HER3 overexpression: a predictive marker for poor prognosis in advanced ALK-positive non-small cell lung cancer treated with ALK inhibitors.

Background: Anaplastic lymphoma kinase (ALK)-targeted tyrosine kinase inhibitors (TKIs) improve patient survival; however, some patients develop ALK-TKI resistance with unidentified mechanisms. We investigated ErbB family and c-MET expression in patients with ALK-positive non-small cell lung cancer (NSCLC) to understand their roles in the ALK-TKI response. Methods: We studied 72 patients with advanced ALK-positive NSCLC with EML4-ALK fusion variant subtyping and immunostaining for c-MET, EGFR, HER2, and HER3 on tissue specimens both pre- (primary) and post-treatment (secondary) with ALK-TKI. We investigated the association of their expression with survival outcomes and assessed the effectiveness of combining ALK and EGFR inhibitors in ALK-positive NSCLC cell lines stimulated with the HER3-specific ligand HRG1. Results: High expression of c-MET, EGFR, HER2, and HER3 was observed in 4.9%, 18.0%, 1.6%, and 25.8% of primary tumors, respectively, and 18.5%, 37.0%, 10.7%, and 35.7% of secondary tumors, respectively. HER3 overexpression in primary tumors showed inferior survival (P=0.132). In the subgroup with EML4-ALK variant 1/2 (V1/V2), HER3 overexpression was significantly associated with inferior survival in both primary and secondary tumors (P=0.022 and P=0.004, respectively). Combination treatment with lorlatinib and erlotinib significantly reduced HRG1-induced activation of RTK signaling in ALK-positive NSCLC cells. Conclusions: HER3 overexpression has potential as a prognostic marker in ALK-positive NSCLCs, including ALK-TKI naïve and treated cases, especially those with EML4-ALK V1/V2. Assessing HER3 expression may be crucial for treatment planning and outcome prediction in these patients.

Development of polysaccharide-complexed nano-sized rice protein dispersion.

The objective of this study was to improve water solubility of the rice protein (RP) by forming complexes with anionic polysaccharides, such as sodium alginate (SA) and xanthan gum (XG). In addition, utilization of the RP complexes as an emulsifier was evaluated. The prepared RP-SA or RP-XG complexes were analyzed by measuring their particle size, ζ-potential, and water solubility as well as by confocal laser scanning microscopy. The formation of a complex between RP-SA and RP-XG improved the water solubility and dispersibility of RP over a wide range of pH values (3, 5, 7, and 9). Confocal fluorescence images showed that the aggregation of RP molecules was prevented by the formation of complexes between RP and polysaccharides. When soybean oil-in-water emulsions were prepared with complexes, RP-SA (ratio 4:1) and RP-XG(ratio 4:1) complex-stabilized emulsions were stable for 4 weeks of storage.

c-MET-positive circulating tumor cells and cell-free DNA as independent prognostic factors in hormone receptor-positive/HER2-negative metastatic breast cancer.

BACKGROUND: Endocrine therapy resistance in hormone receptor-positive/HER2-negative (HR+/HER2-) breast cancer (BC) is a significant clinical challenge that poses several unmet needs in the management of the disease. This study aimed to investigate the prognostic value of c-MET-positive circulating tumor cells (cMET+ CTCs), ESR1/PIK3CA mutations, and cell-free DNA (cfDNA) concentrations in patients with hormone receptor-positive (HR+) metastatic breast cancer (mBC). METHODS: Ninety-seven patients with HR+ mBC were prospectively enrolled during standard treatment at Samsung Medical Center. CTCs were isolated from blood using GenoCTC® and EpCAM or c-MET CTC isolation kits. PIK3CA and ESR1 hotspot mutations were analyzed using droplet digital PCR. CfDNA concentrations were calculated using internal control copies from the ESR1 mutation test. Immunocytochemistry was performed to compare c-MET overexpression between primary and metastatic sites. RESULTS: The proportion of c-MET overexpression was significantly higher in metastatic sites than in primary sites (p = 0.00002). Survival analysis showed that c-MET+ CTC, cfDNA concentration, and ESR1 mutations were significantly associated with poor prognosis (p = 0.0026, 0.0021, and 0.0064, respectively) in HR+/HER2- mBC. By contrast, EpCAM-positive CTC (EpCAM+ CTC) and PIK3CA mutations were not associated with progression-free survival (PFS) in HR+/HER2- mBC. Multivariate analyses revealed that c-MET+ CTCs and cfDNA concentration were independent predictors of PFS in HR+/HER2- mBC. CONCLUSIONS: Monitoring c-MET+ CTC, rather than assessing c-MET expression in the primary BC site, could provide valuable information for predicting disease progression, as c-MET expression can change during treatment. The c-MET+ CTC count and cfDNA concentration could provide complementary information on disease progression in HR+ /HER2- mBC, highlighting the importance of integrated liquid biopsy.

Modulation of warm temperature-sensitive growth using a phytochrome B dark reversion variant, phyB[G515E], in Arabidopsis and rice.

INTRODUCTION: Ambient temperature-induced hypocotyl elongation in Arabidopsis seedlings is sensed by the epidermis-localized phytochrome B (phyB) and transduced into auxin biosynthesis via a basic helix-loop-helix transcription factor, phytochrome-interacting factor 4 (PIF4). Once synthesized, auxin travels down from the cotyledons to the hypocotyl, triggering hypocotyl cell elongation. Thus, the phyB-PIF4 module involved in thermosensing and signal transduction is a potential genetic target for engineering warm temperature-insensitive plants. OBJECTIVES: This study aims to manipulate warm temperature-induced elongation of plants at the post-translational level using phyB variants with dark reversion, the expression of which is subjected to heat stress. METHODS: The thermosensitive growth response of Arabidopsis was manipulated by expressing the single amino acid substitution variant of phyB (phyB[G515E]), which exhibited a lower dark reversion rate than wild-type phyB. Other variants with slow (phyB[G564E]) or rapid (phyB[S584F]) dark reversion or light insensitivity (phyB[G767R]) were also included in this study for comparison. Warming-induced transient expression of phyB variants was achieved using heat shock-inducible promoters. Arabidopsis PHYB[G515E] and PHYB[G564E] were also constitutively expressed in rice in an attempt to manipulate the heat sensitivity of a monocotyledonous plant species. RESULTS: At an elevated temperature, Arabidopsis seedlings transiently expressing PHYB[G515E] under the control of a heat shock-inducible promoter exhibited shorter hypocotyls than those expressing PHYB and other PHYB variant genes. This warm temperature-insensitive growth was related to the lowered PIF4 and auxin responses. In addition, transgenic rice seedlings expressing Arabidopsis PHYB[G515E] and PHYB[G564E] showed warm temperature-insensitive shoot growth. CONCLUSION: Transient expression of phyB variants with altered dark reversion rates could serve as an effective optogenetic technique for manipulating PIF4-auxin-mediated thermomorphogenic responses in plants.

Color-Tuning Mechanism of the Lit Form of Orange Carotenoid Protein.

Orange carotenoid protein (OCP) of photosynthetic cyanobacteria binds to ketocarotenoids noncovalently and absorbs excess light to protect the host organism from light-induced oxidative damage. Herein, we found that mutating valine 40 in the α3 helix of Gloeocapsa sp. PCC 7513 (GlOCP1) resulted in blue- or red-shifts of 6-20 nm in the absorption maxima of the lit forms. We analyzed the origins of absorption maxima shifts by integrating X-ray crystallography, homology modeling, molecular dynamics simulations, and hybrid quantum mechanics/molecular mechanics calculations. Our analysis suggested that the single residue mutations alter the polar environment surrounding the bound canthaxanthin, thereby modulating the degree of charge transfer in the photoexcited state of the chromophore. Our integrated investigations reveal the mechanism of color adaptation specific to OCPs and suggest a design principle for color-specific photoswitches.

Awe promotes moral expansiveness via the small-self.

The experience of awe has been shown to challenge how people think about themselves and the world around them, linking them to something greater than themselves. We investigated whether this emotional experience of awe may also challenge the boundaries of our moral consideration, leading to a generalized expansion in our moral worlds. Across five studies (N = 990), we examined whether awe might promote moral expansiveness; that is, increased moral concern across a broad range of entities (e.g., out-groups, animals, plants, environments). Cross-sectional Studies 1a and 1b, found dispositional awe was related to greater moral expansiveness. Experimental Studies 2 and 3, using video-induced awe, found consistent indirect effects on moral expansiveness, via self-reported awe and the small-self sense of vastness. Experimental Study 4, using Virtual Reality induced awe, found those in the awe condition (vs. control) reported greater moral expansiveness, and this was fully mediated by the small-self sense of vastness. Our findings show awe expands our sense of connectedness to the broader world, and through this, increases the breath of our moral concern.

Overexpression of the NEK9-EG5 axis is a novel metastatic marker in pathologic stage T3 colon cancer.

NEK9 is a key player in the NEK9-EG5 axis for microtubule polymerization, chromosome alignment, and mitosis. In present study, we investigated the altered expression of the NEK9, EG5 and acetyl-α-tubulin as well as common epithelial-mesenchymal transition (EMT) markers (E-cadherin, vimentin, claudin-1, and ß-catenin) through the immunohistochemistry analysis of 138 patients with pathologic T3 (pT3) stage colon cancers, and evaluated their metastatic potential. NEK9 expression showed an association with distant metastasis (P = 0.032) and was an independent predictive factor for distant metastasis (HR = 3.365, P < 0.001) by multivariate analysis, which was more significant than either the regional nodal metastasis (HR = 2.496, P = 0.007) or lymphovascular invasion (HR = 2.090, P = 0.153). Positive correlations were observed between NEK9 and EG5 or acetyl-α-tubulin (r = 0.236 and P = 0.007; r = 0.181 and P = 0.038, respectively) and concordant overexpression of the NEK9-EG5 axis was further confirmed in colon cancer cell lines. These findings collectively suggest that the overexpression of the NEK9-EG5 axis is present and associated with distant metastasis in colon cancer. These biomarkers might be useful for predicting metastatic potential among the patients with pT3 colon cancers.

Cancer-Specific Sequences in the Diagnosis and Treatment of NUT Carcinoma.

PURPOSE: NUT carcinoma (NC) is a solid tumor caused by the rearrangement of NUTM1 that usually develops in midline structures, such as the thorax. No standard treatment has been established despite high lethality. Thus, we investigated whether targeting the junction region of NUTM1 fusion breakpoints could serve as a potential treatment option for NC. Materials and Methods: We designed and evaluated a series of small interfering RNAs (siRNAs) targeting the junction region of BRD4-NUTM1 fusion (B4N), the most common form of NUTM1 fusion. Droplet digital polymerase chain reaction using the blood of patients was also tested to evaluate the treatment responses by the junction sequence of the B4N fusion transcripts. RESULTS: As expected, the majority of NC fusion types were B4N (12 of 18, 67%). B4N fusion-specific siRNA treatment on NC cells showed specific inhibitory effects on the B4N fusion transcript and fusion protein without affecting the endogenous expression of the parent genes, resulting in decreased relative cell growth and attenuation of tumor size. In addition, the fusion transcript levels in platelet-rich-plasma samples of the NC patients with systemic metastasis showed a negative correlation with therapeutic effect, suggesting its potential as a measure of treatment responsiveness. CONCLUSION: This study suggests that tumor-specific sequences could be used to treat patients with fusion genes as part of precision medicine for a rare but deadly disease.

Analysis of PIK3CA Mutation Concordance and Frequency in Primary and Different Distant Metastatic Sites in Breast Cancer.

PURPOSE: The purpose of this study was to investigate the concordance rate of PIK3CA mutations between primary and matched distant metastatic sites in patients with breast cancer and to verify whether there are differences in the frequency of PIK3CA hotspot mutations depending on the metastatic sites involved. MATERIALS AND METHODS: Archived formalin-fixed paraffin-embedded (FFPE) specimens of primary breast and matched distant metastatic tumors were retrospectively obtained for 49 patients. Additionally, 40 archived FFPE specimens were independently collected from different breast cancer metastatic sites, which were limited to three common sites: the liver, brain, and lung. PIK3CA mutations were analyzed using droplet digital PCR, including hotspots involving exons 9 and 20. RESULTS: After analysis of 49 breast tumors with matched metastasis sites, 87.8% showed concordance in PIK3CA mutation status. According to PIK3CA hotspot mutation testing in 89 cases of breast cancer metastatic sites, the proportion of PIK3CA mutations at sites of metastasis involving the liver, brain, and lung was 37.5%, 28.6%, and 42.9%, respectively, which did not result in statistical significance. CONCLUSION: The high concordance of PIK3CA mutation status between primary and matched metastasis sites suggests that metastatic sites, regardless of the metastatic organ, could be considered sample sources for PIK3CA mutation testing for improved therapeutic strategies in patients with metastatic breast cancer.

Unearthing novel fusions as therapeutic targets in solid tumors using targeted RNA sequencing.

Detection of oncogenic fusion genes in cancers, particularly in the diagnosis of uncertain tumors, is crucial for determining effective therapeutic strategies. Although novel fusion genes have been discovered through sequencing, verifying their oncogenic potential remain difficult. Therefore, we evaluated the utility of targeted RNA sequencing in 165 tumor samples by identifying known and unknown fusions. Additionally, by applying additional criteria, we discovered eight novel fusion genes that are expected to process oncogenicity. Among the novel fusion genes, RAF1 fusion genes were detected in two cases. PTPRG-RAF1 fusion led to an increase in cell growth; while dabrafenib, a BRAF inhibitor, reduced the growth of cells expressing RAF1. This study demonstrated the utility of RNA panel sequencing as a theragnostic tool and established criteria for identifying oncogenic fusion genes during post-sequencing analysis.

A Polytherapy Strategy Using Vincristine and ALK Inhibitors to Sensitise EML4-ALK-Positive NSCLC.

The oncogenic fusion of EML4-ALK is present in about 4-6% of non-small cell lung cancer (NSCLC). A targeted approach with ALK tyrosine kinase inhibitors (TKIs) has been proven highly effective in ALK-positive NSCLC patients. However, despite the initial responses, the outcome of the treatment is variable. Previous studies have shown that the differential response depends in part on the type of EML4-ALK variant. Here, we examined the combination of ALK inhibitors and microtubule poison, vincristine, in cells expressing EML4-ALK V1 and V3, the two most common variants in NSCLC. We showed that combination therapy of ALK-TKIs with vincristine had anti-proliferative effects and blocked RAS/MAPK, PI3K/AKT and JAK/STAT3 signalling pathways in EML4-ALK V1 but not V3 cells. Our results demonstrate that high levels of tubulin acetylation are associated with poor response to vincristine in EML4-ALK V3 cells. Additionally, we demonstrated differences in microtubule stability between the two EML4-ALK fusions. EML4-ALK V3 cells exhibited dynamic microtubules that confer poor response to vincristine compared to V1 cells. Hence, we suggested that the portion of EML4 in the fusion has an important role for the outcome of the combination treatment.

Production of IL-31 in CD45RO+CLA+H4R+ T Cells in Atopic Dermatitis.

IL-31 is involved in pruritus in atopic dermatitis (AD) and the pathogenesis of AD. However, the mechanism of IL-31 production is not fully understood. We sought to investigate the association between CD45RO+CLA+H4R+ T cells and IL-31 production. Immunofluorescence studies were performed retrospectively on punch-biopsy specimens from five people with AD and three healthy controls. In addition, blood samples were collected prospectively from eight patients with AD and eight healthy controls for sorting CD45RO+CLA+H4R+ T cells. There was no overlap of patients between the biopsy group and the blood sampling group. Sorted cells were stimulated with 4-methylhistamine (4MH), and the level of IL-31 was measured by an enzyme-linked immunosorbent assay. Immunofluorescence showed co-localization of H4R and IL-31 in lesional AD skin but not in normal skin of healthy controls. The proportion of CLA+H4R+ T cells among CD3+CD45RO+ lymphocytes was 18.3 ± 6.2% in patients with AD and 11.2 ± 7.6% in healthy controls. In the AD group, production of IL-31 by CD45RO+CLA+H4R+ T cells increased from 32.4 ± 13.3 pg/mL to 47.5 ± 18.7 pg/mL by 4MH stimulation after 24 h (p < 0.001). However, in the control group, production of IL-31 was 20.1 ± 10.6 pg/mL without and 22.1 ± 9.3 pg/mL with 4MH stimulation (p > 0.05). According to our study, CD45RO+CLA+H4R+ T cells are an important source of IL-31 in AD, and may be a target for treatment of IL-31-induced pruritus.

Implant complications in bruxism patients.

Bruxism is defined as a parafunctional activity during sleep or while awake that includes locking and grinding of teeth and clenching. It generates excessive occlusal force that may lead to implant failure. Therefore, diagnosis of bruxism and providing specific protocols such as occlusal splint and/or injection of botulinum toxin before implant installation are important to prevent increases the risk of implant failure in bruxism patients.

Epigallocatechin-3-gallate Can Prevent Type 2 Human Papillomavirus E7 from Suppressing Interferon-Stimulated Genes.

Human papillomavirus (HPV) in high-risk groups is known to suppress the type I interferon (IFN) signaling pathway leading to the transcription of interferon-stimulated genes (ISGs), which have many antiviral functions. However, the effects of HPV on the action of various ISGs in low-risk groups are not fully understood. We aimed to investigate whether antiviral ISGs are expressed in transfected keratinocytes with type 2 HPV (HPV-2) E7. The mRNA and protein expressions of ISGs and type I IFN signaling pathway components were evaluated by quantitative real-time polymerase chain reaction, western blot, immunofluorescence, and/or immunohistochemistry. Compared with normal skin, mRNA expression of all ISGs in HPV-2 positive cutaneous warts was significantly decreased (p < 0.05). In comparison with empty vector transfection, E7 transfection significantly down-regulated the mRNA and protein expressions of ISGs and type I IFN signaling pathway components, which were significantly up-regulated by E7 siRNA transfection (p < 0.05). Interestingly, epigallocatechin-3-gallate (EGCG) pretreatment up-regulated the mRNA and protein expressions of ISGs and type I IFN signaling pathway components, which were significantly down-regulated by E7 transfection (p < 0.05). Our results demonstrate that EGCG is a potential candidate for cutaneous wart prevention.

Type 2 human papillomavirus E7 attenuates E-cadherin expression in human keratinocytes.

Human papillomaviruses (HPVs) are known to utilize the down-regulation of epithelial (E)-cadherin, a major component of adherens junctions of keratinocytes, to evade host immune surveillance in high-risk group. However, the effects of HPV on the function of E-cadherin in low-risk groups remain unknown. We investigated whether type 2 HPV (HPV-2) E7 could induce alterations in E-cadherin expression in transiently transfected keratinocytes and cell lines expressing HPV-2 E7. To examine the expression pattern of E-cadherin in cutaneous warts and normal skin samples, immunohistochemical analysis was performed. Quantitative real-time polymerase chain reactions, luciferase assays, western blot, immunocytochemistry, and electron microscopy were used to evaluate the mRNA and protein expression levels of E-cadherin in normal human epidermal keratinocytes transfected with HPV-2 E7 plasmid DNA or E7-specific siRNA and in E7-expressing cell lines. E-cadherin expression levels in HPV-2 positive cutaneous warts were significantly decreased compared to those in normal skin (p < 0.05). Similarly, the mRNA and protein expression levels of E-cadherin in E7 transiently transfected cells were significantly decreased compared to those in empty vector-transfected cells. The decreases were restored by transfection with E7-specific siRNA (p < 0.05). Likewise, cell lines expressing E7 showed a decreased expression of E-cadherin. When the cells were cultured in low attachment plates, cell-to-cell aggregation was inhibited. Taken together, our data suggest that HPV-2 E7, the causative agent of cutaneous warts, could mediate the transcriptional repression of E-cadherin.

TM4SF4 and LRRK2 Are Potential Therapeutic Targets in Lung and Breast Cancers through Outlier Analysis.

PURPOSE: To find biomarkers for disease, there have been constant attempts to investigate the genes that differ from those in the disease groups. However, the values that lie outside the overall pattern of a distribution, the outliers, are frequently excluded in traditional analytical methods as they are considered to be 'some sort of problem.' Such outliers may have a biologic role in the disease group. Thus, this study explored new biomarker using outlier analysis, and verified the suitability of therapeutic potential of two genes (TM4SF4 and LRRK2). MATERIALS AND METHODS: Modified Tukey's fences outlier analysis was carried out to identify new biomarkers using the public gene expression datasets. And we verified the presence of the selected biomarkers in other clinical samples via customized gene expression panels and tissue microarrays. Moreover, a siRNA-based knockdown test was performed to evaluate the impact of the biomarkers on oncogenic phenotypes. RESULTS: TM4SF4 in lung cancer and LRRK2 in breast cancer were chosen as candidates among the genes derived from the analysis. TM4SF4 and LRRK2 were overexpressed in the small number of samples with lung cancer (4.20%) and breast cancer (2.42%), respectively. Knockdown of TM4SF4 and LRRK2 suppressed the growth of lung and breast cancer cell lines. The LRRK2 overexpressing cell lines were more sensitive to LRRK2-IN-1 than the LRRK2 under-expressing cell lines. CONCLUSION: Our modified outlier-based analysis method has proved to rescue biomarkers previously missed or unnoticed by traditional analysis showing TM4SF4 and LRRK2 are novel target candidates for lung and breast cancer, respectively.

[Opening Status of the Korea Midwifery Birthing Centers and Development of Midwifery Practice Guideline].

PURPOSE: This study was to investigate the operational status of the midwifery birthing centers (MBCs) and midwives' job status (Phase 1) and to develop midwifery practice guidelines (MPG) (Phase 2) in Korea. METHODS: In the first phase, the subjects were 15 midwives who operated 11 of 14 MBCs that were opened as of August 2018. The questionnaire consisted of items to measure the operational status of the MBC and midwives' job status. In the second phase, the MPG was developed from literature review, interviews with five midwives opening their MBCs, surveys with 74 midwives, and a validity evaluation conducted by seven experts. RESULTS: The distribution of operating MBCs was five in Gyunggi-do, two each in Seoul and Incheon, one each in Busan, Chungcheongbuk-do, Gyeongsangbuk-do, Gyeongsangnam-do and Jeju-do. The mean age of midwives was 54.3 and all were female. In 2017, a total of 762 births including 81 homebirths were performed by midwives. The job performance was highest in the order of neonatal care 3.81, childbirth care 3.56, and postpartal care 3.53, respectively. The MPG included seven areas of prenatal care, childbirth care, postpartal care, neonatal care, primary health care, law/ethics, and administration, with 56 tasks and 166 task elements. CONCLUSION: This study provides the valid basic data for the operational status of the MBC and the midwives' job status. The MPG describes the midwife's job and may be used as basic data for preparing policies for the development of midwifery practice in Korea.

Spectral and photochemical diversity of tandem cysteine cyanobacterial phytochromes.

The atypical trichromatic cyanobacterial phytochrome NpTP1 from Nostoc punctiforme ATCC 29133 is a linear tetrapyrrole (bilin)-binding photoreceptor protein that possesses tandem-cysteine residues responsible for shifting its light-sensing maximum to the violet spectral region. Using bioinformatics and phylogenetic analyses, here we established that tandem-cysteine cyanobacterial phytochromes (TCCPs) compose a well-supported monophyletic phytochrome lineage distinct from prototypical red/far-red cyanobacterial phytochromes. To investigate the light-sensing diversity of this family, we compared the spectroscopic properties of NpTP1 (here renamed NpTCCP) with those of three phylogenetically diverged TCCPs identified in the draft genomes of Tolypothrix sp. PCC7910, Scytonema sp. PCC10023, and Gloeocapsa sp. PCC7513. Recombinant photosensory core modules of ToTCCP, ScTCCP, and GlTCCP exhibited violet-blue-absorbing dark-states consistent with dual thioether-linked phycocyanobilin (PCB) chromophores. Photoexcitation generated singly-linked photoproduct mixtures with variable ratios of yellow-orange and red-absorbing species. The photoproduct ratio was strongly influenced by pH and by mutagenesis of TCCP- and phytochrome-specific signature residues. Our experiments support the conclusion that both photoproduct species possess protonated 15E bilin chromophores, but differ in the ionization state of the noncanonical "second" cysteine sulfhydryl group. We found that the ionization state of this and other residues influences subsequent conformational change and downstream signal transmission. We also show that tandem-cysteine phytochromes present in eukaryotes possess similar amino acid substitutions within their chromophore-binding pocket, which tune their spectral properties in an analogous fashion. Taken together, our findings provide a roadmap for tailoring the wavelength specificity of plant phytochromes to optimize plant performance in diverse natural and artificial light environments.

IRS2 Amplification as a Predictive Biomarker in Response to Ceritinib in Small Cell Lung Cancer.

Small cell lung cancer (SCLC) is a fast-growing and malignant cancer that responds well to chemotherapy; however, the survival rate is less than 15% after 2 years of diagnosis. Therefore, novel therapeutic agents for treating SCLC patients need to be evaluated. This study aims to identify the therapeutic targets based on the comprehensive genomic profiling of SCLC patients. Among the molecular-profiled SCLC samples obtained using targeted sequencing, the array-based comparative genomic hybridization (array CGH) identified focal insulin receptor substrate 2 (IRS2) amplification in the SCLC patients. IRS2 amplification was confirmed in 5% of 73 SCLC patients. To determine whether IRS2 amplification could act as a therapeutic target, we generated a patient-derived xenograft (PDX) model and subsequently screened 43 targeted agents using the PDX-derived cells (PDCs). Ceritinib significantly inhibited the cell growth and impaired the tumor sphere formation in IRS2-expressing PDCs. Its effects were confirmed in various in vitro assays and were further validated in the mouse xenograft models. In this study, we present that IRS2 amplification and/or expression serve as preclinical implications for a novel therapeutic target in SCLC progression. Furthermore, we suggest that insulin-like growth factor-1 (IGF-1) receptor inhibitor-based therapy could be used for treating SCLC with IRS2 amplification.

Peptide transporter2 (PTR2) enhances water uptake during early seed germination in Arabidopsis thaliana.

KEY MESSAGE: PTR2 in Arabidopsis thaliana is negatively regulated by ABI4 and plays a key role in water uptake by seeds, ensuring that imbibed seeds proceed to germination. Peptide transporters (PTRs) transport nitrogen-containing substrates in a proton-dependent manner. Among the six PTRs in Arabidopsis thaliana, the physiological role of the tonoplast-localized, seed embryo abundant PTR2 is unknown. In the present study, a molecular physiological analysis of PTR2 was conducted using ptr2 mutants and PTR2CO complementation lines. Compared with the wild type, the ptr2 mutant showed ca. 6 h delay in testa rupture and consequently endosperm rupture because of 17% lower water content and 10% higher free abscisic acid (ABA) content. Constitutive overexpression of the PTR2 gene under the control of the Cauliflower mosaic virus (CaMV) 35S promoter in ptr2 mutants rescued the mutant phenotypes. After cold stratification, a transient increase in ABA INSENSITIVE4 (ABI4) transcript levels during induction of testa rupture was followed by a similar increase in PTR2 transcript levels, which peaked prior to endosperm rupture. The PTR2 promoter region containing multiple CCAC motifs was recognized by ABI4 in electrophoretic mobility shift assays, and PTR2 expression was repressed by 67% in ABI4 overexpression lines compared with the wild type, suggesting that PTR2 is an immediate downstream target of ABI4. Taken together, the results suggest that ABI4-dependent temporal regulation of PTR2 expression may influence water status during seed germination to promote the post-germinative growth of imbibed seeds.