Regulated signal transduction in discrete microdomains of the cell surface is an attractive hypothesis for achieving spatial and temporal specificity in signaling. A procedure for purifying caveolae separately from other similarly buoyant microdomains including those rich in glycosylphosphatidylinositol-anchored proteins has been developed (Schnitzer, J. E., McIntosh, D. P., Dvorak, A. M., Liu, J., and Oh, P. (1995) Science 269, 1435-1439) and used here to show that caveolae contain many signaling molecules including select kinases (platelet-derived growth factor (PDGF) receptors, protein kinase C, phosphatidylinositol 3-kinase, and Srclike kinases), phospholipase C, sphingomyelin, and even phosphoinositides. More importantly, two different techniques reveal that caveolae function as signal transducing subcompartments of the plasma membrane. PDGF rapidly induces phosphorylation of endothelial cell plasmalemmal proteins residing in caveolae as detected by membrane subfractionation and confocal immunofluorescence microscopy. This PDGF signaling cascade is halted when the caveolar compartment is disassembled by filipin. Finally, in vitro kinase assays show that caveolae contain most of the intrinsic tyrosine kinase activity of the plasma membrane. As signal transducing organelles, caveolae organize a distinct set of signaling molecules to permit direct regionalized signal transduction within their boundaries.

Background Bromodomain and extra-terminal domain proteins are promising epigenetic anticancer drug targets. This first-in-human study evaluated the safety, recommended phase II dose, pharmacokinetics, pharmacodynamics, and preliminary antitumor activity of the bromodomain and extra-terminal domain inhibitor molibresib (GSK525762) in patients with nuclear protein in testis (NUT) carcinoma (NC) and other solid tumors. Methods This was a phase I and II, open-label, dose-escalation study. Molibresib was administered orally once daily. Single-patient dose escalation (from 2 mg/d) was conducted until the first instance of grade 2 or higher drug-related toxicity, followed by a 3 + 3 design. Pharmacokinetic parameters were obtained during weeks 1 and 3. Circulating monocyte chemoattractant protein-1 levels were measured as a pharmacodynamic biomarker. Results Sixty-five patients received molibresib. During dose escalation, 11% experienced dose-limiting toxicities, including six instances of grade 4 thrombocytopenia, all with molibresib 60–100 mg. The most frequent treatment-related adverse events of any grade were thrombocytopenia (51%) and gastrointestinal events, including nausea, vomiting, diarrhea, decreased appetite, and dysgeusia (22%–42%), anemia (22%), and fatigue (20%). Molibresib demonstrated an acceptable safety profile up to 100 mg; 80 mg once daily was selected as the recommended phase II dose. Following single and repeat dosing, molibresib showed rapid absorption and elimination (maximum plasma concentration: 2 hours; t1/2: 3–7 hours). Dose-dependent reductions in circulating monocyte chemoattractant protein-1 levels were observed. Among 19 patients with NC, four achieved either confirmed or unconfirmed partial response, eight had stable disease as best response, and four were progression-free for more than 6 months. Conclusions Once-daily molibresib was tolerated at doses demonstrating target engagement. Preliminary data indicate proof-of-concept in NC.

Desensitization plays an important role in the rapid termination of G-protein signaling pathways. This process, which involves phosphorylation by a G-protein-coupled receptor kinase (GRK) followed by arrestin binding, has been studied extensively in the rod photoreceptor cell of the mammalian retina. In contrast, less is known regarding desensitization in cone photoreceptor cells, which occurs more rapidly than in rod cells. Recently, our laboratory has cloned a novel GRK family member, GRK7, from the retina of a cone-dominant mammal, the 13-lined ground squirrel. Here we report the cloning of GRK7 from rod-dominant pig and human retinas, suggesting that this kinase plays a role in human visual signaling. Because GRK1 (rhodopsin kinase), the GRK that mediates rhodopsin desensitization in the rod cell, is reportedly expressed in both rods and cones, a detailed comparison of the localization of the two kinases is a necessary step toward determining their potential roles in cone visual signaling. Immunocytochemical analysis using antibodies selective for these two GRKs unexpectedly demonstrated species-specific differences in GRK7 and GRK1 expression in cones. In pigs and dogs, cones express only GRK7, whereas in mice and rats, we detected only GRK1 in cones. These results suggest that either GRK7 or GRK1 may participate in cone opsin desensitization, depending on the expression pattern of the kinases in different species. In contrast, GRK7 and GRK1 are coexpressed in monkey and human cones, suggesting that coordinate regulation of desensitization by both kinases may occur in primates.

Purpose: Enhancer of zeste homolog 2 (EZH2) activity is dysregulated in many cancers.Patients and Methods: This phase I study determined the safety, maximum-tolerated dose (MTD), pharmacokinetics, and pharmacodynamics of the intravenously administered, highly selective EZH2 inhibitor, GSK2816126, (NCT02082977). Doses of GSK2816126 ranged from 50 to 3,000 mg twice weekly, and GSK2816126 was given 3-weekson/1-week-off in 28-day cycles. Eligible patients had solid tumors or B-cell lymphomas with no available standard treatment regimen.Results: Forty-one patients (21 solid tumors, 20 lymphoma) received treatment. All patients experienced !1 adverse event (AE). Fatigue [22 of 41 (53.7%)] and nausea [20 of 41 (48.8%)] were the most common toxicity. Twelve (32%) patients experienced a serious AE. Dose-limiting elevated liver transaminases occurred in 2 of 7 patients receiving 3,000 mg of GSK2816126; 2,400 mg was therefore established as the MTD. Following intravenous administration of 50 to 3,000 mg twice weekly, plasma GSK2816126 levels decreased biexponentially, with a mean terminal elimination half-life of approximately 27 hours. GSK2816126 exposure (maximum observed plasma concentration and area under the plasmatime curve) increased in a dose-proportional manner. No change from baseline in H3K27me3 was seen in peripheral blood mononuclear cells. Fourteen of 41 (34%) patients had radiological best response of stable disease, 1 patient with lymphoma achieved a partial response, 21 of 41 (51%) patients had progressive disease, and 5 patients were unevaluable for antitumor response.Conclusions: The MTD of GSK2816126 was established at 2,400 mg, but the dosing method and relatively short half-life limited effective exposure, and modest anticancer activity was observed at tolerable doses.