kpt-276 and selinexor

Selective Inhibitor of Nuclear Export (SINE) compounds are a family of small-molecules that inhibit nuclear export through covalent binding to cysteine 528 (Cys528) in the cargo-binding pocket of Exportin 1 (XPO1/CRM1) and promote cancer cell death. Selinexor is the lead SINE compound currently in phase I and II clinical trials for advanced solid and hematological malignancies. In an effort to understand selinexor-XPO1 interaction and to establish whether cancer cell response is a function of drug-target engagement, we developed a quantitative XPO1 occupancy assay. Biotinylated leptomycin B (b-LMB) was utilized as a tool compound to measure SINE-free XPO1. Binding to XPO1 was quantitated from SINE compound treated adherent and suspension cells in vitro, dosed ex vivo human peripheral blood mononuclear cells (PBMCs), and PBMCs from mice dosed orally with drug in vivo. Evaluation of a panel of selinexor sensitive and resistant cell lines revealed that resistance was not attributed to XPO1 occupancy by selinexor. Administration of a single dose of selinexor bound XPO1 for minimally 72 hours both in vitro and in vivo. While XPO1 inhibition directly correlates with selinexor pharmacokinetics, the biological outcome of this inhibition depends on modulation of pathways downstream of XPO1, which ultimately determines cancer cell responsiveness.

Topics: Acrylamides; Acrylates; Active Transport, Cell Nucleus; Animals; Antibiotics, Antineoplastic; Biotinylation; Cell Line, Tumor; Cell Nucleus; Cell Survival; Cells, Cultured; Drug Evaluation, Preclinical; Exportin 1 Protein; Fatty Acids, Unsaturated; HCT116 Cells; Humans; Hydrazines; Karyopherins; Leukocytes, Mononuclear; Mice; Molecular Structure; Receptors, Cytoplasmic and Nuclear; Reproducibility of Results; Thiazoles; Triazoles

In previous studies we demonstrated that targeting the nuclear exporter protein exportin-1 (CRM1/XPO1) by a selective inhibitor of nuclear export (SINE) compound is a viable therapeutic strategy against Non-Hodgkin Lymphoma (NHL). Our studies along with pre-clinical work from others led to the evaluation of the lead SINE compound, selinexor, in a phase 1 trial in patients with CLL or NHL (NCT02303392). Continuing our previous work, we studied combinations of selinexor-dexamethasone (DEX) and selinexor-everolimus (EVER) in NHL. Combination of selinexor with DEX or EVER resulted in enhanced cytotoxicity in WSU-DLCL2 and WSU-FSCCL cells which was consistent with enhanced apoptosis. Molecular analysis showed enhancement in the activation of apoptotic signaling and down-regulation of XPO1. This enhancement is consistent with the mechanism of action of these drugs in that both selinexor and DEX antagonize NF-κB (p65) and mTOR (EVER target) is an XPO1 cargo protein. SINE compounds, KPT-251 and KPT-276, showed activities similar to CHOP (cyclophosphamide-hydroxydaunorubicin-oncovin-prednisone) regimen in subcutaneous and disseminated NHL xenograft models in vivo. In both animal models the anti-lymphoma activity of selinexor is enhanced through combination with DEX or EVER. The in vivo activity of selinexor and related SINE compounds relative to 'standard of care' treatment is consistent with the objective responses observed in Phase I NHL patients treated with selinexor. Our pre-clinical data provide a rational basis for testing these combinations in Phase II NHL trials.

Topics: Acrylamides; Active Transport, Cell Nucleus; Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Cell Line, Tumor; Cell Survival; Cyclophosphamide; Dexamethasone; Dose-Response Relationship, Drug; Doxorubicin; Drug Synergism; Everolimus; Exportin 1 Protein; Humans; Hydrazines; Karyopherins; Lymphoma, Non-Hodgkin; Mice, Inbred ICR; Mice, SCID; Oxadiazoles; Prednisone; Protein Kinase Inhibitors; Receptors, Cytoplasmic and Nuclear; Signal Transduction; Thiazoles; Time Factors; TOR Serine-Threonine Kinases; Transcription Factor RelA; Triazoles; Tumor Burden; Vincristine; Xenograft Model Antitumor Assays

Glioblastoma (GBM) is poorly responsive to current chemotherapy. The nuclear transporter exportin 1 (XPO1, CRM1) is often highly expressed in GBM, which may portend a poor prognosis. Here, we determine the efficacy of novel selective inhibitors of nuclear export (SINE) specific to XPO1 in preclinical models of GBM.. Seven patient-derived GBM lines were treated with 3 SINE compounds (KPT-251, KPT-276, and Selinexor) in neurosphere culture conditions. KPT-276 and Selinexor were also evaluated in a murine orthotopic patient-derived xenograft (PDX) model of GBM. Cell cycle effects were assayed by flow cytometry in vitro and immunohistochemistry in vivo. Apoptosis was determined by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and caspase 3/7 activity assays.. Treatment of GBM neurosphere cultures with KPT-276, Selinexor, and KPT-251 revealed dose-responsive growth inhibition in all 7 GBM lines [range of half-maximal inhibitory concentration (IC50), 6-354 nM]. In an orthotopic PDX model, treatment with KPT-276 and Selinexor demonstrated pharmacodynamic efficacy, significantly suppressed tumor growth, and prolonged animal survival. Cellular proliferation was not altered with SINE treatment. Instead, induction of apoptosis was apparent both in vitro and in vivo with SINE treatment, without overt evidence of neurotoxicity.. SINE compounds show preclinical efficacy utilizing in vitro and in vivo models of GBM, with induction of apoptosis as the mechanism of action. Selinexor is now in early clinical trials in solid and hematological malignancies. Based on these preclinical data and excellent brain penetration, we have initiated clinical trials of Selinexor in patients with relapsed GBM.

Topics: Acrylamides; Active Transport, Cell Nucleus; Animals; Antineoplastic Agents; Apoptosis; Brain Neoplasms; Cell Cycle Checkpoints; Cell Differentiation; Cell Line, Tumor; Dose-Response Relationship, Drug; Exportin 1 Protein; Glioblastoma; Humans; Hydrazines; Karyopherins; Macaca fascicularis; Male; Mice; Oxadiazoles; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; Thiazoles; Treatment Outcome; Triazoles; Xenograft Model Antitumor Assays

Survivin, which is highly expressed and promotes cell survival in diffuse malignant peritoneal mesothelioma (DMPM), exclusively relies on exportin 1 (XPO1/CRM1) to be shuttled into the cytoplasm and perform its anti-apoptotic function. Here, we explored the efficacy of Selective Inhibitors of Nuclear Export (SINE), KPT-251, KPT-276 and the orally available, clinical stage KPT-330 (selinexor), in DMPM preclinical models. Exposure to SINE induced dose-dependent inhibition of cell growth, cell cycle arrest at G1-phase and caspase-dependent apoptosis, which were consequent to a decrease of XPO1/CRM1 protein levels and the concomitant nuclear accumulation of its cargo proteins p53 and CDKN1a. Cell exposure to SINE led to a time-dependent reduction of cytoplasmic survivin levels. In addition, after an initial accumulation, the nuclear protein abundance progressively decreased, as a consequence of an enhanced ubiquitination and proteasome-dependent degradation. SINE and the survivin inhibitor YM155 synergistically cooperated in reducing DMPM cell proliferation. Most importantly, orally administered SINE caused a significant anti-tumor effect in subcutaneous and orthotopic DMPM xenografts without appreciable toxicity. Overall, we have demonstrated a marked efficacy of SINE in DMPM preclinical models that may relay on the interference with survivin intracellular distribution and function. Our study suggests SINE-mediated XPO1/CRM1 inhibition as a novel therapeutic option for DMPM.

Topics: Acrylamides; Active Transport, Cell Nucleus; Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cyclin-Dependent Kinase Inhibitor p21; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Exportin 1 Protein; G1 Phase Cell Cycle Checkpoints; Humans; Hydrazines; Inhibitor of Apoptosis Proteins; Karyopherins; Lung Neoplasms; Mesothelioma; Mesothelioma, Malignant; Mice; Mice, SCID; Neoplasm Proteins; Oxadiazoles; Peritoneal Neoplasms; Real-Time Polymerase Chain Reaction; Receptors, Cytoplasmic and Nuclear; Survivin; Thiazoles; Triazoles; Tumor Suppressor Protein p53