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Allogeneic hematopoietic stem cell transplantation (SCT) regimens incorporating the lymphocytotoxic CD52 antibody alemtuzumab demonstrate efficient engraftment and reduced graft-versus-host disease (GVHD). However, these protocols substantially impair posttransplantation antiviral and antitumor immunity. To accelerate immune reconstitution after alemtuzumab-based reduced-intensity SCT, we administered prophylactic CD8-depleted donor lymphocyte infusions (DLIs) starting on days 60 and 120 after transplantation. DLIs were processed in an immunomagnetic good manufacturing practice depletion procedure resulting in a 2.5- to 6-log reduction in CD8 T cells. Of 23 high-risk patients with hematologic malignancies, 11 received a total of 21 CD8-depleted DLIs. Five patients developed transient grade I acute GVHD following transfer. Only 2 patients with HLA-C–mismatched donors showed grade II and III acute GVHD and subsequently progressed to limited chronic GVHD. Following DLIs, 4 patients with declining hematopoietic donor chimerism converted to full chimeras. A 2.1-fold median increase of circulating CD4 T cells was observed within 2 weeks after infusion. Non-DLI patients did not show a comparable rise in CD4 counts. Four patients demonstrated enhanced frequencies of cytomegalovirus-specific CD4 and CD8 T cells following transfer. Our results suggest that prophylactic CD8-depleted DLIs accelerate immune reconstitution after lymphodepleted HLA-matched SCT and carry a low risk of inducing severe GVHD.

Human PF4 is a heparin-binding chemokine known to be capable of inhibiting endothelial cell proliferation and angiogenesis. To explore the biological mechanisms responsible for this action, we investigated the effect of PF4 on epidermal growth factor (EGF)-stimulated human umbilical vein endothelial cells (HUVEC), a model system in which stimulation is essentially independent of interaction with cell-surface glycosaminoglycans. Based on previous findings that PF4 blocks endothelial cell cycle entry and progression into S phase, we studied the molecular mechanism(s) of PF4 interference with cell cycle machinery. PF4 treatment of EGF-stimulated HUVEC caused a decrease in cyclin E–cyclin-dependent kinase 2 (cdk2) activity with resulting attenuation of retinoblastoma protein phosphorylation. PF4-dependent downregulation of cyclin E-cdk2 activity was associated with increased binding of the cyclin-dependent kinase inhibitor, p21Cip1/WAF1, to the cyclin E-cdk2 complex. Analysis of total cellular p21Cip1/WAF1 showed that in the presence of PF4, p21Cip1/WAF1 levels were sustained at time points when p21Cip1/WAF1 was no longer detectable in cells stimulated by EGF in the absence of PF4. These findings indicate that PF4 inhibition of HUVEC proliferation in response to EGF is associated with impaired downregulation of p21Cip1/WAF1 and provide the first evidence for interference with cell cycle mechanisms by a chemokine.

Although thrombocytopenia associated with the use of histamine H2 receptor (H2R) antagonists has been described, a drug-dependent, platelet-reactive antibody has not previously been identified in such cases. We studied serum from a patient who developed acute, severe thrombocytopenia after exposure to the H2 receptor antagonist, ranitidine, and identified an antibody that reacted with normal platelets in the presence of this drug at pharmacologic concentrations. In flow cytometric and immunoprecipitation studies, the antibody was shown to be specific for the glycoprotein Ib/IX complex (GPIb/IX). From the pattern of monoclonal antibody (MoAb) inhibition and the reactions of antibody with Chinese hamster ovary (CHO) cells transfected with GPIX and GPIbβ, we found that the patient's antibody is specific for an epitope on GPIX close to, or identical with a site recognized by the MoAb SZ1 that is a common target for antibodies induced by quinine and quinidine, drugs structurally unrelated to ranitidine. These findings provide evidence that immune thrombocytopenia can be caused by sensitivity to an H2 R antagonist and suggest that the SZ1 binding site on GPIX may be a common target for drug-induced antibodies. Further studies of the epitope for which SZ1 is specific may provide clues to the mechanism(s) by which drugs promote tight binding of antibody to a membrane glycoprotein and cause platelet destruction in patients with drug sensitivity.

Background Carfilzomib is a novel second generation proteasome-inhibitor with significant anti-MM activity and favorable toxicity profile. In a recent phase 1/2 study in relapsed/refractory patients (pts) a weekly schedule of carfilzomib in combination with dexamethasone showed to be effective (overall response rate of 77%) and safe (ASCO 2015). The ongoing phase 3 ARROW study is comparing once- with twice-weekly carfilzomib. In the newly diagnosed setting, no data are available on weekly carfilzomib. We designed a phase 1/2 study of weekly carfilzomib in combination with cyclophosphamide and dexamethasone (wCCyd) for newly diagnosed MM pts. Results of the dose-escalation phase 1 portion of study were previously reported (Palumbo A et al, Blood 2014), the maximum tolerated dose of weekly carfilzomib was established as 70 mg/m2. Here we report efficacy and safety results of the phase 2 portion of the study. Methods Newly diagnosed pts ineligible for autologous stem-cell transplantation due to age or co-morbidities were enrolled in the phase 2 portion of the study. Pts received IV carfilzomib at the maximum tolerated dose 70 mg/m2 on days 1, 8, 15 combined with oral cyclophosphamide at 300 mg/m2 on days 1, 8, 15 and oral dexamethasone at 40 mg on days 1, 8, 15, 22, in 28-daycycles. After the completionof 9 cycles, pts received 28-day maintenance cycles with carfilzomib at 70 mg/m2 on days 1, 8, 15 until disease progression or intolerance. The primary objectives were to determine the efficacy and safety of wCCyd. The secondary objectives included the evaluation of time to progression, progression-free survival, time to next therapy and overall survival. Response was assessed according to the modified International Uniform Response Criteria. Adverse events (AEs) were graded following NCI-CTCAE v4. Results As of July 15, 2015, 47 newly diagnosed MM pts were enrolled in the phase 2 portion of the study. Median age was 72 years, 23% of pts were older than 75 years, 30% had ISS stage III, 34% had unfavorable FISH profile [t(4;14) or t (14;16) or del17p or amp1]. Toxicityand response data were available in 40 pts, who completed atleast the first cycle; 7 pts were still receiving their first cycle of treatment. Pts received a median of 6 cycles (range 1-9). Overall, 80% of pts achieved at least a partial response, 60% at least a very good partial response, and 28% a near complete response. Responses improved over time (Table 1). During the study, 9 pts progressed or died, the progression-free survival at 1 year was 75%. Grade (G) 3-4 drug-related adverse events included neutropenia (22%, 9 pts), thrombocytopenia (7%, 3 pts), infection (10%, 4 pts), acute pulmonary edema (5%, 2 pts), creatinine increase (5%, 2 pts), fever (2.5%, 1 pt), fatigue (2.5%, 1 pt) and headache (2.5%, 1 pt). G1-2 hypertension was reported in 6 pts (15%). No peripheral neuropathy was reported. Overall, the wCCyd regimen was well tolerated, 4 pts (10%) required carfilzomib dose-reduction (G3 hematologic toxicities [2 pts], G3 headache [1 pt] and G2 fatigue [1 pt]) and 9 pts (22%) required treatment discontinuation due to adverse events (2 infections, 1 acute pulmonary edema, 1 creatinine increase, 1 fever, 1 pt condition, 1 second tumor, 1 pericardial effusion, 1 sudden death). Conclusions This is the first prospective study evaluating once-weekly carfilzomib in treatment-naïve MM. wCCyd therapy appears safe and effective in newly diagnosed MM pts. Responses became deeper with subsequent cycles and toxicities were manageable. The response rate observed with weekly carfilzomib compares favorably with similar studies with standard twice-weekly carfilzomib infusion. Updated results will be presented at the meeting. Table 1. 2nd cycle 6th cycle 9th cycle Complete Response 17% 26% 33% At least near Complete Response 29% 39% 40% At least Very Good Partial Response 66% 82% 87% At least Partial Response 86% 87% 87% Disclosures Bringhen: Janssen-Cilag, Celgene, Novartis: Honoraria; Onyx: Consultancy; Merck Sharp & Dohme: Membership on an entity's Board of Directors or advisory committees. Off Label Use: Use off-label of drugs for the dose and/or schedule and/or association. Larocca:Janssen-Cilag, Celgene: Honoraria. Offidani:Janssen-Cilag, Celgene, Sanofi, Amgen, Mundipharma: Honoraria. Gaidano:Celgene, Onyx: Membership on an entity's Board of Directors or advisory committees. Boccadoro:Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Onyx Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Consultancy, Membership on an entity's Board of Directors or advisory committees. Sonneveld:Janssen-Cilag, Celgene, Onyx, Karyopharm: Honoraria, Research Funding; novartis: Honoraria. Palumbo:Celgene, Millennium Pharmaceuticals, Amgen, Bristol-Myers Squibb, Genmab, Janssen-Cilag, Onyx Pharmaceuticals: Consultancy, Honoraria; Novartis, Sanofi Aventis: Honoraria.

Essential thrombocythemia (ET) is a myeloid neoplasm characterized by platelet activation and thrombotic risk. Aspirin (ASA) is the standard therapy to normal platelet hyperaggregation and to prevent the thrombosis. It is reported that thrombocythaemic patients are ASA insensitive. It is debated if inherited thrombophilia increases the thrombocythemic platelet activation and, hence, the ASA platelet insensitivity. Therefore, we evaluated βFibrinogen G-455-A gene polymorphism, as thrombophilic molecular mutation associated with increased platelet aggregation, platelet count, β-thromboglobulin (β-TG) and platelet factor 4(PF4) as markers of platelet activation, fibrinogen (Fg), platelet functional activity (PFA), as indicator of ASA platelet sensitivity, clot formation time (CFT) and the maximum clot firmness (MCF), as indicators of aspirinated platelet contribution to clot firmness. We studied 40 patients (24 men, 16 women; mean age 56 years, range 37-77) with ET according to WHO criteria. The mean duration of disease was 11 years. All patients were on ASA 100 mg once daily. The βFibrinogen G455-A genotype was determined using a commercialized polymerase chain reaction kit with sequence-specific primers. Platelets were measured by automated analyzer. β-TG and PF4 were determined by ELISA. PFA, CFT and MCF were measured by Platelet Function Analyzer (PFA-100) and by ROTEM delta, respectively. All patients had heterozygous βFibrinogen G455-A. The mean platelet count was 441±72x109/L. All patients had normal Fg (244±47 mg/dl) high β-TG and PF4 (244±15 IU/ml vs 20±11 IU/ml and 162±56 IU/ml vs 6±2 IU/ml, respectively) (p Disclosures No relevant conflicts of interest to declare.

The effects of interferon-alpha (IFN-alpha) on clinical and serologic manifestations of mixed cryoglobulinemia (MC) were investigated by randomized, crossover-controlled trial in 26 patients. The trial alternated 6 months with and 6 months without IFN-alpha therapy (2 x 10(6) IU daily for a month, then every other day for 5 months). In 22 patients, pretreatment steroid dosage remained unchanged during the study. Six patients dropped out (three because of side effects), whereas another 20 patients experienced a significant improvement of purpura (P < .02) and serum transaminases (P < .005) during IFN-alpha treatment. The presence of clinical improvement was supported by the outcome measurements of several immunologic parameters. In particular, serum cryoglobulins were significantly reduced (P < .04) during IFN- alpha therapy. A rebound phenomenon of clinical and serologic parameters was observed after IFN-alpha discontinuation. In addition, no variations were recorded during 6 months without therapy. Hepatitis C virus (HCV) RNA was detected in 91% (20/22) of our MC patients; in 2/13 cases HCV RNA was no longer detectable in serum samples after IFN- alpha therapy. Thus, IFN-alpha could be considered as treatment for MC in patients with HCV seropositivity.

Background Carfilzomib is a novel second generation proteasome-inhibitor with significant anti MM activity and favorable toxicity profile, including very limited neurotoxicity and neutropenia. This Phase I/II study was designed to determine the maximum tolerated dose (MTD) of once weekly carfilzomib combined with cyclophosphamide-dexamethasone (wCCd) and to assess safety and efficacy of this combination in elderly patients with newly diagnosed MM. Here we report the first findings from the Phase I dose-escalation and expansion portions. Enrolment in the phase II portion is ongoing. Methods In the Phase I, the standard 3+3 dose-escalation scheme was adopted, with Carfilzomib as the only escalating agent starting at 45 mg/m2 (level 0), maximal planned dose 70 mg/m2 (level 2), and 36 mg/m2, if needed (level -1), given IV on days 1, 8, 15 in 28-day cycles. Oral cyclophosphamide was administered at 300 mg/m2 on days 1, 8, 15 and oral dexamethasone at 40 mg on days 1, 8, 15, 22 for all dose levels. Dose escalation of Carfilzomib was based on dose-limiting toxicities (DLTs) occurring in cycle 1. After completion of 9 cycles, patients receive 28-day maintenance cycles with Carfilzomib (days 1, 8, 15) at the maximum tolerated dose (MTD) defined by the Phase I study until disease progression or intolerance. The objectives were to determine the MTD and assess activity and safety. Results As of June 15, 2014, 28 newly diagnosed MM patients were enrolled. Median age was 74 years, 29% of patients were older than 75 years, 36% had ISS stage III, 24% had unfavorable FISH profile [t(4;14) or t (14;16) or del17p]. Twelve patients were enrolled in the Phase I portion of the study. At dose level 0 (Carfilzomib 45 mg/m2) no DLT was reported; at dose level 1 (Carfilzomib 56 mg/m2), 1 of 6 patients experienced DLT, consisting of grade 3 creatinine increase; at dose level 2 (Carfilzomib 70 mg/m2) no DLT occurred. The MTD of weekly Carfilzomib was thus established as 70 mg/m2. Toxicity and response data are available for 25 patients, who have completed at least the first cycle; 3 patients are currently receiving their first cycle of treatment. Grade 3-4 drug-related adverse events occurred in less than 15% of patients and included neutropenia (12%, 3 patients), anemia (12%, 3 patients), acute pulmonary edema (8%, 2 patients), pulmonary embolism (4%, 1 patient), creatinine increase (4%, 1 patient), nausea (4%, 1 patient), and fatigue (4%, 1 patient). No peripheral neuropathy was observed. Overall, the wCCd regimen was well tolerated, 3 patients (12%) required Carfilzomib dose reduction (grade 3 creatinine increase, grade 3 transaminase increase and grade 2 fever) and 3 patients (12%) required drug discontinuation due to adverse events (2 acute pulmonary edemas and 1 creatinine increase). Patients received a median of 5 cycles (range 1-9). After 4 induction cycles, 83% of patients achieved at least partial response, 39% at least very good partial response, and 22% complete response. Responses improved over time, as shown in table 1. During the study, only 2 patients progressed and 1 patient died, due to acute pulmonary edema considered probably related to treatment. Conclusions This is the first prospective study evaluating once weekly carfilzomib in treatment-naïve MM. wCCd therapy appears safe and effective in newly diagnosed MM patients. Responses became deeper with subsequent cycles and toxicities were manageable. The response rate observed with weekly carfilzomib compares favorably with similar studies with standard twice weekly carfilzomib infusion. Updated results will be presented at the meeting. Table 1 2nd cycle 4th cycle 6th cycle Complete Response 5% 22% 27% At least Very Good Partial Response 9% 39% 63% At least Partial Response 73% 83% 91% Disclosures Palumbo: Celgene: Consultancy, Honoraria; Janssen-Cilag: Consultancy, Honoraria; Millennium Pharmaceuticals, Inc.: Consultancy, Honoraria; Onyx Pharmaceuticals: Consultancy, Honoraria; Array BioPharma: Honoraria; Amgen: Consultancy, Honoraria; Sanofi: Honoraria; Genmab A/S: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria. Off Label Use: Use off-label of Carfilzomib (proteasome inhibitor).. Bringhen:Merck Sharp & Dohme: Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria; Janssen and Cilag: Honoraria; Celgene: Honoraria; Onyx: Consultancy. Larocca:Janssen Cilag: Honoraria; Celgene: Honoraria. Cavallo:Onyx: Honoraria; Janssen-Cilag: Honoraria; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Boccadoro:Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees; Onyx: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees. Gaidano:Onyx: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Sonneveld:Millenium: Honoraria, Research Funding; Onyx: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding.

Background: Solitary plasmacytoma (SP) is a rare form of plasma cell dyscrasia that represents 2.8-5% of all plasma cell disorders. It is characterized by a localized accumulation of neoplastic monoclonal plasma cells, without evidence of systemic disease, such as multiple myeloma (MM). It can be classified into two groups according to the localization: SP of the bone (SBP) and extramedullary plasmacytoma (EMP). The common presentation of the SBP is in the axial skeleton, whereas the EMP is usually seen in the head and neck. The clinical course and prognosis are quite different between these two entities. Here we present the clinical features, treatment strategies and relative prognostic factors of a series of SP patients followed at a single center over four decades. Methods: A retrospective study was carried out in 53 patients with SP referred at our Institution between 1976 and 2012. Patients’ characteristics are shown in Table 1. The median follow-up was 8.9 years (range 0.5-38.1). Thirty-five patients had SBP and 18 patients EMP. The median age was 56.8 years (range 27-80). The male to female ratio was 1.18:1 and 2.6:1 in SBP and EMP, respectively. The vertebral column (48% of cases) and the upper respiratory tract (50% of cases) were the most common sites of involvement SBP and EMP, respectively. Serum M-protein was detected in 62.8% and 27.7% of the SBP and EMP cases (P=0.0156). The tumor sizes were larger in the SBP group than in the EMP group (P=0.0039). Treatment consisted of local radiotherapy (n=26), combined radiotherapy + chemotherapy (n=15), surgery alone (n=4) and chemotherapy alone (n=8). The radiation dose was available for 35 patients and the median radiation dose administered was 41 Gy (range 21-88 Gy). Results: All patients were evaluable for treatment response and outcome. We found no differences between treatment approaches in terms of response rate, progression-free survival (PFS) and overall survival (OS). The local control rate was 94.3% and there was no statistically significant difference between the two groups. Progression to MM was recorded in 20/35 (57.1%) patients with SBP and in 1/18 (5.5%) patients with EMP. The median time to progression was 2.5 years (range 0.6-11). The 5-year PFS rate for all patients was 66.3% (95% CI: 53.9-81.4). Patients with SBP showed a worse PFS rate than EMP patients (53.0% vs 88.5%), with a statistically significant difference (P=0.0003). The total 5-year OS rate was 78.4% (95% CI: 67.3-91.3). Patients with SBP had a significantly worse OS rate than patient with EMP (71.9% vs 88.2%, respectively) (P=0.0290). Univariate analysis revealed that SBP and a larger tumor size (≥5 cm) were adverse prognostic factors affecting PFS (P=0.0027 and P=0.0498, respectively). SBP had an adverse effect also on OS (P=0.0405). In multivariate analysis, bone localization was the only independent worse prognostic factor for both PFS (P=0.0041) and OS (P=0.0216). Conclusions: Patients with SBP have a significantly worse prognosis than patients with EMP, underlining that they are indeed two different entities within the spectrum of plasma cells dyscrasias. The use of modern testing is mandatory to better characterize the patients’ risks of progression and to exclude an occult MM, especially in SBP. There is also a need for further prospective studies with large series of patients evaluating SBP and EMP separately, to elucidate the role of prognostic factors and treatment options for these conditions. | Characteristics | Patient (number) | | --------------------------------------------------------------------------------------------------------------- | -------------------------------------------------------------------------------- | | Sex, mean (%) | | Male Female | 33 (62.3) 20 (37.7) | | Age (years) | | Median (range) ≤ 60 years, mean (%) | 56.81 (27-80) 33 (62.3) | | Histological type | | SBP mean, % EMP mean, % | 35 (66) 18 (34) | | Tumor size at Diagnosis, number (%) | | ≤5 cm >5 cm N/A | 19 (35.8) 20 (37.7) 14 (26.5) | | Serum M protein (%) | | Positive Negative | 27 (50.9) 26 (49.1) | | Urinary B-J protein | | Positive Negative | 6 (11.3) 47 (88.7) | | SBP Anatomic site | | Vertebral column Long tubal bone Sternum Ribs Craniofacial bone Scapula | 17 (48) 5 (14) 4 (11) 3 (8.5) 3 (8.5) 3 (8.5) | | EMP Anatomic site | | Upper respiratory tract Paranasal sinus Lymph nodes GI tract testis | 9 (50) 4 (22) 2 (11) 2 (11) 1 (6) | Table 1 Disclosures Petrucci: Jansse-Cilag: Honoraria; Celgene: Honoraria; Sanofi: Honoraria; Bristol-Myers Squibb: Honoraria.

Background: Bendamustine is effective as first-line treatment of multiple myeloma (MM) patients, as well as in heavily pre-treated relapsed/refractory patients. The combination of bendamustine with new drugs has been investigated in patients who have exhausted other treatment options. Aims: This single arm study was conducted, after the approval of our Ethic’s Committee, to assess the activity and safety of bendamustine and dexamethasone in heavily pre-treated relapsed/refractory MM patients in our Home Care Unit program. Patients and Methods: Between May 2012 and March 2013, 8 relapsed/refractory frail MM patients were treated with bendamustine (60 mg/m2 days 1, 8, 15) and dexamethasone (20 mg days 1, 8, 15, 22) for up to nine 28-day cycles. Based on the frailty of our patient’s population, the use of other new agents was avoided. All patients were followed by our Home Care Unit in view of the advanced stage of the disease and of the patients’ inability to attend our day hospital unit. All patients had received a median of 4 (range 2-6) previous lines of therapy including alkylating and new drugs, such as thalidomide (25%), lenalidomide (100%) and bortezomib (100%). One patient received, as salvage treatment, a second autologous stem cell transplant (ASCT) followed by a sibling allogenic transplantation. Six patients (75%) were refractory to bortezomib and 5 (62,5%) to lenalidomide, 4 (50%) were double refractory. Main pre-treatment characteristics were: median time from diagnosis 82.1 months (range 14.2- 255.6), median age 76 years (range 53-83), ECOG performance status 2-3, median Hb level of 10.5 g/dl (range 9.0-13.0), median platelet count of 227.000/mm3 (range 58.000-378.000); all patient had bone lytic lesions and 1 had an extramedullary plasmacytoma of the right thigh; 1 patient had a moderate renal failure (creatinine clearance 44ml/min). Stable disease (SD) was considered a valid therapeutic goal in this advanced cohort of patients. Results: After a median number of 6 cycles administered (range 1-9), among the 7 patients evaluable for response (1 patient was withdrawn early due to death) 75% achieved a clinical benefit (≥ SD) and in particular: 1 very good partial response (VGPR), 3 partial response (PR), 1 minor response (MR) and 1 SD. The patient with extramedullary plasmacytoma had a PR confirmed by the mass reduction. Four patients had an early discontinuation of treatment due to: infections in 3 cases (2 fatal) and a heart failure, not related to treatment, in 1. The median time to best response was 4 months (range 1-6). Median time to progression was 10.8 months (range 5.0-15.2) and the median progression-free survival (PFS) was 9.1 months (range 0.6-15.2). Infectious complication was the primary and major side effect in 6/8 (75%) patients, two of which of grade 5, one grade 3 and three grade 2 according to CTCAE ver. 4. Myelotoxicity was acceptable: only one grade 3-4 anemia, thrombocytopenia and neutropenia. The other non-hematological side effect was nausea in all patients (grade 2), which resolved with antiemetic prophylaxis. Conclusions: Bendamustine and dexamethasone administered at home has proven effective, with a favorable balance in terms of cost/effectiveness, in our cohort of high risk and advanced MM patients in poor clinical conditions. The response rate and PFS are encouraging in this setting of patients and this approach should be considered as a valid option for relapsed/refractory MM, including patients double refractory to lenalidomide and bortezomib. In view of the acceptable and manageable toxicity, this combination is feasible in the framework of a Home Care Unit program. Disclosures Off Label Use: Use off-label of Bendamustine (alkylating). Petrucci:Celgene: Honoraria; Janssen-Cilag: Honoraria; Sanofi: Honoraria; Bristol Meyer-Sqibb: Honoraria.

Hepatitis C virus (HCV) may be associated with the mixed cryoglobulinemia syndrome and other B-cell lymphoproliferative disorders (LPDs). The t(14;18) translocation may play a pathogenetic role. Limited data are available regarding the effects of antiviral therapy on rearranged B-cell clones. We evaluated the effects of interferon and ribavirin on serum, B-lymphocyte HCV RNA, and t(14; 18) in 30 HCV+, t(14;18)+ patients without either mixed cryoglobulinemia syndrome or other LPDs. The t(14;18) translocation was analyzed by both bcl-2/JH polymerase chain reaction and bcl-2/JH junction sequencing in peripheral blood mononuclear cells in all patients. Fifteen untreated patients with comparable characteristics served as controls. Throughout the study, the presence or absence of both t(14;18) and HCV RNA sequences were, in most cases, associated in the same cell samples. At the end of treatment, t(14;18) was no longer detected in 15 patients (50%) with complete or partial virologic response, whereas it was persistently detected in nonresponders (P < .05), as well as in 14 of 15 control patients. In 4 responder patients, t(14;18) and HCV RNA sequences were no longer detected in blood cells after treatment, but were again detected after viral relapse; the same B-cell clones were involved in the pretreatment and posttreatment periods. In conclusion, this study suggests that antiviral therapy may induce regression of t(14;18)–bearing B-cell clones in HCV+ patients and that this phenomenon may be related, at least in part, to the antiviral effect of therapy. This in turn suggests that antiviral treatment may help prevent or treat HCV-related LPDs.

Background The current treatment for newly diagnosed elderly multiple myeloma (MM) patients, not eligible for transplant, induces approximately 30% near-complete response/complete response (nCR/CR). Carfilzomib is a novel, irreversible proteasome-inhibitor with significant activity and favourable toxicity profile, including very low rates of peripheral neuropathy and neutropenia. We evaluated efficacy and safety of the combination carfilzomib-cyclophosphamide-dexamethasone (CCd) in elderly newly diagnosed MM patients. Methods The Bryant and Day two-stage design was used to evaluate both efficacy and safety. Patients received oral cyclophosphamide (300 mg/m2 on days 1,8,15), oral dexamethasone (40 mg on days 1, 8, 15, 22) and iv carfilzomib administered over 30 minutes (20 mg/m2 on days 1, 2, and 36 mg/m2 on days 8, 9, 15, 16, cycle 1; 36 mg/m2 on days 1, 2, 8, 9, 15, 16, cycles 2-9) every 28 days for 9 cycles, followed by maintenance with iv carfilzomib (36 mg/m2 on days 1, 2, 15, 16) every 28 days until progression or intolerance. Results Enrollment is complete (58 pts): median age was 71 years, 28% of patients were older than 75 years, 40% had ISS stage III, 35% had unfavorable FISH profile [t(4;14) or t (14;16) or del17p] and 31% are frail, defined according to Charlson co-morbidity index (≥2), geriatric assessment score ADL ( Conclusions The CCd regimen is highly active, showing rapid and deep responses, reaching after 9 cycles, 64% (at least nCR) and 24% sCR, further improving approximately 10-15% during maintenance. These responses compare favorably with the best frontline regimens, showing a doubling in nCR rate. It is well tolerated with limited grade 3-4 AEs, only 11% of patients required drug discontinuation due to AEs. An update will be presented at the meeting. Disclosures: Bringhen: Onyx: Consultancy. Sonneveld:Onyx: Honoraria, Research Funding. Palumbo:Amgen: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Janssen Pharmaceuticals: Consultancy, Honoraria; Millenium: Consultancy, Honoraria; Onyx: Consultancy, Honoraria.

Background The introduction of new agents has substantially changed the management of patients with multiple myeloma (MM). PATIENTS AND METHODS: We evaluated retrospectively 69 symptomatic newly diagnosed transplant-ineligible MM patients treated at our Institute, between 2004 and 2012, with Melphalan and Prednisone (MP) plus Thalidomide (MPT; 23 patients) or plus Bortezomib (MPV; 30 patients) or plus Lenalidomide (MPR; 16 patients). There were 37 men and 32 woman, median age was 73 years (range 65-84) with 20 patients >75 years, 19 (27.5%) were in stage III according to ISS, 12 (17%) had renal failure (creatinine >1.5 mg/dl at baseline), 7 (10%) an underlying diabetes mellitus and 36 (52%) a cardiovascular disease. Melphalan was given at 9 mg/m2 and Prednisone at 60 mg/m2 orally on days 1-4; Bortezomib at 1.3 mg/m2 intravenously on days 1, 4, 8, 11, 22, 25, 29, 32 for the first 4 cycles and thereafter on days 1, 8, 15, 22; Lenalidomide at 10 mg on days 1-21 and Thalidomide 100 mg/daily were administered orally. All patients received antibacterial prophylaxis; thromboprophylaxis and antiviral prophylaxis were administered to patients treated with IMIDs and Bortezomib, respectively. Aims To evaluate the safety and efficacy within the MPT-, MPV- and MPR-treated groups. Results The overall response rates ( ≥ partial response), according to the IMWG criteria, were 20/23 (87%) in the MPT group, 26/30 (87%) in the MPV group, 11/16 (68%) in the MPR group (including 3, 9, 2 very good partial response and 2, 3, 2 complete response/near complete response, respectively). The median PFS was 29 months (95% CI: 18-NA months) for patients who received MPT, 24 months (95% CI: 20-32 months) with MPV and 21.5 months (95% CI: 17-NA months) with MPR, with no significant differences between the three regimens. Among the 69 patients, the overall grade 3-4 hematologic and non-hematologic toxicities were 36% and 43%, respectively. The most common non-hematologic toxicities included all grades of peripheral neuropathy (14% with MPV, 7.2% with MPT and none with MPR), grade 3-4 infections (7.2% with MPV, 4.3% with MPT and 2.9 with MPR) and grade 3-4 cardiovascular disease (4.3% with MPT, 1.4% with MPV and none with MPR). With the use of thromboprophylaxis in all IMID-treated patients, we observed only one deep vein thrombosis in one patient treated with MPT. Conclusions MPT, MPV and MPR are highly active and well tolerated regimens for previously untreated MM patients. The rates of treatment-associated thrombocytopenia and neutropenia were similar between the 3 different regimens and proved transient, predictable and manageable; few patients required supportive care. The results obtained using MP plus one of the recently developed drugs confirm that MPT, MPV and MPR should be considered the standard of care in the frontline treatment of newly diagnosed transplant-ineligible MM patients. Disclosures: Petrucci: Janssen and Celgene: Honoraria; Bristol-Myers Squibb: Membership on an entity’s Board of Directors or advisory committees.

Human PF4 is a heparin-binding chemokine known to be capable of inhibiting endothelial cell proliferation and angiogenesis. To explore the biological mechanisms responsible for this action, we investigated the effect of PF4 on epidermal growth factor (EGF)-stimulated human umbilical vein endothelial cells (HUVEC), a model system in which stimulation is essentially independent of interaction with cell-surface glycosaminoglycans. Based on previous findings that PF4 blocks endothelial cell cycle entry and progression into S phase, we studied the molecular mechanism(s) of PF4 interference with cell cycle machinery. PF4 treatment of EGF-stimulated HUVEC caused a decrease in cyclin E-cyclin-dependent kinase 2 (cdk2) activity with resulting attenuation of retinoblastoma protein phosphorylation. PF4-dependent downregulation of cyclin E-cdk2 activity was associated with increased binding of the cyclin-dependent kinase inhibitor, p21(Cip1/WAF1), to the cyclin E-cdk2 complex. Analysis of total cellular p21(Cip1/WAF1) showed that in the presence of PF4, p21(Cip1/WAF1) levels were sustained at time points when p21(Cip1/WAF1) was no longer detectable in cells stimulated by EGF in the absence of PF4. These findings indicate that PF4 inhibition of HUVEC proliferation in response to EGF is associated with impaired downregulation of p21(Cip1/WAF1) and provide the first evidence for interference with cell cycle mechanisms by a chemokine.

Age is an important prognostic parameter, especially in patients with advanced high-grade non-Hodgkin's lymphoma (HG-NHL) who require more intensive and extensive therapy for any possible chance of cure. We investigated the potential of granulocyte colony-stimulating factor (G-CSF) for reducing myelotoxicity, which is the most important dose-limiting factor for chemotherapy. Between March 1993 and June 1995, 158 previously untreated patients 60 years and older with HG-NHL were included in a cooperative randomized comparative trial and treated with a combination therapy including VNCOP-B (cyclophosphamide, mitoxantrone, vincristine, etoposide, bleomycin, and prednisone) with or without G-CSF. G-CSF was administered at 5 microg/kg/d throughout the treatment starting on day 3 of every week for 5 consecutive days. Of the 158 patients registered for the trial, 149 patients were evaluable: 77 received VNCOP-B plus G-CSF and 72 received VNCOP-B alone. The overall response rate was 81.5%, with complete response in 59%: 60% in the VNCOP-B plus G-CSF group, and 58% in the VNCOP-B group. At 30 months (median 24 months), 68% of all complete responders were alive without disease in the G-CSF group and 65% in the control group. Neutropenia occurred in 18 out of 77 (23%) of the G-CSF treated patients and in 40 out of 72 (55.5%) of the controls (P = .00005). Clinically relevant infections occurred in 4 out of 77 (5%) of the G-CSF group and in 15 out of 72 (21%) of the controls (P = .004). The delivered dose intensity was higher in patients receiving G-CSF (95% v 85%), but the difference was not statistically significant. Our data show that VNCOP-B is a feasible and effective regimen in elderly HG-NHL patients, and that the use of G-CSF reduces infection and neutropenia rates without producing any significant modifications to the dose intensity, CR rate, and relapse-free survival curve.

To the Editor: The possible association between hepatitis C virus (HCV) infection and B-cell–type non-Hodgkin's lymphoma (NHL) has previously been suggested.[1-4][1] HCV tropism for hepatocytes and also, according to recent data, for the salivary gland ductular cells[5][2] may imply the existence

Abstract 200 Background: In a multicenter phase 3 randomized trial, VMPT-VT was superior to VMP for response rates, progression-free survival and time to next treatment (Palumbo A, et al. J Clin Oncol 2010). Here we report an updated analysis on survival after 4 years of follow-up. Methods: Patients (N=511) were randomly assigned to receive nine 6-week cycles of VMPT-VT (induction: bortezomib 1.3 mg/m2, d 1, 4, 8, 11, 22, 25, 29, 32, cycles 1–4, d 1, 8, 22, 29, cycles 5–9; melphalan 9 mg/m2 d 1–4, prednisone 60 mg/m2, d 1–4, thalidomide 50 mg d 1–42; maintenance: bortezomib 1.3 mg/m2 every 14 days and thalidomide 50 mg/day up to 2 years) or VMP alone. After the inclusion of 139 patients, the protocol was amended: both VMPT-VT and VMP induction schedules were changed to nine 5-week cycles and bortezomib schedule was modified to weekly administration (1.3 mg/m2 d 1,8,15,22, all cycles). Results: After a median follow-up of 47.2 months, median OS was not reached in the VMPT-VT arm and was 58.2 months in the VMP arm; 5-year OS rates were 59.3% and 45.9%, respectively (HR 0.74, p=0.04), with 26% reduced risk of death for patients receiving VMPT-VT (Figure-panel A). This benefit was more evident in patients younger than 75 years (5-year rates 67.8% for VMPT-VT vs 49.9% for VMP, HR 0.63, p=0.01, Figure-panel B) and in patients in complete response (CR) after induction (5-year rates 81.4% for VMPT-VT vs 48.2% for VMP, HR 0.38, p=0.006, Figure-panel C) while no significant differences were evident in patients with standard- or high-risk features detected by FISH (HR 0.99, p=0.99). A 1-year landmark analysis for patients completing induction was performed: the 4-year OS was 64.6% in the VMPT-VT group and 49.7% in the VMP group, with 33% reduced the risk of death for patients receiving VT maintenance (HR 0.67, p=0.02). Forty-nine percent of VMPT-VT and 70% of VMP patients relapsed and received subsequent salvage therapies; there was no difference in survival from relapse in the two groups (2-year OS rates 40.7% vs 50.2%,HR 1.11, p=0.54). The median duration of VT maintenance was 23.8 months. During VT maintenance 7% of patients experienced grade 3–4 peripheral neuropathy, 5% grade 3–4 hematological toxicity, 3% grade 3–4 infection and 12% discontinued due to adverse events. Second primary malignancies were reported in 7/254 patients in the VMPT-VT group and 7/257 patients in the VMP group. These corresponded to incidence rates of 0.9 and 1.05 per 100 patient-years, respectively, and were consistent with background incidence rates in the general population (aged 65–74 years 1.9, aged ≥ 75 years 2.3, SEER database). Conclusions: VMPT-VT significantly prolonged OS compared with VMP, especially in patients younger than 75 years and in patients achieving CR after induction. In patients 67–75 years of age, VMPT-VT reduced the risk of death by 37% and it should be considered a new standard of care. Disclosures: Palumbo: Celgene: Advisory Board, Advisory Board Other, Consultancy, Honoraria; Janssen: Advisory Board Other, Consultancy, Honoraria. Bringhen:Janssen: Honoraria; Celgene: Honoraria. Gentilini:Janssen: Honoraria; Celgene: Honoraria. Patriarca:Janssen: Honoraria. Guglielmelli:Janssen: Honoraria; Celgene: Honoraria. Musto:Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding. Petrucci:Janssen: Honoraria; Celgene: Honoraria. Boccadoro:Janssen: Consultancy, Research Funding, Scientific Advisory Board Other; Celgene: Consultancy, Research Funding, Scientific Advisory Board, Scientific Advisory Board Other.

Abstract 3029 Background. In a recent phase 3 trial, bortezomib–melphalan – prednisone–thalidomide followed by maintenance treatment with bortezomib–thalidomide (VMPT-VT) demonstrated superior efficacy compared with VMP. Peripheral neuropathy (PN) was the most important dose limiting toxicity. To decrease neurologic toxicities, the protocol was amended and patients in both arms received once-weekly instead of the initial twice-weekly bortezomib infusions. This post-hoc analysis assessed the impact of bortezomib dose-modification schedule on clinical outcomes and safety. Methods. Patients (N=511) older than 65 years were randomized to receive nine 6-week cycles of VMPT-VT (N=254; induction:V 1.3 mg/m2, d 1, 4, 8, 11, 22, 25, 29, 32, cycles 1–4, d 1, 8, 22, 29, cycles 5–9; M 9 mg/m2 d 1–4, P 60 mg/m2, d 1–4, T 50 mg d 1–42; maintenance: V 1.3 mg/m2 every 14 days and T 50 mg/day) or VMP (N=257) alone. In March 2007, the protocol was amended: both VMPT-VT and VMP induction schedules were changed to nine 5-week cycles and bortezomib schedule was modified to weekly administration (1.3 mg/m2 d 1,8,15,22, all cycles). Patients receiving VMPT-VT and VMP were pooled together and stratified according to the once-weekly or twice-weekly infusion modality; analyses were also conducted for patients receiving VMP only, to eliminate the influence of thalidomide and of maintenance on efficacy and safety. Results. Patients were evaluated in intention-to-treat: 372 patients received once-weekly and 139 twice-weekly bortezomib infusion. Patient characteristics were similar in the two groups, median age was 71 years. The efficacy data did not appear to be affected by the bortezomib schedule. Overall response rates were 85% with once weekly and 86% with twice- weekly schedule (P = .78), including CR rates of 30% and 35% (P = .27).Three-year PFS was 50% in the once-weekly and 47% in the twice-weekly group (P = 1.00), and 3-year OS was 88% and 89%, respectively (P = .54). Similar outcome was seen in the analyses restricted to VMP patients: CR rates were 23% with once-weekly and 27% with twice-weekly schedule (P = .54), 3-years PFS was 46% in once-weekly and 39% (P = .86) in twice-weekly group and 3-years OS was 87% and 89% (P = .47), respectively. The incidence of grade 3/4 hematologic toxicity was similar in the two groups (44% vs 45%, P = .83), but severe thrombocytopenia was slightly less common in the once-weekly patients (19% vs 26%, P = .08).The incidence of non-hematologic grade 3/4 adverse events was significantly reduced in the once-weekly: 35% vs 51% (P = .003). Grade 3/4 gastrointestinal events (6% vs 11%, P = .08), severe systemic events (4% vs 7%, P = .09) and grade 3/4 dermatologic events (2% vs 7%, P = .006) were less frequent in patients receiving once-weekly bortezomib. There was a significantly reduced overall incidence of grade 3/4 PN (8% vs 28%, P < .001) in the once-weekly group. The median time to onset of grade 3/4 sensory PN was 4.3 months in the once-weekly group and 3.2 months in the twice-weekly group (P = .10). The cumulative incidence of sensory PN appeared to plateau after 12 months of therapy in both groups. Rates of discontinuations (5% versus 15%) and dose reductions (15% versus 41%) due to PN were also significantly lower in the once-weekly group (P < .001). These results were reflected in analysis restricted to VMP patients, in which the incidence of grade 3/4 PN (7% vs 29%, P < .001), the discontinuation rate (4% vs 16%, P = 0.002), and the dose reductions rate (15% vs 41% P < 0.001) were significantly lower in once-weekly group. Despite the cumulative planned dose being lower in the once-weekly group (46.8 vs 67.6 mg/m2), the delivered cumulative dose of bortezomib was similar in the two groups (39.4 mg/m2 vs 40.1 mg/m2). No association of PN with age or other baseline characteristics was outlined. The only significant factor influencing the incidence of PN was the reduction of bortezomib infusion from twice- to once-weekly (p Conclusion. These results demonstrate that 1. both once-weekly and twice-weekly schedules in combination with MP ± thalidomide are highly effective in patients ≥ 65 years; 2. once-weekly schedule significantly reduced the incidence of PN and decreased the rate of discontinuation, resulting in similar cumulative bortezomib doses in the two groups; 3. the improvement in the safety profile was not associated with any reduction in the efficacy. Disclosures: Bringhen: Celgene: Honoraria; Janssen Cilag: Honoraria. Leoni:Celgene: Honoraria; Janssen Cilag: Honoraria. Patriarca:Celgene: Honoraria; Janssen Cilag: Honoraria; Roche: Honoraria; Merck: Membership on an entity's Board of Directors or advisory committees. Guglielmelli:Celgene: Honoraria; Janssen Cilag: Honoraria. Elice:Celgene: Honoraria; Novatis: Honoraria. Boccadoro:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen-Cilag: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Palumbo:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Abstract 2370 Regardless of the treatment applied, the outcome for patients with refractory hematological malignancies is extremely poor. For haploidentical stem cell transplantation (haploSCT), which has been evaluated especially in high-risk AML patients lacking an HLA-identical donor, registry data show an overall survival below 10% for AML not transplanted in remission. In order to improve immune competence and to reduce the risk of relapse, we have combined the early transfer of NK cells with the haploSCT approach. PATIENTS AND METHODS: Twenty-five patients (AML = 16, ALL = 5, CML = 2, Hodgkin = 1, MDS = 1) received a haploSCT followed by transfer of purified CD56+CD3-NK cells at day +2. Conditioning consisted of total body irradiation, thiotepa, fludarabine, and OKT3. NK cells were isolated from the CD34- fraction using an automated two-step procedure of CD3+ depletion and subsequent CD56+ selection. No other immunosuppression was routinely delivered. RESULTS: Patients received a mean of 13.3 × 106 CD34+ cells/kg (range 6.12–26.97) with 2.89 × 104/kg (0.95-7.4) contaminating CD3+ cells. A mean of 9.8 × 106 CD56+CD3- NK cells/kg (range 1.61–32.2) was adoptively transferred. Most of the patients developed early GVHD of the skin (median onset day=12), which promptly resolved after short term treatment with steroids and CSA. The main cause of death consisted of infections (n = 10), chronic GvHD (n= 2), and relapse (n = 4). After a median follow-up of 1442 days (3.9 years) (range 0.1 to 7.1 years) 9 of 25 patients are alive and in complete remission resulting in a 2-year overall survival (OS) estimate of 29%. Out of 16 high-risk patients with AML, 10 had refractory or active disease prior to transplantation. For a matched pair analysis, these patients were matched for age, disease status, conditioning regimen and year of transplantation using the EBMT database. AML patients treated with haploSCT plus NK cell transfer had a superior 2-year overall survival (40 vs 11%, 95% CI) in comparison to matched controls with haploSCT only (p=0.02) CONCLUSIONS: HaploSCT plus NK cells is feasible and shows promising survival rates for a group of patients lacking other treatment options. Best results were achieved in patients with AML not only in remission but also with refractory disease. Disclosures: No relevant conflicts of interest to declare.

Abstract 3887 Poster Board III-823 Background Peripheral neuropathy (PN) is a non-hematologic side effect frequently reported in elderly patients treated with bortezomib-melphalan-prednisone (VMP). To address this issue, both bortezomib-melphalan-prednisone-thalidomide (VMPT) and VMP dosing regimens were changed; and bortezomib schedule was modified from twice weekly to weekly administration. Aims To determine incidence and risk factors of bortezomib-associated PN in twice weekly or weekly bortezomib infusion schedules. Methods Patients (N=511) older than 65 years were randomly assigned to receive VMPT followed by maintenance with bortezomib and thalidomide or VMP. Initially, patients were treated with nine 6-week cycles of VMPT (induction: bortezomib 1.3 mg/m2 days 1,4,8,11,22,25,29,32 in cycles 1-4 and days 1,8,22,29 in cycles 5-9; melphalan 9 mg/m2 days 1-4; prednisone 60 mg/m2 days 1-4 and thalidomide 50 mg days 1-42; maintenance: bortezomib 1.3 mg/m2 every 15 days and thalidomide 50 mg/day as maintenance) or VMP (bortezomib, melphalan and prednisone at the same doses and schedules previously described without maintenance). In March 2007, the protocol was amended: both VMPT and VMP induction schedules were changed to nine 5-week cycles and bortezomib schedule was modified to weekly administration (1.3 mg/m2 days 1,8,15,22 in cycles 1-9). Baseline grade ≥ 2 PN was an exclusion criteria. Results 254 VMPT patients and 257 VMP patients were evaluated in intention-to-treat: 141 patients received twice weekly infusion of bortezomib and 370 once weekly. The overall incidence of PN was 37% in the VMPT patients and 27% in the VMP patients (p=0.01) while the grade ≥ 3 was quite similar (8% and 5%. p=0.19). When VMPT and VMP groups were combined, the incidence of PN was significantly higher in patients who received twice weekly infusion of bortezomib: the incidence of all grade PN was 45% in the twice weekly group and 27% in the once weekly group (p=0.0002), including a grade ≥ 3 PN incidence of 16% and 3% (p Conclusion The weekly infusion of bortezomib significantly decreased incidence of PN, discontinuation rate and dose-reduction rate without significant reduction of PFS. The addition of low-dose thalidomide to VMP did not increase the incidence of grade 3-4 PN. An update of these data and correlation between PN and clinical outcome will be presented at the meeting. Disclosures: Bringhen: Celgene: Honoraria; Janssen-Cilag: Honoraria. Boccadoro:Janssen-Cilag: Research Funding, consultancy and advisory committees; Celgene: Research Funding, consultancy and advisory committees; Pharmion: Research Funding, consultancy and advisory committees. Palumbo:Janssen-Cilag: Honoraria; Celgene: Honoraria.