POS-492 PREDICTING SURVIVAL FACTORS IN PATIENTS DIAGNOSED MULTIPLE MYELOMA WITH RENAL IMPAIREMENT :MONOCENTRIC STUDY ABOUT 53 CASES

Abstract SNS-032, formerly BMS-387032, is a highly selective and potent inhibitor of cyclin-dependent kinases (CDK) 2, 7 and 9. CDK2 and CDK7 are involved in cell cycle regulation. CDK7, along with CDK9, regulate RNA polymerase (Pol) II-dependent transcription. Temporary inhibition of RNA Pol II-dependent transcription by SNS-032 has significant effects on short half-life transcripts and proteins, particularly survival factors, cell cycle regulatory proteins, and cytokines that are critical for the survival of malignant B-cells in chronic lymphocytic leukemia (CLL) and multiple myeloma (MM). A phase 1 dose-escalation study in patients with MM and CLL is ongoing with separate dose escalations for each indication. The study is designed to evaluate safety, pharmacokinetics (PK) and preliminary evidence of activity of a loading dose (LD) followed by a 6 hour infusion of SNS-032 given weekly for 3 consecutive weeks of each 28-day cycle. Dose and schedule aim to maintain for 6 hours threshold plasma concentrations of 115 ng/mL (in vitro IC90) and higher. The study incorporates an exploratory analysis of potential pharmacodynamic (PD) biomarkers such as decreased phosphorylation of RNA Pol II C-terminal domain to demonstrate inhibition of CDK7 and CDK9, and decreased expression of survival factors to indicate transcriptional inhibition. Methods: Previously treated patients with advanced CLL or MM, measurable disease, and ECOG status 0–1 were eligible. Increasing doses of SNS-032 given as an LD followed by a 6 hr infusion were evaluated. The total starting dose was 15 mg/m2 comprised of a LD of 5 mg/m2 followed by 10 mg/m2 over 6 hr with dose escalation by modified Fibonacci. PD studies of target modulation were performed on peripheral blood mononuclear cells (PBMC) obtained pre- and post-dose. Direct target modulation or downstream effects of target inhibition were evaluated. Results: 35 patients have been treated to date, 18 MM patients and 17 CLL patients. Median age was 61 (range 45–82), with 13 females and 24 males. Median number of prior therapies was 5 (range: 1–11). MM patients have received total doses of 15 – 75 mg/m2. No drug-related dose limiting toxicities (DLTs) or objective responses have been reported thus far in MM. CLL patients have received total doses of 15 –100 mg/m2. No drug-related DLTs were observed through the 50 mg/m2 dose cohort. At 75 mg/m2, concentrations of SNS- 032 exceeded IC90 (mean maximum concentration during the 6 hr infusion was 261 ± 45 ng/mL). Evidence of biochemical tumor lysis syndrome (TLS) was observed in all CLL patients treated at this dose level. One patient experienced a DLT of vascular leak syndrome and failure to receive all 3 cycle 1 doses. One CLL patient has been treated thus far at 100 mg/m2. This patient experienced TLS with a DLT of elevated liver function enzymes for >48 hr and received only 2 of 3 doses in cycle 1. No objective responses have been observed. PD analyses showed evidence of decreased Mcl-1 or XIAP in several patients. Conclusions: The mechanism of action of SNS-032 supports testing this agent in B-cell malignancies such as MM and CLL. A pharmacologically-derived dose regimen that sustains IC90 SNS-032 concentrations or higher for 6 hr is being studied; target levels were achieved and exceeded in cohort 5 (75 mg/m2) for both MM and CLL. No DLTs or objective responses have been observed thus far in MM. AEs and DLTs related to mild to moderate TLS were observed in CLL patients at 75 mg/m2 and higher. No objective responses have been observed. Preliminary evidence of target-specific PD modulation has been demonstrated. Enrollment in this trial is continuing.

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Lenalidomide and Bortezomib: Targeting Osteoclastogenesis, Osteoclast Survival Factors, and Bone Remodeling Markers in Multiple Myeloma.

Blood ◽

10.1182/blood.v110.11.1184.1184 ◽

2007 ◽

Vol 110 (11) ◽

pp. 1184-1184 ◽

Cited By ~ 1

Author(s):

Iris Breitkreutz ◽

Marc S. Raab ◽

Sonia Vallet ◽

Teru Hideshima ◽

Noopur Raje ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Resorption ◽

Bone Remodeling ◽

Flow Cytometric Analysis ◽

Tartrate Resistant Acid Phosphatase ◽

Αvβ3 Integrin ◽

Dependent Manner ◽

Survival Factors ◽

Bone Remodeling Markers ◽

Osteolytic Bone Disease

Abstract Osteolytic bone disease in Multiple Myeloma (MM) is caused by enhanced osteoclast (OCLs) activation and inhibition of osteoblast function. Lenalidomide and bortezomib have shown promising anti-MM effects, and bortezomib has inhibitory effects on OCLs. However, the effect of lenalidomide on OCLs in MM and how bortezomib interferes with osteoclastogenesis is unknown. Here we investigated the effect of lenalidomide and bortezomib on human OCLs. Peripheral blood mononuclear cells (PBMC) from MM patients were stimulated with receptor activator of NFk-B ligand (RANKL) (50ng/ml) and M-CSF (25ng/ml) for two weeks to induce OCL formation, in the presence or absence of lenalidomide or bortezomib. OCLs were identified by flow cytometric analysis using anti-αVβ3 integrin. Lenalidomide and bortezomib inhibited OCL differentiation indicated by a decrease in αVβ3-integrin (lenalidomide at 0μM: median 69.3%; range 28.9 – 89.0%; at 2μM: median 50.4%; range 21.5 – 64.2%; at 10μM: median 39.2%; range 33.6 – 47.5%) (bortezomib at 0nM: median 69.3%; range 28.9 – 89.0%; at 2nM: median 35.0%; range 11.0 – 79.0%; at 5nM: median 11.5%; range 5.5 – 8.8%; p<0.05). Tartrate-resistant acid phosphatase (TRAP) staining was performed to identify OCLs and confirm OCL activity. Both drugs decreased TRAP -positive cells (lenalidomide at 0μM: median 70.5%; range 50.0 – 84.0%; at 2μM: median 47.0%; range 31.0 – 77.0%; at 10μM: median 32.5%; range 14.0 – 44.0%; p<0.05) (bortezomib at 0nM: median 70.5%; range 50.0 – 84.0%; at 2nM: median 26.0%; range 11.0 – 39.0%; p<0.01; at 5nM: median 17.0%; range 13.0 – 34.0%; p<0.001). To assess bone resorption activity, OCL were cultured with dentine discs, in the presence or absence of lenalidomide and bortezomib, followed by light microscopic analysis. Both lenalidomide and bortezomib inhibited bone resorption in a dose-dependent manner. Using western blot analysis, we identified PU.1 and pERK as major targets of lenalidomide, and NFATc1 as a target of bortezomib, resulting in inhibition of osteoclastogenesis. Furthermore, downregulation of cathepsin K, essential for resorption of the bone collagen matrix, was also noted. We demonstrated a significant decrease of growth and survival factors including MIP-1α, IL-6, B-cell activation factor (BAFF), and a proliferation-inducing ligand (APRIL). Importantly, in serum from patients with refractory of relapsed MM treated with lenalidomide, the essential bone remodeling factor RANKL, as well as the RANKL/Osteoprotegerin (OPG) ratio, were significantly reduced. The median OPG at baseline was significantly lower (median 8.8pg/ml, range 7.7–12.6pg/ml) than after treatment (median 10.4pg/ml, range 8.5–15pg/ml) (p<0.05). Lenalidomide also significantly inhibited the secretion of RANKL in patient’s serum (baseline: median 17pg/ml, range 9.0–36pg/ml; two months after therapy: median 4.2pg/ml, range 2.5–36pg/ml) (p<0.05). The ratio of RANKL/OPG was significantly higher before than after therapy (baseline: median 2.2pg/ml, range 1.1–3.0pg/ml; two months after therapy: median 0.5pg/ml, range 0.3–2.6pg/ml) (p<0.05). We conclude that both agents specifically target key factors in osteoclastogenesis, thereby inhibiting OCL differentiation and function.

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Lenalidomide inhibits osteoclastogenesis, survival factors and bone-remodeling markers in multiple myeloma

Leukemia ◽

10.1038/leu.2008.174 ◽

2008 ◽

Vol 22 (10) ◽

pp. 1925-1932 ◽

Cited By ~ 147

Author(s):

I Breitkreutz ◽

M S Raab ◽

S Vallet ◽

T Hideshima ◽

N Raje ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Remodeling ◽

Survival Factors ◽

Bone Remodeling Markers

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CD28 Regulates Mitochondrial Metabolism in Long Lived Plasma Cells for Survival

Blood ◽

10.1182/blood.v124.21.570.570 ◽

2014 ◽

Vol 124 (21) ◽

pp. 570-570 ◽

Cited By ~ 1

Author(s):

Adam T Utley ◽

Cheryl H Rozanski ◽

Louise M Carlson ◽

Kelvin P Lee

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

T Cell ◽

Plasma Cell ◽

Mitochondrial Respiration ◽

Plasma Cells ◽

Binding Domain ◽

Mitochondrial Metabolism ◽

Survival Factors ◽

Adaptor Molecule

Abstract Sustained humoral immunity is dependent upon the ability of plasma cells to produce antigen specific antibody titers over a long period of time. Whether fighting against pathogens such as Ebola, Influenza, or the common cold, the continual presence of neutralizing antibodies in circulation is critical for an effective humoral immune response. When activated by antigen, B cells differentiate into a short lived plasma cell (SLPC) pool and reside in secondary lymphoid organs such as the spleen for days to weeks before dying by apoptosis. However, in the absence of constant antigenic presence there are examples of continual production of antigen-specific antibodies in the human body. To explain this, a different subset of long lived plasma cells (LLPC) has been proposed wherein the plasma cells compete for space in specialized bone marrow niches. These cells are not intrinsically long lived, but rather depend upon extrinsic survival factors for their persistence. Many of the survival factors for LLPCs in the bone marrow are shared with their malignant counterpart, e.g. multiple myeloma (MM). Our work centers on the elucidation of mechanisms by which both MM and LLPCs survive in the bone marrow microenvironment. Recently our lab has demonstrated a cell intrinsic role for CD28 signaling in the survival of both LLPCs and MM. CD28 is known as the canonical T cell co-stimulatory molecule and is required for effector T cell metabolic fitness. Under nutrient deprivation and chemotherapeutic challenge, CD28 is able to induce survival of LLPCs and MM cells respectively. However, the molecular and metabolic pathways that govern this prosurvival effect are not well understood. Here we demonstrate that CD28 induces mitochondrial respiration in bone marrow resident LLPCs but not in splenic SLPCs by staining with a mitochondrial-specific dye that is taken up in proportion to the mitochondrial membrane potential. A major byproduct of mitochondrial respiration is the production of reactive oxygen species (ROS). As a result of increased mitochondrial metabolism through the electron transport chain, CD28 is able to induce ROS in LLPCs but not in SLPCs. This is somewhat counterintuitive, in that ROS are well-characterized as cell damaging agents. Fascinatingly, mitochondrial respiration dependent ROS production downstream of CD28 is required for the prosurvival effect seen in LLPCs. Mechanistically, CD28-mediated production of ROS drive NFkB translocation as seen by ImageStream technology, which goes on to drive Blimp1 expression, the master transcriptional regulator of plasma cell identity. Utilizing a luciferase expressing plasmid and including different lengths of the Blimp1 promoter, we show that the CD28 responsive element lies from 4500 to 7500 base pairs from the transcriptional start site. Furthermore, we are able to demonstrate by CHIP that NFkB binds directly to the Blimp1 promoter. In order to understand why this occurs in MM and bone marrow resident LLPCs but not splenic derived SLPCs, we made use of in silico and genetic approaches to discover how the cells differentially signal through CD28. We demonstrate that the Grb2-Vav binding domain in the cytoplasmic tail of CD28 is critical for its prosurvival signal. Vav is known to bind the major adaptor molecule Slp-76. Using transcriptomic analysis we demonstrate that in humans, the major adaptor molecule Slp-76 is highly expressed in LLPCs but not SLPCs. A major downstream target of Slp-76 is PLC-g1 which is phosphorylated by CD28 activation in LLPCs but not SLPCs. To demonstrate that signals emanating from Slp-76 drive LLPC survival, we made use of Slp-76 mutant mice wherein the Vav binding domain is mutated. These mice had lower LLPC numbers and could not transduce a CD28 prosurvival signal. Furthermore, genetic knockdown of Slp-76 diminished the ability of CD28 to induce Blimp1 upregulation. Altogether these data suggest that CD28, through a Grb2-Vav-Slp-76 signal induces mitochondrial respiration dependent production of ROS. These ROS go on to activate NFkB mediated induction of Blimp1, thereby reinforcing the plasma cell phenotype for survival and function. This knowldedge will augment our ability to create effective vaccines as well as disrupt antibody-mediated autoimmunity and multiple myeloma progression in patients. Disclosures No relevant conflicts of interest to declare.

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BAFF and APRIL as osteoclast-derived survival factors for myeloma cells: a rationale for TACI-Fc treatment in patients with multiple myeloma

Leukemia ◽

10.1038/sj.leu.2404228 ◽

2006 ◽

Vol 20 (7) ◽

pp. 1313-1315 ◽

Cited By ~ 74

Author(s):

M Abe ◽

S Kido ◽

M Hiasa ◽

A Nakano ◽

A Oda ◽

...

Keyword(s):

Multiple Myeloma ◽

Myeloma Cells ◽

Survival Factors

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Erratum: Lenalidomide inhibits osteoclastogenesis, survival factors and bone-remodeling markers in multiple myeloma

Leukemia ◽

10.1038/leu.2008.216 ◽

2008 ◽

Vol 22 (10) ◽

pp. 1973-1973 ◽

Cited By ~ 2

Author(s):

I Breitkreutz ◽

M S Raab ◽

S Vallet ◽

T Hideshima ◽

N Raje ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Remodeling ◽

Survival Factors ◽

Bone Remodeling Markers

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Proteasome inhibitors as therapeutics

Essays in Biochemistry ◽

10.1042/bse0410205 ◽

2005 ◽

Vol 41 ◽

pp. 205-218

Author(s):

Constantine S. Mitsiades ◽

Nicholas Mitsiades ◽

Teru Hideshima ◽

Paul G. Richardson ◽

Kenneth C. Anderson

Keyword(s):

Multiple Myeloma ◽

Drug Class ◽

Proteasome Inhibitors ◽

Cell Cycle Regulators ◽

Current State ◽

Intracellular Mechanism ◽

Ubiquitin Proteasome ◽

Proteasome Pathway ◽

Hodgkin's Lymphomas ◽

Clinical Research Data

The ubiquitin–proteasome pathway is a principle intracellular mechanism for controlled protein degradation and has recently emerged as an attractive target for anticancer therapies, because of the pleiotropic cell-cycle regulators and modulators of apoptosis that are controlled by proteasome function. In this chapter, we review the current state of the field of proteasome inhibitors and their prototypic member, bortezomib, which was recently approved by the U.S. Food and Drug Administration for the treatment of advanced multiple myeloma. Particular emphasis is placed on the pre-clinical research data that became the basis for eventual clinical applications of proteasome inhibitors, an overview of the clinical development of this exciting drug class in multiple myeloma, and a appraisal of possible uses in other haematological malignancies, such non-Hodgkin's lymphomas.

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