Targeting Cancer at the Nuclear Pore.

Nuclear protein transport regulates the function of cellular proteins in a compartmentalization-dependent fashion. Proteins require an active form of transport through the nuclear pore that involves specialized transporters belonging to the karyopherin family (Fig 1). The import of the majority of the cytosolic proteins is regulated by importins through nuclear localization signal recognition. The export of nuclear proteins is almost exclusively mediated through exportin 1 (XPO1), also known as chromosome maintenance region 1 (CRM1), through the recognition of a conserved nuclear export signal (NES). Nuclear export is a crucial aspect in the biology of any protein because mislocalization causes their functional inactivation. Most of the transcription factors and tumor suppressor proteins (TSPs), such as p53, IkB, p27, Rb, prostate apoptosis response-4 (PAR-4), and others, require nuclear retention and sequence-specific alignment to DNA to induce gene expression changes and transcriptional regulation of targets. It is logical to assume that the aberrant expression of nuclear transporters would result in unusual import or export of proteins, leading to their functional inactivation. Supporting this, there is ample evidence that cancer cells harbor an unusually higher expression of XPO1, making it an attractive therapeutic target to restore the proper localization of TSPs. XPO1/CRM1 can transport more than 200 target proteins. Many of these cargoes are TSPs and transcription factors that can affect themajority of cancer hallmarks, driving the interest in the pharmaceutical industry as a target worth pursuing. The first agent, leptomycin B (LMB), a natural product (originally developed as an antifungal compound), could specifically inhibit CRM1 export function. LMB covalently binds to the NES recognizing Cys528 amino acid in XPO1/CRM1, thereby effectively blocking its target recognition. Nevertheless, LMB showed numerous secondary effects and proved to be highly toxic and, therefore, was discontinued from a single phase I clinical trial. Later, some modified forms of LMB (eg, leptomycin A) with better toxicity profiles were also tested preclinically. Ratjadone is another natural product that works as a CRM1 inhibitor in a fashion similar to LMB. In parallel, CBS9106 was developed as a potent CRM1 inhibitor with a mechanism distinct from that of LMB. Around 2010, a series of specific inhibitors of nuclear export (SINEs) developed at Karyopharm Therapeutics (KPT) inNatick,Massachusetts (now located inNewton,MA), were presented at the American Society of Hematology annual meeting. These agents were shown to effectively block nuclear export function by binding to the Cys528 amino acid in the NES-recognizing domain of XPO1. Unlike irreversibly binding LMB, the SINE compounds bind Cys528 in XPO1 in a slowly reversible fashion. Their specificity has been validated using CRISPR/Cas9 genome editing where a single mutation in Cys528 renders the drug ineffective. These agents showed cancer cell selectivity and broad activity against a spectrum of solid tumors and hematologic cell lines as single agents. Numerous groups in heme models have demonstrated that SINE could synergize with chemotherapeutic agents such as cyclophosphamide, doxorubicin, vincristine, and prednisone; BRAF inhibitors; mammalian target of rapamycin inhibitors; and dexamethasone. Studies also showed synergy with gemcitabine and platinum compounds in solid tumor models (reviewed in Turner et al). SINEs could also suppress cancer stemness and reverse epithelial-to-mesenchymal transition, which is an event considered critical to metastasis. These multimodel analyses ushered the SINE compound selinexor into multiple phase I and II clinical trials. The two recent articles published in Journal of Clinical Oncology (JCO) present the results of multi-institutional clinical studies on selinexor. The first JCO article, by Abdul-Razak et al, presents results from a multicenter phase I study that evaluated selinexor (KPT-330) and determined the recommended phase II dose. The study enrolled 189 patients with advanced solid tumors who received selinexor (3 to 85 mg/m) in either 3or 4-week continuous cycles. The study used gene expression to evaluate preand post-treatment levels of XPO1 mRNA in patient-derived leukocytes. The tumor biopsies were also examined by immunohistochemistry for changes in markers consistent with XPO1 inhibition. The main outcomes include the observation of dose-dependent elevations in XPO1 mRNA in leukocytes up to a dose level of 28 mg/m before reaching a plateau. In line with the proposedmechanism of action of the drug, paired tumor biopsies showed nuclear accumulation of key TSPs, reduction of cell proliferation, and induction of apoptosis. Among 157 patients evaluable for response, one complete and six partial responses were observed (n5 7, 4%), with 27 patients (17%) achieving stable disease for $ 4 months. In the second JCO article, Gounder et al evaluated pharmacodynamic changes in tumor biopsies of 54 patients treated with oral selinexor administered twice per week (administered on days 1 and 3) at one of three doses (30 mg/m, 50 mg/m, or a flat dose of 60mg) either continuously or on a schedule of 3 weeks on and 1week off. Pharmacokinetic analysis was performed under fasting and feeding states (lowv high-fat content) and using various formulations of selinexor (tablet, capsule, or suspension). Most strikingly, there