The effect of selinexor on prostaglandin synthesis in virus-positive Merkel cell carcinoma cell lines.

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine skin cancer with high rates of metastasis and mortality. In vitro studies suggest that selinexor (KPT-330), an inhibitor of exportin 1, may be a targeted therapeutic option for MCC. This selective inhibitor prevents the transport of oncogenic mRNA out of the nucleus. Of note, 80% of MCC tumors are integrated with Merkel cell polyomavirus (MCPyV), and virally encoded tumor-antigens, small T (sT) and large T (LT) mRNAs may require an exportin transporter to relocate to the cytoplasm and modulate host tumor-suppressing pathways. To explore selinexor as a targeted therapy for MCC, we examine its ability to inhibit LT and sT antigen expression in vitro and its impact on the prostaglandin synthesis pathway. Protein expression was determined through immunoblotting and quantified by densitometric analysis. Statistical significance was determined with t-test. Treatment of MCPyV-infected cell lines with selinexor resulted in a significant dose-dependent downregulation of key mediators of the prostaglandin synthesis pathway. Given the role of prostaglandin synthesis pathway in MCC, our findings suggest that selinexor, alone or in combination with immunotherapy, could be a promising treatment for MCPyV-infected MCC patients who are resistant to chemotherapy and immunotherapy.

The nuclear export protein exportin-1 in solid malignant tumours: From biology to clinical trials.

BACKGROUND: Exportin-1 (XPO1), a crucial protein regulating nuclear-cytoplasmic transport, is frequently overexpressed in various cancers, driving tumor progression and drug resistance. This makes XPO1 an attractive therapeutic target. Over the past few decades, the number of available nuclear export-selective inhibitors has been increasing. Only KPT-330 (selinexor) has been successfully used for treating haematological malignancies, and KPT-8602 (eltanexor) has been used for treating haematologic tumours in clinical trials. However, the use of nuclear export-selective inhibitors for the inhibition of XPO1 expression has yet to be thoroughly investigated in clinical studies and therapeutic outcomes for solid tumours. METHODS: We collected numerous literatures to explain the efficacy of XPO1 Inhibitors in preclinical and clinical studies of a wide range of solid tumours. RESULTS: In this review, we focus on the nuclear export function of XPO1 and results from clinical trials of its inhibitors in solid malignant tumours. We summarized the mechanism of action and therapeutic potential of XPO1 inhibitors, as well as adverse effects and response biomarkers. CONCLUSION: XPO1 inhibition has emerged as a promising therapeutic strategy in the fight against cancer, offering a novel approach to targeting tumorigenic processes and overcoming drug resistance. SINE compounds have demonstrated efficacy in a wide range of solid tumours, and ongoing research is focused on optimizing their use, identifying response biomarkers, and developing effective combination therapies. KEY POINTS: Exportin-1 (XPO1) plays a critical role in mediating nucleocytoplasmic transport and cell cycle. XPO1 dysfunction promotes tumourigenesis and drug resistance within solid tumours. The therapeutic potential and ongoing researches on XPO1 inhibitors in the treatment of solid tumours. Additional researches are essential to address safety concerns and identify biomarkers for predicting patient response to XPO1 inhibitors.

Adverse events reporting of XPO1 inhibitor - selinexor: a real-word analysis from FAERS database.

As the world's first oral nuclear export inhibitor, selinexor is increasingly being used in clinical applications for malignant tumors. However, there is no extensive exploration on selinexor's adverse events (ADEs), necessitating a real-word assessment of its clinical medication safety. FAERS data (July 2019-June 2023) were searched for selinexor ADE reports across all indications. Use the system organ class (SOC) and preferred terms (PT) from the medical dictionary for regulatory activities (MedDRA) to describe, categorize, and statistic ADEs. Disproportionality analysis was employed through calculation of reporting odds ratio (ROR) and proportional reporting ratio (PRR). Based on total of 4392 selinexor related ADE reports as the primary suspect (PS), of which 2595 instances were severe outcomes. The predominant ADEs included gastrointestinal disorders, myelosuppression symptoms, and various nonspecific manifestations. 124 signals associated with selinexor ADE were detected, and 10 of these top 15 signals were not included into the instructions. Our study provides real-world evidence regarding the drug safety of selinexor, which is crucial for clinicians to safeguard patients' health.

A first-in-class inhibitor of HSP110 to potentiate XPO1-targeted therapy in primary mediastinal B-cell lymphoma and classical Hodgkin lymphoma.

BACKGROUND: Primary mediastinal B-cell lymphoma (PMBL) and classical Hodgkin lymphoma (cHL) are distinct hematological malignancies of B-cell origin that share many biological, molecular, and clinical characteristics. In particular, the JAK/STAT signaling pathway is a driver of tumor development due to multiple recurrent mutations, particularly in STAT6. Furthermore, the XPO1 gene that encodes exportin 1 (XPO1) shows a frequent point mutation (E571K) resulting in an altered export of hundreds of cargo proteins, which may impact the success of future therapies in PMBL and cHL. Therefore, targeted therapies have been envisioned for these signaling pathways and mutations. METHODS: To identify novel molecular targets that could overcome the treatment resistance that occurs in PMBL and cHL patients, we have explored the efficacy of a first-in-class HSP110 inhibitor (iHSP110-33) alone and in combination with selinexor, a XPO1 specific inhibitor, both in vitro and in vivo. RESULTS: We show that iHSP110-33 decreased the survival of several PMBL and cHL cell lines and the size of tumor xenografts. We demonstrate that HSP110 is a cargo of XPO1wt as well as of XPO1E571K. Using immunoprecipitation, proximity ligation, thermophoresis and kinase assays, we showed that HSP110 directly interacts with STAT6 and favors its phosphorylation. The combination of iHSP110-33 and selinexor induces a synergistic reduction of STAT6 phosphorylation and of lymphoma cell growth in vitro and in vivo. In biopsies from PMBL patients, we show a correlation between HSP110 and STAT6 phosphorylation levels. CONCLUSIONS: These findings suggest that HSP110 could be proposed as a novel target in PMBL and cHL therapy.

XPO1 blockade with KPT-330 promotes apoptosis in cutaneous T-cell lymphoma by activating the p53-p21 and p27 pathways.

Dysregulated nuclear-cytoplasmic trafficking has been shown to play a role in oncogenesis in several types of solid tumors and hematological malignancies. Exportin 1 (XPO1) is responsible for the nuclear export of several proteins and RNA species, mainly tumor suppressors. KPT-330, a small molecule inhibitor of XPO1, is approved for treating relapsed multiple myeloma and diffuse large B-cell lymphoma. Cutaneous T-cell lymphoma (CTCL) is an extranodal non-Hodgkin lymphoma with an adverse prognosis and limited treatment options in advanced stages. The effect of therapeutically targeting XPO1 with KPT-330 in CTCL has not been established. We report that XPO1 expression is upregulated in CTCL cells. KPT-330 reduces cell proliferation, induces G1 cell cycle arrest and apoptosis. RNA-sequencing was used to explore the underlying mechanisms. Genes associated with the cell cycle and the p53 pathway were significantly enriched with KPT-330 treatment. KPT-330 suppressed XPO1 expression, upregulated p53, p21WAF1/Cip1, and p27Kip1 and their nuclear localization, and downregulated anti-apoptotic protein (Survivin). The in vivo efficacy of KPT-330 was investigated using a bioluminescent xenograft mouse model of CTCL. KPT-330 blocked tumor growth and prolonged survival (p < 0.0002) compared to controls. These findings support investigating the use of KPT-330 and next-generation XPO1 inhibitors in CTCL.

Selinexor in multiple myeloma.

INTRODUCTION: Selinexor, an XPO1 inhibitor, has emerged as a promising therapeutic option in the challenging landscape of relapsed/refractory multiple myeloma (RRMM). AREAS COVERED: This article provides a review of selinexor, with a focus on available clinical studies involving MM patients and its safety profile. Clinical trials, such as STORM and BOSTON, have demonstrated its efficacy, particularly in combination regimens, showcasing notable overall response rates (ORR) and prolonged median progressionfree survival (mPFS). Selinexor's versatility is evident across various combinations, including carfilzomibdexamethasone (XKd), lenalidomidedexamethasone (XRd), and pomalidomidedexamethasone (XPd), with efficacy observed even in tripleclass refractory and highrisk patient populations. However, challenges, including resistance mechanisms and adverse events, necessitate careful management. Realworld evidence also underscores selinexor's effectiveness in RRMM, though dose adjustments and supportive measures remain crucial. Ongoing trials are exploring selinexor in diverse combinations and settings, including pomalidomidenaïve patients and postautologous stem cell transplant (ASCT) maintenance. EXPERT OPINION: The evolving landscape of selinexor's role in the sequencing of treatment for RRMM, its potential in highrisk patients, including those with extramedullary disease, as revealed in the most recent international meetings, and ongoing investigations signal a dynamic era in myeloma therapeutics. Selinexor emerges as a pivotal component in multidrug strategies and innovative combinations.

Selinexor for the treatment of patients with relapsed or refractory multiple myeloma.

OBJECTIVE: Multiple myeloma cells resist standard therapies due to overexpression of the transport protein, exportin 1. Selinexor is a novel drug that targets the Exportin 1 protein in these cells. DATA SOURCE: A comprehensive search was done, and data showing the efficacy and safety of selinexor in relapsed/refractory multiple myeloma was collected using PubMed, Google Scholar, and clincialtrials.gov. DATA SUMMARY: Results from the clinical trials STORM, BOSTON, and STOMP were included. Parts I and II of the STORM trial revealed a progression-free survival (PFS) of 4.7 and 3.7 months, a median duration of response of 6.2 and 4.4 months, and an overall survival of 7.3 and 8.4 months, respectively. BOSTON trial's SVd arm (selinexor, bortezomib, and dexamethasone) had a median follow-up period of 13.2 months and an mPFS of 13.93 months. The Vd arm (bortezomib and dexamethasone) had a median follow-up duration of 16.5 months and an mPFS of 9.46 months. The STOMP trial is still active and has limited data available. The SKd arm (selinexor, carfilzomib, and dexamethasone) reported an overall response rate of 66.7% in patients with triple refractory multiple myeloma, and 82% in patients with high-risk cytogenetics. The SPd arm (selinexor, pomalidomide, and dexamethasone) shows an overall response rate of 54.30% in pomalidomide naïve-nonrefractory, 35.70% in pomalidomide refractory and 60% in those dosed at RP2D. SRd arm (selinexor, lenalidomide, and dexamethasone) shows an overall response rate of 91.7% in lenalidomide naïve and 12.5% in lenalidomide refractory patients. SVd (selinexor, bortezomib, and dexamethasone) arm reported an overall response rate of 63% in all patients while the SDd arm (selinexor, daratumumab, and dexamethasone) showed an overall response rate of 73%. CONCLUSION: To improve the outcome of patients with relapsed/refractory multiple myeloma, it is critical to develop new therapies, assess potential therapeutic synergies, and overcome drug resistance by determining the efficacy of multiple myeloma therapies across multiple disease subgroups.

Nuclear export of circular RNA.

Circular RNAs (circRNAs), which are increasingly being implicated in a variety of functions in normal and cancerous cells1-5, are formed by back-splicing of precursor mRNAs in the nucleus6-10. circRNAs are predominantly localized in the cytoplasm, indicating that they must be exported from the nucleus. Here we identify a pathway that is specific for the nuclear export of circular RNA. This pathway requires Ran-GTP, exportin-2 and IGF2BP1. Enhancing the nuclear Ran-GTP gradient by depletion or chemical inhibition of the major protein exporter CRM1 selectively increases the nuclear export of circRNAs, while reducing the nuclear Ran-GTP gradient selectively blocks circRNA export. Depletion or knockout of exportin-2 specifically inhibits nuclear export of circRNA. Analysis of nuclear circRNA-binding proteins reveals that interaction between IGF2BP1 and circRNA is enhanced by Ran-GTP. The formation of circRNA export complexes in the nucleus is promoted by Ran-GTP through its interactions with exportin-2, circRNA and IGF2BP1. Our findings demonstrate that adaptors such as IGF2BP1 that bind directly to circular RNAs recruit Ran-GTP and exportin-2 to export circRNAs in a mechanism that is analogous to protein export, rather than mRNA export.

Dual Inhibition of CDK4/6 and XPO1 Induces Senescence With Acquired Vulnerability to CRBN-Based PROTAC Drugs.

BACKGROUND & AIMS: Despite the increasing number of treatment options available for liver cancer, only a small proportion of patients achieve long-term clinical benefits. Here, we aim to develop new therapeutic approaches for liver cancer. METHODS: A compound screen was conducted to identify inhibitors that could synergistically induce senescence when combined with cyclin-dependent kinase (CDK) 4/6 inhibitor. The combination effects of CDK4/6 inhibitor and exportin 1 (XPO1) inhibitor on cellular senescence were investigated in a panel of human liver cancer cell lines and multiple liver cancer models. A senolytic drug screen was performed to identify drugs that selectively killed senescent liver cancer cells. RESULTS: The combination of CDK4/6 inhibitor and XPO1 inhibitor synergistically induces senescence of liver cancer cells in vitro and in vivo. The XPO1 inhibitor acts by causing accumulation of RB1 in the nucleus, leading to decreased E2F signaling and promoting senescence induction by the CDK4/6 inhibitor. Through a senolytic drug screen, cereblon (CRBN)-based proteolysis targeting chimera (PROTAC) ARV-825 was identified as an agent that can selectively kill senescent liver cancer cells. Up-regulation of CRBN was a vulnerability of senescent liver cancer cells, making them sensitive to CRBN-based PROTAC drugs. Mechanistically, we find that ubiquitin specific peptidase 2 (USP2) directly interacts with CRBN, leading to the deubiquitination and stabilization of CRBN in senescent liver cancer cells. CONCLUSIONS: Our study demonstrates a striking synergy in senescence induction of liver cancer cells through the combination of CDK4/6 inhibitor and XPO1 inhibitor. These findings also shed light on the molecular processes underlying the vulnerability of senescent liver cancer cells to CRBN-based PROTAC therapy.

XPO1 inhibition sensitises CLL cells to NK cell mediated cytotoxicity and overcomes HLA-E expression.

The first-in-class inhibitor of exportin-1 (XPO1) selinexor is currently under clinical investigation in combination with the BTK inhibitor ibrutinib for patients with chronic lymphocytic leukaemia (CLL) or non-Hodgkin lymphoma. Selinexor induces apoptosis of tumour cells through nuclear retention of tumour suppressor proteins and has also recently been described to modulate natural killer (NK) cell and T cell cytotoxicity against lymphoma cells. Here, we demonstrate that XPO1 inhibition enhances NK cell effector function against primary CLL cells via downregulation of HLA-E and upregulation of TRAIL death receptors DR4 and DR5. Furthermore, selinexor potentiates NK cell activation against CLL cells in combination with several approved treatments; acalabrutinib, rituximab and obinutuzumab. We further demonstrate that lymph node associated signals (IL-4 + CD40L) inhibit NK cell activation against CLL cells via upregulation of HLA-E, and that inhibition of XPO1 can overcome this protective effect. These findings allow for the design of more efficacious combination strategies to harness NK cell effector functions against CLL.

Exportin(g) precision oncology beyond genomics with XPO1 inhibition.

We urgently need to identify new druggable targets for pediatric cancers. Coutinho et al.1 identified and validated XPO1 aberrant activity in patients with Wilms tumor (WT) and malignant rhabdoid tumor (MRT) using public databases and report an early proof-of-concept bench-to-bedside translation of XPO1 inhibition in pediatric WT and MRT.

P2RY2-AKT activation is a therapeutically actionable consequence of XPO1 inhibition in acute myeloid leukemia.

Selinexor is a first-in-class inhibitor of the nuclear exportin XPO1 that was recently approved by the US Food and Drug Administration for the treatment of multiple myeloma and diffuse large B-cell lymphoma. In relapsed/refractory acute myeloid leukemia (AML), selinexor has shown promising activity, suggesting that selinexor-based combination therapies may have clinical potential. Here, motivated by the hypothesis that selinexor's nuclear sequestration of diverse substrates imposes pleiotropic fitness effects on AML cells, we systematically catalog the pro- and anti-fitness consequences of selinexor treatment. We discover that selinexor activates PI3Kγ-dependent AKT signaling in AML by upregulating the purinergic receptor P2RY2. Inhibiting this axis potentiates the anti-leukemic effects of selinexor in AML cell lines, patient-derived primary cultures and multiple mouse models of AML. In a syngeneic, MLL-AF9-driven mouse model of AML, treatment with selinexor and ipatasertib outperforms both standard-of-care chemotherapy and chemotherapy with selinexor. Together, these findings establish drug-induced P2RY2-AKT signaling as an actionable consequence of XPO1 inhibition in AML.

CRM1 Promotes Capsid Disassembly and Nuclear Envelope Translocation of Adenovirus Independently of Its Export Function.

After receptor-mediated endocytosis and endosomal escape, adenoviral capsids can travel via microtubule organizing centers to the nuclear envelope. Upon capsid disassembly, viral genome import into nuclei of interphase cells then occurs through nuclear pore complexes, involving the nucleoporins Nup214 and Nup358. Import also requires the activity of the classic nuclear export receptor CRM1, as it is blocked by the selective inhibitor leptomycin B. We have now used artificially enucleated as well as mitotic cells to analyze the role of an intact nucleus in different steps of the viral life cycle. In enucleated U2OS cells, viral capsids traveled to the microtubule organizing center, whereas their removal from this complex was blocked, suggesting that this step required nuclear factors. In mitotic cells, on the other hand, CRM1 promoted capsid disassembly and genome release, suggesting a role of this protein that does not require intact nuclear envelopes or nuclear pore complexes and is distinct from its function as a nuclear export receptor. Similar to enucleation, inhibition of CRM1 by leptomycin B also leads to an arrest of adenoviral capsids at the microtubule organizing center. In a small-scale screen using leptomycin B-resistant versions of CRM1, we identified a mutant, CRM1 W142A P143A, that is compromised with respect to adenoviral capsid disassembly in both interphase and mitotic cells. Strikingly, this mutant is capable of exporting cargo proteins out of the nucleus of living cells or digitonin-permeabilized cells, pointing to a role of the mutated region that is not directly linked to nuclear export. IMPORTANCE A role of nucleoporins and of soluble transport factors in adenoviral genome import into the nucleus of infected cells in interphase has previously been established. The nuclear export receptor CRM1 promotes genome import, but its precise function is not known. Using enucleated and mitotic cells, we showed that CRM1 does not simply function by exporting a crucial factor out of the nucleus that would then trigger capsid disassembly and genome import. Instead, CRM1 has an export-independent role, a notion that is also supported by a mutant, CRM1 W142A P143A, which is export competent but deficient in viral capsid disassembly, in both interphase and mitotic cells.

Chromosome Region Maintenance 1 (XPO1/CRM1) as an Anticancer Target and Discovery of Its Inhibitor.

Chromosome region maintenance 1 (CRM1) is a major nuclear export receptor protein and contributes to cell homeostasis by mediating the transport of cargo from the nucleus to the cytoplasm. CRM1 is a therapeutic target comprised of several tumor types, including osteosarcoma, multiple myeloma, gliomas, and pancreatic cancer. In the past decade, dozens of CRM1 inhibitors have been discovered and developed, including KPT-330, which received FDA approval for multiple myeloma (MM) and diffuse large B-cell lymphoma (DLBCL) in 2019 and 2020, respectively. This review summarizes the biological functions of CRM1, the current understanding of the role CRM1 plays in cancer, the discovery of CRM1 small-molecule inhibitors, preclinical and clinical studies on KPT-330, and other recently developed inhibitors. A new CRM1 inhibition mechanism and structural dynamics are discussed. Through this review, we hope to guide the future design and optimization of CRM1 inhibitors.

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.

Bench to bedside radiosensitizer development strategy for newly diagnosed glioblastoma.

Glioblastoma is the most common primary brain malignancy and carries with it a poor prognosis. New agents are urgently needed, however nearly all Phase III trials of GBM patients of the past 25 years have failed to demonstrate improvement in outcomes. In 2019, the National Cancer Institute Clinical Trials and Translational Research Advisory Committee (CTAC) Glioblastoma Working Group (GBM WG) identified 5 broad areas of research thought to be important in the development of new herapeutics for GBM. Among those was optimizing radioresponse for GBM in situ. One such strategy to increase radiation efficacy is the addition of a radiosensitizer to improve the therapeutic ratio by enhancing tumor sensitivity while ideally having minimal to no effect on normal tissue. Historically the majority of trials using radiosensitizers have been unsuccessful, but they provide important guidance in what is required to develop agents more efficiently. Improved target selection is essential for a drug to provide maximal benefit, and once that target is identified it must be validated through pre-clinical studies. Careful selection of appropriate in vitro and in vivo models to demonstrate increased radiosensitivity and suitable bioavailability are then necessary to prove that a drug warrants advancement to clinical investigation. Once investigational agents are validated pre-clinically, patient trials require consistency both in terms of planning study design as well as reporting efficacy and toxicity in order to assess the potential benefit of the drug. Through this paper we hope to outline strategies for developing effective radiosensitizers against GBM using as models the examples of XPO1 inhibitors and HDAC inhibitors developed from our own lab.

TNPO2 variants associate with human developmental delays, neurologic deficits, and dysmorphic features and alter TNPO2 activity in Drosophila.

Transportin-2 (TNPO2) mediates multiple pathways including non-classical nucleocytoplasmic shuttling of >60 cargoes, such as developmental and neuronal proteins. We identified 15 individuals carrying de novo coding variants in TNPO2 who presented with global developmental delay (GDD), dysmorphic features, ophthalmologic abnormalities, and neurological features. To assess the nature of these variants, functional studies were performed in Drosophila. We found that fly dTnpo (orthologous to TNPO2) is expressed in a subset of neurons. dTnpo is critical for neuronal maintenance and function as downregulating dTnpo in mature neurons using RNAi disrupts neuronal activity and survival. Altering the activity and expression of dTnpo using mutant alleles or RNAi causes developmental defects, including eye and wing deformities and lethality. These effects are dosage dependent as more severe phenotypes are associated with stronger dTnpo loss. Interestingly, similar phenotypes are observed with dTnpo upregulation and ectopic expression of TNPO2, showing that loss and gain of Transportin activity causes developmental defects. Further, proband-associated variants can cause more or less severe developmental abnormalities compared to wild-type TNPO2 when ectopically expressed. The impact of the variants tested seems to correlate with their position within the protein. Specifically, those that fall within the RAN binding domain cause more severe toxicity and those in the acidic loop are less toxic. Variants within the cargo binding domain show tissue-dependent effects. In summary, dTnpo is an essential gene in flies during development and in neurons. Further, proband-associated de novo variants within TNPO2 disrupt the function of the encoded protein. Hence, TNPO2 variants are causative for neurodevelopmental abnormalities.

PSEN1-selective gamma-secretase inhibition in combination with kinase or XPO-1 inhibitors effectively targets T cell acute lymphoblastic leukemia.

BACKGROUND: T cell acute lymphoblastic leukemia (T-ALL) is a high-risk subtype that comprises 10-15% of childhood and 20-25% of adult ALL cases. Over 70% of T-ALL patients harbor activating mutations in the NOTCH1 signaling pathway and are predicted to be sensitive to gamma-secretase inhibitors. We have recently demonstrated that selective inhibition of PSEN1-containing gamma-secretase complexes can overcome the dose-limiting toxicity associated with broad gamma-secretase inhibitors. In this study, we developed combination treatment strategies with the PSEN1-selective gamma-secretase inhibitor MRK-560 and other targeted agents (kinase inhibitors ruxolitinib and imatinib; XPO-1 inhibitor KPT-8602/eltanexor) for the treatment of T-ALL. METHODS: We treated T-ALL cell lines in vitro and T-ALL patient-derived xenograft (PDX) models in vivo with MRK-560 alone or in combination with other targeted inhibitors (ruxolitinib, imatinib or KPT-8602/eltanexor). We determined effects on proliferation of the cell lines and leukemia development and survival in the PDX models. RESULTS: All NOTCH1-signaling-dependent T-ALL cell lines were sensitive to MRK-560 and its combination with ruxolitinib or imatinib in JAK1- or ABL1-dependent cell lines synergistically inhibited leukemia proliferation. We also observed strong synergy between MRK-560 and KPT-8602 (eltanexor) in all NOTCH1-dependent T-ALL cell lines. Such synergy was also observed in vivo in a variety of T-ALL PDX models with NOTCH1 or FBXW7 mutations. Combination treatment significantly reduced leukemic infiltration in vivo and resulted in a survival benefit when compared to single treatment groups. We did not observe weight loss or goblet cell hyperplasia in single drug or combination treated mice when compared to control. CONCLUSIONS: These data demonstrate that the antileukemic effect of PSEN1-selective gamma-secretase inhibition can be synergistically enhanced by the addition of other targeted inhibitors. The combination of MRK-560 with KPT-8602 is a highly effective treatment combination, which circumvents the need for the identification of additional mutations and provides a clear survival benefit in vivo. These promising preclinical data warrant further development of combination treatment strategies for T-ALL based on PSEN1-selective gamma-secretase inhibition.

Selinexor, a novel selective inhibitor of nuclear export, reduces SARS-CoV-2 infection and protects the respiratory system in vivo.

The novel coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the recent global pandemic. The nuclear export protein (XPO1) has a direct role in the export of SARS-CoV proteins including ORF3b, ORF9b, and nucleocapsid. Inhibition of XPO1 induces anti-inflammatory, anti-viral, and antioxidant pathways. Selinexor is an FDA-approved XPO1 inhibitor. Through bioinformatics analysis, we predicted nuclear export sequences in the ACE-2 protein and confirmed by in vitro testing that inhibition of XPO1 with selinexor induces nuclear localization of ACE-2. Administration of selinexor inhibited viral infection prophylactically as well as therapeutically in vitro. In a ferret model of COVID-19, selinexor treatment reduced viral load in the lungs and protected against tissue damage in the nasal turbinates and lungs in vivo. Our studies demonstrated that selinexor downregulated the pro-inflammatory cytokines IL-1ß, IL-6, IL-10, IFN-γ, TNF-α, and GMCSF, commonly associated with the cytokine storm observed in COVID-19 patients. Our findings indicate that nuclear export is critical for SARS-CoV-2 infection and for COVID-19 pathology and suggest that inhibition of XPO1 by selinexor could be a viable anti-viral treatment option.

Structure-Guided Design of the First Noncovalent Small-Molecule Inhibitor of CRM1.

Nuclear export factor chromosome region maintenance 1 (CRM1) is an attractive anticancer and antiviral drug target that spurred several research efforts to develop its inhibitor. Noncovalent CRM1 inhibitors are desirable, but none is reported to date. Here, we present the crystal structure of yeast CRM1 in complex with S109, a substructure of CBS9106 (under clinical test). Superimposition with the LFS-829 (another covalent CRM1 inhibitor) complex inspired the design of a noncovalent CRM1 inhibitor. Among nine synthesized compounds, noncovalent CRM1 inhibitor 1 (NCI-1) showed a high affinity to human and yeast CRM1 in the absence or presence of GST-bound Ras-related nuclear protein (RanGTP). Unlike covalent inhibitors, the crystal structure showed that NCI-1 is bound in the "open" nuclear export signal (NES) groove of CRM1, simultaneously occupying two hydrophobic pockets. NCI-1 additionally inhibited the nuclear export and proliferation of cells harboring the human CRM1-C528S mutant. Our work opens up the avenue of noncovalent CRM1 inhibitor development toward a more potent, less toxic, and broad-spectrum anticancer/antiviral therapy.