1.9 v 3.7, respectively [P < .0001 for both]). In multivariate analysis, PB classification was also highly prognostic. There is no survival difference between the patients who were classified as PB by both reviewers versus patients classified as PB by only one reviewer. We conclude that PB MM is a discrete entity associated with more aggressive disease and shortened survival. Tumor cell rasmutations and increased sIL-6R may contribute to a higher proliferation rate and reduced survival. There were significant improvements in response and progression with the addition of HiCy and rIFNα2 to VBMCP, but the numbers were small and improved survival could not be shown.

Abstract Background: Patients with newly diagnosed multiple myeloma is at increased risk of thrombotic events, primarily venous thrombosis. The risk is significantly increased in patients receiving certain therapeutic regimens such as the combination of thalidomide and dexamethasone. Deep vein thrombosis, especially if complicated by pulmonary embolism, can result in death. It is not clear if venous thrombotic events can result in compromised survival in this group of patients. Methods: We studied patients enrolled in the Eastern Co-operative Group trial (E1A00) that randomized patients with newly diagnosed myeloma to single agent dexamethasone or thalidomide and dexamethasone. Patients with grade 3 or greater thrombotic events (DVT, or myocardial ischemia, or stroke) were identified. Wilcoxon rank sum test (for continuous variables) or Fisher’s exact test (for categorical variables) was used to compare age, baseline M-protein, hemoglobin, WBC, platelets count, creatinine, beta2-microglobulin, or ECOG PS between patients with and without thrombotic events. Results: Among the 207 patients enrolled, 36 patients (17.4%) had grade 3 or greater thrombotic events with any treatment attribution, of whom, 27 patients had DVT, 3 patients had an MI and 6 pts had a stroke. However, in only 31 of these patients were the event considered treatment-related (possible, probable, or definite) and among these 24 had a DVT. The baseline characteristics did not differ between the patients with thrombosis and those without, irrespective of its relationship to treatment. Similarly, the baseline characteristics did not diff for those with venous or arterial thrombosis. The overall survival was similar among the patients developing a DVT compared to those without a thrombotic event, when all DVT events were considered irrespective of the treatment attribution (Figure). However, majority of the patients with an arterial event (3 patients with MI and 6 with stroke) had a short overall survival. Conclusion: Venous thrombosis remains a common complication in patients with myeloma, especially when treated with thalidomide and dexamethasone. While occurrence of thrombosis clearly contributes to morbidity and often leads to dose reductions and missed doses, it does not appear to have an adverse impact on the overall survival of these patients. Figure Figure

The plasma cell labeling index (PCLI) and serum beta 2-microglobulin (beta 2M) are independent prognostic factors in multiple myeloma (MM). Recently, levels of thymidine kinase (TK) and C-reactive protein (CRP) have been shown to have prognostic value. We studied 107 patients with newly diagnosed myeloma to determine whether TK and CRP values added prognostic information not already available using the PCLI and beta 2M. Univariate survival analysis showed prognostic significance for the PCLI, TK, beta 2M, age, serum albumin, and CRP. Multivariate analysis showed that only PCLI and beta 2M have independent prognostic significance. The survival curves were better separated using the PCLI and beta 2M than with other combinations of variables. Among nine patients under age 65 with low PCLI and low beta 2M, eight were alive almost 6 years after starting chemotherapy. These good-risk patients could not be identified by standard clinical features. Although creatinine and calcium were normal, other features such as bone lesions, osteoporosis, fracture, and anemia were present and stage distribution was similar to other patients in the study. In conclusion, PCLI and beta 2M measured at diagnosis are independent prognostic factors. They must be considered when interpreting the results of clinical trials and should be helpful in counseling patients and in designing new trials. When the PCLI and beta 2M values are known, the TK and CRP values do not add useful additional prognostic information.

Abstract Abstract 3981 Background: Patients with multiple myeloma (MM) can present with life threatening complications including lethal infections during the disease course. The mortality is high during the early period following diagnosis, mostly as a result of complications of the disease or treatment. It is not clear if we can identify patients at the highest risk of early death based on the presenting clinical and laboratory features. Methods: From among 1545 patients seen at our institution between Jan 1999 and Dec 2008, we identified 265 patients who died within 12 months of diagnosis. For each of these patients we identified two “controls” who had at least 12 months of follow up, were alive at the time of last follow up and were closest to the ‘case' patient in terms of time of diagnosis. We performed logistic regression using the clinical and laboratory features, to identify parameters that predict for 12-month mortality and to determine the best cutoffs. Results: This study included 265 patients as cases and 530 controls. The gender distribution was similar in the two groups. We examined the impact of age, performance status (PS), hemoglobin, platelets, serum creatinine, calcium, LDH, albumin, B2M, free light chain difference, plasma cell labeling index, ISS stage, risk category and exposure to novel agents upfront. Based on the results of univariate and then multivariate analysis – age> 72, ECOG PS>2, Calcium >11.3 and ISS stage 3 were found to be significant. We developed the risk score using 1 point for each adverse factor, Age> 72, PS > 2, ISS 3, and Calcium > 11.3. The odds ratio of patients dying in the first year was 2.7, 9.2 and 37 in the presence of one, two and three or more risk factors compared to none of the risk factors. We then looked at the impact of the initial therapy on outcome. 279 patients (35%) received one of the novel agents (thalidomide, lenalidomide or bortezomib) as initial therapy. The odds ratio for dying in the first year was 0.35 for patients receiving a novel agent compared to the rest. The impact of the novel agent was independent of the risk score, with the risk score predicting outcome in both groups. Conclusions: In newly diagnosed MM advanced age, poor performance status, ISS stage and high calcium were associated with early mortality. If eligible all patients should get the benefit of use of novel agent upfront. Identifying these patients at the time of diagnosis is important and should help develop better risk-adapted strategies. Disclosures: No relevant conflicts of interest to declare.

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.