p-290 the new drug partner for combination therapy in ... · the new drug partner for combination...
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P-290 The New Drug Partner for Combination Therapy in Multiple Myleoma : Preclinical & Clinical Development of ACY-1215, a Selective Histone Deacetylase (HDAC) 6 Inhibitor
Jones S1, Raje N2, Hideshima T3, Lonial S4, Supko J2, Quayle S1, Yang M1, Tamang D1, Wheeler C1 & Anderson K3
Acetylon Pharmaceuticals, Boston, MA1, Massachusetts General Hospital, Boston, MA2, Dana-Farber Cancer Institute, Boston, MA3, Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA4
Abstract HDACs are a family of enzymes that regulate critical cellular functions. First generation, non-selective, HDAC inhibitors are active in multiple myeloma (MM) in combination with standard therapies, but are associated with severe fatigue, gastrointestinal effects and myelosuppression. ACY-1215 is the first selective HDAC6 inhibitor (HDAC6i)1 to reach clinical trials and data support the well-tolerated safety profile observed in preclinical studies. HDAC6 acts in the cytoplasm of MM cells regulating aggresome formation, an alternative route for degradation of excessive and misfolded proteins2. Knockdown of HDAC6 significantly increased the activity of the proteasome inhibitors (PIs) bortezomib and carfilzomib in MM cells. ACY-1215 recapitulates these results and shows synergistic cell killing with PIs providing the rationale for inhibiting the two major pathways of protein degradation in the clinic to treat multiple myeloma. The inhibition of HDAC6 correlates with preferential increase in acetylated tubulin. Further, ACY-1215 synergizes with immunomodulatory agents (IMiDs) in the presence or absence of dexamethasone to induce apoptosis in MM cells. The monotherapy experience with ACY-1215 in the clinic demonstrates few treatment related adverse events and is associated with stable disease up to 10 cycles of treatment3. Based on the preclinical and early clinical data ACY-1215 has entered phase 1b combination trials with bortezomib (ACY-100) and with lenalidomide (ACE-MM-101) in patients with relapsed (R) and relapsed/refractory (RR) MM.
misfolded waste protein
protein aggregates (toxic)
26S proteasome
aggresome
Zolinza®, panobinostat,
ACY-1215
microtubule autophagy
Velcade®, Kyprolis®, NPI0052, MLN9708, ONX 0912
lysosome
HDAC6
HDAC6
HDAC6
ubiquitin Ub Ub
Ub Ub Ub Ub
Ub Ub
dynein Ub Ub
dynein Ub Ub
Ub Ub Ub
Ub
Figure 1. HDAC6 regulates acetylation of cytoplasmic α-tubulin, an essential component of aggresome formation an alternative pathway to proteasome degradation to remove unfolded or misfolded proteins for example in MM cells2.
Background The rate of relapse in MM remains high despite novel therapies such as the proteasome inhibitor bortezomib. Non-selective HDAC inhibitors (panobinostat4 and vorinostat5) in combination with bortezomib have demonstrated activity in patients with relapsed and refractory MM. However, these inhibitors target multiple HDACs, including HDAC6 and Class 1 enzymes, and are associated with significant adverse effects such as severe fatigue, nausea, vomiting, diarrhea, and myelosuppression.
HDAC6 Genetic Knockdown Potentiates PI Activity
HDAC6
Ac-tub.
Ac-H3K9
GAPDH
shRNA C HDAC6
Figure 2. RPMI8226 cells were infected with scrambled or HDAC6 pLKO.1 lentiviral shRNA constructs. After puromycin selection, cells were incubated (48h) with indicated concentrations of of carfilzomib (left). Cell growth was assessed by MTT assay. p-value was calculated by student-t test and p<0.05*. Cell lysates were analyzed by western (right) blot with the indicated antibodies. Similar results were observed with bortezomib (data not shown).
HDAC6i Synergize with PI in MM Cell Killing
HDAC6i Synergize with IMiDs in MM Cell Killing
Grade 3 & 4 Possibly Related AEs for ACY-1215 -/+ Bortezomib
Figure 3. RPMI8226 cells were incubated (48h) with ACY-1215 (0.5-8.5 µM) (left) or ACY-775 (0.5-30 µM) (right) with carfilzomib (2-7 nM). Cell viability was assessed by MTS reduction assay. The fraction affected (Fa: x-axis) for each combination was calculated and the combination index (CI: y-axis) generated by CalcuSyn software. CI<1 (red circle) indicates synergistic MM cell killing. Similar results were observed with bortezomib or using MM.1S cells (data not shown).
HDAC6i + PI Increases Apoptosis & Suppresses Tumor Growth
Figure 4. RPMI8226 cells (left) were cultured (16h) with DMSO (C), ACY-1215 (2 µM) +/- carfilzomib (10 nM). Whole cell lysates were subjected to immunoblotting using indicated antibodies. RPMI8226 cells (1x107) were implanted SQ in CB-17 SCID mice (n=10/group). ACY-1215 (IP; days 1-5, 50 mg/kg) and carfilzomib (IV; 1 or 2 mg/kg, days 1 & 2) were dosed for 4 weeks. Significant differences in tumor volume were assessed by 2-Way ANOVA with a Tukey post-hoc test (right).
HDAC6i with IMiDS Increase Apoptosis & Decrease c-Myc
Figure 5. MM.1S cells were cultured (48h) with lenalidomide (left) or pomalidomide (right) +/- ACY-1215. Cell growth was assessed by MTT assay. Combination index (CI) was calculated using CompuSyn software.
Ac-a-tub.
GAPDH
1215(-)
1215(0.5)
1215(1)
1215(2)
- + - + - + - +
Myc
Casp.-3
PARP
Len (1)Dex(50) Myc
PARP
Casp-3
1215 (3) - + - + - + - +
C Len(1)
Dex(50)
LenDex
GAPDH
XIAP
Figure 6. MM.1S cells were cultured (48h) with lenalidomide (1 µM) and ACY-1215 (Left: 0.5, 1 or 2 µM; right: 3 µM) +/- dexamethasone (50 nM). Whole cell lysates were subjected to immunoblotting using indicated antibodies.
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0 nM 2.5 nM 5 nM 7.5 nM 10 nM 12.5 nM
% c
ontr
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Carfilzomib
RPMI8226 + Carfilzomib
comtrol-sh HDAC6-sh
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C 1215 C 1215CFZ (-) CFZ (+)
Cleaved-caspase-8
Caspase-9
Caspase-3
Caspase-7
PARP
GAPDH
AIF
Caspase-10
CF
CF
CF
CF
References
0 10 20 300
300
600
900 VehicleCarfilzomib (1 mg/kg)
ACY-1215 (50 mg/kg) +Carfilzomib (1 mg/kg)
ACY-1215 (50 mg/kg)Carfilzomib (2 mg/kg)
***
* p<0.0001 vs. Vehicle
Days
Tumo
r Volu
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m3 )
Study Design ACY-100 (ACY-1215 + Bortezomib + Dex)
Figure 7. Phase 1a patients received single oral daily doses of ACY-1215 for 5 consecutive days every 2 weeks of a 3 week cycle. Phase 1b patients receive ACY-1215 as above with IV bortezomib week 1: days 1 & 4 and week 2: days 8 & 11 and with dexamethasone week 1: days 1,2, 4 & 5 and week 2: days 8, 9, 11 & 12. Twice daily dosing of ACY-1215 will be examined if ≥ 160 mg of ACY-1215 QD is tolerated in combination with bortezomib. Patients have R or RR MM and prior IMiD and PI. Patient Demographics, Disease Characteristics & Responses
Table 1. As of January 22 2013, 23 patients have been enrolled and one patient is in screening. Two patients remain on study (9+ & 1+ cycles) and all other patients have withdrawn due to disease progression (18), patient request (2) or investigator discretion (1).
Treatment Emergent AEs (≥15%) for ACY-1215 Monotherapy
Pharmacokinetics & Pharmacodynamics of ACY-1215 Figure 8. Mean plasma ACY-1215 concentration-time profiles on day 1 and in the presence of bortezomib (insert). ACY-1215 was rapidly absorbed Tmax ~1h with t1/2 ~3h and no evidence of accumulation (day 1 vs day 11, data not shown). Dose proportional increase in Cmax and exposure was observed with doses of 40-160mg and exposures at ≥160 mg were similar. Pharmacokinetics of ACY-1215 were similar in combination with bortezomib.
Figure 9. Mean fold change in acetylated tubulin (Ac-tub) and histone levels in blood cells compared to pre-dose levels at 1 and 4h post-dose on day 1. Ac-tub levels increased with increasing dose of ACY-1215. Maximal levels of Ac-tub correlated with Tmax. All patients in Cohorts 3 – 4 had detectable increase in Ac-tub. In Cohorts 1 and 2 there were one and two patients, respectively, with a detectable increase in Ac-tub correlating with higher plasma levels of drug. Minimal increase in Ac-histones was observed except in two patients in Cohort 4.
Study Design ACE-MM-101 (ACY-1215 + Lenalidomide + Dex)
Figure 10. Patients receive single oral daily dose of ACY-1215 for 5 consecutive days every 2 weeks and lenalidomide daily for 3 weeks with dexamethasone on days 1, 8, 15 & 22 of a 4 week cycle. If this schedule is tolerated ACY-1215 dosing will be increased to 5 consecutive days every 3 weeks to overlap with the dosing schedule for lenalidomide. Twice daily dosing of ACY-1215 will be explored if the 3 week dose schedule is tolerated. Patients have R or RR MM and at least one prior Rx
Acknowledgements
Grade 3 & 4 Possibly Related AEs for ACY-1215 + Lenalidomide
Summary
Patient Demographics, Disease Characteristics & Responses
1. Santo L, et al. Blood 2012; 119: 2579 2. Hideshima T. et al. Clin Cancer Res. 2005;11: 8530 & Hideshima T, et al. Proc Natl Acad Sci 2005; 102: 8567 3. Raje N, et al. Blood (ASH Annual Meeting), Nov 2012; 120: 4061 4. Catley L, et al. Blood 2006; 108: 3441 & Richardson P, et al, Blood (ASH Annual Meeting) Nov 2012; 120: 1852 5. Siegel S, et al. Blood (ASH Annual Meeting), Nov 2011; 118: 480
Nonclinical Results
• HDAC6 genetic knockdown potentiates PI activity in MM cells • HDAC6i synergize with PIs in MM cell killing in vitro and in vivo suppressing tumor growth • ACY-1215 potentiate IMiDs (+/- dexamethasone) cell killing and decrease c-Myc levels
Clinical Results
• ACY-1215 is well tolerated as monotherapy & experience in combination is encouraging • Doses of ≥ 160 mg achieved Cmax > 1 µM (therapeutic levels in MM animal models) • All patients at doses of ≥ 160 mg showed preferential increase in Ac-tub vs Ac-histones • Stable disease has been achieved in 6/15 heavily pretreated patients in monotherapy • Disease responses observed in combination therapy with bortezomib or lenalidomide • Dose escalation is ongoing - combination trials are planned with carfilzomib or pomalidomide
HDAC6i in combination with PIs or IMiDs may provide a well-tolerated treatment for MM
Table 5. As of January 22 2013, 5 out of 6 patients in the first two cohorts have at least a PR.
ACY-100 Clinical InvestigatorsSagar Lonial, MD (Lead Investigator) Winship Cancer Institute
Noopur Raje, MD Massachusetts General Hospital
Robert Orlowski, PhD, MD MD Anderson Cancer Center
Hari Parameswaran, MD Medical College of Wisconsin
Sundar Jagannath, MD Mt. Sinai School of Medicine
Dan Vogl, MD Abramson Cancer Center
ACE-MM-101 Clinical InvestigatorsNoopur Raje, MD & Anuj Mahindra, MD Massachusetts General Hospital
William Bensinger, MD Fred Hutchinson Cancer Research Center
Peter Voorhees, MD University of North Carolina
Jesus Berdeja, MD Sarah Cannon Research Institute