XPO1 Inhibitor Research Articles

Selinexor Enhances NK Cell Activation Against Malignant B Cells via Downregulation of HLA-E.

Selinexor is an FDA approved selective inhibitor of the nuclear export protein exportin-1 (XPO1) and causes specific cancer cell death via nuclear accumulation of tumor suppressor proteins. Design of rational studies for the use of selinexor in combination with other therapeutic agents, such as immunotherapies, requires a fundamental understanding of the effects of selinexor on the immune system. One important emerging area of immunotherapy are natural killer (NK) cell based therapeutics. NK cell function is tightly regulated by a balance of signals derived from multiple activating and inhibitory receptors. Thus in cancer, up-regulation of stress ligands recognised by activating receptors or down-regulation of HLA class I recognised by inhibitory receptors can result in an anti-cancer NK cell response. Changes in XPO1 function therefore have the potential to affect NK cell function through shifting this balance. We therefore sought to investigate how selinexor may affect NK cell function. Selinexor pre-treatment of lymphoma cells significantly increased NK cell mediated cytotoxicity against SU-DHL-4, JeKo-1 and Ramos cells, concurrent with increased CD107a and IFNγ expression on NK cells. In addition, selinexor enhanced ADCC against lymphoma cells coated with the anti-CD20 antibodies rituximab and obinutuzumab. In probing the likely mechanism, we identified that XPO1 inhibition significantly reduced the surface expression of HLA-E on lymphoma cell lines and on primary chronic lymphocytic leukemia cells. HLA-E binds the inhibitory receptor NKG2A and in accordance with this, selinexor selectively increased activation of NKG2A+ NK cells. Our data reveals that selinexor, in addition to its direct cytotoxic activity, also activates an anti-cancer immune response via disruption of the inhibitory NKG2A:HLA-E axis.

Inhibition of XPO-1 Mediated Nuclear Export through the Michael-Acceptor Character of Chalcones.

The nuclear export receptor exportin-1 (XPO1, CRM1) mediates the nuclear export of proteins that contain a leucine-rich nuclear export signal (NES) towards the cytoplasm. XPO1 is considered a relevant target in different human diseases, particularly in hematological malignancies, tumor resistance, inflammation, neurodegeneration and viral infections. Thus, its pharmacological inhibition is of significant therapeutic interest. The best inhibitors described so far (leptomycin B and SINE compounds) interact with XPO1 through a covalent interaction with Cys528 located in the NES-binding cleft of XPO1. Based on the well-established feature of chalcone derivatives to react with thiol groups via hetero-Michael addition reactions, we have synthesized two series of chalcones. Their capacity to react with thiol groups was tested by incubation with GSH to afford the hetero-Michael adducts that evolved backwards to the initial chalcone through a retro-Michael reaction, supporting that the covalent interaction with thiols could be reversible. The chalcone derivatives were evaluated in antiproliferative assays against a panel of cancer cell lines and as XPO1 inhibitors, and a good correlation was observed with the results obtained in both assays. Moreover, no inhibition of the cargo export was observed when the two prototype chalcones 9 and 10 were tested against a XPO1-mutated Jurkat cell line (XPO1C528S), highlighting the importance of the Cys at the NES-binding cleft for inhibition. Finally, their interaction at the molecular level at the NES-binding cleft was studied by applying the computational tool CovDock.

Enhanced p53 Activation By Dual Inhibition of MDM2 and XPO1 Disrupts MYC Transcriptional Program and Restores Sensitivity to BCL-2 Inhibition in Ven/HMA Resistant AML

Enhanced p53 Activation By Dual Inhibition of MDM2 and XPO1 Disrupts MYC Transcriptional Program and Restores Sensitivity to BCL-2 Inhibition in Ven/HMA Resistant AML

Anti-tumor activity of the pan-RAF inhibitor TAK-580 in combination with KPT-330 (selinexor) in multiple myeloma.

RAS/RAF/MEK/ERK pathway inhibitors exhibit significant anti-tumor effects against various tumor types, including multiple myeloma (MM), and they are predicted to play a pivotal role in precision medicine. The XPO1 inhibitor KPT-330 has also exhibited promising efficacy in combination with other novel drugs in treating relapsed/refractory MM (RRMM). In this study, we explored the anti-tumor effects of a combination of the pan-RAF inhibitor TAK-580 and KPT-330. Importantly, TAK-580 enhanced KPT-330-induced cytotoxicity and apoptosis in human myeloma cell lines and primary myeloma cells from RRMM patients. Moreover, TAK-580 and KPT-330 synergistically inhibited nuclear phospho-FOXO3a and enhanced cytoplasmic phospho-FOXO3a in MM cells, leading to cytoplasmic enhanced Bim expression and finally apoptosis. This indicates that TAK-580 enhances KPT-330-induced cytotoxicity and apoptosis primarily via the FOXO3a-Bim axis. In addition, TAK-580 enhanced the cytotoxicity of KPT-330 against MM cells even in the presence of IGF-1. Taken together, our results demonstrate that a combination of pan-RAF inhibitor and XPO1 inhibitor is a potential therapeutic option in treating MM.

Effects of Cytogenetic Risk on Outcomes in Multiple Myeloma Treated with Selinexor, Bortezomib, and Dexamethasone (XVd)

Effects of Cytogenetic Risk on Outcomes in Multiple Myeloma Treated with Selinexor, Bortezomib, and Dexamethasone (XVd)

Selinexor Enhances NK Cell Activation Against Lymphoma Cells Via Downregulation of HLA-E

Introduction: Natural killer (NK) cells are powerful immune effectors which induce direct cytotoxicity, promote adaptive immune responses and mediate antibody dependent cellular cytotoxicity (ADCC). Enhancement of NK cell activity against cancer is currently the focus of intense research efforts and strategies include CAR-NK, stimulatory antibodies, cytokines and checkpoint inhibitors.Upregulation of exportin-1 (XPO1) is common in human cancers and high expression is negatively associated with survival in various cancers including diffuse large B cell lymphoma (DLBCL). Targeted inhibition of XPO1 by the selective inhibitor selinexor leads to cancer cell death via accumulation of tumour suppressor proteins in the nucleus, dysregulation of growth regulatory proteins and blockade of oncogene protein translation. The therapeutic efficacy of XPO1 inhibition has led to FDA approval of the oral XPO1 inhibitor selinexor for the treatment of multiple myeloma and DLBCL. The effect of selinexor on NK cell activity has not previously been investigated and was therefore addressed in this study.Methods: The B lymphoma cell lines JeKo-1, SU-DHL-4 and Ramos were incubated with selinexor (50-2000nM) for 18 hours before analysis. Flow cytometry was used to assess cell surface expression of activating and inhibitory ligands for NK cells. For NK based assays, peripheral blood derived NK cells were isolated from healthy donors and incubated with IL-15 (1ng/ml) overnight prior to co-culture with target lymphoma cells for a further 4 hours. Cytotoxicity was assessed using propidium iodide staining of target cells and degranulation of NK cells was assessed by measurement of CD107a. Whole blood samples from colorectal cancer patients (n=11) at pre-treatment and 3 weeks post selinexor monotherapy were assessed by flow cytometry for CD45+CD3-CD19-CD56+ NK cells.Results: Selinexor pre-treatment of target lymphoma cells significantly increased NK cell mediated cytotoxicity against SU-DHL-4 (2.2 Fold increase, p<0.01), JeKo-1 (2 Fold increase, p<0.01) and Ramos (1.7 Fold increase, p<0.01) cells. In accordance with this, selinexor pre-treatment of target cells also increased the activation of NK cells against SU-DHL-4, JeKo-1 and Ramos cells as measured by CD107a expression in both CD56 bright and CD56 dim NK cell sub-groups.To identify the mechanism behind this, we measured expression of activating and inhibitory ligands for NK cells on SU-DHL-4 cells after incubation with selinexor. No significant changes in expression of activating ligands (MICA/B, ULBP-2/5/6, ULBP-1, Vimentin, B7H6, CD54) were evident. In contrast, selinexor significantly (p<0.001) reduced the surface expression of HLA-E on SU-DHL-4 cells by 50%. Selinexor mediated downregulation of HLA-E was also evident in Ramos (60% reduction, p<0.001) and JeKo-1 cells (20% reduction, p<0.01). HLA-E binds the ITIM containing receptor NKG2A, a key inhibitory receptor for NK cells and subsets of T cells. In accordance with this, selinexor pre-treatment of SU-DHL-4 cells selectively increased NKG2A+ NK cell activation (p<0.01) following co-culture.To examine the effect of selinexor on NK cells in patients, we assessed the proportion of NK cells in the peripheral blood of 11 colorectal cancer patients at pre-treatment and three weeks post selinexor monotherapy. % NK cells of CD45+ peripheral blood lymphocytes following treatment with selinexor was increased 2-fold (Median 5% pre-treatment vs 10% post selinexor). In addition, increased abundance of the less mature and less cytotoxic CD56 bright subset of NK cells was associated with poor response to therapy (Median 4% responders (n=3) vs 20% non-responders (n=8)). Larger patient datasets are required to confirm these effects and this analysis is currently ongoing. The effect of selinexor on NK cells in patients with lymphoma is also currently under investigation.Conclusions:The NKG2A:HLA-E axis is a novel immune checkpoint target and our data identifies that selinexor sensitises lymphoma cells to NK cell mediated killing via disruption of this interaction. In addition, we provide initial evidence that NK cells may be associated with clinical response to selinexor. This data indicates that NK cells may contribute to the therapeutic efficacy of selinexor and that selinexor may synergise with NK cell targeted therapies for the treatment of lymphoma. DisclosuresWalker: Karyopharm Therapeutics: Current Employment, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Johnson: Morphosys: Honoraria; Kymera: Honoraria; Kite Pharma: Honoraria; Incyte: Honoraria; Genmab: Honoraria; Celgene: Honoraria; Bristol-Myers: Honoraria; Epizyme: Consultancy, Research Funding; Boehringer Ingelheim: Consultancy; Novartis: Honoraria; Takeda: Honoraria; Oncimmune: Consultancy; Janssen: Consultancy. Cragg: BioInvent International: Consultancy, Research Funding; GSK: Research Funding; UCB: Research Funding; iTeos: Research Funding; Roche: Research Funding. Forconi: Novartis: Honoraria; Roche: Honoraria; Janssen: Consultancy, Honoraria, Speakers Bureau; AbbVie: Consultancy, Honoraria, Speakers Bureau; Gilead: Research Funding. Landesman: Karyopharm Therapeutics: Current Employment, Current equity holder in publicly-traded company. Blunt: Karyopharm Therapeutics: Research Funding.

Venetoclax Combined with Dose-Adjusted R-EPOCH (VR-DA-EPOCH) As Treatment of Richter's Syndrome: A Real-World Study

Background: To evaluate the efficacy and safety of venetoclax, rituximab plus dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin (VR-DA-EPOCH) in Richter's syndrome (RS), we conducted a single-arm, retrospective, observational, real-world study in our center.Methods: Patients who had history of CLL/SLL were diagnosed as RS by biopsy during treatment or watch and wait strategy. VR-DA-EPOCH was given as follow, venetoclax was administered with accelerated ramp-up from 20 mg per day to 400 mg per day, d1-10 during cycle 1, 400 mg daily on day1-10 of cycle 2-6, rituximab 375 mg/m 2 on day 0 of cycle 1 and 500 mg/m 2 on day 0 of cycle 2-6, plus etoposide (50 mg/m 2,day1-4), vincristine (0.4 mg/m 2 day 1-4) or vindesine 3 mg/m 2 day 1 , doxorubicin (10 mg/m 2 day 1-4), prednisone (60 mg/m 2, day 1-5), cyclophosphamide (750 mg/m 2 day 5), 21 days per cycle，dose adjustment on the basis of nadir ANC and platelet count are as previously reported by Wison WH. Response assessment was conducted after 2 or 3 cycles by enhanced CT or PET/CT and after 6 cycles (EOT) by PET/CT according to 2014 Lugano criteria. Minimal residual disease (MRD) of CLL cell in peripheral blood (PB) and bone marrow (BM) was detected after 2 or 3 and 6 cycles by flow cytometry. uMRD was defined as less than 1 CLL cell per 10 4 leukocytes.Results: 7 RS patients were enrolled in Pukou CLL Center from 10/2019 to 7/2021 and the last follow up was 07/25/2021. The median age was 52 years old. Unmutated IGHV, complex karyotype (CK) and TP53 deletion and/or mutation was detected in 100% (6/6), 20% (1/5) and 40% (2/5) patients, respectively. 5 patients received at least one prior line (range: 1-5) treatment for CLL/SLL, with 4 patients received ibrutinib as last prior therapy and one patient previously exposed to venetoclax. 2 patients were diagnosed as RS during watch and wait. The median duration from diagnoses or previous treatment for CLL/SLL to RS was 12 months (range: 3-14). All patients underwent lymph node (n=6) or bone biopsy(n=1) at the site of SUVmax or secondary SUV uptake (unaccessible for SUVmax) by PET/CT and was confirmed as transformed to non-GCB type of diffuse large B-cell lymphoma (DLBCL). Furthermore, 4 of 4 (100%) available patients were confirmed as clonal-related RS by detecting IGHV gene usage. 3 patients acquired CK, and 2 patients appeared BTK C481S mutation. 7 patients completed at least 2 cycles and were available for efficacy and safety assessment. Overall response rate (ORR) was 100% after 2 or 3 cycles, and CR rate (CRR) was 60% after 6 cycles in 5 patients who completed 6 cycles. 2 patients experience disease progression (PD) after cycle 2 and cycle 4 respectively, with one ceased after the addition of brentuximab and the other received CD20 UCAR-T and progressed 3 months later, transit to allo-hemapoietic stem cell transplant (allo-HCT). One patient received auto-hemapoietic stem cell transplant (auto-HCT) and CD19-CAR-T as consolidation and remains in CR, one patient experience PD after 6 cycles and attained CR with chidamide , programmed death-1 (PD-1) inhibitor Sintilimab and XPO1 inhibitor Selinexor, bridging to allo-HCT and remains in CR. Another patient who achieved CR after 6 cycles progressed 6 months later and ceased within one month. 2 patients transformed without previous treatment wait for final evaluation. 60% (3/5) patients attained MRD negativity both in the PB and BM after 6 cycles. The median tolerable dose was 60% (50%-70%) of standard EPOCH and the median dose of venetoclax was 400mg daily for 7 days each cycle. No tumor lysis syndrome (TLS) happened during venetoclax ramp-up. The most common grade 3 or 4 adverse effects (AEs) was agranulocytic fever (6/7, 85.7%), thrombocytopenia (3/7, 42.9%) and sepsis (2/7, 28.6%). 3 (42.9%) patients discontinued venetoclax due to severe AEs and the median duration of discontinuation was 3 days. 85.7% (6/7) had venetoclax dose reduction or interruption due to grade 3 or 4 neutropenia.Conclusion: VR-DA-EPOCH showed high response rate, impressive CR with uMRD and manageable toxicity in patients with RS, even with previous exposure to new target drugs. VR-DA-EPOCH could be recommended as an effective treatment choice for RS, CAR-T or allo-HCT should be considered as subsequent strategy for long term disease control. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Efficacy and Safety of Selinexor-Containing Regimens in Patients with Multiple Myeloma Previously Treated with Anti-CD38 Monoclonal Antibodies (αCD38 mAb)

Efficacy and Safety of Selinexor-Containing Regimens in Patients with Multiple Myeloma Previously Treated with Anti-CD38 Monoclonal Antibodies (αCD38 mAb)

Novel Non-Immunologic Agents for Relapsed and Refractory Multiple Myeloma: A Review Article.

Simple SummaryImmunotherapy-based treatments have brought many new options in the treatment of multiple myeloma; still, the disease will inevitably relapse as it remains incurable. The development of non-immunologic drugs is therefore still needed, particularly for patients with advanced myeloma who become refractory to most of the drugs available, including immunotherapy. Non-immunologic agents have proven effective in the field for the past 20 years, notably with the advent of cytotoxic drugs and, subsequently, targeted therapy. In this review, we summarize the information currently available on novel generations of non-immunologic agents:proteasome inhibitors, immunomodulatory agents, anti-BCL-2/MCL-1, anti-XPO1, peptide–drug conjugates, and targeted drugs in early-stage development.Novel treatments are needed to address the lack of options for patients with relapsed or refractory multiple myeloma. Even though immunotherapy-based treatments have revolutionized the field in recent years, offering new opportunities for patients, there is still no curative therapy. Thus, non-immunologic agents, which have proven effective for decades, are still central to the treatment of multiple myeloma, especially for advanced disease. Building on their efficacy in myeloma, the development of proteasome inhibitors and immunomodulatory drugs has been pursued, and has led to the emergence of a novel generation of agents (e.g., carfilzomib, ixazomib, pomalidomide). The use of alkylating agents is decreasing in most treatment regimens, but melflufen, a peptide-conjugated alkylator with a completely new mechanism of action, offers interesting opportunities. Moreover, with the identification of novel targets, new drug classes have entered the myeloma armamentarium, such as XPO1 inhibitors (selinexor), HDAC inhibitors (panobinostat), and anti-BCL-2 agents (venetoclax). New pathways are still being explored, especially the possibility of a mutation-driven strategy, as biomarkers and targeted treatments are increasing. Though multiple myeloma is still considered incurable, the treatment options are expanding and are progressively becoming more diverse, largely because of the continuous development of non-immunologic agents.

Relapsed/Refractory Multiple Myeloma in 2020/2021 and Beyond.

Simple SummaryDuring the last two decades, collaborative translational studies utilizing novel methodologies have dramatically advanced our understanding of multiple myeloma (MM) pathophysiology and revolutionized derived treatment strategies. Nevertheless, MM remains an incurable disease, with the vast majority of patients relapsing due to evolving genetic alterations within tumor cell clones as well as the pressure of the immunosuppressive bone marrow (BM) microenvironment. Therefore, continuous translational efforts are needed to further increase our understanding of MM biology, develop rationally derived drugs and to thereby improve patient outcome.Despite the challenges imposed by the COVID-19 pandemic, exciting therapeutic progress continues to be made in MM. New drug approvals for relapsed/refractory (RR)MM in 2020/2021 include the second CD38 monoclonal antibody, isatuximab, the first BCMA-targeting therapy and first-in-class antibody–drug conjugate (ADC) belantamab mafodotin, the first BCMA-targeting CAR T cell product Idecabtagen-Vicleucel (bb2121, Ide-Cel), the first in-class XPO-1 inhibitor selinexor, as well as the first-in-class anti-tumor peptide-drug conjugate, melflufen. The present introductory article of the Special Issue on “Advances in the Treatment of Relapsed and Refractory Multiple Myeloma: Novel Agents, Immunotherapies and Beyond” summarizes the most recent registration trials and emerging immunotherapies in RRMM, gives an overview on latest insights on MM genomics and on tumor-induced changes within the MM microenvironment, and presents some of the most promising rationally derived future therapeutic strategies.

Identification of nuclear export inhibitor-based combination therapies in preclinical models of triple-negative breast cancer

An estimated 284,000 Americans will be diagnosed with breast cancer in 2021. Of these individuals, 15–20% have basal-like triple-negative breast cancer (TNBC), which is known to be highly metastatic. Chemotherapy is standard of care for TNBC patients, but chemoresistance is a common clinical problem. There is currently a lack of alternative, targeted treatment strategies for TNBC; this study sought to identify novel therapeutic combinations to treat basal-like TNBCs. For these studies, four human basal-like TNBC cell lines were utilized to determine the cytotoxicity profile of 1363 clinically-used drugs. Ten promising therapeutic candidates were identified, and synergism studies were performed in vitro. Two drug combinations that included KPT-330, an XPO1 inhibitor, were synergistic in all four cell lines. In vivo testing of four basal-like patient-derived xenografts (PDX) identified one combination, KPT-330 and GSK2126458 (a PI3K/mTOR inhibitor), that decreased tumor burden in mice significantly more than monotherapy with either single agent. Bulk and single-cell RNA-sequencing, immunohistochemistry, and analysis of published genomic datasets found that XPO1 was abundantly expressed in human basal-like TNBC cell lines, PDXs, and patient tumor samples. Within basal-like PDXs, XPO1 overexpression was associated with increased proliferation at the cellular level. Within patient datasets, XPO1 overexpression was correlated with greater rates of metastasis in patients with basal-like tumors. These studies identify a promising potential new combination therapy for patients with basal-like breast cancer.

2MO XPO1 inhibition strongly sensitizes to first-line and second-line therapy in small cell lung cancer

Small cell lung cancer (SCLC) is an exceptionally aggressive disease comprising 13% of all lung cancer cases. With limited treatment options that typically result in transient responses, SCLC is responsible for approximately 250,000 deaths globally per year. The recent addition of immunotherapy to first-line platinum-based doublet chemotherapy shows only limited benefit in a small subset of patients. Major hurdles to improving SCLC treatment include development of rapid chemoresistance and ineffective second-line therapies.

Reversible disruption of XPO1-mediated nuclear export inhibits respiratory syncytial virus (RSV) replication

Respiratory syncytial virus (RSV) is the primary cause of serious lower respiratory tract disease in infants, young children, the elderly and immunocompromised individuals. Therapy for RSV infections is limited to high risk infants and there are no safe and efficacious vaccines. Matrix (M) protein is a major RSV structural protein with a key role in virus assembly. Interestingly, M is localised to the nucleus early in infection and its export into the cytoplasm by the nuclear exporter, exportin-1 (XPO1) is essential for RSV assembly. We have shown previously that chemical inhibition of XPO1 function results in reduced RSV replication. In this study, we have investigated the anti-RSV efficacy of Selective Inhibitor of Nuclear Export (SINE) compounds, KPT-335 and KPT-185. Our data shows that therapeutic administration of the SINE compounds results in reduced RSV titre in human respiratory epithelial cell culture. Within 24 h of treatment, RSV replication and XPO1 expression was reduced, M protein was partially retained in the nucleus, and cell cycle progression was delayed. Notably, the effect of SINE compounds was reversible within 24 h after their removal. Our data show that reversible inhibition of XPO1 can disrupt RSV replication by affecting downstream pathways regulated by the nuclear exporter.

XPO1/CRM1 is a promising prognostic indicator for neuroblastoma and represented a therapeutic target by selective inhibitor verdinexor

BackgroundHigh-risk neuroblastoma patients have a 5-year survival rate of less than 50%. It’s an urgent need to identify new therapeutic targets and the appropriate drugs. Exportin-1 (XPO1), also known as chromosomal region maintenance 1, plays important roles in the progression of tumorigenesis. However, the prognostic and therapeutic values of XPO1 in neuroblastoma have not been reported.MethodsCorrelations between XPO1 expression level and clinical characteristics were analyzed using the Neuroblastoma Research Consortium (NRC) dataset and tissue microarray analysis. Cell proliferation assays, colony formation assays, apoptosis assays, cell cycle analysis were performed to analyze the anti-tumor effects of verdinexor (KPT-335) in vitro. Western blot and mRNA sequencing were performed to explore underlying mechanism. In vivo anti-tumor effects of verdinexor were studied in a neuroblastoma xenograft model.ResultsHigher XPO1 levels were associated with advanced stage and poor prognosis in neuroblastoma patients. The specific inhibitor of XPO1 verdinexor suppressed the neuroblastoma cell growth both in vitro and in vivo. Specifically, inhibition of XPO1 suppressed the neuroblastoma cell proliferation and induced cell apoptosis by nuclear accumulation of FOXO1 and RB1 in the neuroblastoma due to the inhibition of the PI3K/AKT pathway, and induced G0/G1 phase cell cycle arrest by activation of P53 function.ConclusionsXPO1 is a promising prognostic indicator for neuroblastoma and a novel target for antitumor treatment with selective inhibitor verdinexor.

Novel Approaches to Treating Relapsed and Refractory Multiple Myeloma with a Focus on Recent Approvals of Belantamab Mafodotin and Selinexor.

Though survival outcomes in multiple myeloma patients have improved drastically over the past few decades, there still remains an ongoing need for effective and tolerable treatment options in the relapsed and refractory space. Encouragingly, there have been three recent FDA approvals for triple-class refractory multiple myeloma, and there is promising ongoing development of additional agents with varying novel mechanisms of action. Here, we will review the most recent data on both belantamab mafodotin, an antibody drug conjugate (ADC) targeting BCMA, and selinexor, a first-in-class selective inhibitor of XPO1, as well as touch on some of the recently published data for other immunotherapies in development, namely bispecific T cell engagers, ADCs, and CAR-T cell therapies.

Antitumor efficacy of XPO1 inhibitor Selinexor in KRAS-mutant lung adenocarcinoma patient-derived xenografts

Gain-of-function Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations occur in 25% of lung adenocarcinomas, and these tumors are challenging to treat. Some preclinical work, largely based on cell lines, suggested KRASmut lung cancers are especially dependent on the nuclear export protein exportin-1 (XPO1), while other work supports XPO1 being a broader cancer dependency. To investigate the sensitivity of KRASmut lung cancers to XPO1 inhibition in models that more closely match clinical tumors, we treated 10 independently established lung cancer patient-derived tumor xenografts (PDXs) with the clinical XPO1 inhibitor, Selinexor. Monotherapy with Selinexor reduced tumor growth in all KRASmut PDXs, which included 4 different codon mutations, and was more effective than the clinical MEK1/2 inhibitor, Trametinib. Selinexor was equally effective in KRASG12C and KRASG12D tumors, with TP53 mutations being a biomarker for a weaker drug response. By mining genome-wide dropout datasets, we identified XPO1 as a universal cancer cell dependency and confirmed this functionally in two KRASWT PDX models harboring kinase drivers. However, targeted kinase inhibitors were more effective than Selinexor in these models. Our findings support continued investigation of XPO1 inhibitors in KRASmut lung adenocarcinoma, regardless of the codon alteration.

Anti Tumor Activity Of Selinexor (KPT-330), A First-In-Class Oral Selective Inhibitor Of Nuclear Export (SINE) XPO1/CRM1 Antagonist In Patients (pts) With Relapsed / Refractory Multiple Myeloma (MM) Or Waldenstrom's Macroglobulinemia (WM)

BackgroundExportin 1 (XPO1/CRM1) is overexpressed in MM and was identified as an essential protein for MM cell growth. The majority of Tumor Suppressor Proteins (TSP) are transported out of the nucleus exclusively by XPO1, leading to their inactivation. Selinexor (KPT-330) is a potent, selective oral inhibitor of XPO1 and shows potent anti myeloma activity in preclinical models. MethodsPatients (pts) with advanced, relapsed/refractory MM or WM were dosed with oral Selinexor (8-10 doses / 4-week cycle) as part of a broad Phase 1 program (NCT 01607892) in advanced hematological malignancies. Detailed pharmacokinetic (PK) and pharmacodynamic (PDn) analyses and tumor biopsies on selected patients were performed. Response evaluation was performed every cycle. All pts in this study had documented progressive disease on study entry and were relapsed/refractory to at least one proteasome inhibitor and one immunomodulating agent. Results17 MM and 2 WM pts with median age 67yrs (range 50-75); ECOG PS 0/1: 6/13; median number of prior regimens: 5 [range 1-13], received Selinexor across 6 dose levels (3 to 30 mg/m2). Five patients experienced drug-related grade 3/4 Adverse Events (AEs), including thrombocytopenia without bleeding (n=4), neutropenia (n=4), impaired renal function (n=1), decreased WBC (n=1) and febrile neutropenia (n=1). The most common grade 1/2 toxicities are gastrointestinal (GI) including nausea (16/19; 84%), anorexia (11/19pts; 58%), vomiting (8/19; 42%), diarrhea (7/19pts; 37%), weight loss (4/19; 21%) and dysgeusia (4/19; 21%). Grade 1/2 study drug related fatigue was also observed in 10/19 or 53% of the patients. No grade ≥3 GI related AEs were observed. These side effects were well managed with supportive care. No clinically significant cumulative drug toxicities have been noted and patients have remained on therapy for >8 months (median duration on therapy 50 days, range 8-274 days). Two pts died during the study, one due to E.coli sepsis and the other due to renal dysfunction, both events deemed by treating investigators to be unrelated to study drug. PK analysis demonstrated total exposure increased with increasing dose, with no accumulation and without affecting half-life (5-7 hrs) or clearance of KPT-330. At 23 mg/m2, exposure (Cmax 289 ng/mL and AUC0-inf 2219 ng*h/mL) was comparable to anti tumor exposure observed in mice and dogs. Significant increases (2-20x) in leukocyte XPO1 mRNA levels (PDn marker) were observed at all doses, with higher doses demonstrating higher levels of XPO1 mRNA. Response was evaluable in 15 MM pts: Partial response (PR) in 1 pt (6.7%) at 35mg/m2, Minimal Response (MR) in 6 pts (40%) at doses of 16.8 to 30mg/m2, Stable Disease (SD) in 5 pts (33%) and Progressive Disease (PD) in 3 pts (20%). One MR and 1 SD were observed in the 2 WM pts. Evaluation of serial bone marrow samples from one patient confirms Selinexor-induced nuclear localization of multiple TSPs as well as reduction in CD138+ MM cells. Dose escalation is ongoing at 35 mg/m2 twice weekly. ConclusionsOral Selinexor treatment is generally well tolerated, with favorable PK and PDn properties. Prolonged disease control and responses are observed in heavily pretreated patients with progressive MM whose disease is relapsed or refractory to available agents. Disclosures:Chen:Johnson & Johnson: Consultancy, Research Funding; Lundbeck: Consultancy; Celgene: Consultancy, Research Funding; GlaxoSmithKline: Research Funding; Roche: Honoraria. Off Label Use: Experimental use of Selinexor, a drug not yet approved. Baz:Celgene, Millennium, BMS, Novartis, Karyopharm, Sanofi: Research Funding. Reece:Janssen: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Millennium Pharmaceuticals: Research Funding; Novartis: Honoraria, Research Funding; Merck: Honoraria, Research Funding; BMS: Research Funding; Otsuka: Honoraria, Research Funding; Onyx: Consultancy. Siegel:Celgene: Honoraria, Speakers Bureau; Millennium: Honoraria, Speakers Bureau; Onyx: Honoraria, Speakers Bureau. Kuruvilla:Seattle Genetics : Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria, Research Funding; Janssen: Honoraria; Celgene: Consultancy, Honoraria; Lundbeck: Consultancy, Honoraria; Karyopharm: Research Funding. Shacham:Karyopharm Therapeutics Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Rashal:Karyopharm Therapeutics Inc.: Employment, Equity Ownership. McCauley:Karyopharm Therapeutics: Employment, Equity Ownership. Saint-Martin:Karyopharm Therapeutics Inc.: Employment, Equity Ownership. McCartney:Karyopharm Therapeutics Inc.: Employment, Equity Ownership. Landesman:Karyopharm Therapeutics: Employment, Equity Ownership, Patents & Royalties. Klebanov:Karyopharm Therapeutics: Employment, Equity Ownership. Pond:Osmozis : Consultancy. Kauffman:Karyopharm Therapeutics Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Mirza:Karyopharm Therapeutics Inc.: Consultancy, Equity Ownership.

Abstract 2383: Mir-7974: An oncogenic miRNA that perpetuates gastric cancer

Abstract The global incidence of gastric cancer exceeds 1 million people per year and has a 5-year survival rate of 31%. Current chemotherapies are not effective because of genetic and epigenetic heterogeneity and late stage detection. Previously, we showed that exportin 1 (XPO1) is a therapeutic target in gastric cancer, as it facilitates the nuclear export of various cancer related tumor suppressor proteins (TSPs) through direct interaction with their nuclear export sequences (NES). XPO1 is upregulated in a variety of tumors including gastric cancer, leading to increased cytoplasmic localization of TSPs supporting malignant transformation and cancer cell proliferation, Knockdown of XPO1 leads to anti-tumor effects and pharmacologic targeting of XPO1 is achievable with selective inhibitors of nuclear export (SINE) compounds. In 2019 and 2020, the first generation XPO1 inhibitor KPT-330 was FDA approved for the use in penta-refractory multiple myeloma and non-Hodgkin's Lymphoma. Besides protein, mRNA and non-coding RNAs are also transported across the nuclear membrane. Non-coding RNA transport is generally attributed to another nuclear export protein, exportin 5 (XPO5), which plays a critical role in microRNA (miRNA) biogenesis. Nevertheless, there is emerging evidence supporting the role of XPO1 in non-coding RNA transport. In order to elucidate the role of XPO1 in non-coding RNA biology, we performed small RNA sequencing on gastric cancer cell lines treated with the XPO1 inhibitor selinexor. Our results revealed statistically significant changes in miR-7974 and miR-129-1-3p. These two miRs are recognized to influence protein modifications and cell survival. Administration of a miR-7974 mimic and a miR-129-1-3p inhibitor reduced proliferation of gastric cancer cells. Next, we evaluated the miR target genes of interest. At the mRNA level, we found upregulation of RasGEF Domain Family Member 1a (RASGEF1A), downregulation of DNA damage-regulated autophagy modulator 1 (DRAM1), downregulation of TIMP2 and downregulation of Cyclin G2 protein encoding gene CCNG2. Western Blots confirmed inhibition of AKT, RAS and Wnt signaling. Our results also showed that miR-7974 overexpression correlated with cellular XPO1 levels through the enhancement of XPO1 and its cargoes RCC1 and RANBP2. Finally inhibition of XPO1 by SINE compounds reversed the observed oncogenic phenotypes caused by these microRNAs. Our results unveil a novel interaction network between XPO1 and small non-coding RNAs and show that XPO1 induced small-noncoding microRNAs influence gastric cancer proliferation, and can be reversed by SINE compounds. This implies that modulation of microRNAs might be explored as a biomarker for XPO1 inhibitor therapy and warrants further investigation. Citation Format: Rachel E. Sexton, Mohammad Najeeb Al-Hallak, Bayan Al-Share, Yiwei Li, Amro Aboukameel, Yosef Landesman, Trinayan Kashyap, Steve Kim, Ramzi M. Mohammad, Philip A. Philip, Asfar S. Azmi. Mir-7974: An oncogenic miRNA that perpetuates gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2383.

Abstract 994: Investigating inhibition of nuclear export in breast cancer cells

Abstract XPO1 (Exportin aka CRM1) is a mediator of nuclear export that is known to be dysregulated in cancers. Selective inhibitors of nuclear export (SINEs) have shown some promise in cancer treatment, but the mechanism is poorly understood in cancer specific subtypes. Here, we investigated XPO1 inhibition in breast cancer by treating a panel of breast cancer cell lines with a SINE (Selinexor) and saw a bimodal response, with most triple-negative cell lines showing sensitivity and some ER+ cell lines showing resistance to the therapy. Changes in the expression of BIRC5 (Survivin) have been previously implicated in XPO1 inhibitor response. Using RT-qPCR experiments we were able to show that after 24 hours of selinexor treatment, the expression of BIRC5 does change in two sensitive cell lines MDA-MB-231 and MCF7, but is not necessarily changed at earlier time points. To more holistically explore the mechanism of XPO1 inhibition induced cell death, we performed RNA-Seq and differential gene expression analysis on these two cell lines at 2, 8, and 24 hours post treatment with Selinexor (compared to vehicle). Our results indicate that several breast cancer and cell cycle associated genes show expression changes as early as 2 hours post-treatment and showed-consistent changes across the time-points. Overall, our analysis sheds light on the mechanism by which XPO1 inhibition induces cell death in breast cancer, which may help elucidate potential biomarkers as well as to nominate rationale combinations to explore with XPO1 inhibition. Citation Format: Robert F. Gruener, Geoffrey L. Greene, R. Stephanie Huang. Investigating inhibition of nuclear export in breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 994.

Abstract 1058: Inhibition of nuclear transport protein XPO1 potentiates the effect of KRASG12C inhibitors

Abstract Introduction: The discovery of compounds that can bind covalently to KRASG12C and lock it in its inactive GDP-bound conformation has opened a window of opportunity to selectively target KRASG12C. Such agents have shown promising results in preclinical tumor models and recently in clinical trials as well. However, factors such as feedback reactivation or bypass of KRAS dependence are known to limit antitumor activity of KRAS inhibitors. This necessitates the need for combination approaches that can potentially sensitize tumors to KRAS inhibitors when co-targeted. In this study, we have tested two potent KRASG12C inhibitors, MRTX1257 and AMG510, on PDAC and NSCLC cell lines both as single agents and also in combination with nuclear transport inhibitor, Selinexor (KPT-330/XPOVIO). Methods: Cell growth inhibition was determined by MTT assay. Drug synergy analysis was performed and the isobolograms were generated using CalcuSyn 2.1 software (Biosoft, Cambridge, UK). Spheroid disintegration assay, clonogenic assay, immunofluorescence assay, and Western blotting were performed by standard methods. Results: MRTX1257 and AMG510, when combined with KPT-330, synergistically inhibited (Combination Index values less than 1) the proliferation of KRASG12C mutant cancer cell lines, namely MiaPaCa-2 (PDAC) and NCI-H2122 (NSCLC). Such an effect was observed neither in KRASWT (NCI-H1650) nor KRASG12D mutant (Panc-1) cells, suggesting that the synergistic effect on cell viability was specific to KRASG12C. Combined treatment of KPT-330 with either MRTX1257 or AMG510 resulted in enhanced disintegration of MiaPaCa-2 and NCI-H2122 spheroids, indicating the efficacy of the drug combinations in a 3-D cell growth model. In addition, similar synergistic action was also seen in a clonogenic assay, where MRTX1257 and AMG510 reduced the number and size of colonies formed by MiaPaCa-2 cells and KPT-330 further augmented this effect. KPT-330 also potentiated the MRTX1257- or AMG510-induced inhibition of KRAS-dependent signaling targets such as ERK1/2 phosphorylation. Mechanistically, we observed that KPT-330 caused nuclear retention of ETS repressor factor (ERF), a transcriptional repressor that is exported upon phosphorylation by ERK. Given that XPO1 transports hundreds of proteins, we are currently evaluating the global changes in RAS pathway proteins using high throughput approaches. Such studies are anticipated to unveil the molecular underpinnings of the effectiveness of this co-targeting approach that would help advance this combination of nuclear transport inhibitor with KRASG12C inhibitors to be tested in the clinic. Conclusion: In conclusion, we have demonstrated that the inhibitor of nuclear transport protein XPO1 has the ability to potentiate the anticancer activity of KRASG12C inhibitors in in vitro preclinical models of PDAC and NSCLC. Citation Format: Husain Y. Khan, Md. Hafiz Uddin, Yue Zhang, Yosef Landesman, Ammar Sukari, Misako Nagasaka, Asfar S. Azmi. Inhibition of nuclear transport protein XPO1 potentiates the effect of KRASG12C inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1058.