RNA molecules with novel functions have revived interest in the accurate prediction of RNA three-dimensional (3D) structure and folding dynamics. However, existing methods are inefficient in automated 3D structure prediction. Here, we report a robust computational approach for rapid folding of RNA molecules. We develop a simplified RNA model for discrete molecular dynamics (DMD) simulations, incorporating base-pairing and base-stacking interactions. We demonstrate correct folding of 150 structurally diverse RNA sequences. The majority of DMD-predicted 3D structures have <4 Å deviations from experimental structures. The secondary structures corresponding to the predicted 3D structures consist of 94% native base-pair interactions. Folding thermodynamics and kinetics of tRNA Phe , pseudoknots, and mRNA fragments in DMD simulations are in agreement with previous experimental findings. Folding of RNA molecules features transient, non-native conformations, suggesting nonhierarchical RNA folding. Our method allows rapid conformational sampling of RNA folding, with computational time increasing linearly with RNA length. We envision this approach as a promising tool for RNA structural and functional analyses.

Fluorescent proteins have proven to be excellent reporters and biochemical sensors with a wide range of applications. In a split form, they are not fluorescent, but their fluorescence can be restored by supplementary protein-protein or protein-nucleic acid interactions that reassemble the split polypeptides. However, in prior studies, it took hours to restore the fluorescence of a split fluorescent protein because the formation of the protein chromophore slowly occurred de novo concurrently with reassembly. Here we provide evidence that a fluorogenic chromophore can self-catalytically form within an isolated N-terminal fragment of the enhanced green fluorescent protein (EGFP). We show that restoration of the split protein fluorescence can be driven by nucleic acid complementary interactions. In our assay, fluorescence development is fast (within a few minutes) when complementary oligonucleotide-linked fragments of the split EGFP are combined. The ability of our EGFP system to respond quickly to DNA hybridization should be useful for detecting the kinetics of many other types of pairwise interactions both in vitro and in living cells.split EGFP ͉ DNA duplex ͉ EGFP reassembly ͉ protein folding ͉ DMD simulations S plit fluorescent proteins are convenient tools to detect specific protein-protein or protein-nucleic acid interactions (1-5). The approach is based on the reassembly of a fluorescent protein from two nonfluorescent fragments driven by additional biomolecular interactions, and it results in restoration of fluorescence. The development of fluorescence, however, usually takes several hours because of the requirement of the de novo formation of the chromophore within the reassembled protein (6). Because this approach provides a slow response, it would clearly be advantageous to accelerate it.A straightforward way to do this would be to use a fragment of a split protein with a preformed chromophore that is not fluorescent per se but is capable of bright fluorescence within a full-size protein. To the best of our knowledge, such a strategy has not been previously accomplished. In this report, we demonstrate the feasibility of an alternative approach based on the nucleic acidsupported fast complementation of EGFP fragments, one of which contains a mature profluorescent chromophore. Results and Discussion Molecular Modeling of Protein Folding: Large EGFP Fragment CanPotentially Form a Chromophore. In this study, we used two fragments of the EGFP, which are linked, in its native structure, by a flexible loop of nine amino acids, residues 153-161 (7, 8). The larger, N-terminal EGFP fragment is known to contain the three amino acids that form a chromophore, which is fluorescent in native, but not in denatured, protein (6, 7). It is also known that this tripeptide chromophore exhibits no fluorescence in a separate large EGFP fragment (2, 4). EGFP chromophore formation is a self-catalytic process requiring correct protein folding (6). We were curious to see whether the N-terminal EGFP fragment (approximately two-third...

Background NF1 is a tumor suppressor that negatively regulates Ras signaling. NF1 mutations occur in lung cancer, but their clinical significance is unknown. We evaluated clinical and molecular characteristics of NF1 mutant lung cancers with comparison to tumors with KRAS mutations. Methods Between July 2013 and October 2014, 591 NSCLC tumors underwent targeted next generation sequencing in a 275 gene panel that evaluates gene mutations and genomic rearrangements. NF1 and KRAS cohorts were identified, with subsequent clinical and genomic analysis. Results Among 591 pts, 60 had NF1 mutations (10%) and 141 (24%) had KRAS mutations. 15 NF1 mutations (25%) occurred with other oncogenic mutations (BRAF (2); ERBB2 (2); KRAS (9); HRAS (1); NRAS (1)). There were 72 unique NF1 variants. NF1 tumor pathology was diverse, including both adenocarcinoma (36, 60%) and squamous cell carcinoma (10, 17%). In contrast, KRAS mutations occurred predominantly in adenocarcinoma (136, 96%). Both mutations were common in former/current smokers. Among NF1 tumors without concurrent oncogenic alterations, TP53 mutations/2-copy deletions occurred more often (33, 65%) than with KRAS mutation (46, 35%) (p<.001). No difference between cohorts was seen with other tumor suppressors. Conclusions NF1 mutations define a unique population of NSCLC. NF1 and KRAS mutations present in similar patient populations, but NF1 mutations occur more often with other oncogenic alterations and TP53 mutations. Therapeutic strategies targeting KRAS activation, including inhibitors of MAP kinase signaling, may warrant investigation in NF1 mutant tumors. Tumor suppressor inactivation patterns may help further define novel treatment strategies.

We live in the genomic era of medicine, where a patient's genomic/molecular data is becoming increasingly important for disease diagnosis, identification of targeted therapy, and risk assessment for adverse reactions. However, decoding the genomic test results and integrating it with clinical data for retrospective studies and cohort identification for prospective clinical trials is still a challenging task. In order to overcome these barriers, we developed an overarching enterprise informatics framework for translational research and personalized medicine called Synergistic Patient and Research Knowledge Systems (SPARKS) and a suite of tools called Oncology Data Retrieval Systems (OncDRS). OncDRS enables seamless data integration, secure and self-navigated query and extraction of clinical and genomic data from heterogeneous sources. Within a year of release, the system has facilitated more than 1500 research queries and has delivered data for more than 50 research studies.

Background-NF1 is a tumor suppressor that negatively regulates Ras signaling. NF1 mutations occur in lung cancer, but their clinical significance is unknown. We evaluated clinical and molecular characteristics of NF1 mutant lung cancers with comparison to tumors with KRAS mutations. Methods-Between July 2013 and October 2014, 591 NSCLC tumors underwent targeted next generation sequencing in a 275 gene panel that evaluates gene mutations and genomic rearrangements. NF1 and KRAS cohorts were identified, with subsequent clinical and genomic analysis. Results-Among 591 pts, 60 had NF1 mutations (10%) and 141 (24%) had KRAS mutations. 15 NF1 mutations (25%) occurred with other oncogenic mutations (BRAF (2); ERBB2 (2); KRAS (9); HRAS (1); NRAS (1)). There were 72 unique NF1 variants. NF1 tumor pathology was diverse, including both adenocarcinoma (36, 60%) and squamous cell carcinoma (10, 17%). In contrast, KRAS mutations occurred predominantly in adenocarcinoma (136, 96%). Both mutations were common in former/current smokers. Among NF1 tumors without concurrent oncogenic alterations, TP53 mutations/2-copy deletions occurred more often (33, 65%) than with KRAS mutation (46, 35%) (p<.001). No difference between cohorts was seen with other tumor suppressors. Conclusions-NF1 mutations define a unique population of NSCLC. NF1 and KRAS mutations present in similar patient populations, but NF1 mutations occur more often with other oncogenic

<div>Abstract<p><b>Purpose:</b><i>NF1</i> is a tumor suppressor that negatively regulates Ras signaling. <i>NF1</i> mutations occur in lung cancer, but their clinical significance is unknown. We evaluated clinical and molecular characteristics of <i>NF1</i> mutant lung cancers with comparison to tumors with <i>KRAS</i> mutations.</p><p><b>Experimental Design:</b> Between July 2013 and October 2014, 591 non–small cell lung cancer (NSCLC) tumors underwent targeted next-generation sequencing in a 275 gene panel that evaluates gene mutations and genomic rearrangements. <i>NF1</i> and <i>KRAS</i> cohorts were identified, with subsequent clinical and genomic analysis.</p><p><b>Results:</b> Among 591 patients, 60 had <i>NF1</i> mutations (10%) and 141 (24%) had <i>KRAS</i> mutations. 15 <i>NF1</i> mutations (25%) occurred with other oncogenic mutations [BRAF (2); ERBB2 (2); KRAS (9); HRAS (1); NRAS (1)]. There were 72 unique <i>NF1</i> variants. <i>NF1</i> tumor pathology was diverse, including both adenocarcinoma (36, 60%) and squamous cell carcinoma (10, 17%). In contrast, <i>KRAS</i> mutations occurred predominantly in adenocarcinoma (136, 96%). Both mutations were common in former/current smokers. Among <i>NF1</i> tumors without concurrent oncogenic alterations, <i>TP53</i> mutations/2-copy deletions occurred more often (33, 65%) than with <i>KRAS</i> mutation (46, 35%; <i>P</i> < 0.001). No difference between cohorts was seen with other tumor suppressors.</p><p><b>Conclusions:</b><i>NF1</i> mutations define a unique population of NSCLC. <i>NF1</i> and <i>KRAS</i> mutations present in similar patient populations, but <i>NF1</i> mutations occur more often with other oncogenic alterations and <i>TP53</i> mutations. Therapeutic strategies targeting <i>KRAS</i> activation, including inhibitors of MAP kinase signaling, may warrant investigation in <i>NF1</i> mutant tumors. Tumor-suppressor inactivation patterns may help further define novel treatment strategies. <i>Clin Cancer Res; 22(13); 3148–56. ©2016 AACR</i>.</p></div>

<div>Abstract<p><b>Purpose:</b><i>NF1</i> is a tumor suppressor that negatively regulates Ras signaling. <i>NF1</i> mutations occur in lung cancer, but their clinical significance is unknown. We evaluated clinical and molecular characteristics of <i>NF1</i> mutant lung cancers with comparison to tumors with <i>KRAS</i> mutations.</p><p><b>Experimental Design:</b> Between July 2013 and October 2014, 591 non–small cell lung cancer (NSCLC) tumors underwent targeted next-generation sequencing in a 275 gene panel that evaluates gene mutations and genomic rearrangements. <i>NF1</i> and <i>KRAS</i> cohorts were identified, with subsequent clinical and genomic analysis.</p><p><b>Results:</b> Among 591 patients, 60 had <i>NF1</i> mutations (10%) and 141 (24%) had <i>KRAS</i> mutations. 15 <i>NF1</i> mutations (25%) occurred with other oncogenic mutations [BRAF (2); ERBB2 (2); KRAS (9); HRAS (1); NRAS (1)]. There were 72 unique <i>NF1</i> variants. <i>NF1</i> tumor pathology was diverse, including both adenocarcinoma (36, 60%) and squamous cell carcinoma (10, 17%). In contrast, <i>KRAS</i> mutations occurred predominantly in adenocarcinoma (136, 96%). Both mutations were common in former/current smokers. Among <i>NF1</i> tumors without concurrent oncogenic alterations, <i>TP53</i> mutations/2-copy deletions occurred more often (33, 65%) than with <i>KRAS</i> mutation (46, 35%; <i>P</i> < 0.001). No difference between cohorts was seen with other tumor suppressors.</p><p><b>Conclusions:</b><i>NF1</i> mutations define a unique population of NSCLC. <i>NF1</i> and <i>KRAS</i> mutations present in similar patient populations, but <i>NF1</i> mutations occur more often with other oncogenic alterations and <i>TP53</i> mutations. Therapeutic strategies targeting <i>KRAS</i> activation, including inhibitors of MAP kinase signaling, may warrant investigation in <i>NF1</i> mutant tumors. Tumor-suppressor inactivation patterns may help further define novel treatment strategies. <i>Clin Cancer Res; 22(13); 3148–56. ©2016 AACR</i>.</p></div>

<p>Supplemental Table 1: Identified NF1 variants; Supplemental Table 2: Tumors with multiple NF1 mutations include splice site, missense, nonsense, and frameshift mutations; Supplemental Table 3: KRAS mutations identified by next-generation sequencing.</p>