Inhibitors targeting BCL-2 apoptotic proteins have significant potential for the treatment of acu... more Inhibitors targeting BCL-2 apoptotic proteins have significant potential for the treatment of acute myeloid leukemia (AML); however, complete responses are observed in only 20% of patients, suggesting that targeting BCL-2 alone is insufficient to yield durable responses. Here, we assessed the efficacy of coadministration of the PI3K/mTOR inhibitor GDC-0980 or the p110β-sparing PI3K inhibitor taselisib with the selective BCL-2 antagonist venetoclax in AML cells. Tetracycline-inducible downregulation of BCL-2 significantly sensitized MV4-11 and MOLM-13 AML cells to PI3K inhibition. Venetoclax/GDC-0980 coadministration induced rapid and pronounced BAX mitochondrial translocation, cytochrome c release, and apoptosis in various AML cell lines in association with AKT/mTOR inactivation and MCL-1 downregulation; ectopic expression of MCL-1 significantly protected cells from this regimen. Combined treatment was also effective against primary AML blasts from 17 patients, including those beari...
The caspase inhibitor BOC-D-fmk does not change HHT/Bort–mediated down-regulation of MCL-1. OCI-L... more The caspase inhibitor BOC-D-fmk does not change HHT/Bort–mediated down-regulation of MCL-1. OCI-LY18 and Carnaval cells were treated with HHT + Bort for 24 h either in the absence or presence of 5 μmol/L BOC-D-fmk. At the end of this period, cells were lysed and subjected to Western blot analysis using the indicated primary antibodies. Each lane was loaded with 25 μg of protein. Blots were stripped and reprobed with antitubulin antibodies to ensure equal loading and transfer of protein. Representative of two separate experiments. (PPTX 100 kb)
HHT inhibits MCL-1 expression through a post-transcriptional mechanism. A. SU-DHL4 and SU-DHL16 c... more HHT inhibits MCL-1 expression through a post-transcriptional mechanism. A. SU-DHL4 and SU-DHL16 cells were treated with HHT for 8 h after which cells were lysed and proteins extracted. Expression of the indicated proteins was determined by Western blotting using the indicated antibodies. B. SU-DHL4 and SU-DHL16 cells were treated with HHT for 8 h after which cells were extracted for mRNA. Relative levels of MCL-1 mRNA/GAPDH were calculated. C. SU-DHL4 and SU-DHL16 cells were pre-treated with actinomycin (2.5 μg/ml) for 30 min and then exposed to HHT 2 h (SU-DHL4 60 nM, SU-DHL16 20 nM) after which cells were lysed and proteins extracted. Expression of the indicated proteins was determined by western blott using the indicated antibodies. D. SU-DHL4 and SU-DHL16 cells were pre-treated with cyclohexamide (5 μg/ml) for 30 min and then exposed to HHT 2 h and 4 h (SU-DHL4 60 nM, SU-DHL16 20 nM) after which cells were lysed and proteins extracted. Expression of the indicated proteins was de...
A regimen combining the Wee1 inhibitor AZD1775 with HDAC inhibitors targets human acute myeloid l... more A regimen combining the Wee1 inhibitor AZD1775 with HDAC inhibitors targets human acute myeloid leukemia cells harboring
Interactions between the inhibitor of apoptosis protein antagonist LCL161 and the histone deacety... more Interactions between the inhibitor of apoptosis protein antagonist LCL161 and the histone deacetylase inhibitor panobinostat (LBH589) were examined in human multiple myeloma (MM) cells. LCL161 and panobinostat interacted synergistically to induce apoptosis in diverse MM cell lines, including those resistant to bortezomib (PS-R). Similar interactions were observed with other histone deacetylase inhibitors (MS-275) or inhibitors of apoptosis protein antagonists (birinapant). These events were associated with downregulation of the noncanonical (but not the canonical) NF-κB pathway and activation of the extrinsic, caspase-8–related apoptotic cascade. Coexposure of MM cells to LCL161/LBH589 induced TRAF3 upregulation and led to TRAF2 and NIK downregulation, diminished expression of BCL-XL, and induction of γH2A.X. Ectopic expression of TRAF2, NIK, or BCL-XL, or short hairpin RNA TRAF3 knock-down, significantly reduced LCL161/LBH589 lethality, as did ectopic expression of dominant-negativ...
2926 Patients with multiple myeloma (MM) undergoing high dose therapy and autologous stem cell tr... more 2926 Patients with multiple myeloma (MM) undergoing high dose therapy and autologous stem cell transplantation (SCT) remain at risk for disease progression. Maintenance therapy may delay recurrence but is associated with toxicities, highlighting the need for alternative strategies for long-term disease control. Malignant plasma cells in MM patients occasionally express highly immunogenic cancer testis antigens (CTA). CTA expression is regulated by DNA methylation, and may be increased by 5-azacitidine (Celgene corp., Summit, NJ) (Aza), a DNA hypomethylating agent. The addition of lenalidomide (Celgene corp.) (L) may augment any ensuing adoptive CTA-specific immune response. These immune effector cells may then be collected and adoptively transferred in a setting of lympho-depletion and minimal residual disease following SCT, serving a maintenance function. To demonstrate the feasibility of this approach, we initiated a phase II clinical trial of Aza administered sequentially with L ...
Histone deacetylase (e.g., vorinostat) and proteasome inhibitors interact synergistically to prom... more Histone deacetylase (e.g., vorinostat) and proteasome inhibitors interact synergistically to promote cell death in various cancer cells, including malignant human hematopoietic cells. The novel, proteasome inhibitor carfilzomib is an attractive candidate for combination regimens due to diminished neurotoxicity, irreversible proteasome inhibition, favorable tolerability profile relative to bortezomib, and preclinical and clinical evidence of activity in bortezomib-resistant cells and patients. Preclinical studies demonstrated synergistic interactions between carfilzomib and vorinostat in human diffuse large B-cell lymphoma (DLBCL) and mantle cell lymphoma (MCL) cells both in vitro and in vivo as well as in bortezomib-resistant cells (Dasmahapatra et al., Blood 115:4478; 2010; Mol Cancer Ther 10:1686, 2001). These preclinical findings prompted a phase 1 trial, using a 3+3 design, with the goal of determining the maximum tolerated dose (MTD)/recommended phase 2 dose (RP2D) for the comb...
3588 Although reports of synergistic interactions between proteasome and histone deacetylase (HDA... more 3588 Although reports of synergistic interactions between proteasome and histone deacetylase (HDAC) inhibitors in acute leukemias have been limited, they are well described in B-cell malignancies (e.g., myeloma and lymphoma). Nevertheless, preclinical findings have shown striking synergism between the HDAC inhibitor belinostat (previously PXD-101) and the proteasome inhibitor bortezomib, administered at low (sub-micromolar) concentrations, in cultured and primary acute myeloid leukemia (AML) and acute lymphocytic leukemia cells (Dai Y et al. Br J Haematol. 2011). These findings prompted initiation of a phase I trial, using a 3+3 design, with the primary objective of determining the maximum tolerated dose (MTD) for the combination of bortezomib and belinostat in patients with relapsed or refractory acute leukemia, myelodysplastic syndrome (MDS), or chronic myelogenous leukemia in blast crisis (CML-BC). To date, 25 patients with the following disease types have been treated: acute leu...
Inhibitor of apoptosis proteins (IAPs e.g., XIAP, cIAP1, cIAP2) inhibit apoptosis through diverse... more Inhibitor of apoptosis proteins (IAPs e.g., XIAP, cIAP1, cIAP2) inhibit apoptosis through diverse mechanisms. Recent attention has focused on novel functions of cIAP1/2, including activation of the canonical and non-canonical NF-κB pathways and inhibition of the extrinsic apoptotic pathway through the ripoptosome. These considerations have stimulated the development of IAP antagonists such as the bivalent IAP inhibitor birinapant (TL-32711). Proteasome inhibitors such as bortezomib are highly active in multiple myeloma (MM), and act, at least in part, by inhibiting NF-κB, raising the possibility of cooperative interactions with IAP antagonists. Synergistic induction of apoptosis was observed in human MM cell lines, including drug-naïve U266 cells, their bortezomib-resistant counterparts (PS-R), and multiple other human MM lines. Birinapant (500 nM) sharply down-regulated cIAP1/2 in MM cells, accompanied by reduced expression of TRAF2, RIP1, and p-IKKβ, as well as robust TRAF3 up-reg...
2931 AZD6244 is a potent, selective, oral, non-ATP competitive small molecule inhibitor of the mi... more 2931 AZD6244 is a potent, selective, oral, non-ATP competitive small molecule inhibitor of the mitogen-activated protein kinase, MEK 1/2 that has shown significant pre-clinical activity in multiple myeloma (MM) cells, both in vitro and in vivo, as well as a favorable clinical profile. The present phase II study was designed to determine the response rate for AZD6244 in patients with relapsed or refractory MM. The study utilized a two-stage Simon design to allow for early termination if there was strong evidence of regimen inactivity. Eligible patients were restricted to those with MM who have had at least 2 prior regimens. AZD6244 capsules (75 mg) were administered orally twice daily continuously for 28 day cycles. Response was evaluated after 3 cycles. To date, 37 patients have been enrolled (13 in the 1st stage and 24 in the 2nd stage). One subject enrolled in the 1st stage was not treated. Gender enrollment was balanced (male/female 18/19). The median age of treated patients was ...
We report the results of a phase 1 dose-escalation study of belinostat and bortezomib in adult pa... more We report the results of a phase 1 dose-escalation study of belinostat and bortezomib in adult patients with acute leukemia or MDS or CML with blast crisis. Thirty-eight patients received IV belinostat days 1-5 and 8-12 with IV bortezomib days 1, 4, 8, and 11 every 21 days. QTc prolongation was the only identified DLT. The RP2Ds were 1.3 mg/m2 bortezomib and 1000 mg/m2 belinostat. One patient with highly refractory MLL-ENL rearranged biphenotypic AML with multiple karyotypic aberrations had a complete pathologic and karyotypic response. One patient with post-MPN AML remained on study with stable disease (SD) for 32 cycles. Whole-exome sequencing revealed no aberrations in the first patient and a hyper-mutator genotype in the second. Eighteen patients had a best response of SD. We conclude that this treatment strategy is feasible but has limited activity in this population. Nevertheless, the factors that predict exceptional responses to this strategy warrant further investigation.
1794 Preclinical studies have demonstrated synergistic interactions between proteasome and histon... more 1794 Preclinical studies have demonstrated synergistic interactions between proteasome and histone deacetylase (HDAC) inhibitors in diverse hematologic malignancies, including those of B-cell origin. We have previously shown that the combination of the proteasome inhibitor bortezomib and the HDAC inhibitor romidepsin, administered at extremely low concentrations (∼3–5 nM), results in a striking increase in apoptosis in primary CLL cells, including cells from patients with CLL refractory to standard treatments (Dai Y et al. Clin Cancer Res. 2008). These findings prompted initiation of a phase I trial, using a 3+3 design, with the primary objective of determining the maximum tolerated dose (MTD) for the combination of bortezomib and romidepsin in patients with relapsed or refractory chronic lymphocyte leukemia/small lymphocytic lymphoma (CLL/SLL), indolent B-cell lymphoma, or peripheral T-cell lymphoma (PTCL). To date, 11 patients have been enrolled and treated. All of the patients tr...
Purpose: This phase I study was conducted to identify the maximum-tolerated dose (MTD) of alvocid... more Purpose: This phase I study was conducted to identify the maximum-tolerated dose (MTD) of alvocidib when combined with vorinostat in patients with relapsed, refractory, or poor prognosis acute leukemia, or refractory anemia with excess blasts-2. Secondary objectives included investigating the pharmacokinetic and pharmacodynamic effects of the combination. Experimental Design: Patients received vorinostat (200 mg orally, three times a day, for 14 days) on a 21-day cycle, combined with 2 different alvocidib administration schedules: a 1-hour intravenous infusion, daily × 5; or a 30-minute loading infusion followed by a 4-hour maintenance infusion, weekly × 2. The alvocidib dose was escalated using a standard 3+3 design. Results: Twenty-eight patients were enrolled and treated. The alvocidib MTD was 20 mg/m2 (30-minute loading infusion) followed by 20 mg/m2 (4-hour maintenance infusion) on days one and eight, in combination with vorinostat. The most frequently encountered toxicities we...
Inhibitors targeting BCL-2 apoptotic proteins have significant potential for the treatment of acu... more Inhibitors targeting BCL-2 apoptotic proteins have significant potential for the treatment of acute myeloid leukemia (AML); however, complete responses are observed in only 20% of patients, suggesting that targeting BCL-2 alone is insufficient to yield durable responses. Here, we assessed the efficacy of coadministration of the PI3K/mTOR inhibitor GDC-0980 or the p110β-sparing PI3K inhibitor taselisib with the selective BCL-2 antagonist venetoclax in AML cells. Tetracycline-inducible downregulation of BCL-2 significantly sensitized MV4-11 and MOLM-13 AML cells to PI3K inhibition. Venetoclax/GDC-0980 coadministration induced rapid and pronounced BAX mitochondrial translocation, cytochrome c release, and apoptosis in various AML cell lines in association with AKT/mTOR inactivation and MCL-1 downregulation; ectopic expression of MCL-1 significantly protected cells from this regimen. Combined treatment was also effective against primary AML blasts from 17 patients, including those beari...
The caspase inhibitor BOC-D-fmk does not change HHT/Bort–mediated down-regulation of MCL-1. OCI-L... more The caspase inhibitor BOC-D-fmk does not change HHT/Bort–mediated down-regulation of MCL-1. OCI-LY18 and Carnaval cells were treated with HHT + Bort for 24 h either in the absence or presence of 5 μmol/L BOC-D-fmk. At the end of this period, cells were lysed and subjected to Western blot analysis using the indicated primary antibodies. Each lane was loaded with 25 μg of protein. Blots were stripped and reprobed with antitubulin antibodies to ensure equal loading and transfer of protein. Representative of two separate experiments. (PPTX 100 kb)
HHT inhibits MCL-1 expression through a post-transcriptional mechanism. A. SU-DHL4 and SU-DHL16 c... more HHT inhibits MCL-1 expression through a post-transcriptional mechanism. A. SU-DHL4 and SU-DHL16 cells were treated with HHT for 8 h after which cells were lysed and proteins extracted. Expression of the indicated proteins was determined by Western blotting using the indicated antibodies. B. SU-DHL4 and SU-DHL16 cells were treated with HHT for 8 h after which cells were extracted for mRNA. Relative levels of MCL-1 mRNA/GAPDH were calculated. C. SU-DHL4 and SU-DHL16 cells were pre-treated with actinomycin (2.5 μg/ml) for 30 min and then exposed to HHT 2 h (SU-DHL4 60 nM, SU-DHL16 20 nM) after which cells were lysed and proteins extracted. Expression of the indicated proteins was determined by western blott using the indicated antibodies. D. SU-DHL4 and SU-DHL16 cells were pre-treated with cyclohexamide (5 μg/ml) for 30 min and then exposed to HHT 2 h and 4 h (SU-DHL4 60 nM, SU-DHL16 20 nM) after which cells were lysed and proteins extracted. Expression of the indicated proteins was de...
A regimen combining the Wee1 inhibitor AZD1775 with HDAC inhibitors targets human acute myeloid l... more A regimen combining the Wee1 inhibitor AZD1775 with HDAC inhibitors targets human acute myeloid leukemia cells harboring
Interactions between the inhibitor of apoptosis protein antagonist LCL161 and the histone deacety... more Interactions between the inhibitor of apoptosis protein antagonist LCL161 and the histone deacetylase inhibitor panobinostat (LBH589) were examined in human multiple myeloma (MM) cells. LCL161 and panobinostat interacted synergistically to induce apoptosis in diverse MM cell lines, including those resistant to bortezomib (PS-R). Similar interactions were observed with other histone deacetylase inhibitors (MS-275) or inhibitors of apoptosis protein antagonists (birinapant). These events were associated with downregulation of the noncanonical (but not the canonical) NF-κB pathway and activation of the extrinsic, caspase-8–related apoptotic cascade. Coexposure of MM cells to LCL161/LBH589 induced TRAF3 upregulation and led to TRAF2 and NIK downregulation, diminished expression of BCL-XL, and induction of γH2A.X. Ectopic expression of TRAF2, NIK, or BCL-XL, or short hairpin RNA TRAF3 knock-down, significantly reduced LCL161/LBH589 lethality, as did ectopic expression of dominant-negativ...
2926 Patients with multiple myeloma (MM) undergoing high dose therapy and autologous stem cell tr... more 2926 Patients with multiple myeloma (MM) undergoing high dose therapy and autologous stem cell transplantation (SCT) remain at risk for disease progression. Maintenance therapy may delay recurrence but is associated with toxicities, highlighting the need for alternative strategies for long-term disease control. Malignant plasma cells in MM patients occasionally express highly immunogenic cancer testis antigens (CTA). CTA expression is regulated by DNA methylation, and may be increased by 5-azacitidine (Celgene corp., Summit, NJ) (Aza), a DNA hypomethylating agent. The addition of lenalidomide (Celgene corp.) (L) may augment any ensuing adoptive CTA-specific immune response. These immune effector cells may then be collected and adoptively transferred in a setting of lympho-depletion and minimal residual disease following SCT, serving a maintenance function. To demonstrate the feasibility of this approach, we initiated a phase II clinical trial of Aza administered sequentially with L ...
Histone deacetylase (e.g., vorinostat) and proteasome inhibitors interact synergistically to prom... more Histone deacetylase (e.g., vorinostat) and proteasome inhibitors interact synergistically to promote cell death in various cancer cells, including malignant human hematopoietic cells. The novel, proteasome inhibitor carfilzomib is an attractive candidate for combination regimens due to diminished neurotoxicity, irreversible proteasome inhibition, favorable tolerability profile relative to bortezomib, and preclinical and clinical evidence of activity in bortezomib-resistant cells and patients. Preclinical studies demonstrated synergistic interactions between carfilzomib and vorinostat in human diffuse large B-cell lymphoma (DLBCL) and mantle cell lymphoma (MCL) cells both in vitro and in vivo as well as in bortezomib-resistant cells (Dasmahapatra et al., Blood 115:4478; 2010; Mol Cancer Ther 10:1686, 2001). These preclinical findings prompted a phase 1 trial, using a 3+3 design, with the goal of determining the maximum tolerated dose (MTD)/recommended phase 2 dose (RP2D) for the comb...
3588 Although reports of synergistic interactions between proteasome and histone deacetylase (HDA... more 3588 Although reports of synergistic interactions between proteasome and histone deacetylase (HDAC) inhibitors in acute leukemias have been limited, they are well described in B-cell malignancies (e.g., myeloma and lymphoma). Nevertheless, preclinical findings have shown striking synergism between the HDAC inhibitor belinostat (previously PXD-101) and the proteasome inhibitor bortezomib, administered at low (sub-micromolar) concentrations, in cultured and primary acute myeloid leukemia (AML) and acute lymphocytic leukemia cells (Dai Y et al. Br J Haematol. 2011). These findings prompted initiation of a phase I trial, using a 3+3 design, with the primary objective of determining the maximum tolerated dose (MTD) for the combination of bortezomib and belinostat in patients with relapsed or refractory acute leukemia, myelodysplastic syndrome (MDS), or chronic myelogenous leukemia in blast crisis (CML-BC). To date, 25 patients with the following disease types have been treated: acute leu...
Inhibitor of apoptosis proteins (IAPs e.g., XIAP, cIAP1, cIAP2) inhibit apoptosis through diverse... more Inhibitor of apoptosis proteins (IAPs e.g., XIAP, cIAP1, cIAP2) inhibit apoptosis through diverse mechanisms. Recent attention has focused on novel functions of cIAP1/2, including activation of the canonical and non-canonical NF-κB pathways and inhibition of the extrinsic apoptotic pathway through the ripoptosome. These considerations have stimulated the development of IAP antagonists such as the bivalent IAP inhibitor birinapant (TL-32711). Proteasome inhibitors such as bortezomib are highly active in multiple myeloma (MM), and act, at least in part, by inhibiting NF-κB, raising the possibility of cooperative interactions with IAP antagonists. Synergistic induction of apoptosis was observed in human MM cell lines, including drug-naïve U266 cells, their bortezomib-resistant counterparts (PS-R), and multiple other human MM lines. Birinapant (500 nM) sharply down-regulated cIAP1/2 in MM cells, accompanied by reduced expression of TRAF2, RIP1, and p-IKKβ, as well as robust TRAF3 up-reg...
2931 AZD6244 is a potent, selective, oral, non-ATP competitive small molecule inhibitor of the mi... more 2931 AZD6244 is a potent, selective, oral, non-ATP competitive small molecule inhibitor of the mitogen-activated protein kinase, MEK 1/2 that has shown significant pre-clinical activity in multiple myeloma (MM) cells, both in vitro and in vivo, as well as a favorable clinical profile. The present phase II study was designed to determine the response rate for AZD6244 in patients with relapsed or refractory MM. The study utilized a two-stage Simon design to allow for early termination if there was strong evidence of regimen inactivity. Eligible patients were restricted to those with MM who have had at least 2 prior regimens. AZD6244 capsules (75 mg) were administered orally twice daily continuously for 28 day cycles. Response was evaluated after 3 cycles. To date, 37 patients have been enrolled (13 in the 1st stage and 24 in the 2nd stage). One subject enrolled in the 1st stage was not treated. Gender enrollment was balanced (male/female 18/19). The median age of treated patients was ...
We report the results of a phase 1 dose-escalation study of belinostat and bortezomib in adult pa... more We report the results of a phase 1 dose-escalation study of belinostat and bortezomib in adult patients with acute leukemia or MDS or CML with blast crisis. Thirty-eight patients received IV belinostat days 1-5 and 8-12 with IV bortezomib days 1, 4, 8, and 11 every 21 days. QTc prolongation was the only identified DLT. The RP2Ds were 1.3 mg/m2 bortezomib and 1000 mg/m2 belinostat. One patient with highly refractory MLL-ENL rearranged biphenotypic AML with multiple karyotypic aberrations had a complete pathologic and karyotypic response. One patient with post-MPN AML remained on study with stable disease (SD) for 32 cycles. Whole-exome sequencing revealed no aberrations in the first patient and a hyper-mutator genotype in the second. Eighteen patients had a best response of SD. We conclude that this treatment strategy is feasible but has limited activity in this population. Nevertheless, the factors that predict exceptional responses to this strategy warrant further investigation.
1794 Preclinical studies have demonstrated synergistic interactions between proteasome and histon... more 1794 Preclinical studies have demonstrated synergistic interactions between proteasome and histone deacetylase (HDAC) inhibitors in diverse hematologic malignancies, including those of B-cell origin. We have previously shown that the combination of the proteasome inhibitor bortezomib and the HDAC inhibitor romidepsin, administered at extremely low concentrations (∼3–5 nM), results in a striking increase in apoptosis in primary CLL cells, including cells from patients with CLL refractory to standard treatments (Dai Y et al. Clin Cancer Res. 2008). These findings prompted initiation of a phase I trial, using a 3+3 design, with the primary objective of determining the maximum tolerated dose (MTD) for the combination of bortezomib and romidepsin in patients with relapsed or refractory chronic lymphocyte leukemia/small lymphocytic lymphoma (CLL/SLL), indolent B-cell lymphoma, or peripheral T-cell lymphoma (PTCL). To date, 11 patients have been enrolled and treated. All of the patients tr...
Purpose: This phase I study was conducted to identify the maximum-tolerated dose (MTD) of alvocid... more Purpose: This phase I study was conducted to identify the maximum-tolerated dose (MTD) of alvocidib when combined with vorinostat in patients with relapsed, refractory, or poor prognosis acute leukemia, or refractory anemia with excess blasts-2. Secondary objectives included investigating the pharmacokinetic and pharmacodynamic effects of the combination. Experimental Design: Patients received vorinostat (200 mg orally, three times a day, for 14 days) on a 21-day cycle, combined with 2 different alvocidib administration schedules: a 1-hour intravenous infusion, daily × 5; or a 30-minute loading infusion followed by a 4-hour maintenance infusion, weekly × 2. The alvocidib dose was escalated using a standard 3+3 design. Results: Twenty-eight patients were enrolled and treated. The alvocidib MTD was 20 mg/m2 (30-minute loading infusion) followed by 20 mg/m2 (4-hour maintenance infusion) on days one and eight, in combination with vorinostat. The most frequently encountered toxicities we...
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Papers by Maciej Kmieciak