Abstract: Abstract 168FN2 CLL cells with del(17p) typically have loss of functional p53, rendering them refractory to chemotherapeutic agents. However, del(17p) CLL cells activated by CD40 ligand (CD154) are induced to express pro-apoptotic factors to overcome resistance to the cytotoxic activity of p53-dependent drugs, such as fludarabine. To examine whether a CD154-based therapeutic strategy can be developed in vivo for del(17p) and/or fludarabine-refractory CLL, a phase 1b clinical study evaluating an autologous cellular gene immunotherapy is being conducted. Autologous CLL cells transduced ex vivo with a replication-defective adenovirus vector encoding a membrane-stable, re-engineered form of CD154 (Ad-ISF35) are administered, followed by standard courses of FCR. Subjects with fludarabine-refractory and/or del(17p) CLL received three IV doses (one dose every two weeks) of 3×108autologous Ad-ISF35-transduced CLL cells. Two weeks following the third dose of Ad-ISF35-transduced cells, subjects receive up to six monthly cycles of FCR. Study endpoints include analysis of safety and efficacy. Nine (9) subjects have been enrolled and treated on study. Median age was 63 (range 48–70). All subjects were del(17p) (range 14–96%), and included treatment naïve (n=4) and previously treated (n=5) subjects. The number of prior treatments range from 0–5, including three subjects that previously received fludarabine-containing regimens. The overall response rate was 67% with 56% of subjects achieving a complete response (CR), including 3 CRu pending bone marrow assessment. Two subjects with a marked percentage del(17p) (range 63–66%) continue to have an ongoing complete response (CR) after a median follow up of 〉 2 years, and no detectable minimal residual disease (MRD) in one subject. Three subjects that showed disease progression were treated with either alemtuzumab (1 subject) or ofatumumab plus high dose methylprednisolone therapy followed by allogeneic stem cell transplant (2 subjects). We observed clinical responses not only after FCR but also after infusion of Ad-ISF35-transduced cell. These ISF35-specific responses included reductions in absolute lymphocyte counts in all subjects (decrease from baseline 4–89%), and decreased lymphadenopathy ( 〉 50% reduction) in 78% of the subjects (decrease from baseline 19–100%). Infusion of Ad-ISF35-transduced cells plus FCR has been well-tolerated. The primary non-hematologic adverse events have been flu-like symptoms following infusion of Ad-ISF35 transduced cells. This includes transient grade I/II fever (89%), fatigue (56%) and chills (56%). The primary hematologic adverse events have been cytopenias following FCR treatment, including grade III/IV neutropenia (33%) and anemia (22%). Grade I/II hypophosphatemia (56%) following ISF35 has been observed and this might be related to increased serum cytokine levels following Ad-ISF35-transduced cell administration. Correlative studies on CLL cells obtained before and after infusions of Ad-ISF35-transduced CLL cells demonstrated that CLL cells prior to treatment were refractory to the cytoxic effects of P53-dependent drugs (e.g. F-ara-A). However, the CLL cells obtained after treatment with Ad-ISF35-transduced CLL had increases of p73, p21 and Bid and became sensitive in vitro to the cytotoxic activity of F-ara-A. We also observed up-regulation of costimulatory molecules (CD80, CD86, CD54) and death receptors (CD95). The majority of subjects developed antibodies against adenovirus with neutralizing activity. However, they did not developed antibodies against human CD154. Subjects also showed increases in TNFα, IL-6 and IL12 after infusion of Ad-ISF35 transduced cells. In conclusion, the combination of Ad-ISF35 transduced CLL cells plus FCR appears to be well-tolerated and highly effective in CLL patients with fludarabine-refractory disease and/or del(17p). The CR rate that we have observed in this high-risk CLL population is higher than those reported in the literature and makes our results very encouraging. Correlative data suggest that Ad-ISF35 promotes upregulation of costimulatory and death receptor molecules as well as pro-apoptotic proteins that may overcome resistance to FCR in vivo. These encouraging data suggest the combination of Ad-ISF35 plus chemoimmunotherapy could offer an effective treatment option for patients who otherwise would be resistant to standard forms of therapy. Disclosures: Cantwell: Memgen, LLC: Employment, Patents & Royalties.

Abstract: Abstract 4422 Background: Nilotinib is a potent, highly selective Bcr-Abl kinase inhibitor approved for newly diagnosed adult patients (pts) with Philadelphia chromosome–positive (Ph+) chronic myeloid leukemia in chronic phase (CML-CP) and for adult pts with imatinib-resistant or -intolerant Ph+ CML-CP and CML-AP (accelerated phase). This ongoing study assesses the change in chronic low-grade (LG) non-heme adverse events (AEs) when pts are switched from imatinib (IM) to nilotinib. Methods: Adult CML-CP pts were eligible for the study if they were treated with imatinib 400 mg/d for ≥3 months (mos) and had imatinib-related Grade 1 or 2 non-heme AEs persisting ≥2 mos or recurring ≥3 times and recurring despite best supportive care. Pts are treated with nilotinib 300 mg twice daily on study for up to 1 year. The primary end point is to measure the improvement of imatinib-related LG non-heme AEs at the end of cycle (EOC) 3 after switching to nilotinib therapy. Disease response was monitored and patient-reported outcomes measured by 2 quality-of-life (QoL) questions and the MD Anderson Symptom Inventory (MDASI)-CML. Results: Thirty-eight pts were enrolled as of the data cut-off date (6/27/11) and were included in this analysis. The median time of nilotinib treatment was 7.2 mos. A total of 155 imatinib-related non-heme AEs were reported at baseline; 113 AEs were Grade 1 and 42 AEs were Grade 2. A total of 30 pts completed EOC 3 by the data cut-off date. These pts accounted for 126 of the baseline imatinib-related LG non-heme AEs (Grade 1 = 93, Grade 2 = 33). The median number of IM-related LG non-heme AEs at baseline was 3 per patient. Twenty-one pts reported 1–4 baseline IM-related AEs, 6 pts reported 5–9 IM-related AEs, and 3 pts reported 10–12 IM-related AEs. Of these AEs, 81 (64%) improved (primary end point) by EOC 3; 71 IM-related AEs resolved (51, 15, 5 resolved by mos 1, 2, 3, respectively) and 10 IM-related AEs decreased from Grade 2 to 1. Forty-two AEs were unchanged across 18 pts (20 of which were reported by 3 pts). Three AEs increased in severity by month 3. Overall, 31 (82%) pts had major molecular response (MMR) at entry. MMR is defined as a 3-log reduction of Bcr-Abl from a standardized baseline (Bcr-Abl ≤0.1% IS). All pts maintained MMR after switching to nilotinib on study. The remaining 7 pts achieved MMR during the study. At baseline, 15 pts had a 4-log reduction in Bcr-Abl (Bcr-Abl ≤0.01% IS) and 7 pts with complete molecular response (CMR = Bcr-Abl ≤0.0032% IS). Twelve additional pts achieved a 4-log reduction on study and 9 went on to achieve CMR. Patients completed 2 global QoL questions and the MDASI-CML questionnaire during the study. The MDASI-CML is a patient-reported outcome measure of symptom burden in patients with CML. These were administered at baseline, EOC 1, EOC 3, and then every 3 mos thereafter while on study. Compared to baseline, 68% and 62% of pts (n=34) reported an improvement in global QoL over the last 24 hours and last 7 days, respectively, by EOC 3. The mean reductions from baseline in MDASI-CML severity score and interference score, and therefore improvement in symptoms, were at EOC 1: 1.2 (n=26) and 1.5 (n=25) and at EOC 3: 1.2 (n=24) and 1.6 (n=23), respectively. Thirteen pts were dose reduced for nilotinib-related AEs and subsequently dose re-escalated if the AEs recovered to Grade 1 or resolved. Twenty-seven Grade 3 AEs occurred in 12 pts; of these, 17 AEs were investigator reported and suspected to be nilotinib related (increased bilirubin, hyperglycemia, hypokalemia, hypophosphatemia, increased lipase, pruritus, bronchitis, dehydration, exfoliative rash, rash erythematous, rash, and arthralgia). No pt had a Grade 4 AE. Most AEs were managed by brief dose interruption. A total of 5 pts discontinued, 4 for AEs, and 1 pt withdrew consent. No QTcF prolongation 〉 500 msec occurred. Conclusions: In this analysis, at EOC 3, 64% of the chronic LG non-heme IM-related AEs showed improvement after switching to nilotinib. Twenty-eight of 30 pts who completed 3 mos on study had at least 1 LG non-heme IM-related AE improve after switching to nilotinib. At least 62% of pts improved in QoL. In addition, an overall improvement of symptoms as measured by MDASI-CML was seen by the reduction of severity scores. Disclosures: Lipton: Novartis Canada: Consultancy, Research Funding, Speakers Bureau. Mauro:Novartis Oncology: Consultancy, Research Funding, Speakers Bureau. Ailawadhi:Novartis Pharmaceuticals: Consultancy, Speakers Bureau. Miller:Novartis: Consultancy, Research Funding, Speakers Bureau. Busque:Bristol-Myers Squibb: Consultancy; Novartis Pharmaceuticals: Consultancy, Research Funding, Speakers Bureau. Akard:Eisai: Speakers Bureau; Bristol Myers-Squibb: Speakers Bureau; Novartis: Speakers Bureau; Millenium: Speakers Bureau; Chemgenex: Consultancy. Pinilla-Ibarz:Novartis Pharmaceuticals: Consultancy, Research Funding, Speakers Bureau; Bristol-Myers Squibb: Consultancy, Research Funding, Speakers Bureau. Ericson:Novartis Pharmaceuticals Corporation: Employment, Equity Ownership. Shah:Novartis Pharmaceuticals: Employment, Equity Ownership. Cortes:Bristol-Myers Squibb: Consultancy, Research Funding; Novartis Pharmaceuticals: Consultancy, Research Funding; Ariad Pharmaceuticals: Consultancy, Research Funding; ChemGenex Pharmaceuticals: Consultancy, Research Funding; Pfizer: Research Funding; Deciphera Pharmaceuticals: Research Funding.

Abstract: Chronic lymphocytic leukemia (CLL) represents 30% of adult leukemia. TCL1 is expressed in ∼ 90% of human CLL. Transgenic expression of TCL1 in murine B cells (Eμ-TCL1) results in mouse CLL. Here we show for the first time that the previously unexplored endoplasmic reticulum (ER) stress response is aberrantly activated in Eμ-TCL1 mouse and human CLL. This includes activation of the IRE-1/XBP-1 pathway and the transcriptionally up-regulated expression of Derlin-1, Derlin-2, BiP, GRP94, and PDI. TCL1 associates with the XBP-1 transcription factor, and causes the dysregulated expression of the transcription factors, Pax5, IRF4, and Blimp-1, and of the activation-induced cytidine deaminase. In addition, TCL1-overexpressing CLL cells manufacture a distinctly different BCR, as we detected increased expression of membrane-bound IgM and altered N-linked glycosylation of Igα and Igβ, which account for the hyperactive BCR in malignant CLL. To demonstrate that the ER stress-response pathway is a novel molecular target for the treatment of CLL, we blocked the IRE-1/XBP-1 pathway using a novel inhibitor, and observed apoptosis and significantly stalled growth of CLL cells in vitro and in mice. These studies reveal an important role of TCL1 in activating the ER stress response in support for malignant progression of CLL.

Abstract: Abstract 2769 Background: Since imatinib was approved in 2001, tyrosine kinase inhibitors (TKIs) have become standard of care for first line treatment of Chronic Myeloid Leukemia (CML). Macrocytosis has been observed in some patients being treated with imatinib but the etiology of this phenomenon is unclear. Altered DNA metabolism resulting from the inhibition of c-kit by imatinib may be an explanation but anecdotal correction of macrocytic anemia after iron replacement has prompted us to explore this further. The correlation between bone marrow iron stores, anemia and macrocytosis has not been extensively studied in this population. Methods: The Total Cancer Care (TCC) database was used to identify all patients treated for CML at Moffitt cancer Center (MCC) between 1992 and 2010. All patients treated with TKI's were evaluated. Bone marrow iron stores, macrocytosis and anemia were recorded from the first records available at MCC while patients were taking TKI's. Iron stores, anemia and macrocytosis were recorded as categorical variables, while hemoglobin was recorded as a continuous variable. Descriptive data were reported and chi square test was used for categorical variables. Statistical analysis was done using SPSS statistical software, version 19. Results: A total of 540 patients were treated for CML at MCC between 1992 and 2010, and 478 received treatment with TKI's. Of these, 53% were male, 47% were female and 70% were under the age of 60 at diagnosis. At the time of data cutoff 68% were still living. Imatinib was the first TKI used in 470 (98.3%) patients, while nilotinib and dasatnib were first line treatment in 6 (1.3%) and 2 (0.4%) patients respectively. Second line TKI's were necessary in 177 (37%) patients. Anemia was present in 274 (57%) of the 478 patients treated with TKI's. Bone marrow iron stores were increased, adequate or decreased/absent in 38 (8%), 85 (18%) and 289 (60%) respectively. Reports on iron stores were missing in 66 (14%) patients. One hundred eighty nine (40%) patients had macrocytosis compared with 278 (58%) who did not. This data was missing in 11 (2%) patients. Data correlating iron stores and anemia were available on 410 patients (Table 1). From them, 178 (43%) patients were anemic. Of these, 34 (19%) had adequate iron stores, 131 (74%) had decreased/absent iron stores and 13 (7%) had increased iron stores. There were 232 patients with available data on iron stores who were not anemia. In this group, iron stores were adequate, decreased/absent and increased in 51 (22%), 156 (67%) and 25 (16%) patients respectively. Data correlating anemia with macrocytosis were available in 467 patients (Table 2) and in this assessment 270 patients were anemic. One hundred nineteen (44%) had a macrocytic anemia while 151 (56%) did not. Of the 197 patients in this evaluation who were not anemic, 70 (36%) had a macrocytosis while 127 (64%) did not. Lastly, an association between iron stores and macrocytosis was looked at, and there were 407 evaluable patients in this group (Table 3). Macrocytosis was noted in 239 patients and of these, bone marrow iron stores were adequate, decreased/absent, and increased in 32 (19%), 120 (71%) and 16 (10%) patients respectively. Of the 168 patients without macrocytosis, iron stores were adequate in 53 (22%) patients, decreased/absent in 164 (69%) patients and increased in 22 (9%) patients. Conclusions: Macrocytic anemia is most often caused by an alteration in DNA metabolism due to nutritional deficiencies or medications. Contrary to what would be expected, out data show that over two-thirds of the patients with macrocytosis had decreased iron stores in their bone marrow. Similarly, nearly half of all anemic patients had a macrocytic anemia which raises the question of whether iron deficiency really is the primary etiology of their anemia. Inhibition of kinases involved in iron metabolism may be involved in the etiology of the macrocytosis seen in these patients. Disclosures: No relevant conflicts of interest to declare.

Abstract: Abstract 4758 Introduction: Among patients with chronic myeloid leukemia (CML), the success of clinical care has traditionally been evaluated in terms of survival, defined clinical events (e.g., transfusions, hospital days), and test results (e.g., major molecular or cytogenetic response). However, some of these measures may not adequately describe whether patients actually show improvements in health-related quality of life (HRQoL). Conceptually, increases in HRQoL are derived from two possible effects of pharmacotherapies: (a) comparative improvement of disease symptoms, and (b) reduction of therapy-related adverse events (AEs). With the advent of tyrosine kinase inhibitors (TKIs), the leading agents in CML treatment, improved morbidity and delayed mortality have been achieved with astonishing success. Moreover, through rigorously conducted research, much has been learned about the tolerability profiles to these agents with careful delineation of AEs. Recently, there has been a growing recognition by the clinical community about the need to examine the impact of TKI-related AEs on HRQoL from the patient's perspective as well as the perspectives of their health care proxies including physicians and family caregivers. As summarized in Figure 1, the triangulation of these three perspectives in cancer care has been shown to provide valuable information that informs clinical decision-making, improves adherence to prescribed treatment regimens, and optimizes care management. Thus, the purpose of this study was to evaluate the CML literature to determine the extent to which the consequences of TKI-related AEs on HRQoL has been explored from the comparative perspectives of physicians, patients and family caregivers. Methods: A systematic literature review of PubMed conducted by three independent reviewers identified 44 articles using combinations of the following broad key words: CML, HRQoL, imatinib, nilotinib, dasatinib, tyrosine kinase inhibitors, adverse events, side effects and toxicity. Articles that were economic focused (n=11), commentaries (n=9), systematic reviews without HRQoL as an endpoint measure (n=7), non-English publications (n=4), studies on pediatric patients or those without CML as a primary diagnosis (n=5), non-TKI specific (n=4) or multiple publications on the same patient population (n=1) were excluded from review. The remaining 3 articles employed different study designs including cross-sectional, longitudinal and randomized clinical trial approaches with HRQoL as a primary endpoint. Collectively, they involved 1,587 CML patients treated with TKIs. Results: The studies were limited to patients in the chronic phase of CML when HRQoL was likely to be at its highest. The average follow-up time was short, ranging between 6 to 18 months with no long-term follow-up. None of the studies provided the comparative perspective of the physician or the primary family caregiver in relation to the patient with regards to either short-term or long-term impact of AEs on HRQoL. Rather, the studies were centered solely on the patient's reported HRQoL secondary to their experience with TKI therapy but none actually linked the type, frequency or severity of AEs to the various dimensions of HRQoL. Therefore, no information was available on which AEs have the greatest effect on what dimension of HRQoL. The most commonly reported AEs that adversely impacted HRQoL included (a) non-hematologic events related to gastroenterology, dermatology, musculoskeletal system and other (e.g., edema); and (b) hematologic events such as neutropenia, thrombocytopenia, and anemia. Across the studies, the most consistently distressing AE experienced by 〉 80% of patients was “lack of energy”. Desired life activities, such as being able to return to work as well as the ability to participate in social and family engagements, were also rated highly in relation to better HRQoL. Conclusions: On the whole, little to no data are available on the temporal impact of TKI-related AEs on the multiple dimensions of HRQoL. This substantial dearth of information in the CML literature is a stark contrast to other types of cancers and therapeutic agents where the myriad of factors that affect the patient's HRQoL have been investigated from the perspectives of key stakeholders in the health care paradigm, resulting in optimal clinical decision-making and improved adherence to prescribed regimens. Disclosures: Pinilla-Ibarz: Novartis Pharmaceuticals Corporation: Consultancy. Navaie:Novartis Pharmaceuticals Corporation: Consultancy. Smith:Novartis Pharmaceuticals Corporation: Consultancy. Beaubrun:Novartis Pharmaceuticals Corporation: Consultancy. McCullough:Novartis Pharmaceuticals Corporation: Consultancy. Hickey:Novartis Pharmaceuticals Corporation: Consultancy. Toseland:Novartis Pharmaceuticals Corporation: Consultancy.

Abstract: Abstract 2007 Introduction: Identification of genetic and molecular markers of high risk disease for chronic lymphocytic leukemia (CLL) have improved risk-stratification of these patients and allow for adapted treatment. Allogeneic hematopoietic cell transplantation (HCT) offers a chance for prolonged disease free survival and there is increasing use of this strategy for CLL. The risk which these factors incur on HCT outcomes remains uncertain. Methods and Patients: We performed a retrospective analysis of all CLL patients who underwent aHCT at the Moffitt Cancer Center between 2003 and 2011. Our IRB approved study was designed to examine the effect of 13 variables upon overall survival (OS) via univariate and multivariate analysis. Secondary outcomes included progression free survival (PFS), non-relapse mortality (NRM), and relapse rate. Variables were: age (years) at HCT of 〈 50 vs. 50–59 vs. 〉 =60; HLA matched related donor (MRD)/matched unrelated donor (MUD) vs. mismatched unrelated donor (MMUD); Chemoresponsive to last therapy (CR/PR) vs. stable disease (SD) vs. progressive disease (PD); fludarabine vs. pentostatin-based conditioning regimen; Karnofsky performance status (KPS) 〈 90%; presence of any acute GVHD; presence of deletion 17p; presence of deletion 11q; 〉 =3 treatment regimens prior to HCT; 〉 =3 enlarged lymph nodes (LN) areas at HCT; presence of 〉 =5 cm LN (bulky disease) at HCT; 〉 =30% CLL bone marrow involvement; HCT specific comorbidity index (HCT-CI) score 〉 = 1. 43 patients, 37% female, were identified. At diagnosis of CLL 19% had B symptoms; 58% had a Rai stage of II-IV; and 35% had bulky disease. Genetic abnormalities present at any time before HCT were: 35% del 17p and 35% del 11p. Two patients had Richter's transformation. Prior to HCT conditioning, 88% had received fludarabine and 63% had received 〉 = 3 regimens. Median age at HCT was 55 (34–65) years, 19% had bulky disease, and 63% had a Karnofsky performance status (KPS) of at least 90%. Disease status at HCT: CR (23%), PR (33%), SD (21%), PD (23%) and the number of LN areas at HCT 〉 =3 in 65% patients. The HCT-CI was 〉 = 1 in 56%; donors were 42% MRD, 35% MUD, and 23% MMUD (including 3 umbilical cord blood); conditioning regimens were 33% fludarabine+busulfan, 37% pentostatin+busulfan, 19% fludarabine+TBI and 11% other. Results: At a median follow-up of 2.6 (range:) years for survivors, median OS and PFS were 46.5 mo and 31.4 mo by Kaplan Meier estimate. By univariate analysis, 5 factors were found predictive of worse OS: age at HCT 〉 =60 (HR=6.1,95%CI:1.5–24.9) vs reference 〈 50; MMUD (HR=3.3;95%CI:1.3–8.1); PD(HR=4.9,95%CI:1.9–12.4) vs reference CR/PR; presence of del 17p (HR=2.7,95%CI:1.1–6.2); and presence of del 11q (HR=5.2,95%CI:2.1–13.0). The same five factors were predictive for PFS compared to reference group: age at HCT 〉 =60 (HR=5.5,95%CI:1.3–22.4) vs reference 〈 50; MMUD (HR=3.6;95%CI:1.5–9.0); PD(HR=5.7,95%CI:2.2–14.4) vs reference CR/PR; presence of del 17p (HR=2.3,95%CI:1.0–5.3); and presence of del 11q (HR=4.2,95%CI:1.7–10.4). The presence of del 11q (n=15) and del 17p (n=15) did not correlate (p=1.0). Median OS was significantly worse for del 17p patients (7 mo vs 46 mo, p=0.0193) or those with del 11q (7 mo vs. 55 mo, p=0.0001). The cumulative incidence of relapse was not different among del 11q (HR=0.82; 95%CI: 0.22 – 3.11) or del 17p patients (HR=0.86; 95%CI: 0.22 – 3.43) while NRM was worse for del 11q patients(HR=5.14; 95%CI: 1.82–14.6). Published data is inconclusive on the influence of genetic risk factors upon outcomes after HCT for CLL. Therefore, we conducted a multivariable analysis recognizing the number of variables in the final model would be relatively large considering the limited number of events. In our model the following three variables were associated with worse OS: MMUD (HR=3.7; 95% CI:1.1–12.6; p=0.035); del 17p (HR=5.1; 95%CI:1.6–16.6; p=0.006); and del 11q (HR=10.9; 95%CI:3.1–38.4; p 〈 0.001). These same 3 variables correlated with worse PFS: MMUD (HR=14.4; 95% CI:3.1–66.7; p=0.001); del 17p (HR=4.4; 95%CI:1.4–14.5; p=0.014); and del 11q (HR=16.0; 95%CI:3.8–68.5; p 〈 0.001). Conclusion: For patients undergoing HCT for CLL, the presence of del 17p OR del 11q predicted for worse OS and PFS; however, the risk of relapse did not appear different. Additional prospective studies should examine the independent effects of these variables in larger cohorts of CLL patients undergoing HCT. Disclosures: No relevant conflicts of interest to declare.

Abstract: While multiple myeloma (MM) remains incurable presently, expanded therapeutic options over the past decade have improved patient survival markedly. Proteasome inhibitors have redefined the treatment paradigm for myeloma, often serving as the backbone of front-line treatment. Histone deacetylase (HDAC) inhibitors (HDI), although only marginally active as single agent therapy in hematological malignancies, have demonstrated an ability to salvage bortezomib responsiveness in refractory patients, prompting heightened interest in this class of targeted therapeutics in myeloma. HDAC’s represent a family of enzymes, currently with 11 known members in the classical HDAC family, and subdivided into 4 sub-classes. HDAC11 is currently the only member of the sub-class IV and, as the newest member of the HDAC family, its impact on B cell lymphopoiesis and myeloma development is only starting to be unveiled. Intriguingly, we show that mice with germ-line silencing of HDAC11 (HDAC11KO mice) exhibit a 50% decrease in plasma cells in both the bone marrow and peripheral blood plasma cell compartments relative to wild-type mice. Consistent with this, Tg-HDAC11-eGFP mice, a transgenic strain engineered to express GFP under control of the HDAC11 promoter (Heinz, N Nat. Rev. Neuroscience 2001) reveals that HDAC11 expression is increased in the plasma cell population and to a lesser extent B1 B cells, as compared to earlier lineage stages. Similar observations based on measurements of HDAC11 mRNA were seen in normal human plasma cells. Significant increases in HDAC11 mRNA expression were observed in 7 of 11 primary human multiple myeloma samples and 11 of 12 human myeloma cell lines as compared to normal plasma cells, further emphasizing the potential relevance of HDAC11 to the underlying pathologic processes driving myeloma development and/or survival. Targeted silencing of HDAC11 in RPMI-8226 cells lines using siRNA results in a modest decrease in cell viability as measured by Annexin/PI staining and detection of activated caspase-3. Quisinostat, a second generation pan-HDI, has previously demonstrated activity against human myeloma cell lines in vitro (Stuhmer, Brit J Haematol, 2010), and suppressed bone destruction in an in vivo murine myeloma model (Deleu, Cancer Res, 2009). We similarly observe dose-dependent survival impairment in 10 human myeloma cell lines when cultured in the presence of quisinostat, with EC50’s consistently in the 1-10nM range. Importantly, quisinostat acts synergistically with proteasome inhibitiors (bortezomib and carfilzomib) in RPMI-8226 cells; more importantly, the degree of synergism is amplified in the RPMI-6226-B25 bortezomib-resistant cell line. Although a clear mechanism of action remains to be elucidated, preliminary data suggests that RPMI-8226 cells exposed to quisinostat appear to exhibit a decrease nuclear, but not cytosolic HDAC11. Collectively, these data illustrate a previously unknown role for HDAC11 in plasma cell differentiation and survival. Increased HDAC11 expression seen in myeloma patient specimens and primary myeloma cell lines highlights the potential of HDAC11 as a therapeutic target. Furthermore, we show that quisinostat, a pan-HDI with selectivity towards HDAC11 at lower dosing, acts synergistically with proteasome inhibitors in vitro in proteasome inhibitor sensitive and resistant cell lines. Future work will focus on further elucidating the role of HDAC11 in myeloma survival and drug response, with particular emphasis on proteasome inhibitors. Disclosures No relevant conflicts of interest to declare.

Abstract: Abstract 840 T-cells are an essential component of immune mediated tumor rejection. Adoptive transfer of T-cells results in a durable anti-tumor response in some patients with hematological malignancies. To further improve the efficacy of T-cell adoptive transfers, a better understanding of the regulatory checkpoints of these cells is needed. Here we show that HDAC11 is a negative regulator of CD8+ T-cell function, thus representing a potential target in adoptive immunotherapy. HDACs are a group of enzymes initially known for their role in deacetylating histones, thereby condensing chromatin structure and repressing gene expression. The known roles of HDACs as epigenetic regulators have recently expanded to include more complex regulatory functions including interactions with non-histone targets. HDAC11 is the most recently identified member of the HDAC family, and is highly expressed in brain, testis and T-cells. Recently, our group reported HDAC11 as a regulator of IL-10 production in antigen presenting cells. To determine the role of HDAC11 in T-cell biology, T-cells from HDAC11 knock out (HDAC11KO) mice were compared to wild-type T-cells in number, function and phenotype. HDAC11KO T-cells had no differences in absolute number or percentages of CD4+ or CD8+ lymphocytes. However CD8+ T-cells were hyper-proliferative upon CD3/CD28 stimulation and produced significantly higher levels of the pro-inflammatory, Tc1 cytokines IL-2, INF-γ, and TNF-α. However, no significant increases in the production of the Tc2 cytokines IL-4, IL-6 or IL-10 were seen. Further investigation of phenotypic differences also revealed that HDAC11KO mice have a larger percentage of central memory CD8+ T-cells. Additionally, HDAC11KO CD8+ T-cells express higher levels of the transcription factor Eomes, a known contributor to central memory cell formation as well as a controller of granzyme B and perforin production in CD8+ T-cells. This Tc1 and central memory-like phenotype translated to delayed tumor progression and survival in vivo in C1498 AML bearing mice treated with adoptively transferred HDAC11KO T-cells, as compared with wild type T-cells. Collectively, we have demonstrated HDAC11 as a negative regulator of CD8+ T-cell function, and a novel potential target to augment the efficacy of adoptive T-cell tumor immunotherapy. Disclosures: No relevant conflicts of interest to declare.

Abstract: Ponatinib is a potent oral pan–BCR-ABL tyrosine kinase inhibitor (TKI) that has demonstrated significant clinical activity in heavily pretreated CP-CML pts. A multivariate analysis of CP-CML pts in the PACE trial found significant associations between major cytogenetic response (MCyR) and higher dose intensity; however, dose reductions and/or interruptions (DR/I) of ponatinib occur often in pts who experience adverse events (AEs). The clinical significance of such DR/I are not well known. Objectives To assess the impact of DR/I and dose intensity of ponatinib on clinical outcomes in pts with CP-CML enrolled in the PACE trial. Methods A total of 270 CP-CML pts were enrolled in this ongoing, phase 2, international, open-label clinical trial. The efficacy population (N=267) was included in this post hoc analysis. Dose reductions were defined as any reduction below the standard 45 mg daily dose; interruptions were defined as a period in which ponatinib was held for ≥3 consecutive days between non-missing doses. Up to 2 reductions (to 30 or 15 mg/day) were permitted for managing AEs. To assess the impact of dose modification on response, pts were grouped according to tertiles of average dose intensity (mg/day), calculated as the cumulative dose divided by treatment exposure. All variables were calculated within 12 mos of the first dose to correspond to the primary outcome measure of MCyR by 12 mos. Secondary efficacy endpoints included complete cytogenetic response (CCyR) and major molecular response (MMR). Responses were assessed every 3 mos. The Cochran–Armitage trend test was used to assess whether response rates increased with higher average dose intensity tertiles; all P-values were 2-sided. Data are as of 01 Apr 2013, with a median follow-up of 20 (0.1–28) mos. Minimum follow-up for pts still on study was 18 mos. Results A total of 209 (78%) pts required DR/I at least once within 12 mos: 172 pts (64%) had at least 1 dose reduction (median time to first dose reduction was 64 days). In pts with 〉 1 dose reduction (n=75, 28%), the median time between the first and second reduction was 91 days. Among pts with a dose reduction at any time, 35% re-escalated to 45 mg daily. Dose interruption was experienced by 199 (75%) pts (median total duration of 35 days). The most common reason for DR/I was thrombocytopenia (33%). For pts with average dose intensity ≤27 mg/day (N=89), 〉 27 to ≤42 mg/day (N=88), and 〉 42 mg/day (N=90), respectively, the median age was 62, 62, and 56 yrs; median time since initial diagnosis was 11, 7, and 6 yrs; each group had received a median of 3 prior TKIs. Among these tertiles, the best response to the most recent dasatinib- or nilotinib-containing regimen was MCyR or better in 21%, 22%, and 35%; CCyR or better in 11%, 14%, and 23%; MMR or better in 1%, 2%, and 6%, respectively. Within 12 mos of the first dose, median duration of treatment exposure was 356 (26–366), 366 (51–366) and 366 (3–366) days, respectively. Twenty-nine pts had 〈 3 mos of treatment exposure. Of those, 11 had a post-baseline cytogenetic assessment (1 had MCyR). Responses to ponatinib according to tertile are presented below (Table) for pts with ≥3 mos follow-up (N=238). MCyR, CCyR, and MMR rates increased with higher average dose intensity. Comparable response rates were seen between pts with average dose intensity 〉 27 to ≤42 mg/day and 〉 42 mg/day. Response rates were lower in pts with average dose intensity ≤27 mg/day; however, these pts still achieved MCyR, CCyR, and MMR rates that substantially exceeded those reported with the most recent dasatinib- or nilotinib-containing regimen. Conclusions Higher dose intensity of ponatinib was associated with higher response rates in this heavily pretreated CP-CML population, but lower dose intensity still led to positive clinical outcomes. It should be noted that higher responses to the most recent dasatinib- or nilotinib-containing regimen were also seen in pts with higher average dose intensity. In summary, these data indicate that although optimal responses were seen with average ponatinib dose intensity 〉 42 mg/day, pts can be effectively managed with dose reduction or interruption if clinically indicated. Disclosures: Pinilla-Ibarz: Novartis, Ariad: Research Funding; Novartis, Ariad, BMS and Pfizer: Speakers Bureau. Cortes:Ariad, Pfizer, Teva: Consultancy; Ariad, BMS, Novartis, Pfizer, Teva: Research Funding. Kim:BMS, Novartis, IL-Yang: Consultancy; BMS, Novartis, Pfizer, ARIAD, IL-Yang: Research Funding; BMS, Novartis, Pfizer, IL-Yang: Honoraria; BMS, Novartis, Pfizer: Speakers Bureau; BMS, Pfizer: Membership on an entity’s Board of Directors or advisory committees. Le Coutre:Novartis: Research Funding; Novartis, BMS, Pfizer: Honoraria. Paquette:Ariad, BMS, Novartis: Consultancy; Ariad, BMS, Novartis: Honoraria; Ariad, BMS, Novartis: Speakers Bureau. Chuah:Novartis, BMS: Honoraria. Nicolini:Novartis, ARIAD, Teva: Consultancy; Novartis, BMS: Research Funding; Novartis, BMS, Teva, Pfizer, ARIAD: Honoraria; Novartis, BMS, TEva: Speakers Bureau; Novartis, ARIAD, Teva, Pfizer: Membership on an entity’s Board of Directors or advisory committees. Apperley:Novartis: Research Funding; Ariad, Bristol Myers Squibb, Novartis, Pfizer, Teva: Honoraria. DeAngelo:Araid, Novartis, BMS: Consultancy. Abruzzese:BMS, Novartis: Consultancy. Rea:BMS, Novartis, Pfizer, Ariad, Teva: Honoraria. Baccarani:ARIAD, Novartis, BMS: Consultancy; ARIAD, Novartis, BMS, Pfizer, Teva: Honoraria; ARIAD, Novartis, BMS, Pfizer, Teva: Speakers Bureau. Muller:Novartis, BMS, ARIAD: Consultancy; Novartis, BMS: Research Funding; Novartis, BMS, ARIAD: Honoraria. Gambacorti-Passerini:Pfizer: Research Funding; Pfizer, BMS: Honoraria. Lustgarten:ARIAD: employees of and own stock/stock options in ARIAD Pharmaceuticals, Inc Other, Employment. Rivera:ARIAD: employees of and own stock/stock options in ARIAD Pharmaceuticals, Inc. Other, Employment. Clackson:ARIAD: employees of and own stock/stock options in ARIAD Pharmaceuticals, Inc Other, Employment. Turner:ARIAD: Employment. Haluska:ARIAD: employees of and own stock/stock options in ARIAD Pharmaceuticals, Inc Other, Employment. Deininger:BMS, ARIAD, NOVARTIS: Consultancy; BMS, NOVARTIS, CELGENE, GILEAD: Research Funding; ARIAD, NOVARTIS: Advisory Boards, Advisory Boards Other. Hochhaus:Ariad, Novartis, BMS, MSD, Pfizer: Research Funding; Novartis, BMS, Pfizer: Honoraria. Hughes:Novartis, BMS, ARIAD: Honoraria, Research Funding. Goldman:ARIAD: Honoraria. Shah:Ariad, Bristol-Myers Squibb: Consultancy, Research Funding. Kantarjian:ARIAD, Novartis, BMS, Phizer: Research Funding. Talpaz:Ariad, BMS, Sanofi, INCYTE: Research Funding; Ariad, Novartis: Speakers Bureau; Ariad, Sanofi, Novartis: Membership on an entity’s Board of Directors or advisory committees.

Abstract: Abstract 3763 Background: Ponatinib is a potent pan-BCR-ABL inhibitor that is active against native and mutated forms of BCR-ABL, including the TKI resistant T315I mutant. The efficacy and safety of ponatinib (45 mg orally once daily) in patients with chronic phase (CP), accelerated phase (AP), or blast phase (BP) chronic myeloid leukemia (CML), or Ph+ acute lymphoblastic leukemia (Ph+ ALL) were evaluated in a pivotal phase 2, international, open-label clinical trial (PACE). Objective: To evaluate the patterns of molecular response in patients treated with ponatinib in the PACE trial. Methods: The PACE trial enrolled 449 patients resistant or intolerant (R/I) to dasatinib or nilotinib or with the T315I BCR-ABL mutation. Patients were assigned to 1 of 6 cohorts according to disease phase (CP-CML, AP-CML, or BP-CML/Ph+ ALL), R/I to dasatinib or nilotinib, and presence of T315I. Mutation and molecular response analyses were performed at a single central laboratory. Molecular responses are reported on the International Scale (BCR-ABLIS; b2a2/b3a2 [p210] transcript only): major molecular response (MMR), ≤0.1%; MR4, ≤0.01%; MR4.5, ≤0.0032%. The trial is ongoing. Data as of 23 July 2012 are reported, with a minimum follow-up of 9 months (median 12 [0.1 to 21] months). Results: The median age was 59 (18 to 94) years. The median time from initial diagnosis to start of ponatinib was 6 (0.3 to 28) years. 96% had received prior imatinib, 84% dasatinib, 65% nilotinib; 8% received 1, 39% received 2, and 53% received all 3 prior approved TKIs. In patients previously treated with dasatinib or nilotinib (N=427), 88% had a history of resistance and 12% were purely intolerant to dasatinib or nilotinib. BCR-ABL mutations were detected in 55% of all patients at baseline: 22.9% T315I only, 26.5% mutations other than T315I, 5.8% mutations in addition to T315I. The most common mutations were T315I (29%), F317L (8%), E255K (4%), F359V (4%), and G250E (3%). Baseline BCR-ABL transcript levels were 〉 10% in 74% (74% in CP-CML) and 〉 1% to ≤10% in 14% (20% in CP-CML). No patients entered the study in MMR. The best response to the most recent dasatinib or nilotinib containing regimen was MMR or better in 4% (3% in CP-CML). Molecular response rates by cohort are shown below for CP-CML and AP-CML. Deep molecular responses, including MR4.5, were observed in both disease phases. Of 16 BP-CML and 3 Ph+ ALL patients with the b2a2/b3a2 transcript and baseline and post-baseline bone marrow assessments, 5 BP-CML and 0 Ph+ ALL patients achieved MMR. In CP-CML, MMR rates for the most frequent mutations other than T315I were 41% F317L, 50% E255K, 31% F359V, 38% G250E. Subgroup analyses in CP-CML indicated significant differences in the MMR rate for patients with T315I only (56%; p 〈 0.001) and mutations in addition to T315I (50%; p=0.0216) vs. no mutation (21%), and for T315I only vs. mutations other than T315I (34%; p=0.0237). These differences are likely due to younger age (median 51 vs. 61 years) and exposure to fewer prior TKIs (median 2 vs. 3) in CP-CML patients with T315I vs. those without T315I. In CP-CML, 53% maintained or achieved BCR-ABLIS ≤10% by 3 months, with a trend towards higher rates in patients receiving fewer prior approved TKIs (1: 81%; 2: 61%; 3: 45%). The MMR rate (cumulative) in CP-CML improved over time: 13% by 3 months, 24% by 6 months, 28% by 9 months. The median time to MMR for CP-CML patients achieving MMR was 6 (2 to 17) months. CP-CML patients with MMR had an estimated probability of remaining in MMR at 6 months and 1 year of 87% and 84%, respectively (Kaplan-Meier). Ponatinib was generally well-tolerated. Data with a minimum follow-up of 12 months will be presented. Conclusions: Ponatinib treatment led to significant and deep molecular responses in this heavily pretreated population. In CP-CML, the MMR rate was ∼10-fold higher than that reported with the most recent dasatinib or nilotinib treatment. Disclosures: Hochhaus: ARIAD, Novartis, Bristol Myers-Squibb, Pfizer, and MSD: Membership on an entity's Board of Directors or advisory committees, Research Funding. Kim:Novartis, Bristol Myers-Squibb, Pfizer, ARIAD, and Il-Yang: Consultancy, Employment, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Pinilla-Ibarz:Novartis, BMS: Research Funding, Speakers Bureau. le Coutre:Novartis and BMS: Honoraria. Paquette:ARIAD: Consultancy. Chuah:Novartis and Bristol Myers-Squibb: Honoraria. Nicolini:Novartis, Bristol Myers Squibb, Pfizer, ARIAD: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Apperley:Novartis, Bristol Myers-Squibb, and ARIAD: Honoraria, Research Funding. Talpaz:Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; BMS: Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees, Research Funding; Millenium: Research Funding; Celgene: Research Funding; ARIAD: Research Funding; Deciphera: Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees. Abruzzese:BMS: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees. Rea:Bristol Myers-Squibb, Novartis, and Teva: Honoraria. Baccarani:ARIAD, NOVARTIS PHARMA, BRISTOL MYERS SQUIBB, PFIZER: Consultancy, Honoraria, Speakers Bureau. Muller:ARIAD: Consultancy. Wong:MolecularMD Corp: Employment, Equity Ownership. Lustgarten:ARIAD Pharmaceuticals, Inc.: Employment, Equity Ownership. Rivera:ARIAD Pharmaceuticals, Inc.: Employment, Equity Ownership. Clackson:ARIAD: Employment, Equity Ownership. Turner:ARIAD: Employment, Equity Ownership. Haluska:ARIAD: Employment, Equity Ownership. Guilhot:ARIAD: Honoraria. Hughes:Novartis, BMS, ARIAD: Honoraria, Research Funding. Goldman:Novartis, Bristol Myers-Squibb, and Amgen: Honoraria. Shah:ARIAD: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Novartis: Consultancy. Kantarjian:Novartis: Consultancy; Pfizer: Research Funding; ARIAD: Research Funding; BMS: Research Funding; Novartis: Research Funding. Cortes:Novartis, BMS, ARIAD, Pfizer, and Chemgenex: Consultancy, Research Funding.