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Both previous and additional genetic knockdown studies reported herein implicate G protein-coupled receptor kinase 6 (GRK6) as a critical kinase required for the survival of multiple myeloma (MM) cells. Therefore, we sought to develop a small molecule GRK6 inhibitor as an MM therapeutic. From a focused library of known kinase inhibitors, we identified two hits with moderate biochemical potencies against GRK6. From these hits, we developed potent (IC50< 10 nM) analogues with selectivity against off-target kinases. Further optimization led to the discovery of an analogue (18) with an IC50value of 6 nM against GRK6 and selectivity against a panel of 85 kinases. Compound 18has potent cellular target engagement and antiproliferative activity against MM cells and is synergistic with bortezomib. In summary, we demonstrate that targeting GRK6 with small molecule inhibitors represents a promising approach for MM and identify 18as a novel, potent, and selective GRK6 inhibitor.

Autologous stem cell transplant (ASCT) is a feasible treatment option for multiple myeloma (MM) patients with renal insufficiency; however, these patients tend to experience higher rates of drug toxicity and transplant-related mortality (TRM) during ASCT. Recent adoption of bortezomib-based induction regimens and dose reduction of melphalan during conditioning may improve outcomes in this population. In this single center retrospective study, we compared the toxicity and survival outcomes of 96 MM patients with renal insufficiency undergoing ASCT between two eras: 1998–2007 and 2008–2016. The proportion of dialysis dependent patients was similar in both groups (49 and 45%). We found no TRM in those transplanted more recently as compared with 13% in the older era of ASCT. There were significantly more high grade (grades 3–4) toxicities in the older era of ASCT including high grade electrolyte abnormalities, mucositis, delirium, and bleeding. Patients transplanted more recently had significantly higher overall response rate (ORR) as well as deeper responses to ASCT (≥VGPR in 79% vs 39%). Progression-free survival (PFS) was prolonged by 26 months in the more recent era compared with the older era. Overall, improvements in treatment regimens have resulted in reduced TRM and toxicities for patients with renal insufficiency undergoing ASCT.

Introduction:

Introduction:

Multiple Myeloma (MM) is a B cell neoplasm characterized by the accumulation of mature plasma cells (PCs) within the bone marrow (BM). Despite treatment advances, MM remains incurable in the vast majority of patients. Disease relapse typically occurs regardless of treatment with proteasome inhibitors (such as bortezomib and carfilzomib), immunomodulatory drugs (such as lenalidomide and pomalidomide), anti-CD38 monoclonal antibodies (such as daratumumab and isatuximab) or myeloablative melphalan and autologous stem cell transplantation, and also occurs following treatment with immunotherapeutics such as bispecific antibodies and/or CAR-T cells (targeting BCMA, FcRH5 or GPRC5D). This failure to achieve reliable cure in MM, despite the routine attainment of deep treatment responses against PCs, indicates the existence of important intra-tumor heterogeneity and the presence of rare drug-resistant MM cells with full tumorigenic capacity.

The molecular target(s) cooperating with proteasome inhibition in multiple myeloma (MM) remain unknown. We therefore measured proliferation in MM cells transfected with 13 984 small interfering RNAs in the absence or presence of increasing concentrations of bortezomib. We identified 37 genes, which when silenced, are not directly cytotoxic but do synergistically potentiate the growth inhibitory effects of bortezomib. To focus on bortezomib sensitizers, genes that also sensitized MM to melphalan were excluded. When suppressed, the strongest bortezomib sensitizers were the proteasome subunits PSMA5, PSMB2, PSMB3, and PSMB7providing internal validation, but others included BAZ1B, CDK5, CDC42SE2, MDM4, NME7, RAB8B, TFE3, TNFAIP3, TNK1, TOP1, VAMP2, and YY1. The strongest hit CDK5also featured prominently in pathway analysis of primary screen data. Cyclin-dependent kinase 5 (CDK5) is expressed at high levels in MM and neural tissues with relatively low expression in other organs. Viral shRNA knockdown of CDK5 consistently sensitized 5 genetically variable MM cell lines to proteasome inhibitors (bortezomib and carfilzomib). Small-molecule CDK5 inhibitors were demonstrated to synergize with bortezomib to induce cytotoxicity of primary myeloma cells and myeloma cell lines. CDK5 regulation of proteasome subunit PSMB5was identified as a probable route to sensitization.

The molecular target(s) cooperating with proteasome inhibition in multiple myeloma (MM) remain unknown. We therefore measured proliferation in MM cells transfected with 13 984 small interfering RNAs in the absence or presence of increasing concentrations of bortezomib. We identified 37 genes, which when silenced, are not directly cytotoxic but do synergistically potentiate the growth inhibitory effects of bortezomib. To focus on bortezomib sensitizers, genes that also sensitized MM to melphalan were excluded. When suppressed, the strongest bortezomib sensitizers were the proteasome subunits PSMA5, PSMB2, PSMB3, and PSMB7 providing internal validation, but others included BAZ1B, CDK5, CDC42SE2, MDM4, NME7, RAB8B, TFE3, TNFAIP3, TNK1, TOP1, VAMP2, and YY1. The strongest hit CDK5 also featured prominently in pathway analysis of primary screen data. Cyclin-dependent kinase 5 (CDK5) is expressed at high levels in MM and neural tissues with relatively low expression in other organs. Viral shRNA knockdown of CDK5 consistently sensitized 5 genetically variable MM cell lines to proteasome inhibitors (bortezomib and carfilzomib). Small-molecule CDK5 inhibitors were demonstrated to synergize with bortezomib to induce cytotoxicity of primary myeloma cells and myeloma cell lines. CDK5 regulation of proteasome subunit PSMB5 was identified as a probable route to sensitization.

A paucity of validated kinase targets in human multiple myeloma has delayed clinical deployment of kinase inhibitors in treatment strategies. We therefore conducted a kinome-wide small interfering RNA (siRNA) lethality study in myeloma tumor lines bearing common t(4;14), t(14;16), and t(11;14) translocations to identify critically vulnerable kinases in myeloma tumor cells without regard to preconceived mechanistic notions. Fifteen kinases were repeatedly vulnerable in myeloma cells, including AKT1, AK3L1, AURKA, AURKB, CDC2L1, CDK5R2, FES, FLT4, GAK, GRK6, HK1, PKN1, PLK1, SMG1, and TNK2. Whereas several kinases (PLK1, HK1) were equally vulnerable in epithelial cells, others and particularly G protein–coupled receptor kinase, GRK6, appeared selectively vulnerable in myeloma. GRK6 inhibition was lethal to 6 of 7 myeloma tumor lines but was tolerated in 7 of 7 human cell lines. GRK6 exhibits lymphoid-restricted expression, and from coimmunoprecipitation studies we demonstrate that expression in myeloma cells is regulated via direct association with the heat shock protein 90 (HSP90) chaperone. GRK6 silencing causes suppression of signal transducer and activator of transcription 3 (STAT3) phosphorylation associated with reduction in MCL1 levels and phosphorylation, illustrating a potent mechanism for the cytotoxicity of GRK6 inhibition in multiple myeloma (MM) tumor cells. As mice that lack GRK6 are healthy, inhibition of GRK6 represents a uniquely targeted novel therapeutic strategy in human multiple myeloma.

A paucity of validated kinase targets in human multiple myeloma has delayed clinical deployment of kinase inhibitors in treatment strategies. We therefore conducted a kinome-wide small interfering RNA (siRNA) lethality study in myeloma tumor lines bearing common t(4;14), t(14;16), and t(11;14) translocations to identify critically vulnerable kinases in myeloma tumor cells without regard to preconceived mechanistic notions. Fifteen kinases were repeatedly vulnerable in myeloma cells, including AKT1, AK3L1, AURKA, AURKB, CDC2L1, CDK5R2, FES, FLT4, GAK, GRK6, HK1, PKN1, PLK1, SMG1, and TNK2. Whereas several kinases (PLK1, HK1) were equally vulnerable in epithelial cells, others and particularly G protein–coupled receptor kinase, GRK6, appeared selectively vulnerable in myeloma. GRK6 inhibition was lethal to 6 of 7 myeloma tumor lines but was tolerated in 7 of 7 human cell lines. GRK6 exhibits lymphoid-restricted expression, and from coimmunoprecipitation studies we demonstrate that expression in myeloma cells is regulated via direct association with the heat shock protein 90 (HSP90) chaperone. GRK6 silencing causes suppression of signal transducer and activator of transcription 3 (STAT3) phosphorylation associated with reduction in MCL1 levels and phosphorylation, illustrating a potent mechanism for the cytotoxicity of GRK6 inhibition in multiple myeloma (MM) tumor cells. As mice that lack GRK6 are healthy, inhibition of GRK6 represents a uniquely targeted novel therapeutic strategy in human multiple myeloma.

As multiple myeloma tumors universally dysregulate cyclin D genes we conducted high-throughput chemical library screens for compounds that induce suppression of cyclin D2 promoter transcription. The top-ranked compound was a natural triterpenoid, pristimerin. Strikingly, the early transcriptional response of cells treated with pristimerin closely resembles cellular responses elicited by proteosome inhibitors, with rapid induction of heat shock proteins, activating transcription factor 3 (ATF3), and CHOP. Enzymatic assays and immunoblotting confirm that pristimerin rapidly (< 90 minutes) and specifically inhibits chymotrypsin-like proteosome activity at low concentrations (< 100 nM) and causes accumulation of cellular ubiquitinated proteins. Notably, cytotoxic triterpenoids including pristimerin inhibit NF-κB activation via inhibition of IKKα or IKKβ, whereas proteosome inhibitors instead suppress NF-κB function by impairing degradation of ubiquitinated IκB. By inhibiting both IKK and the proteosome, pristimerin causes overt suppression of constitutive NF-κB activity in myeloma cells that may mediate its suppression of cyclin D. Multiple myeloma is exquisitely sensitive to proteosome or NF-κB pathway inhibition. Consistent with this, pristimerin is potently and selectively lethal to primary myeloma cells (IC50 < 100 nM), inhibits xenografted plasmacytoma tumors in mice, and is synergistically cytotoxic with bortezomib—providing the rationale for pharmaceutical development of triterpenoid dual-function proteosome/NF-κB inhibitors as therapeutics for human multiple myeloma and related malignancies.

As multiple myeloma tumors universally dysregulate cyclin D genes we conducted high-throughput chemical library screens for compounds that induce suppression of cyclin D2 promoter transcription. The top-ranked compound was a natural triterpenoid, pristimerin. Strikingly, the early transcriptional response of cells treated with pristimerin closely resembles cellular responses elicited by proteosome inhibitors, with rapid induction of heat shock proteins, activating transcription factor 3 (ATF3), and CHOP. Enzymatic assays and immunoblotting confirm that pristimerin rapidly (< 90 minutes) and specifically inhibits chymotrypsin-like proteosome activity at low concentrations (< 100 nM) and causes accumulation of cellular ubiquitinated proteins. Notably, cytotoxic triterpenoids including pristimerin inhibit NF-κB activation via inhibition of IKKα or IKKβ, whereas proteosome inhibitors instead suppress NF-κB function by impairing degradation of ubiquitinated IκB. By inhibiting both IKK and the proteosome, pristimerin causes overt suppression of constitutive NF-κB activity in myeloma cells that may mediate its suppression of cyclin D. Multiple myeloma is exquisitely sensitive to proteosome or NF-κB pathway inhibition. Consistent with this, pristimerin is potently and selectively lethal to primary myeloma cells (IC50< 100 nM), inhibits xenografted plasmacytoma tumors in mice, and is synergistically cytotoxic with bortezomib—providing the rationale for pharmaceutical development of triterpenoid dual-function proteosome/NF-κB inhibitors as therapeutics for human multiple myeloma and related malignancies.

Introduction

Introduction

Introduction:Autologous stem cell transplantation (ASCT) is standard therapy for selected patients with newly diagnosed multiple myeloma (MM). Studies in MM and lymphoma have suggested that ability to mobilize and collect a higher yield of CD34 +cells predicts for improved survival outcomes, perhaps reflecting better bone marrow reserve (Bolwell 2007, Raschle 2011). We aimed to validate this hypothesis by correlating high CD34 +cell collection (“supermobilizers”) and survival outcomes in a large myeloma cohort with long follow-up.

Prica: Kite Gilead: Honoraria; Astra-Zeneca: Honoraria. Chen: Astrazeneca: Membership on an entity's Board of Directors or advisory committees; Beigene: Membership on an entity's Board of Directors or advisory committees; 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; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy. Crump: Novartis: Membership on an entity's Board of Directors or advisory committees; Kyte/Gilead: Membership on an entity's Board of Directors or advisory committees; Epizyme: Research Funding; Roche: Research Funding. Kridel: Gilead Sciences: Research Funding. Kuruvilla: AbbVie: Honoraria; Incyte: Honoraria; Medison Ventures: Honoraria; Merck: Honoraria; Novartis: Honoraria; Gilead: Honoraria; Seattle Genetics: Honoraria; Karyopharm: Honoraria, Other: Data and Safety Monitoring Board; Roche: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; AstraZeneca: Honoraria, Research Funding; TG Therapeutics: Honoraria; Amgen: Honoraria; Pfizer: Honoraria; BMS: Honoraria; Antengene: Honoraria. Reece: Celgene: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; BMS: Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Sanofi: Honoraria; Millennium: Research Funding; Karyopharm: Consultancy, Research Funding; Amgen: Consultancy, Honoraria; GSK: Honoraria.

Prica: Astra-Zeneca: Honoraria; Kite Gilead: Honoraria. Reece: Takeda: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria; Millennium: Research Funding; Sanofi: Honoraria; BMS: Honoraria, Research Funding; Karyopharm: Consultancy, Research Funding; GSK: Honoraria. Trudel: Amgen: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; GlaxoSmithKline: Consultancy, Honoraria, Research Funding; BMS/Celgene: Consultancy, Honoraria, Research Funding; Genentech: Research Funding; Roche: Consultancy; Sanofi: Honoraria; Pfizer: Honoraria, Research Funding. Chen: Beigene: Membership on an entity's Board of Directors or advisory committees; Astrazeneca: Membership on an entity's Board of Directors or advisory committees; 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; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy.

Reece: Millennium: Research Funding; Sanofi: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria; Takeda: Consultancy, Honoraria, Research Funding; Karyopharm: Consultancy, Research Funding; GSK: Honoraria; BMS: Honoraria, Research Funding. Trudel: Amgen: Honoraria, Research Funding; BMS/Celgene: Consultancy, Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Genentech: Research Funding; Sanofi: Honoraria; Pfizer: Honoraria, Research Funding; GlaxoSmithKline: Consultancy, Honoraria, Research Funding; Roche: Consultancy. Prica: Astra-Zeneca: Honoraria; Kite Gilead: Honoraria. Chen: Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS: Consultancy, Membership on an entity's Board of Directors or advisory committees; Astrazeneca: Membership on an entity's Board of Directors or advisory committees; Beigene: Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy.

D-cyclins are regulators of cell division that act in a complex with cyclin-dependent kinases to commit cells to a program of DNA replication. D-cyclins are overexpressed in many tumors, including multiple myeloma and leukemia, and contribute to disease progression and chemoresistance. To better understand the role and impact of D-cyclins in hematologic malignancies, we conducted a high throughput screen for inhibitors of the cyclin D2 promoter and identified the drug cyproheptadine. In myeloma and leukemia cells, cyproheptadine decreased expression of cyclins D1, D2, and D3 and arrested these cells in the G0/G1 phase. After D-cyclin suppression, cyproheptadine induced apoptosis in myeloma and leukemia cell lines and primary patient samples preferentially over normal hematopoietic cells. In mouse models of myeloma and leukemia, cyproheptadine inhibited tumor growth without significant toxicity. Cyproheptadine-induced apoptosis was preceded by activation of the mitochondrial pathway of caspase activation and was independent of the drug's known activity as an H1 histamine and serotonin receptor antagonist. Thus, cyproheptadine represents a lead for a novel therapeutic agent for the treatment of malignancy. Because the drug is well tolerated and already approved in multiple countries for clinical use as an antihistamine and appetite stimulant, it could be moved directly into clinical trials for cancer.

D-cyclins are regulators of cell division that act in a complex with cyclin-dependent kinases to commit cells to a program of DNA replication. D-cyclins are overexpressed in many tumors, including multiple myeloma and leukemia, and contribute to disease progression and chemoresistance. To better understand the role and impact of D-cyclins in hematologic malignancies, we conducted a high throughput screen for inhibitors of the cyclin D2 promoter and identified the drug cyproheptadine. In myeloma and leukemia cells, cyproheptadine decreased expression of cyclins D1, D2, and D3 and arrested these cells in the G0/G1phase. After D-cyclin suppression, cyproheptadine induced apoptosis in myeloma and leukemia cell lines and primary patient samples preferentially over normal hematopoietic cells. In mouse models of myeloma and leukemia, cyproheptadine inhibited tumor growth without significant toxicity. Cyproheptadine-induced apoptosis was preceded by activation of the mitochondrial pathway of caspase activation and was independent of the drug's known activity as an H1 histamine and serotonin receptor antagonist. Thus, cyproheptadine represents a lead for a novel therapeutic agent for the treatment of malignancy. Because the drug is well tolerated and already approved in multiple countries for clinical use as an antihistamine and appetite stimulant, it could be moved directly into clinical trials for cancer.

The oncogene c-maf is frequently overexpressed in multiple myeloma cell lines and patient samples and contributes to increased cellular proliferation in part by inducing cyclin D2 expression. To identify regulators of c-maf, we developed a chemical screen in NIH3T3 cells stably overexpressing c-maf and the cyclin D2 promoter driving luciferase. From a screen of 2400 off-patent drugs and chemicals, we identified glucocorticoids as c-maf–dependent inhibitors of cyclin D2 transactivation. In multiple myeloma cell lines, glucocorticoids reduced levels of c-maf protein without influencing corresponding mRNA levels. Subsequent studies demonstrated that glucocorticoids increased ubiquitination-dependent degradation of c-maf and up-regulated ubiquitin C mRNA. Moreover, ectopic expression of ubiquitin C recapitulated the effects of glucocorticoids, demonstrating regulation of c-maf protein through the abundance of the ubiquitin substrate. Thus, using a chemical biology approach, we identified a novel mechanism of action of glucocorticoids and a novel mechanism by which levels of c-maf protein are regulated by the abundance of the ubiquitin substrate.

The oncogene c-maf is frequently overexpressed in multiple myeloma cell lines and patient samples and contributes to increased cellular proliferation in part by inducing cyclin D2 expression. To identify regulators of c-maf, we developed a chemical screen in NIH3T3 cells stably overexpressing c-maf and the cyclin D2 promoter driving luciferase. From a screen of 2400 off-patent drugs and chemicals, we identified glucocorticoids as c-maf–dependent inhibitors of cyclin D2 transactivation. In multiple myeloma cell lines, glucocorticoids reduced levels of c-maf protein without influencing corresponding mRNA levels. Subsequent studies demonstrated that glucocorticoids increased ubiquitination-dependent degradation of c-maf and up-regulated ubiquitin C mRNA. Moreover, ectopic expression of ubiquitin C recapitulated the effects of glucocorticoids, demonstrating regulation of c-maf protein through the abundance of the ubiquitin substrate. Thus, using a chemical biology approach, we identified a novel mechanism of action of glucocorticoids and a novel mechanism by which levels of c-maf protein are regulated by the abundance of the ubiquitin substrate.

Bcl family members Bcl-2, Bcl-xL, and Mcl-1, are frequently expressed and implicated in the survival of myeloma cells. Obatoclax (GX015-070) is a novel, small-molecule antagonist of the BH3-binding groove of the Bcl family of proteins. We show that GX015-070 inhibits the binding of Bak to Mcl-1, up-regulates Bim, induces cytochrome crelease, and activates capase-3 in human myeloma cell lines (HMCLs), confirming the predicted mechanism of action. Consequently, GX015-070 potently inhibited the viability of 15 of 16 HMCLs (mean IC50of 246 nM), including those resistant to melphalan and dexamethasone. In combination studies, GX015-070 enhanced the antimyeloma activity induced by melphalan, dexamethasone, or bortezomib. Sensitivity to GX015-070 correlated with the absence or near absence of Bcl-xL. Coculture with interleukin-6 or adherence to bone marrow stroma conferred modest resistance; however, it did not overcome GX015-070–induced cytotoxicity. Of importance, GX015-070 as a single agent induced potent cytotoxic responses against patient-derived tumor cells. GX015-070 inhibited normal bone marrow–derived colony formation; however, cytotoxicity to human blood lymphocytes was not observed. Taken together, these studies describe a novel BH3 mimic with selectivity for Mcl-1, and support the therapeutic application of GX015-070 for diverse neoplasias including multiple myeloma.

Bcl family members Bcl-2, Bcl-xL, and Mcl-1, are frequently expressed and implicated in the survival of myeloma cells. Obatoclax (GX015-070) is a novel, small-molecule antagonist of the BH3-binding groove of the Bcl family of proteins. We show that GX015-070 inhibits the binding of Bak to Mcl-1, up-regulates Bim, induces cytochrome c release, and activates capase-3 in human myeloma cell lines (HMCLs), confirming the predicted mechanism of action. Consequently, GX015-070 potently inhibited the viability of 15 of 16 HMCLs (mean IC50 of 246 nM), including those resistant to melphalan and dexamethasone. In combination studies, GX015-070 enhanced the antimyeloma activity induced by melphalan, dexamethasone, or bortezomib. Sensitivity to GX015-070 correlated with the absence or near absence of Bcl-xL. Coculture with interleukin-6 or adherence to bone marrow stroma conferred modest resistance; however, it did not overcome GX015-070–induced cytotoxicity. Of importance, GX015-070 as a single agent induced potent cytotoxic responses against patient-derived tumor cells. GX015-070 inhibited normal bone marrow–derived colony formation; however, cytotoxicity to human blood lymphocytes was not observed. Taken together, these studies describe a novel BH3 mimic with selectivity for Mcl-1, and support the therapeutic application of GX015-070 for diverse neoplasias including multiple myeloma.

Recombinant streptococcal pyrogenic exotoxin C (SPE-C) is a potent superantigen that stimulates Vβ2-bearing human T cells, but is inactive in mice. SPE-C binds with high affinity to both human HLA-DR and murine I-E molecules, but not to murine I-A molecules in a zinc-dependent fashion. Competition binding studies with other recombinant toxins revealed that SPE-C lacks the generic low affinity major histocompatibility complex (MHC) class II α-chain binding site common to all other bacterial superantigens. Despite this, SPE-C cross-links MHC class II to induce homotypic aggregation of class II–bearing B cells. Nondenaturing sodium dodecyl sulfate electrophoresis and size exclusion chromatography revealed that both wild-type and recombinant SPE-C exist in a stable dimer at neutral or alkaline pH. These data support a recent crystal structure of SPE-C and reveal yet another mechanism by which bacterial superantigens ligate and cross-link MHC class II.

Although melphalan 200 mg/m2(Mel 200) followed by ASCT has been a standard approach for eligible multiple myeloma (MM) patients (pts) for over 25 years, efforts to improve transplant results continue, particularly in high-risk (HR) pts. One approach has been to augment the high-dose (HD) regimen by adding agents to Mel or performing “tandem” ASCT using Mel each time. The first MCRN Canadian national trial (MCRN-001) was designed to optimize the results of ASCT by using bortezomib (BTZ)-based induction, augmented HD chemotherapy with busulfan + melphalan (BuMel) before ASCT, and lenalidomide (Len) maintenance post-ASCT. We now report longer-term outcomes from this study, including results in HR pts and the incidence of secondary primary malignancies (SPMs).

Chen: Celgene: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Amgen: Honoraria. Kridel:Gilead Sciences: Research Funding. Kukreti:Celgene: Honoraria; Amgen: Honoraria; Takeda: Honoraria. Kuruvilla:Celgene: Honoraria; Astra Zeneca: Honoraria; Seattle Genetics: Consultancy; Amgen: Honoraria; Roche: Consultancy; Karyopharm: Consultancy; Gilead: Consultancy; Abbvie: Consultancy; BMS: Consultancy; Roche: Research Funding; Janssen: Research Funding; Merck: Consultancy; Gilead: Honoraria; BMS: Honoraria; Karyopharm: Honoraria; Janssen: Honoraria; Roche: Honoraria; Seattle Genetics: Honoraria; Novartis: Honoraria; Merck: Honoraria. Reece:Otsuka: Research Funding; Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; BMS: Research Funding. Tiedemann:Amgen: Honoraria; Novartis: Honoraria; Takeda: Honoraria; Celgene: Honoraria; BMS: Honoraria; Janssen: Honoraria. Trudel:Pfizer: Honoraria; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Honoraria; Janssen: Honoraria, Research Funding; Astellas: Research Funding; Genentech: Research Funding; Sanofi: Honoraria. Prica:Janssen: Honoraria; Celgene: Honoraria.

Reece: Otsuka: Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding. White:Takeda: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Amgen: Consultancy, Honoraria. Venner:Janssen: Honoraria; Amgen: Honoraria, Research Funding; Takeda: Honoraria; Celgene: Honoraria; Sanofi: Honoraria; J&J: Research Funding. Stakiw:Novartis: Honoraria, Speakers Bureau; Celgene: Honoraria, Speakers Bureau; Sanofi: Honoraria; Amgen: Honoraria, Speakers Bureau; Roche: Research Funding; BMS: Honoraria; Janssen: Honoraria, Research Funding, Speakers Bureau; Lundbeck: Honoraria. Sebag:Amgen: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees. Comeau:Celgene: Consultancy; Janssen: Consultancy; Takeda: Consultancy; Seattle Genetics: Consultancy; Novartis: Consultancy. Song:Celgene: Honoraria, Research Funding; Janssen: Honoraria; Amgen: Honoraria; Takeda: Honoraria. Roy:Celgene: Consultancy, Honoraria, Research Funding; Sanofi Canada: Research Funding; ExCellThera: Patents & Royalties: Royalties from sales of UM171, Research Funding; Amgen Canada: Honoraria; Janssen Canada: Honoraria. Louzada:Amgen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Bayer: Honoraria. McCurdy:Celgene: Honoraria; Janssen: Honoraria. Kukreti:Celgene: Honoraria; Amgen: Honoraria; Takeda: Honoraria. Trudel:Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Honoraria; Astellas: Research Funding; Genentech: Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Honoraria; Janssen: Honoraria, Research Funding; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Honoraria. Prica:Celgene: Honoraria; Janssen: Honoraria. Tiedemann:Celgene: Honoraria; Amgen: Honoraria; BMS: Honoraria; Janssen: Honoraria; Novartis: Honoraria; Takeda: Honoraria. Chen:Amgen: Honoraria; Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding.The combination of busulfan IV and melphalan conditioning for ASCT in myeloma is off-label.

Chen: Celgene: Honoraria, Research Funding; Amgen: Honoraria; Janssen: Honoraria, Research Funding. Prica:Celgene: Honoraria; Janssen: Honoraria. Reece:Otsuka: Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Merck: Research Funding. Tiedemann:Takeda: Honoraria; Novartis: Honoraria; Amgen: Honoraria; BMS: Honoraria; Janssen: Honoraria; Celgene: Honoraria. Trudel:Astellas: Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Honoraria; Genentech: Research Funding; Sanofi: Honoraria; Pfizer: Honoraria; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Honoraria, Research Funding. Kukreti:Celgene: Honoraria; Amgen: Honoraria; Takeda: Honoraria.

Background:

Introduction

Tiedemann: Novartis: Consultancy; Celgene: Consultancy; Bristol-Myers Squibb: Consultancy; Janssen: Consultancy; Amgen: Consultancy; Takeda: Consultancy; Merck: Consultancy; Roche: Consultancy. Chen:Amgen: Honoraria.

Background:Autologous stem cell transplant (ASCT) for multiple myeloma (MM) can be feasible and safe in the outpatient setting and has become standard of care at many centers. Limited literature, however, exists on the types of infectious complications, including specific pathogenic isolates and resistance patterns in the outpatient (OP) setting and whether these differ from the inpatient (IP) setting. This creates uncertainty and variation among local treatment guidelines on antibiotic prophylaxis and treatment of infections in the outpatient population.

Background

Introduction:

Background

Background:In the era prior to the availability of novel agents, we have previously reported that the median progression-free survival (PFS) and overall survival (OS) of high-risk (HR) multiple myeloma (MM) patients (pts) undergoing a single autologous stem cell transplant (ASCT) was only 9.9 and 18.3 months, respectively (Chang H et al., Bone Marrow Transplant, 36(2005), 793). The current study expands our experience in newly diagnosed HR pts and focuses on the outcomes in pts treated with single or tandem ASCT.

Crump: Celgene: Consultancy; Janssen-Ortho: Consultancy; Seattle Genetics: Consultancy; Roche: Consultancy. Kukreti:Lundbeck: Honoraria; Celgene: Honoraria; Amgen: Honoraria. Kuruvilla:Abbvie: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Merck: Honoraria; BMS: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Amgen: Honoraria; Roche Canada: Consultancy, Honoraria, Research Funding; Seattle Genetics: Consultancy, Honoraria; Lundbeck: Honoraria. Tiedemann:Celgene: Honoraria; Janssen: Honoraria. Chen:Takeda: Research Funding; Celgene: Honoraria, Research Funding; Janssen: Honoraria, Research Funding. Prica:Celgene: Honoraria; Janssen: Honoraria.

Reece: Takeda: Consultancy, Honoraria, Research Funding; Otsuka: Honoraria, Research Funding; Merck: Research Funding; BMS: Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding. White:Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria. Venner:Takeda: Honoraria; Celgene: Honoraria, Research Funding; J+J: Research Funding; Janssen: Honoraria; Amgen: Honoraria. Stakiw:Roche: Research Funding; BMS: Honoraria; Novartis: Honoraria, Speakers Bureau; Amgen: Honoraria, Speakers Bureau; Celgene: Honoraria, Speakers Bureau; Jansen: Honoraria, Speakers Bureau. Sebag:Celgene: Honoraria; Novartis: Honoraria; Janssen: Honoraria. Comeau:Seattle Genetics: Consultancy; Celgene: Consultancy; Janssen: Consultancy; Takeda: Consultancy. Song:Otsuka: Honoraria; Janssen: Honoraria; Celgene: Honoraria, Research Funding. Louzada:Pfizer: Honoraria; Bayer: Honoraria; Celgene: Consultancy, Honoraria; Janssen: Consultancy, Honoraria. McCurdy:Celgene: Honoraria. Kukreti:Celgene: Honoraria. Trudel:Glaxo Smith Kline: Honoraria, Research Funding; Celgene: Honoraria; Novartis: Honoraria; Oncoethix: Research Funding. Prica:Janssen: Honoraria; Celgene: Honoraria. Tiedemann:Novartis: Honoraria; Takeda Oncology: Honoraria; Janssen: Honoraria; Celgene: Honoraria; Amgen: Honoraria; BMS Canada: Honoraria. Chen:Takeda: Research Funding; Celgene: Honoraria, Research Funding; Janssen: Honoraria, Research Funding.

Cote: Takeda: Honoraria; Janssen: Honoraria. Reece:Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; BMS: Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Merck: Research Funding; Otsuka: Honoraria, Research Funding. Chen:Takeda: Research Funding; Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding. Prica:Celgene: Honoraria; Janssen: Honoraria. Tiedemann:Janssen: Honoraria; Celgene: Honoraria. Trudel:Amgen: Honoraria; Oncoethix: Research Funding; BMS: Honoraria; Glaxo Smith Kline: Honoraria, Research Funding; Novartis: Consultancy, Honoraria; Celgene: Consultancy, Equity Ownership, Honoraria. Kukreti:Lundbeck: Honoraria; Celgene: Honoraria; Amgen: Honoraria.

Chen: Takeda: Research Funding; Celgene: Honoraria, Research Funding; Janssen: Honoraria, Research Funding. Kukreti:Amgen: Honoraria; Celgene: Honoraria; Lundbeck: Honoraria. Prica:Janssen: Honoraria; Celgene: Honoraria. Tiedemann:Celgene: Honoraria; Janssen: Honoraria; BMS Canada: Honoraria; Novartis: Honoraria; Amgen: Honoraria; Takeda Oncology: Honoraria. Trudel:Celgene: Honoraria; Glaxo Smith Kline: Honoraria, Research Funding; Oncoethix: Research Funding; Novartis: Honoraria. Reece:Janssen: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Merck: Research Funding; BMS: Honoraria, Research Funding; Otsuka: Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

Prica: Janssen: Honoraria; Celgene: Honoraria. Chen:Celgene: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Takeda: Research Funding. Reece:Janssen: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Merck: Research Funding; Novartis: Honoraria, Research Funding; Otsuka: Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; BMS: Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding. Trudel:Glaxo Smith Kline: Honoraria, Research Funding; Celgene: Honoraria; Novartis: Honoraria; Oncoethix: Research Funding. Tiedemann:Takeda Oncology: Honoraria; Celgene: Honoraria; Janssen: Honoraria; Novartis: Honoraria; Amgen: Honoraria; BMS Canada: Honoraria. Kukreti:Celgene: Honoraria; Lundbeck: Honoraria; Amgen: Honoraria.

Background:A second (salvage) ASCT has frequently been offered to MM patients with relapsed disease who experience benefit from the first procedure. We have previously reported that pts undergoing a salvage ASCT in the era of VAD or thalidomide (thal) have a median progression-free survival (PFS) of 19 months (mos). The best results were observed in pts who experienced ≥ 2 year benefit after their first ASCT (Jimenez-Zepeda VH et al. Biol Blood Marrow Transplant2012; 18: 773-9). However, the utility of this approach after the introduction of novel chemotherapeutic agents–such as bortezomib (BTZ)–remains unclear. Initially, provincial funding for BTZ in Ontario was provided only for relapsed disease. However, in 2007, the combination of either BTZ + dexamethasone (BTZ-dex) or cyclophosphamide, BTZ + dex (CyBorD) was adopted as the standard induction regimen for newly diagnosed pts before ASCT performed as part of first-line therapy. We now examine the results of salvage ASCT in our centre after the availability of BTZ.

MRD negativity has become an important goal of the initial treatment of MM pts. Our phase 2 multi-center clinical trial, conducted in 10 major Canadian transplant centers, was designed to increase the MRD negativity rate after ASCT by using conditioning with 2 high-dose alkylating agents followed by len maintenance. In addition to conventional response criteria, this trial evaluated serial bone marrow aspirate (BMA) samples for MRD analysis by 8-color multiparameter flow cytometry (MFC) along with serum Hevylite™ assays of the involved HLC that were obtained before and after ASCT and during maintenance therapy.