Kaufmann, Scott H.

Abstract Introduction: The therapy of relapsed/refractory AML remains poor and new treatment options are sorely needed. The Leukemia Committee of ECOG devised a randomized phase II trial of 3 novel regimens to determine overall response rates could be improved. Methods: Patients between the ages of 18 and 70 years with a prior diagnosis of AML who relapsed <1 year after initial achievement of complete remission (CR) or who were refractory to initial induction therapy (<2 courses) or to first reinduction after a relapse (<1 course) were eligible for this trial. Eligible patients were randomized equally to one of three regimens described in Table 1 below. The primary endpoint was achievement of a morphologic complete remission (CR) or a CR with incomplete blood count recovery (CRi). A 2 stage statistical design was planned with 18 patients (assuming 16 eligible) to be accrued in the first stage. If at least 3 CR+CRi were seen among the 16 eligible patients, an additional 19 patients (assuming 17 eligible) would be entered in the second stage. If 8 or more CR+CRi were seen in the 33 eligible patients, this would be considered evidence of a promising response rate. Results: This study activated October 16, 2008 and closed August 2, 2013 with a total of 92 patients accrued (91 eligible). The eligibility criteria were amended during the study to lower the upper age limit from 70 to 65 years because 6 treatment related deaths occurred in the initial 27 patients accrued to the FLAM regimen. All of these patients died of septic shock or multi-organ failure and 5 of the 6 died within 1 month of the start of therapy. Most of these patients were over age 65 ( 53, 62, 67, 68, 69 and 69 years). At the first stage analysis, of the first 16 eligible patients accrued to each arm there were 2 CR and 2 CRi in the CT arm, 2 CR and 2 CRi in the FLAM arm and 2 CR in the sirolimus-MEC arm. Thus, the sirolimus-MEC arm was closed to accrual with a total of 20 eligible patients and second stage accrual continued for the CT and FLAM arms. Among the 35, 36 and 20 eligible patients accrued to arms A, B and C, respectively, there were 2 CR and 3 CRi in the CT arm, 6 CR and 4 CRi in the FLAM arm, and 2 CR and 1 CRi in the sirolimus-MEC arm. All 10 CR or CRi of the FLAM arm were among the first 33 eligible patients, therefore FLAM was the only regimen to achieve our target. The overall response rates for arms A, B and C were 14% (90% CI 7.1% - 34.7%), 28% (90% CI 16.1% - 43.0%) and 15% (90% CI 4.2% - 34.4%), respectively. Among the 92 patients accrued, there were 5, 10 and 2 grade 5 events within 30 days of the end of therapy in arms A, B and C, respectively. The vast majority of these deaths were attributed to infection and multi-organ failure except two which were secondary to disease. Conclusion: In this high risk group of AML patients the FLAM regimen showed an encouraging response rate. A randomized phase II trial has also recently been completed exploring its use in the front-line setting with newly diagnosed high-risk AML. While the FLAM regimen was excessively toxic in elderly patients with relapsed or refractory disease, it is a suitable option for younger patients in this setting. Arm A: CT Carboplatin 150 mg/m2/day over 24 hours by continuous infusion IV for 5 days (days 1-5) Topotecan 1.6 mg/m2/day over 24 hours by continuous infusion IV for 5 days (days 1-5) Arm B: FLAM Flavopiridol 30 mg/m2 for ½ hour followed by 60 mg/m2 over 4 hours IV per day for 3 days Cytarabine 667 mg/m2 over 24 hours by continuous infusion IV for 3 days (days 6-8) Mitoxantrone 40 mg/m2 IV over 1-2 hours (day 9) Arm C: Sirolimus-MEC Sirolimus 12 mg by mouth on day 1, followed by 4 mg by mouth daily for 8 days (days 2-9) Mitoxantrone 8 mg/m2/day over 15 minutes IV for 5 days (days 4-8) Etoposide 100 mg/m2/day over 1 hour IV for 5 days (days 4-8) Cytarabine 1000 mg/m2/day over 3 hours IV for 5 days (days 4-8) Table 2. Responses Table 1. Chemotherapy Regimens Regimen n Median age (range), years Responses (%) Deaths (first 30 days) (%) CT 35 55 (23-67) 5 (14%) 5 (14%) FLAM 36 62 (19-69) 10 (28%) 10 (28%) Sirolimus-MEC 20 52 (22-68) 3 (15%) 2 (10%) Disclosures Off Label Use: Sirolimus-potentiate chemotherapy Carboplatin-chemotherapy for AML treatment Topotecan-chemotherapy for AML treatment.

Abstract Abstract 3634 PARP is activated in response to DNA single strand (SS) breaks and is pivotal to the base excisional repair pathway for chemotherapy-damaged DNA. The orally bioavailable PARP inhibitor V delays DNA repair and potentiates the cytotoxicity of multiple classes of chemotherapy drugs including topoisomerase I inhibitors and platinating agents in leukemia cell lines. A previous clinical trial of T+C yielded promising results in adults with refractory acute leukemia at the maximum tolerated dose (MTD) of T 1.6 mg/m2/d + C 150 mg/m2/d by intravenous continuous infusion (IVCI) × 120 hrs. We are conducting a Phase I dose-escalation trial of V given orally twice daily Days 1–8 with T+C given by 120 hr IVCI Days 3–7. Deaths due to myocardial infarct and splenic infarcts with ascites occurred in one pt each when C 150 mg/m2 was added to V 10 mg BID and T 1.3 mg/m2, resulting in adjustment of the dose escalation schema for C and T.Dose levelVeliparib Oral Daily Days 1–8Topotecan Continuous IV Days 3–7Carboplatin Continuous IV Days 3–7Clinical Responses/#Pts1A10 mg p.o. BID1.0 mg/m2/day—1/3(CRi)1B10 mg p.o. BID1.3 mg/m2/day—1/3(CR)2A10 mg p.o. BID1.3 mg/m2/day150 mg/m2 IV0/42C10 mg p.o. BID1.0 mg/m2 IV120 mg/m2 IV2/6(CRi, PR)2D10 mg p.o BID1.2 mg/m2 IV120 mg/m2 IV1/6(PR)2E10 mg p.o. BID1.2 mg/m2 IV150 mg/m2 IV2/6(CR, CRi)3C20 mg p.o. BID1.0 mg/m2 IV120 mg/m2 IV0/63D20 mg p.o BID1.2 mg/m2 IV120 mg/m2 IV2/6(CR, PR)3E20 mg p.o. BID1.2 mg/m2 IV150 mg/m2 IV2/6(CR, CR)440 mg p.o. BID1.2 mg/m2 IV150 mg/m2 IV2/6(CR, CR)5§80 mg p.o. BID1.2 mg/m2 IV150 mg/m2 IV2/4(CR, CRi) Of 56 pts treated at doses of V 10–80 mg orally BID Days1–8 and T 1.0–1.2 mg/m2/d + C 120–150 mg/m2/d for 120 hrs IVCI Days 3–7, 25 (45%) have experienced > grade 3 non-hematologic toxicity, 6(11%) have had grade 4 toxicity and 1 (4%) died from sepsis with multi-organ failure. Overall response rate is 27% (8CR, 4 CRi, 3 PR), with 7 in relapsed AML, 7 in refractory AML and 1 refractory ALL. Pharmacokinetic (PK) studies in pt plasma, marrow supernatant, and marrow blasts demonstrated that V is detectable in marrow supernatant and cells, does not accumulate with multiple dose administrations in plasma but does in marrow, and is not altered by T or C administration. Nuclear staining for phosphorylated histone H2AX (γH2AX) demonstrated that V 10 mg BID increased DNA damage by > 2-fold in 11/22 (50%) on Day 1 and 13/18 (72 %) on Day 4 after 24 hrs of T+C. A PAR ELISA demonstrated that V suppresses PAR levels to <50% of pretreatment values in 30/41 (73%) blood and 3/4 patients at V 40mg per day dose level had greater than 80% suppression of PAR. The MTD has not been reached and dose escalation is ongoing. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 1531 Checkpoint kinase (Chk1) is a serine-threonine kinase that is activated via phosphorylation in response to DNA damage and is critical to the regulation of cell cycle progression. Ara-C triggers sequential activation of ATR kinase and Chk1 ex vivo to induce the S phase slowing that accompanies Ara-C treatment. Inhibition of Chk1 may abrogate S phase slowing, thereby preventing repair of Ara-C-induced DNA damage and potentiating the antitumor activity of Ara-C. The Hsp90 inhibitor tanespimycin (17-AAG) has been shown to enhance the cytotoxicity of Ara-C in part through Chk1 downregulation (RA Mesa et al., Blood 2005). While a phase I trial of Ara-C plus 17-AAG resulted in decreased blast cell Chk1 phosphorylation, drug toxicities precluded the clinical use of tanespimycin (SH Kaufmann et al., Haematologica, in press 2011). We have now tested the selective Chk1 inhibitor SCH 900776 (SCH 776) in combination with Ara-C in a Phase I dose-escalation trial in 24 adult patients with relapsed and refractory acute leukemias. AML (21) ALL (2) CML-BC (1) Male/Female 9/12 1/1 1/0 Age (range) 57 (23–73) 30, 63 36 Previous Ara-C 19 0 1 Relapsed 8 1 0 No. Prior CRs 2 (1–3) 2 0 Prior Stem Cell Transplant 6 0 0 Refractory 13 1 1 No. Prior Regimens 2 (1–4) 3 3 Prior Stem Cell Transplant 1 0 0 Secondary AML 8 0 0 MDS/MPD 6 0 0 Treatment-Related 2 0 0 Adverse Genetics 13 1 1 Single cytogenetics 3 1 0 Complex cytogenetics 11 0 1 Patients received Ara-C 667 mg/m2 on days 1–3 and 10–12 via 72-hour IV infusion along with SCH 776 on days 2, 3, 11, and 12 over 15 min IV starting at 10 mg/m2 (n=3) escalated to 20 mg/m2 (n=3), 40 mg/m2 (n=6), 56 mg/m2 (n=6), and flat dose of 140 mg (n=6) administered over 30 min IV. Maximal administered dose was achieved at 140 mg due to grade 3 prolonged QTc interval (n=1) and grade 3 hand-foot skin reaction (n=1). None of the expected mucosal or marrow toxicities of timed sequential Ara-C were exacerbated by SCH 776. Complete tumor clearance from day 14 bone marrow was detected in 13/24 (54%). CR plus CRi occurred in 7 patients (29%), with 6 of those CRs occurring at 40 mg/m2 or higher (response rate 40% for ≥ dose level 3). Examination of sequential samples of marrow blasts harvested pretreatment (day 0), 24 hours after initial Ara-C infusion prior to SCH 776 (day 2), and 2 hours after the second SCH 776 administration (day 3) demonstrated increased phosphorylation of Chk1 at Ser317 or Ser345 beginning at 40 mg/m2 consistent with the presence of unrepaired damage and further ATR activation. Pharmacokinetic (PK) data in each cohort showed lack of significant accumulation of SCH 776 in plasma. We plan for a randomized Phase II trial with 100 mg flat dose (or equivalent of 56 mg/m2) of SCH 900776 to assess any contributions to net Ara-C cytotoxicity and efficacy in relapsed and refractory acute leukemias. Disclosures: Gore: Celgene: Consultancy, Equity Ownership, Research Funding. Loechner:Merck Research Laboratories: Employment. Horowitz:Merck Research Labs: Employment, stock. Karp:Schering Merck: Research Funding.

Abstract Abstract 400FN2 Patients with primary and secondary myelofibrosis (MF) have a worse prognosis than those with essential thrombocytosis (ET) or polycythemia vera (PV). MF and chronic myelomonocytic leukemia (CMML) share chromosomal and genetic lesions such as ASXL1 and JAK2 mutations, but effective treatments are lacking. The aim of the study was to identify perturbed pathways in MPN and CMML that were amenable to targeted molecular therapies. In a large genomic analysis of MPN cases, we identified recurrent deletions of multiple DNA repair genes, including BRCA2 and Fanconi anemia (FANC) proteins, which have been associated with poly (ADP-ribose) polymerase (PARP) inhibitor sensitivity. Neoplastic cells from these patients are hypersensitive to PARP inhibitors in vitro and may provide a therapeutic strategy to improve treatment response for patients with advanced MPNs. Genomic screens utilizing SNP-A karyotyping were performed on primary cells from 144 patients with either ET, PV, or MF (90 were previously reported, McDevitt et al., ASH 2009). We focused on the potential functional relevance of recurrent genomic deletions, including BRCA2, by performing a GO analysis using the EASE software application (Hosack et al. 2003). This revealed statistically significant (p≤0.05) enrichment of 13 biological pathways. Notably, genes involved in cell cycle regulation and DNA repair (DR) were enriched in MF loss regions (p=0.007) as compared to regions lost in ET and PV. Several genes directly involved in DR including BRCA2, XRCC2, and several members of the FANC pathway, also critical for DNA crosslink repair, were deleted or present in copy neutral LOH regions. DNA methyltransferase inhibitors are being tested clinically in MF patients, and both cell cycle and DNA repair genes have been identified as epigenetically silenced in hematological malignancies. We therefore screened for BRCA1, BRCA2, FANC-C, L, and N promoter CpG island methylation in a series of 48 MPN samples. No methylation was identified in BRCA2 and FANC members, suggesting a potential haploinsufficiency mechanism. Methylation of BRCA1 was identified in a small number of cases and is under further study in a larger cohort to investigate the effects on BRCA1 expression, DR and sensitivity to PARP inhibitors. Based on these genetic and epigenetic findings, we then focused on a functional analysis of the homologous recombination (HR) pathway. Leukocytes from normal controls and MPN patients were exposed to ionizing radiation (IR) and stained for Rad51 foci, a marker of HR activity. Six of 15 MPN/CMML samples (40%), but none of the controls failed to form IR-induced Rad51 foci, suggesting impaired HR. Of the samples that demonstrated defects in HR [MPN-U, MF, CMML (2), MDS/MPN-U, and CML], there was no significant correlation between the MPN subtype, the JAK2 mutation status, or the specific metaphase karyotype, but the most sensitive case in our series exhibited a deletion of Ch13q that spanned the BRCA2 gene. We then examined sensitivity of MPN vs. normal myeloid progenitors to the PARP inhibitors veliparib and/or olaparib in methylcellulose (MC)-based colony forming assays. Failure to form Rad51 foci with irradiation was statistically correlated with PARP inhibitor sensitivity (p = 0.021). When an expanded series of cases were then analyzed for PARP inhibitor sensitivity using the MC colony assay, lower IC50s for both veliparib and olaparib were observed in MPN cases (N= 25, IC50S=2.69uM vs. 11.2uM, N=15 IC50S=.64uM vs. 1.27uM, respectively) relative to normal controls (p =.0003, p =.05, via Wilconon rank-sum test respectively). Two cases in this series were treated on a clinical trial combining the ABT-888 PARP inhibitor with salvage chemotherapy consisting of carboplatin and topotecan (Karp et al., ASH 2010). Both patients, one with sAML evolving out of PPV-MF, and a second with accelerating MPN-U, had failed four prior therapies but achieved complete clearance of bone marrow blasts at day 14. Interestingly, cells from this MPN-U patient (13q-) were the most sensitive to PARP inhibition in vitro. Additional accrual and correlative evaluations are underway and will be updated at the meeting. Collectively, these results demonstrate that a subset of MPN and CMML patients have intrinsic genetic and epigenetic defects affecting the FANC and HR pathways, providing pre-clinical rationale for the use of PARP inhibitors in the treatment of aggressive MPN and CMML. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 970 The mammalian target of rapamycin, mTOR, is a highly conserved serine/threonine kinase known to play a role in regulating mRNA translation, cell cycle progression, cell proliferation and apoptosis. As a downstream effector of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, mTOR is a component of two distinct complexes, TORC1 and TORC2. While TORC1 facilitates cell cycle progression from G1 into S phase by phosphorylating p70S6 kinase and eukaryotic initiation factor 4E binding protein 1 (4E-BP1), TORC2 catalyzes the activating phosphorylation of Akt on Ser473, providing a feedback loop for further activation of mTOR. Phase II trials have shown activity of the TORC1-selective inhibitor rapamycin and its analogs in a wide range of lymphoma subtypes. The purpose of this study was to evaluate the anti-proliferative and pro-apoptotic effects of the dual TORC1/TORC2 inhibitor OSI-027 in human neoplastic lymphoid cells in vitro. MTS assays demonstrated that OSI-027 inhibited proliferation in a wide range of lymphoid lines, including SeAx (Sezary syndrome), DoHH2 (large cell lymphoma), RL (follicular lymphoma) and Jurkat (T cell ALL), as well as clinical lymphoma and T cell ALL samples, with IC50 values ranging from 0.078 to 10 μM. Propidium iodide staining followed by flow cytometry for subdiploid cells revealed induction of apoptosis within 48 h of treatment with OSI-027 (but not rapamycin) in SeAx, DoHH2, and Jurkat cells. Examination of Jurkat variants with alterations in key proteins involved in the death receptor versus mitochondrial pathway revealed diminished apoptotic responses to OSI-027 when Bcl-2 was overexpressed or caspase 9 was silenced, indicating involvement of the mitochondrial pathway. Immunoblotting for Bcl-2 family members revealed upregulation of Bim and Puma after a 48-hour exposure to OSI-027 but not rapamycin. This upregulation was also seen at the mRNA level, with a 12- to 20-fold increase in Puma mRNA and 4- to 12-fold induction of Bim mRNA. Small interfering RNA (siRNA)-mediated knockdown of Bim and Puma significantly diminished the apoptotic response to OSI-027. Because the Foxo3a transcription factor has been implicated in Bim and Puma expression and is known to be activated when Akt is inhibited, we next examined whether Bim and Puma induction was Foxo3a-dependent. Luciferase reporter assays showed that OSI-027 activated the full-length Puma and Bim promoters and that this activation was diminished when the Foxo3a binding sites were deleted or mutated. In addition, OSI-027 induced nuclear translocation of Foxo3a, while Foxo3a siRNA diminished OSI-027-induced apoptosis in Jurkat cells. Collectively, these results indicate that OSI-027 inhibits proliferation and induces apoptosis in a wide range of neoplastic lymphoid cells through a process that involves Foxo3a-mediated upregulation of Bim and Puma. These results also suggest that dual inhibition of TORC1 and TORC2 may be an effective treatment strategy in lymphoid malignancy. Disclosures: Barr: OSI Pharmaceuticals: Employment. Witzig:Novartis and Celgene: Patents & Royalties, Research Funding, Served on advisory boards with Novartis and Celgene – both uncompensated with compensation to Mayo Clinic.

AbstractTopotecan (TPT) is a topoisomerase I (topo I) poison that has shown promising antineoplastic activity in solid tumors and acute leukemia. In the present study, a band depletion assay was used to evaluate the ability of TPT to stabilize topo I-DNA adducts in human leukemia cell lines and in clinical leukemia samples ex vivo. This assay showed that 50% of the cellular topo I in HL-60 human myelomonocytic leukemia cells became covalently bound to DNA at an extracellular TPT concentration of 4 μmol/L. In contrast, in 13 clinical specimens of human leukemia harvested before treatment of patients with TPT, the TPT concentration required to stabilize 50% of the cellular topo I in topo I-DNA complexes ranged from 3 to greater than 100 μmol/L (median, 30 μmol/L). Flow microfluorimetry showed that cellular TPT accumulation varied over only a twofold range and failed to provide evidence for transport-mediated resistance in the clinical samples. These observations raise the possibility that formation of topo I-DNA adducts is diminished in many specimens of refractory/relapsed acute leukemia by a mechanism that might alter topo I sensitivity to TPT.

Abstract Abstract 772 The mammalian target of rapamycin (mTOR) plays a crucial role in proliferative and anti-apoptotic signals in various lymphomas. The mTORC1 inhibitors, such as the rapamycin analogs temsirolimus and everolimus, have achieved a 38% overall response rate in heavily pretreated mantle cell lymphoma (MCL); however, most responses are partial, and many patients are resistant. One mechanism for resistance to mTORC1 inhibitors is that mTORC2 remains unaffected (Blood 2009 Oct 1; 114 (14): 2926–2935). We hypothesize that combined mTORC1/mTORC2 inhibition will be more effective in MCL. We tested the dual mTORC1/mTORC2 inhibitor OSI-027 (OSI Pharmaceuticals) on MCL cell lines and patient samples that were demonstrated to have a high amount of mTORC2 (Rictor and its target p-AKTS473) and mTORC1 (Raptor and its targets pS6 and p4E-BP1) targets. Dual inhibition through OSI-027 inhibited pAKTSer473, a direct target of mTORC2 without any obvious effect on AKTTh308 in these cells. On the other hand, the mTORC1 inhibitor rapamycin had no effect on pAKTSer473. Interestingly, both drugs effectively suppressed phosphorylation of S6; only OSI-027 decreased phosphorylation of 4E-BP1Th37/46. We next assessed the effect of OSI-027 on MCL proliferation and apoptosis and showed that dual inhibition of mTORC1/mTORC2 through OSI-027 caused growth arrest and apoptosis in a dose-dependent manner in both MCL patient cells treated ex vivo and established MCL cell lines; however, we did not see any significant effect of rapamycin on apoptosis. Recent studies have shown that AKT has direct effects on some members of the apoptosis pathway such as forkhead transcription factors (FOXO), caspase 9, and BAD. OSI-027 treatment significantly inhibited p-FOXO3AT32 with some inhibitory effect on pBADS136. These data suggest that FOXO3A is an important target of AKT through which OSI-027 is exerting its apoptotic effects. There are several potential mechanisms whereby OSI-027 prevents AKTS473 phosphorylation: 1) deactivation of kinases upstream of AKT; 2) activation of phosphatases down-stream of AKT. OSI-027 treatment of MCL cells produced no change in phosphorylated PI3K p85/p55 or PDK1 protein. These data eliminate the possibility of involvement of the PI3K pathway in OSI-027 mediated AKT dephosphorylation. We did find that OSI-027 mediated deactivation of AKTS473 is, at least in part, phosphatase dependent. The PH domain leucine-rich repeat protein phosphatase (PHLPP) family of phosphatases have been shown to directly dephosphorylate and inactivate AKTS473. To determine if AKT and PHLPP1 or PHLPP2 are associated in MCL cells, we immunoprecipitated AKT from MCL cells and probed for association with endogenous PHLPP1 and PHLPP2. All the MCL lines do express endogenous PHLPP1 and PHLPP2; however, they are not complexed to AKT. Further studies using the AKT isoforms AKT1 and AKT2 found AKT2 to be associated with PHLPP1; there was no association of AKT1 or AKT2 to PHLPP2. To investigate if OSI-027 enhanced the AKT2-PHLPP1 interaction, we repeated the co-immunoprecipitation assay in the OSI-027 treated MCL cell lines and demonstrated that OSI-027 increased the interaction of AKT2-PHLPP1. Further we showed that OSI-027 exposure did not alter the expression level of PHLPP1 and PHLPP2 proteins. In summary the dual inhibition of mTORC1/mTORC2 by OSI-027 in MCL is more effective than mTORC1 inhibition by rapamycin. We present the mechanism that OSI-027 induces apoptosis in MCL cell lines by inhibition of Rictor-AKT-FOXO3A signaling, and this is mediated in part by activation of the phosphatase PHLPP1. These findings indicate that simultaneously targeting mTORC1 and mTORC2 may be effective in patients with MCL. Disclosures: Barr: OSI Pharmaceuticals: Employment.

Abstract Background: Cytosine arabinoside (ara-C) remains the backbone of most regimens for the treatment of AML. The incorporation of ara-C into DNA activates checkpoint kinase 1 (Chk1) causing replication fork slowing, the accumulation of cells in S phase, and diminished ara-C cytotoxicity. The selective Chk1 inhibitor MK-8776 abrogates ara-C-induced S-phase arrest, thereby enhancing ara-C-induced apoptosis in vitro.1 A phase I trial demonstrated the safety of MK-8876 in combination with infusional ara-C and correlative studies demonstrated increased ara-C-induced H2Ax phosphorylation in AML cells, consistent with unrepaired DNA damage.2 Herein, we report data from a phase II trial comparing ara-C plus MK8876 with ara-C alone. Methods: Patients with relapsed or refractory AML were randomized to receive either ara-C 2 gm/m2 via continuous infusion (CIV) over 72 hours on days 1 and 10 with MK-8776 100 mg flat dose infused over 30 minutes on days 2, 3, 11, and 12 (combo); or ara-C alone on the same schedule (control). The primary end point was response rate (CR+CRi). Adverse events (AEs) for all patients were reported according to the NCI-CTCAE version 4.0. Correlative studies were performed on blood and marrow samples collected prior to treatment on days 1, 2 and 3 and at one hour after MK-8776 infusion (PB only). Marrow and circulating blasts were analyzed using semi-quantitative flow cytometric determinations of blast cell gH2AX expression, a marker of DNA damage. Results: Thirty-two patients were treated at 4 sites with 14 assigned to the combo arm and 18 to the control arm. The study was stopped early due to lack of drug availability. The median age was 60 (range 29-72) and 17 (53%) were male. Nine (28%) had refractory AML, 16 (50%) were in first relapse, and 7 (22%) were in second relapse. Eleven (34%) had undergone a prior bone marrow transplant (BMT). Risk stratification classified 1 (3%) patient as favorable risk disease, 11 (34%) as intermediate-1, 8 (25%) as intermediate-2, and 11 (34%) as adverse risk. There were 5 (36%) responses (CR/CRi) and 1 (7%) partial response (PR) in the combo arm, and 8 (44%) responses and 1 (6%) PR in the control arm. Median survival did not differ significantly between the two groups (5.9 months in the combo arm vs. 4.2 months in the control arm). Sub-group analysis showed increased responses in the combo arm for the patients in first relapse (50% vs. 40%) and those with a prior BMT (50% vs. 29%). Most non-hematologic grade 3/4 treatment-related AEs were similar in the two groups including febrile neutropenia (64% vs. 61%), infectious complications (50% vs. 50%), and sepsis (21% vs. 17%). Patients in the combo arm experienced more QT prolongation (21% vs. 0%), hypertension (21% vs. 0%), and GI toxicities (21% vs. 0%); while those in the control arm had increased renal (0% vs. 11%) and liver toxicity (7% vs. 17%). Two deaths were attributed to treatment, and both were in the control arm. Eight combo patients had serial PB samples and 2 had serial marrow samples studied by flow cytometry. There was an increase in the percentage of gH2AX+ CD34+ cells on Day 2 directly after MK-8876 infusion (16.9% prior and 36.4% at one hour), which occurred to a lesser extent on Day 3 after MK-8876 infusion (12.7% prior and 15.6% at one hour). Three of eight patients demonstrated a sustained increase in gH2AX+ CD34+ from Day 2 to Day 3, and all three achieved either a PR or CR. Conclusion: This is the first phase II study building on prior work suggesting that inhibition of Chk1 would increase response to ara-C in patients with relapsed or refractory AML. The study confirmed that treatment with the combination is feasible with similar toxicities between the treatment and control groups. The response rates were similar between the two groups with improved responses in patients receiving the combo in first relapse and following BMT. Correlative studies support the finding that responding patients show a sustained increase in DNA damage and this may serve as a biomarker in future studies. Chk1 remains a potential target to improve treatment outcomes with ara-C. References: 1. Schenk EL et al. Clin Cancer Res. 2012 Oct 1; 18(19):5364-73. 2. Karp JE et al. Clin Cancer Res. 2012 Dec 15; 18(24):6723-31. Disclosures No relevant conflicts of interest to declare.

Abstract Abstract 3276 PARP is activated in response to DNA single strand (SS) breaks and is pivotal to the base excisional repair pathway for chemotherapy-damaged DNA. The orally bioavailable PARP inhibitor V delays DNA repair and potentiates the cytotoxicity of multiple classes of chemotherapy drugs including topoisomerase I inhibitors and platinating agents in leukemia cell lines. A previous clinical trial of T+C yielded promising results in adults with refractory acute leukemia at the maximum tolerated dose (MTD) of T 1.6 mg/m2/d + C 150 mg/m2/d by intravenous continuous infusion (IVCI) × 120 hrs. We are conducting a Phase I dose-escalation trial of V given orally twice daily Days 1–8 with T+C given by 120 hr IVCI Days 3–7. Acute Myelogenous Leukemia (AML, 33 patients) Acute Lymphoblastic Leukemia (ALL, 4 patients) Median Age (range) 570 (40-75) 55 (42-62) Relapsed 8 No. Prior CRs 2 (1-3) Prior Stem Cell Transplant 2 0 Refractory 25 4 No. Primary Refractory 10 1 No. Prior Regimens 3 (1-4) 3 (1-4) Prior Stem Cell Tranpslant 3 1 Secondary AML 12 Anecedent Hematologic Dx 6 Treatment-Related 6 Cytogenetics Intermediate 10 2 Adverse: Single/Complex 7/16 2 Ph+ (1 with complex) BRCA-1/BRCA-2 1 (mutation)/ 1 (13q14 deletion) 0 Deaths due to myocardial infarct and splenic infarcts with ascites occurred in one pt each when C 150 mg/m2 was added to V 10 mg BID and T 1.3 mg/m2, resulting in adjustment of the dose escalation schema for C and T. Of 27 pts treated at doses of V 10–20 mg orally BID Days1-8 and T 1.0–1.2 mg/m2/d + C 120–150 mg/m2/d for 120 hrs IVCI Days 3–7, 8 (30%) have experienced > grade 3 non-hematologic toxicity, 4 (15%) have had grade 4 toxicity and 1 (4%) died from sepsis with multi-organ failure. Overall response rate is 22% (2 CR, 1 CRi, 3 PR), with 4 in relapsed AML, 1 each in refractory AML and refractory ALL. Pharmacokinetic (PK) studies in pt plasma, marrow supernatant, and marrow blasts demonstrated that V does not accumulate with multiple dose administrations in plasma but does in marrow, is not altered by T or C administration, and is detectable in marrow supernatant and cells. Nuclear staining for phosphorylated histone H2AX (γH2AX) demonstrated that V 10 mg BID increased DNA damage by > 2-fold in 11/22 (50%) on Day 1 and 13/18 (72 %) on Day 4 after 24 hrs of T+C. A PAR ELISA demonstrated that V suppresses PAR levels to <50% of pretreatment values in 21/29 (72%) blood and 9/13 (69%) marrow samples. The MTD has not been reached and dose escalation is ongoing. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 287 Purpose: Farnesyltransferases are key cellular enzymes involved in the prenylation of proteins. Prenylated proteins are important for malignant cell growth. The purpose of this phase II study (NCT00082888) was to assess tumor response and toxicity associated with the oral farnesyltransferase inhibitor tipifarnib (R115777, Johnson and Johnson) in patients with relapsed aggressive, indolent, or uncommon lymphoma. The uncommon group included low incidence lymphoma types such as Hodgkin lymphoma (HL) and T-cell non-Hodgkin lymphoma- cutaneous T-cell lymphoma (CTCL) and peripheral T-cell lymphoma (PTCL). Patients and Methods: Patients with relapsed or refractory non-Hodgkin (NHL) or Hodgkin (HL) were eligible if they had measurable disease, performance status ≤2, platelet count ≥75,000 × 10(6)/L, an absolute neutrophil count ≥1000 × 10(6)/L and a creatinine ≤2 × upper limit of normal. Patients initially received tipifarnib 300 mg twice daily, days 1–21 every 28 days. The trial was conducted by the University of Iowa/Mayo Clinic Lymphoma SPORE. Results: Ninety-three patients with lymphoma (42 aggressive, 15 indolent, and 36 uncommon) enrolled in the trial between March 2004 and November 2008. The median age was 62 years (range, 18–91). The median number of prior therapies was 5 (range, 1–17). The overall response rate (ORR) was 20% (19/93) with 6% (6/93) complete responses (CR) and 14% (13/93) partial responses (PR). The ORR was 17%, 7%, and 31% for the aggressive, indolent, and uncommon groups, respectively (Table). Among the 19 responders were 7 diffuse large B-cell lymphoma (DLBCL), 7 T-cell NHL, 1 follicular grade II, and 4 HL. The overall median response duration for the 19 responders was 7.5 months (mean, 12.8; range, 1.8 – 48.1 months). The median response duration was 11.3 months (mean, 10.1; range, 1.8 – 18.5 months), 2 months, and 7.5 months (mean, 15.5; range, 1.9 – 48.1 months) for the aggressive (7), indolent (1), and uncommon (11) groups, respectively. Five patients in the uncommon group are still receiving treatment with a range of treatment duration between 25 – 60+ months. Among the 5 still receiving treatment 4 responded and 1 is stable. The range of response duration is 15.4, 17.7, 43.1, and 48.1 months. These five patients (2 with HL, 1 anaplastic large cell cutaneous, and 2 PTCL) are currently receiving doses of 400 (1), 200 (2), and 100 (2) mg twice daily. The median time to progression for all patients was 3.6 months (95% CI: 2.1 – 4.5 months). The grade 3,4 toxicities observed were primarily fatigue (10%) and reversible myelosuppression with 11% anemia, 37% neutropenia, and 32% thrombocytopenia. Correlative studies demonstrated tipifarnib-induced upregulation of Bim, a marker of tipifarnib action in lymphoma cell lines, in 7 of 9 paired lymphoma biopsies. Conclusions: Tipifarnib has activity in lymphoma, with responses in patients with heavily pretreated DLBCL, HL and T-cell types, but little activity in follicular NHL. This agent has an excellent toxicity profile and targets a novel pathway. Further studies in combination with other agents are warranted for patients with DLBCL, HL or T-cell NHL. Disclosures: No relevant conflicts of interest to declare.