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Introduction NRG1 rearrangements produce chimeric ligands that subvert the ERBB pathway to drive tumorigenesis. A better understanding of the signaling networks that mediate transformation by NRG1 fusions is needed to inform effective therapeutic strategies. Unfortunately, this has been hampered by a paucity of patient-derived disease models that faithfully recapitulate this molecularly defined cancer subset. Methods Patient-derived xenograft (PDX) and cell line models were established from NRG1-rearranged lung adenocarcinoma samples. Transcriptomic, proteomic, and biochemical analyses were performed to identify activated pathways. Efficacy studies were conducted to evaluate HER3- and MTOR-directed therapies. Results We established a pair of PDX and cell line models of invasive mucinous lung adenocarcinoma (LUAD) (LUAD-0061AS3, SLC3A2-NRG1), representing the first reported paired in vitro and in vivo model of NRG1-driven tumors. Growth of LUAD-0061AS3 models was reduced by the anti-HER3 antibody GSK2849330. Transcriptomic profiling revealed activation of the MTOR pathway in lung tumor samples with NRG1 fusions. Phosphorylation of several MTOR effectors (S6 and 4EBP1) was higher in LUAD-0061AS3 cells compared with human bronchial epithelial cells and the breast cancer cell line MDA-MB-175-VII (DOC4-NRG1 fusion). Accordingly, LUAD-0061AS3 cells were more sensitive to MTOR inhibitors than MDA-MB-175-VII cells and targeting the MTOR pathway with rapamycin blocked growth of LUAD-0061AS3 PDX tumors in vivo. In contrast, MDA-MB-175-VII breast cancer cells had higher MAPK pathway activation and were more sensitive to MEK inhibition. Conclusions We identify the MTOR pathway as a candidate vulnerability in NRG1 fusion-positive lung adenocarcinoma that may warrant further preclinical evaluation, with the eventual goal of finding additional therapeutic options for patients in whom ERBB-directed therapy fails. Moreover, our results uncover heterogeneity in downstream oncogenic signaling among NRG1-rearranged cancers, possibly tumor type-dependent, the therapeutic significance of which requires additional investigation.

Summary Background RET rearrangements are found in 1–2% of non-small-cell lung cancers. Cabozantinib is a multikinase inhibitor with activity against RET that produced a 10% overall response in unselected patients with lung cancers. To assess the activity of cabozantinib in patients with RET -rearranged lung cancers, we did a prospective phase 2 trial in this molecular subgroup. Methods We enrolled patients in this open-label, Simon two-stage, single-centre, phase 2, single-arm trial in the USA if they met the following criteria: metastatic or unresectable lung cancer harbouring a RET rearrangement, Karnofsky performance status higher than 70, and measurable disease. Patients were given 60 mg of cabozantinib orally per day. The primary objective was to determine the overall response (Response Criteria Evaluation in Solid Tumors version 1.1) in assessable patients; those who received at least one dose of cabozantinib, and had been given CT imaging at baseline and at least one protocol-specified follow-up timepoint. We did safety analyses in the modified intention-to-treat population who received at least one dose of cabozantinib. The accrual of patients with RET -rearranged lung cancer to this protocol has been completed but the trial is still ongoing because several patients remain on active treatment. This study was registered with ClinicalTrials.gov, number NCT01639508. Findings Between July 13, 2012, and April 30, 2016, 26 patients with RET -rearranged lung adenocarcinomas were enrolled and given cabozantinib; 25 patients were assessable for a response. KIF5B-RET was the predominant fusion type identified in 16 (62%) patients. The study met its primary endpoint, with confirmed partial responses seen in seven of 25 response-assessable patients (overall response 28%, 95% CI 12–49). Of the 26 patients given cabozantinib, the most common grade 3 treatment-related adverse events were lipase elevation in four (15%) patients, increased alanine aminotransferase in two (8%) patients, increased aspartate aminotransferase in two (8%) patients, decreased platelet count in two (8%) patients, and hypophosphataemia in two (8%) patients. No drug-related deaths were recorded but 16 (62%) patients died during the course of follow-up. 19 (73%) patients required dose reductions due to drug-related adverse events. Interpretation The reported activity of cabozantinib in patients with RET -rearranged lung cancers defines RET rearrangements as actionable drivers in patients with lung cancers. An improved understanding of tumour biology and novel therapeutic approaches will be needed to improve outcomes with RET-directed targeted treatment. Funding Exelixis, National Institutes of Health and National Cancer Institute Cancer Center Support Grant P30 CA008748.

BACKGROUND RET rearrangements are targetable, oncogenic lung cancer drivers. While previous series have shown durable clinical benefit with pemetrexed-based therapies in ALK- and ROS1-rearranged lung cancers, the benefits of pemetrexed-based treatments in patients with RET-rearranged lung cancers relative to other genomic subsets have not previously been explored. PATIENTS AND METHODS A retrospective review of patients with pathologically confirmed stage IIIB/IV lung adenocarcinomas and evidence of a RET, ROS1, or ALK rearrangement, or a KRAS mutation was conducted. Patients were eligible if they received treatment with pemetrexed alone or in combination. The primary outcome of progression-free survival (PFS), and secondary outcomes of overall response rate (ORR, RECIST v1.1), time to progression (TTP), and time to treatment discontinuation were compared between RET-rearranged and groups of ROS1-rearranged, ALK-rearranged, and KRAS-mutant lung cancers. RESULTS We evaluated 104 patients. Patients with RET-rearranged lung cancers (n = 18) had a median PFS of 19 months [95% confidence interval (CI) 12-not reached (NR)] that was comparable with patients with ROS1- (23 months, 95% CI 14-NR, n = 10) and ALK-rearranged (19 months, 95% CI 15-36, n = 36) lung cancers, and significantly improved compared with patients with KRAS-mutant lung cancers (6 months, 95% CI 5-9, P < 0.001, n = 40). ORR (45%), median TTP (20 months, 95% CI 17-NR), and median time to treatment discontinuation (21 months, 95% CI 6-NR) in patients with RET-rearranged lung cancers were not significantly different compared with patients with ALK- and ROS1-rearranged lung cancers, and improved compared with patients with KRAS-mutant lung cancers. CONCLUSION Durable benefits with pemetrexed-based therapies in RET-rearranged lung cancers are comparable with ALK- and ROS1-rearranged lung cancers. When selecting therapies for patients with RET-rearranged lung cancers, pemetrexed-containing regimens should be considered.

Type 2 diabetes is associated with many health complications making it imperative to understand the mechanisms responsible for insulin resistance. An epidemic prevalence has thrust obesity to the forefront as a major risk factor for type 2 diabetes and in this article we focus on recent attempts to develop models of obesity and diabetes. These models have provided new insights into the pathogenesis of insulin resistance in obesity and thus, enhance future potential for drug discovery.

Insulin stimulates glucose uptake in skeletal muscle cells and fat cells by promoting the rapid translocation of GLUT4 glucose transporters to the plasma membrane. Recent work from our laboratory supports the concept that insulin also stimulates the intrinsic activity of GLUT4 through a signaling pathway that includes p38 MAPK. Here we show that regulation of GLUT4 activity by insulin develops during maturation of skeletal muscle cells into myotubes in concert with the ability of insulin to stimulate p38 MAPK. In L6 myotubes expressing GLUT4 that carries an exofacial myc-epitope (L6-GLUT4myc), insulin-stimulated GLUT4myc translocation equals in magnitude the glucose uptake response. Inhibition of p38 MAPK with SB203580 reduces insulin-stimulated glucose uptake without affecting GLUT4myc translocation. In contrast, in myoblasts, the magnitude of insulin-stimulated glucose uptake is significantly lower than that of GLUT4myc translocation and is insensitive to SB203580. Activation of p38 MAPK by insulin is considerably higher in myotubes than in myoblasts, as is the activation of upstream kinases MKK3/MKK6. In contrast, the activation of all three Akt isoforms and GLUT4 translocation are similar in myoblasts and myotubes. Furthermore, GLUT4myc translocation and phosphorylation of regulatory sites on Akt in L6-GLUT4myc myotubes are equally sensitive to insulin, whereas glucose uptake and phosphorylation of regulatory sites on p38 MAPK show lower sensitivity to the hormone. These observations draw additional parallels between Akt and GLUT4 translocation and between p38 MAPK and GLUT4 activation. Regulation of GLUT4 activity by insulin develops upon muscle cell differentiation and correlates with p38 MAPK activation by insulin.

Participation of p38 mitogen-activated protein kinase (p38) in insulin-induced glucose uptake was suggested using pyridinylimidazole p38 inhibitors (e.g. SB203580). However, the role of p38 in insulin action remains controversial. We further test p38 participation in glucose uptake using a dominant-negative p38 mutant and two novel pharmacological p38 inhibitors related to but different from SB203580. We present the structures and activities of the azaazulene pharmacophores A291077 and A304000. p38 kinase activity was inhibited in vitro by A291077 and A304000 (IC(50) = 0.6 and 4.7 microm). At higher concentrations A291077 but not A304000 inhibited JNK2alpha (IC(50) = 3.5 microm). Pretreatment of 3T3-L1 adipocytes and L6 myotubes expressing GLUT4myc (L6-GLUT4myc myotubes) with A291077, A304000, SB202190, or SB203580 reduced insulin-stimulated glucose uptake by 50-60%, whereas chemical analogues inert toward p38 were ineffective. Expression of an inducible, dominant-negative p38 mutant in 3T3-L1 adipocytes reduced insulin-stimulated glucose uptake. GLUT4 translocation to the cell surface, immunodetected on plasma membrane lawns of 3T3-L1 adipocytes or on intact L6-GLUT4myc myotubes, was not altered by chemical or molecular inhibition of p38. We propose that p38 contributes to enhancing GLUT4 activity, thereby increasing glucose uptake. In addition, the azaazulene class of inhibitors described will be useful to decipher cellular actions of p38 and JNK.

The precise mechanisms underlying insulin-stimulated glucose transport still require investigation. Here we assessed the effect of SB203580, an inhibitor of the p38 MAP kinase family, on insulin-stimulated glucose transport in 3T3-L1 adipocytes and L6 myotubes. We found that SB203580, but not its inactive analogue (SB202474), prevented insulin-stimulated glucose transport in both cell types with an IC50 similar to that for inhibition of p38 MAP kinase (0.6 μm). Basal glucose uptake was not affected. Moreover, SB203580 added only during the transport assay did not inhibit basal or insulin-stimulated transport. SB203580 did not inhibit insulin-stimulated translocation of the glucose transporters GLUT1 or GLUT4 in 3T3-L1 adipocytes as assessed by immunoblotting of subcellular fractions or by immunofluorescence of membrane lawns. L6 muscle cells expressing GLUT4 tagged on an extracellular domain with a Myc epitope (GLUT4myc) were used to assess the functional insertion of GLUT4 into the plasma membrane. SB203580 did not affect the insulin-induced gain in GLUT4myc exposure at the cell surface but largely reduced the stimulation of glucose uptake. SB203580 had no effect on insulin-dependent insulin receptor substrate-1 phosphorylation, association of the p85 subunit of phosphatidylinositol 3-kinase with insulin receptor substrate-1, nor on phosphatidylinositol 3-kinase, Akt1, Akt2, or Akt3 activities in 3T3-L1 adipocytes. In conclusion, in the presence of SB203580, insulin caused normal translocation and cell surface membrane insertion of glucose transporters without stimulating glucose transport. We propose that insulin stimulates two independent signals contributing to stimulation of glucose transport: phosphatidylinositol 3-kinase leads to glucose transporter translocation and a pathway involving p38 MAP kinase leads to activation of the recruited glucose transporter at the membrane.