Resistance mediated by alternative receptor tyrosine kinases in FGFR1-amplified lung cancer.

Fibroblast growth factor receptor 1 (FGFR1) amplification has been identified in 10-20% of patients with squamous non-small-cell lung cancer. Preclinical models showed promising activity of specific FGFR inhibitors, but early clinical trials showed that only a small fraction of patients with FGFR1-amplified lung cancer responded to FGFR inhibitors. These unsatisfactory results were partly explained by heterogeneous amplicons around the 8p11 genomic region, leading to false-positive amplification results. Furthermore, discrepancies in the gene amplification and protein expression of FGFR1 were also reported. In this study, we identified the roles of alternative receptor tyrosine kinases (RTKs) in FGFR1-amplified lung cancer. These alternative RTKs dominantly activate phosphoinositide 3-kinase-AKT signaling and also mitigate sustained inhibition of mitogen-activated protein kinase signaling by FGFR inhibitors. The rebound activation of extracellular signal-regulated kinase phosphorylation was associated with sensitivity to the drugs. Combinatorial inhibition of alternative RTKs and FGFR1 was required to suppress both AKT and extracellular signal-regulated kinase phosphorylation and to induce key pro-apoptotic proteins BIM and p53 upregulated modulator of apoptosis (PUMA). Furthermore, even in FGFR inhibitor-sensitive NCI-H1581 lung cancer cells, MET-expressing clones were already detectable at a very low frequency before resistance induction. Selection of these pre-existing subclones resulted in FGFR inhibitor resistance because of the activation of AKT and extracellular signal-regulated kinase by MET signaling that was mediated by GRB2 associated binding protein 1 (GAB1). These results suggest that incomplete suppression of key survival signals led to intrinsic and acquired resistance to FGFR inhibitors. Our results may help explain the low clinical response rates to FGFR inhibitors in FGFR1-amplified lung cancer.

Combined inhibition of KRASG12C and mTORC1 kinase is synergistic in non-small cell lung cancer.

Current KRASG12C (OFF) inhibitors that target inactive GDP-bound KRASG12C cause responses in less than half of patients and these responses are not durable. A class of RASG12C (ON) inhibitors that targets active GTP-bound KRASG12C blocks ERK signaling more potently than the inactive-state inhibitors. Sensitivity to either class of agents is strongly correlated with inhibition of mTORC1 activity. We have previously shown that PI3K/mTOR and ERK-signaling pathways converge on key cellular processes and that inhibition of both pathways is required for inhibition of these processes and for significant antitumor activity. We find here that the combination of a KRASG12C inhibitor with a selective mTORC1 kinase inhibitor causes synergistic inhibition of Cyclin D1 expression and cap-dependent translation. Moreover, BIM upregulation by KRASG12C inhibition and inhibition of MCL-1 expression by the mTORC1 inhibitor are both required to induce significant cell death. In vivo, this combination causes deep, durable tumor regressions and is well tolerated. This study suggests that the ERK and PI3K/mTOR pathways each mitigate the effects of inhibition of the other and that combinatorial inhibition is a potential strategy for treating KRASG12C-dependent lung cancer.

Inhibition of non-homologous end joining mitigates paclitaxel resistance resulting from mitotic slippage in non-small cell lung cancer.

Mitotic slippage, which enables cancer cells to bypass cell death by transitioning from mitosis to the G1 phase without undergoing normal cytokinesis, is one likely mechanism of paclitaxel (PTX) resistance. DNA double-strand breaks (DSBs) in the G1 phase are mainly repaired through non-homologous end joining (NHEJ). Therefore, inhibiting NHEJ could augment the PTX-induced cytotoxicity by impeding the repair of PTX-induced DSBs during the G1 phase following mitotic slippage. We aimed to evaluate the effects of NHEJ inhibition on mitotic slippage after PTX treatment in non-small cell lung cancer (NSCLC). H1299, A549, H1975, and H520 NSCLC cell lines were employed. In addition, A-196 and JQ1 were used as NHEJ inhibitors. H1299 cells were PTX-resistant and exhibited an increased frequency of mitotic slippage upon PTX treatment. NHEJ inhibitors significantly augmented the PTX-induced cytotoxicity, DSBs, and apoptosis in H1299 cells. The newly generated PTX-resistant cells were even more prone to mitotic slippage following PTX treatment and susceptible to the combined therapy. Docetaxel further demonstrated synergistic effects with the NHEJ inhibitor in PTX-resistant cells. NHEJ inhibition may overcome intrinsic or acquired PTX resistance resulting from mitotic slippage by synergistically increasing the cytotoxic effects of antimitotic drugs in NSCLC.

WEE1 inhibition enhances the antitumor immune response to PD-L1 blockade by the concomitant activation of STING and STAT1 pathways in SCLC.

Small cell lung cancers (SCLCs) have high mutational burden but are relatively unresponsive to immune checkpoint blockade (ICB). Using SCLC models, we demonstrate that inhibition of WEE1, a G2/M checkpoint regulator induced by DNA damage, activates the STING-TBK1-IRF3 pathway, which increases type I interferons (IFN-α and IFN-ß) and pro-inflammatory chemokines (CXCL10 and CCL5), facilitating an immune response via CD8+ cytotoxic T cell infiltration. We further show that WEE1 inhibition concomitantly activates the STAT1 pathway, increasing IFN-γ and PD-L1 expression. Consistent with these findings, combined WEE1 inhibition (AZD1775) and PD-L1 blockade causes remarkable tumor regression, activation of type I and II interferon pathways, and infiltration of cytotoxic T cells in multiple immunocompetent SCLC genetically engineered mouse models, including an aggressive model with stabilized MYC. Our study demonstrates cell-autonomous and immune-stimulating activity of WEE1 inhibition in SCLC models. Combined inhibition of WEE1 plus PD-L1 blockade represents a promising immunotherapeutic approach in SCLC.

Development of compact inductive energy storage pulsed-power generator driven by 13 kV SiC-MOSFET.

A compact inductive energy storage (IES) pulsed-power generator that is driven by a novel 13 kV silicon carbide (SiC)-MOSFET is developed and molded into a compact modified TO-268. In this article, the switching characteristics required for IES pulsed-power generator development are evaluated. The maximum slew rates at MOSFET turn-on and turn-off are 157 and 129 kV/µs, respectively, at an input voltage of 10 kV. The maximum current flow from the drain to the source terminal is limited to 128 A during short-circuit switching. The on-resistance between the drain and source terminals increases during the SiC-MOSFET's on state. It increases with the voltage and its minimum value is 1.07 Ω. These characteristics show that the device is suitable for use as an opening switch because of its low on-resistance and rapid large-current cutoff at high operating voltages. The characteristics of an IES pulsed-power generator composed of a SiC-MOSFET, a capacitor, and a pulsed transformer with a turn ratio of 5:15 are also evaluated. The output voltage peak and full width at half maximum reach 31.4 kV and 55 ns, respectively, at a charging voltage of 1100 V. The maximum energy transfer efficiency is 50.2% of the input energy with a load resistance of 2.5 kΩ. The results show that the MOSFET has excellent potential to support the development of a compact plasma generation system that offers better performance pulsed-power generators driven by semiconductor devices.

Distinct dependencies on receptor tyrosine kinases in the regulation of MAPK signaling between BRAF V600E and non-V600E mutant lung cancers.

BRAF is one of the most frequently mutated genes across a number of different cancers, with the best-characterized mutation being V600E. Despite the successes of treating BRAF mutant V600E lung cancer with BRAF pathway inhibitors, treatment strategies targeting tumors with non-V600E mutations are yet to be established. We studied cellular signaling differences between lung cancers with different BRAF mutations and determined their sensitivities to BRAF pathway inhibitors. Here, we observed that MEK inhibition induced feedback activation of the receptor tyrosine kinase (RTK) EGFR, and in some cases the RTK FGFR, resulting in transient suppression of ERK phosphorylation in BRAF non-V600E, but not BRAF V600E, mutant cells. Furthermore, we found that both EGFR and FGFR activated the MEK/ERK pathway, despite the presence of BRAF non-V600E mutations with elevated kinase activity. Moreover, in BRAF non-V600E mutants with impaired kinase activities, EGFR had even greater control over the MEK/ERK pathway, essentially contributing completely to the tonic mitogen-activated protein kinase (MAPK) signal. Accordingly, the combination of MEK inhibitor with EGFR inhibitor was effective at shrinking tumors in mouse model of BRAF non-V600E mutant lung cancer. Furthermore, the results were recapitulated with a clinically relevant dual inhibitor of EGFR and RAF, BGB-283. Overall, although BRAF V600E mutant cells are sensitive to BRAF inhibition, non-V600E mutant cancer cells are reliant on RTKs for their MAPK activation and inhibiting both MEK and RTKs are necessary in these cancers. Our findings provide evidence of critical survival signals in BRAF non-V600E mutant cancers, which could pave the way for effective treatment of these cancers.

Epithelial-to-Mesenchymal Transition Antagonizes Response to Targeted Therapies in Lung Cancer by Suppressing BIM.

Purpose: Epithelial-to-mesenchymal transition (EMT) confers resistance to a number of targeted therapies and chemotherapies. However, it has been unclear why EMT promotes resistance, thereby impairing progress to overcome it.Experimental Design: We have developed several models of EMT-mediated resistance to EGFR inhibitors (EGFRi) in EGFR-mutant lung cancers to evaluate a novel mechanism of EMT-mediated resistance.Results: We observed that mesenchymal EGFR-mutant lung cancers are resistant to EGFRi-induced apoptosis via insufficient expression of BIM, preventing cell death despite potent suppression of oncogenic signaling following EGFRi treatment. Mechanistically, we observed that the EMT transcription factor ZEB1 inhibits BIM expression by binding directly to the BIM promoter and repressing transcription. Derepression of BIM expression by depletion of ZEB1 or treatment with the BH3 mimetic ABT-263 to enhance "free" cellular BIM levels both led to resensitization of mesenchymal EGFR-mutant cancers to EGFRi. This relationship between EMT and loss of BIM is not restricted to EGFR-mutant lung cancers, as it was also observed in KRAS-mutant lung cancers and large datasets, including different cancer subtypes.Conclusions: Altogether, these data reveal a novel mechanistic link between EMT and resistance to lung cancer targeted therapies. Clin Cancer Res; 24(1); 197-208. ©2017 AACR.

Key roles of EMT for adaptive resistance to MEK inhibitor in KRAS mutant lung cancer.

KRAS is frequently mutated in a variety of cancers including lung cancer. Whereas the mitogen-activated protein kinase (MAPK) is a well-known effector pathway of KRAS, blocking this pathway with MEK inhibitors is relatively ineffective. One major contributor to limited efficacy is attributed to the reactivation of MAPK signal following MEK inhibition by multiple feedback mechanisms. In a recent study, we have identified that epithelial-to-mesenchymal transition defines feedback activation of receptor tyrosine kinase signaling following MEK inhibition in KRAS mutant lung cancer. In epithelial-like cells, this feedback was mediated by ERBB3. In contrast, in mesenchymal-like cells, the feedback was attributed to the fibroblast growth factor receptor 1 (FGFR1) pathway. FGFR1 was dominantly expressed in mesenchymal-like cells: suppression of SPRY proteins by MEK inhibition relieved negative feedback control of basal FGFR-FRS2 function, resulting in reactivation of MAPK signaling via FGFR1. Therapeutically, the combination of MEK inhibitor trametinib with an FGFR inhibitor induced tumor regressions in tumor xenografts derived from mesenchymal-like KRAS mutant cancer cell lines as well as a patient derived xenograft model with a representative mesenchymal phenotype. Collectively, feedback activation of MAPK by FGFR1 signaling mitigates the effect of MEK inhibitor in mesenchymal-like KRAS mutant lung tumors, and combinations of clinically available FGFR1 inhibitors and MAPK inhibitors constitute a therapeutic approach to treat these cancers effectively.

Improvements of visual function and outer retinal morphology following spontaneous regression of cancer in anti-recoverin cancer-associated retinopathy.

PURPOSE: To report an anti-recoverin antibody-positive cancer-associated retinopathy (anti-recoverin CAR) patient with remarkable improvements of visual function and outer retinal morphology following spontaneous regression of cancer. OBSERVATIONS: A 65-year-old woman with small cell lung carcinoma developed progressive, bilateral vision loss with diffuse loss of the ellipsoid zone at the macula on optical coherence tomography and marked reduced responses of a- and b-waves on electroretinography. Western blot analysis led to a diagnosis of anti-recoverin CAR. The visual function and outer retinal morphology gradually improved following spontaneous regression of the cancer and the initiation of systemic corticosteroid. Subsequent intermittent chemotherapy and continuation of corticosteroid maintained reduction of the cancer and prevented the recurrence of CAR, with preservation of improvements of the visual function and macular outer retinal morphology. CONCLUSIONS AND IMPORTANCE: These results suggest that requirement for obtaining good visual prognosis in CAR patients is to make the cancer regress prior to falling into photoreceptor apotosis.

Recurrence of renal cell carcinoma diagnosed using contralateral adrenal biopsy with endoscopic ultrasound-guided fine-needle aspiration.

A 76-year-old female in whom a renal cell carcinoma (RCC) lesion was resected 19 years previously presented to our hospital with cognitive dysfunction. Magnetic resonance imaging and computed tomography revealed nodules in the brain, lung, adrenal gland and a pelvic osteolytic lesion. To identify the primary cancer site, the present study performed endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) of the left adrenal lesion. Consequently, the pathological findings of the tissue obtained by EUS-FNA were similar to those of the previous nephrectomy specimen, revealing that the adrenal lesion was the recurrence of RCC. The majority of the metastatic lesions in the patient were reduced in size by the multiple kinase inhibitor, pazopanib. Contralateral adrenal metastasis of RCC is rare and the use of EUS-FNA in the diagnosis of adrenal lesions remains to be elucidated. This is a rare case of adrenal lesion, diagnosed by EUS-FNA. Therefore, EUS-FNA is considered to be a useful diagnostic modality of adrenal metastases from unidentified primary tumor types.

Epithelial-to-Mesenchymal Transition Defines Feedback Activation of Receptor Tyrosine Kinase Signaling Induced by MEK Inhibition in KRAS-Mutant Lung Cancer.

UNLABELLED: KRAS is frequently mutated in lung cancer. Whereas MAPK is a well-known effector pathway of KRAS, blocking this pathway with clinically available MAPK inhibitors is relatively ineffective. Here, we report that epithelial-to-mesenchymal transition rewires the expression of receptor tyrosine kinases, leading to differential feedback activation of the MAPK pathway following MEK inhibition. In epithelial-like KRAS-mutant lung cancers, this feedback was attributed to ERBB3-mediated activation of MEK and AKT. In contrast, in mesenchymal-like KRAS-mutant lung cancers, FGFR1 was dominantly expressed but suppressed by the negative regulator Sprouty proteins; MEK inhibition led to repression of SPRY4 and subsequent FGFR1-mediated reactivation of MEK and AKT. Therapeutically, the combination of a MEK inhibitor (MEKi) and an FGFR inhibitor (FGFRi) induced cell death in vitro and tumor regressions in vivo These data establish the rationale and a therapeutic approach to treat mesenchymal-like KRAS-mutant lung cancers effectively with clinically available FGFR1 and MAPK inhibitors. SIGNIFICANCE: Adaptive resistance to MEKi is driven by receptor tyrosine kinases specific to the differentiation state of the KRAS-mutant non-small cell lung cancer (NSCLC). In mesenchymal-like KRAS-mutant NSCLC, FGFR1 is highly expressed, and MEK inhibition relieves feedback suppression of FGFR1, resulting in reactivation of ERK; suppression of ERK by MEKi/FGFRi combination results in tumor shrinkage. Cancer Discov; 6(7); 754-69. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 681.

Spontaneous regression of small cell lung cancer combined with cancer associated retinopathy.

Spontaneous regression (SR) is defined as the complete or partial disappearance of disease without anticancer treatments. We report a case of SR of small cell lung cancer (SCLC) combined with cancer associated retinopathy (CAR). A 65-year-old woman was admitted to our hospital to examine abnormal shadows of the lung with visual loss. She was diagnosed with SCLC associated with CAR. Subsequent chest X-ray and CT scan showed partial regression of both primary tumor and lymph node metastasis without anticancer treatment. Recoverin antigen was present on the tumor cells and anti-recoverin antibody was observed in the patient's serum. Activation of recoverin-specific antitumor cytotoxic T lymphocyte (CTL) was observed in this patient. SCLC was considered to reduce spontaneously by the activation of recoverin-specific antitumor CTL. To the best of our knowledge, this is the first report of SR in SCLC combined with CAR.