Single Hot Spot Research Articles

Abstract 3917: Double PIK3CA mutations in cis enhance oncogene activation and sensitivity to PI3K alpha inhibitors in breast cancer

Abstract Activating mutations in PIK3CA, the gene coding for the catalytic subunit (p110α) of phosphoinositide 3-kinase (PI3K), are the most frequent oncogenic alterations in all cancers, including estrogen receptor-positive (ER+) breast cancer. There are many distinct oncogenic PIK3CA mutations including major hotspot mutations (e.g. E542K, E545K, H1047R) which are predictive of response to PI3Kα inhibitors. While responses to PI3Kα inhibitors can be variable, some patients with single hotspot mutant tumors derive a deep and prolonged clinical benefit. In a comprehensive analysis of the genomic data available in breast cancer, we observed double PIK3CA mutations and investigated their potential biological relevance and potential correlation with sensitivity to PI3Kα inhibitors. We detected double PIK3CA mutations in 10-15% of all PIK3CA-mutant cancers including breast cancer. Double PIK3CA mutations in breast cancer are defined by a pattern of co-expression of a major hotspot mutation associated with a recurrent second-site PIK3CA mutation (‘minor mutation’). We have found that these double PIK3CA mutations are compound, that is in cis on the same allele, using single molecule real time sequencing (SMRT-seq) of fresh breast tumor samples. Double compound PIK3CA mutations result in increased PI3K activity and downstream signaling compared to single hotspot mutants in nontransformed cells and in ER+ breast cancer cells. Double compound mutations also increase cell proliferation and xenograft growth compared to single hotspot mutants. Biochemical experiments using recombinant PI3K compound mutant protein complexes reveal a combination mechanism of action, with simultaneous PI3K destabilization and increased membrane binding compared to single hotspot mutants, leading to increased kinase activity. Importantly, these compound mutations predict for increased sensitivity to PI3Kα inhibitors compared to single hotspot mutants in both preclinical models and also in patients with breast cancer. Together, our data support that double compound PIK3CA mutations enhance the oncogenicity of single hotspot PIK3CA mutations and this increased dependency results in increased sensitivity to PI3Kα inhibitors compared to single hotspot mutations. We propose that double compound activating mutations are an additional mechanism of oncogenic transformation in addition to other known mechanisms such as gene amplification, point mutation, or gene translocation. We speculate that double compound mutant PIK3CA may function as a clinical biomarker of increased sensitivity to PI3K-directed targeted therapies and may improve the therapeutic window of PI3K inhibitors in ER+ breast cancer and other PIK3CA-mutant tumor histologies. Citation Format: Neil Vasan, Pedram Razavi, Jared Johnson, Hong Shao, Hardik Shah, Alesia Antoine, Erik Ladewig, Alexander Gorelick, Eneda Toska, Guotai Xu, Abiha Kazmi, Matthew T. Chang, Barry S. Taylor, Maura N. Dickler, Komal Jhaveri, Raul Rabadan, Ed Reznik, Melissa L. Smith, Robert Sebra, Lewis C. Cantley, Maurizio Scaltriti, Jose Baselga. Double PIK3CA mutations in cis enhance oncogene activation and sensitivity to PI3K alpha inhibitors in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3917.

Numerical analysis of knocking characteristics and heat release under different turbulence intensities in a gasoline engine

In this work, different knocking characteristics and heat release have been numerically studied in a downsized spark-ignition engine, with addressing turbulence intensity and spark-ignition timing. The results show that knock intensity mainly depends on the heat release of hot-spots, and the heat release is not only associated with unburned mass fraction but also closely related to subsequent autoignition development processes, e.g. end-gas autoignition mode. Under low turbulence intensity, local mixture autoignition is induced by a single hot-spot, accompanied by an autoignition reaction wave propagation; whereas under high turbulence intensity, a nearly homogeneous autoignition event characterizing quasi constant-volume combustion is observed. Meanwhile, compared with the single hot-spot autoignition, the homogeneous one can lead to more intensive local overpressure and thus stronger knock intensity. Further analysis on the underlying reasons shows that the higher heat release rate under homogeneous autoignition conditions is the main reason for the stronger knock intensity. Besides, the dependency of knock intensity and heat release on unburned mass fraction is found effective only for the same autoignition scenarios. Current study can provide insights into knocking suppression with consideration of intake system optimization in modern downsized gasoline engines.

Comparative analysis of the structure of the tectonosphere of the Conrad and Afanasy Nikitin Rises by geophysical data (the Indian Ocean)

A two-dimensional density modeling is performed for profiles crossing the seamounts of the Conrad Rise (Lena, Ob and Marion Dufresne), which are located in the southwestern part of the Indian Ocean and the Afanasy Nikitin Rise, which is located in the central part of the Indian Ocean. According to the assumptions of a number of researchers, the seamount Marion Dufresne and the Afanasy Nikitin Rise were formed as a result of the hotspot 83–73 million years ago. The results of two-dimensional density modeling showed a similar structure of the crust and lithosphere, which confirms the probability of their formation 83–73 million years ago as a result of the action of a single hotspot. Also on the basis of the obtained results, the assumption is confirmed that the seamounts Lena and Ob of the Conrad Rise were formed afterwards due to the continuation of the action of the hotspot under the Antarctic Plate.

Sources vs processes: Unraveling the compositional heterogeneity of rejuvenated-type Hawaiian magmas

Spatial isotope variations in oceanic island basalts hold clues to the geochemical structure of mantle plumes. This signal is however susceptible to contributions from non-plume sources and modifications by physical processing in the upper mantle, i.e. source hybridization and segregation through gradients in melting conditions. A comprehensive survey of lava isotope systematics is therefore necessary to reveal spatiotemporal patterns at the scale of a single hotspot and disentangle plume sources from upper mantle physical and chemical contributions. We present Hf and Pb isotope data on 72 previously characterized onland and submarine lavas from the islands of Ni‘ihau and Kaua‘i, and from the North Arch Volcanic Field. These lavas cover the entire ∼ 5 Ma eruptive history of Ni‘ihau and Kaua‘i, allowing for a detailed assessment of the temporal geochemical evolution of Hawaiian volcanism. Furthermore, the broad lateral coverage of volcanic activity offered by these lavas constrains the spatial variability of sources across the volcanic chain, allowing investigation of the role of an upper asthenosphere contribution to plume volcanism. Early, shield-building (tholeiitic) lavas from Kaua‘i and Ni‘ihau overlap in Nd–Hf–Sr–Pb isotopes, contrasting with the across-plume asymmetry seen in younger shield lavas (i.e. the LOA-KEA double chain), and their isotope systematics suggest sampling of both LOA and KEA components. In contrast, late Kaua‘i and Ni‘ihau rejuvenated lavas do not overlap in isotope composition. They share common isotope characteristics with rejuvenated lavas from North Arch, Ka‘ula and O‘ahu, and together form two isotopically distinct, southwestern and northeastern groups. The two groups have steep slopes (>1.7) in a εNd − εHf plot and require distinct depleted components. The high εHf for a given εNd, high 87Sr/86Sr, and trace element systematics of the southwestern group (Ni‘ihau, Ka‘ula and O‘ahu) indicate a depleted component intrinsic to the plume. In contrast, lavas from the northeastern group (Kaua‘i and North Arch) indicate a source component isotopically identical to Pacific MORB. The shift from a spatially broad and homogeneous plume source in the shield stage of Kaua‘i and Ni‘ihau (6–3.5 Ma) to a bilaterally heterogeneous source during the rejuvenated stage (<3.5 Ma) is synchronous with the emergence of the LOA and KEA geochemical double chain in younger Hawaiian shield lavas. Our data reveal a shift to Pacific asthenosphere contribution to Kaua‘i volcanism through time, while Ni‘ihau exclusively samples the plume. These observations are inconsistent with a scenario where plume geochemical structure is the primary control on spatial heterogeneities in lava chemistry, and instead point to a more complex and dynamic interaction between plume, Pacific asthenosphere and lithosphere. Therefore, deep mantle structure might not be the primary control on the spatial distribution of isotope systematics in oceanic island lavas, making a link between the latter and contrasts imaged in the lower mantle through seismic tomography tenuous.

Design of anapole mode electromagnetic field enhancement structures for biosensing applications.

The design of an all-dielectric nanoantenna based on nonradiating "anapole" modes is studied for biosensing applications in an aqueous environment, using FDTD electromagnetic simulation. The strictly confined electromagnetic field within a circular or rectangular opening at the center of a cylindrical silicon disk produces a single point electromagnetic hotspot with up to 6.5x enhancement of |E|, for the 630-650 nm wavelength range, and we can increase the value up to 25x by coupling additional electromagnetic energy from an underlying PEC-backed substrate. We characterize the effects of the substrate design and slot dimensions on the field enhancement magnitude, for devices operating in a water medium.

RNA-guided piggyBac transposition in human cells.

Safer and more efficient methods for directing therapeutic genes to specific sequences could increase the repertoire of treatable conditions. Many current approaches act passively, first initiating a double-stranded break, then relying on host repair to uptake donor DNA. Alternatively, we delivered an actively integrating transposase to the target sequence to initiate gene insertion. We fused the hyperactive piggyBac transposase to the highly specific, catalytically dead SpCas9-HF1 (dCas9) and designed guide RNAs (gRNAs) to the CCR5 safe harbor sequence. We introduced mutations to the native DNA-binding domain of piggyBac to reduce non-specific binding of the transposase and cause the fusion protein to favor binding by dCas9. This strategy enabled us, for the first time, to direct transposition to the genome using RNA. We showed that increasing the number of gRNAs improved targeting efficiency. Interestingly, over half of the recovered insertions were found at a single TTAA hotspot. We also found that the fusion increased the error rate at the genome-transposon junction. We isolated clonal cell lines containing a single insertion at CCR5 and demonstrated long-term expression from this locus. These vectors expand the utility of the piggyBac system for applications in targeted gene addition for biomedical research and gene therapy.

Where to Cross Over? Defining Crossover Sites in Plants.

It is believed that recombination in meiosis serves to reshuffle genetic material from both parents to increase genetic variation in the progeny. At the same time, the number of crossovers is usually kept at a very low level. As a consequence, many organisms need to make the best possible use from the one or two crossovers that occur per chromosome in meiosis. From this perspective, the decision of where to allocate rare crossover events becomes an important issue, especially in self-pollinating plant species, which experience limited variation due to inbreeding. However, the freedom in crossover allocation is significantly limited by other, genetic and non-genetic factors, including chromatin structure. Here we summarize recent progress in our understanding of those processes with a special emphasis on plant genomes. First, we focus on factors which influence the distribution of recombination initiation sites and discuss their effects at both, the single hotspot level and at the chromosome scale. We also briefly explain the aspects of hotspot evolution and their regulation. Next, we analyze how recombination initiation sites translate into the development of crossovers and their location. Moreover, we provide an overview of the sequence polymorphism impact on crossover formation and chromosomal distribution.

Hotspot enumeration of CD8+ tumor-infiltrating lymphocytes using digital image analysis in triple-negative breast cancer yields consistent results

Tumor-infiltrating lymphocytes (TILs) have emerged as prognostic in triple-negative breast cancer (TNBC). We aimed to assess the consistency of hotspot placement and TIL enumeration among multiple pathologists. Additionally, we assessed hotspot TIL count consistency by comparing hotspot counts in 3 separate locations within a single whole-tissue section. Anti-CD8 immunohistochemistry was performed on a representative section from 66 cases of primary TNBC, which were then scanned as whole-slide images. Quantification of the tissue area and combined stromal and intratumoral CD8+ TILs was performed using digital image analysis (DIA) within 2.2 mm-diameter circle hotspots. TIL counts were quantified as absolute counts and densities (absolute count/tissue area in micrometers2). For each case, 6 pathologists placed a single hotspot, defined as an area with the subjectively highest CD8+ immunoreactivity, within the tumor bed. Separately for each case, a single pathologist placed hotspots in 3 different locations within a single tumor section. Intraclass correlation coefficients (ICCs) were generated following TIL enumeration via DIA. ICCs for single hotspot placement by 6 pathologists were 0.96 for density and 0.97 for absolute counts, respectively. In 32% of cases (21/66), all the hotspots placed by the 6 pathologists were in the same location. When evaluating hotspots in 3 different locations within a tumor, the ICC was 0.95 for both density and absolute counts. Hotspot evaluation by DIA is a reproducible method for CD8+ TIL quantification, and the use of hotspots may reduce TIL count variation caused by intratumoral TIL heterogeneity.

A New Paradigm for Radiation-Induced Toxicity Analysis: Space Based Normal Tissue Complication Probability Modeling

An increasing awareness have been triggered in the RO community regarding the value of including the spatial information of dose distributions within the analysis of radiation-induced toxicity (RIT). Up to now, statistical inference on spatial signature of RIT has been addressed [Acosta 2013, Dean 2016], but no insight on actual NTCP models including full dose spatial info has been achieved. The purpose of this study is to propose a new formalism to fill this gap of knowledge and to develop a space based NTCP (SpNTCP). A SpNTCP approach was devised assuming the availability of a training set of N patients’ dose distributions (sampled on m voxels each) classified according to a binary RIT. A logistic regression on the dose values is performed voxelwise, and, accordingly, a collection of m RIT probabilities is computed on each voxel dose for a test dose distribution. Similarly, a collection of m weights is obtained as the inverse of each logistic Confidence Interval (CI) length. The SpNTCP is then computed as the weighted mean of the m RIT probabilities. The SpNTCP was first demonstrated on two sets of synthetic dose maps (with different and equal DVHs, respectively), classified by thresholding the generalized Equivalent Uniform Dose (gEUD) simulated for given Radio-Sensitivity Map (RSM) and n volume-effect parameter. Moreover, an estimate of the RSM was reconstructed by computing the previous SpNTCP on a new set of hot spots evenly spanning the field of view. The performance was evaluated on a cohort of 98 RT Hodgkin lymphoma survivors classified for lung fibrosis (18 events). For this purpose, each dose map was normalized to a common anatomical reference via a log-diffeomorphic demons registration tool [Palma 2016]. Lyman-Kutcher-Burman (LKB) models were trained for comparison. Asymptotic values of learning curves show that the SpNTCP model outperformed LKB one, with an increasing model bias at lower n values (Table). On the real patients, the area under the ROC curve of the SpNTCP was 0.72 (CI: [0.62-0.81]), significantly higher than 0.61 (CI: [0.51-0.70]) obtained by the LKB (p<0.002).Abstract 197; TableAccuracy of modelsDifferent DVHsEqual DVHsnSpNTCPLKBSpNTCPLKB.1.77.81.77.5.25.85.77.81.5.5.90.76.85.51.93.78.88.5 Open table in a new tab The proposed SpNTCP model, differently from LKB, proved able to distinguish the toxicity effects of given dose distributions in presence of inhomogeneous RSM, even for equal DVHs. Interestingly, our approach also proved able to map a posteriori the organ RSM, by calculating the SpNTCPs on a set of dose distributions with a single hot spot, whose position spans the FOV. As expected, the performance of the SpNTCP model, at least in this preliminary format, decreases as the volume effect decreases. Of note, the SpNTCP modeling of real lung RIT was more accurate than LKB, seemingly in agreement with the evidences of inhomogeneous RSM of the lungs. In conclusion, we believe that the proposed germinal idea could pave the way toward a new NTCP modeling philosophy.

Plasmon-Driven Photocatalysis Leads to Products Known from E-beam and X-ray-Induced Surface Chemistry.

Plasmonic metal nanostructures can concentrate incident optical fields in nanometer-sized volumes, called hot spots. This leads to enhanced optical responses of molecules in such a hot spot but also to chemical transformations, driven by plasmon-induced hot carriers. Here, we employ tip-enhanced Raman spectroscopy (TERS) to study the mechanism of these reactions in situ at the level of a single hot spot. Direct spectroscopic measurements reveal the energy distribution of hot electrons, as well as the temperature changes due to plasmonic heating. Therefore, charge-driven reactions can be distinguished from thermal reaction pathways. The products of the hot-carrier-driven reactions are strikingly similar to the ones known from X-ray or e-beam-induced surface chemistry despite the >100-fold energy difference between visible and X-ray photons. Understanding the analogies between those two scenarios implies new strategies for rational design of plasmonic photocatalytic reactions and for the elimination of photoinduced damage in plasmon-enhanced spectroscopy.

Thermosynechococcaceae as a source of thermostable C-phycocyanins: properties and molecular insights

This study describes characterisation of three thermophilic strains of cyanobacteria belonging to Thermosynechococcaceae and their utilisation as a source of thermostable C-phycocyanin. Three strains NIES 2133, NIES 2134 and PCC 6715 have been cultivated in two types of growth media and two temperatures: 45 and 50 °C and assessed for C-phycocyanin productivity. Among these strains Synechococcus lividus sp. PCC 6715, phylogenetically closer to Thermosynechococcus sp. clade than other Synechococcaceae, has been selected for subsequent study to characterise its C-phycocyanin. The strain showed faster growth rates than the other two and appeared to utilise different bicarbonate uptake mechanism based on SbtA, whilst BCT1 mechanism was found in NIES2133 and 2134. Higher growth rates translated to larger overall C-phycocyanin productivity. C-phycocyanin has been purified using chromatographic methods and its thermostability and pH stability has been assessed against that of Spirulina. Results have shown that PCC 6715 C-phycocyanin is one of the most stable proteins of this type reported to date. It exhibits 90% stability after 5-hour incubation at 50 °C and about 70% stability after 2-week incubation at the same temperature. PCC 6715 phycocyanins show high stability at acidic pH at both 4 °C and 50 °C, whilst at 50 °C maintain 100% stability during 4-hour incubation at pH 4.0 to 8.0. PCC 6715 phycocyanin shows good long-term stability characteristics and could be considered as a thermostable replacement for Spirulina phycocyanin. Analysis of molecular model of PCC 6715 C-phycocyanins and crystal structures of other phycocyanins reveals interesting pattern of amino acid substitutions that are present in thermophilic proteins that could have significant impact on their significantly higher thermostability. Phycocyanins from thermostable cyanobacteria exhibit evolutionarily conserved amino acid substitutions localised in the single structural hotspot of the protein that result in additional stabilising interactions within and between protein subunits when compared to proteins from mesophilic counterparts.

Abstract LB-149: Targeting shared hotspot cancer mutations with a Listeria monocytogenes immunotherapy induce potent anti-tumor immunity

Abstract Introduction: Virtually all tumors contain somatic mutations that can result in novel antigenic sequences that may be targeted by the host cellular immune response. Some of these mutations occur in preferential regions of specific genes commonly referred to as hotspot mutations. Hotspot mutations are commonly shared by cancer patients both within and across multiple tumor types. These hotspot mutations often confer loss or gain of function contributing to oncogenesis, which makes them promising therapeutic targets. One such mutation commonly found in several human tumor types is an aspartic acid substitution for glycine at position 12 (G12D) in KRAS. This same mutation occurs in the CT26 murine colorectal tumor model. To determine if expression of the KRAS G12D sequence in a bacterial immunotherapy vector can control tumor growth in the CT26 murine model, the Advaxis Listeria monocytogenes (Lm)-based platform was engineered to express a 21-amino acid KRAS sequence peptide containing the G12D mutation (Lm-Hot KRAS_G12D). In addition, we evaluated control of tumor growth using an ADXS-HOT construct (ADXS-503) that expresses multiple shared human hotspot and tumor-associated antigens, including the G12D KRAS. The ADXS-HOT clinical program is comprised of several Lm-based immunotherapies designed to target multiple shared hotspot and tumor-associated antigens commonly found in specific cancer types. In this study, we demonstrate control of tumor growth in a mouse model by targeting a commonly shared hotspot mutation using an Lm-based immunotherapy. Results: We show that the Lm-HOT KRAS_G12D therapy significantly delayed tumor growth and improved long-term survival in the murine CT26 colon carcinoma model. This response was associated with an increase in the frequency of tumor infiltrating antigen-specific CD8 T cells and γδ T cells within the tumor microenvironment and a decrease in the frequency of intratumoral regulatory T cells (Tregs). Furthermore, tumor-specific CD8 T cells displayed lower expression of exhaustion markers as well as increased functionality upon restimulation. Interestingly, our proprietary ADXS-503 (a clinical ADXS-HOT construct) which includes KRAS G12D as one of its multiple targets, was also capable of significantly suppressing tumor growth in the CT26 tumor model. Conclusion: These results suggest that our ADXS-HOT platform is a promising approach to target shared hotspot mutations. That ADXS Lm constructs targeting a single hotspot mutation can significantly control tumor growth whether it is in a single or multi-target construct. These data describe an exciting translatable discovery with the potential for broad utility across multiple tumor types and patients who share common hotspot mutations. Citation Format: Daniel Villarreal, Brandon Coder, Susan Armington, Andrew L'Huillier, Cristina Mottershead, Elena Filippova, Nithya Thambi, Kim Ramos, David Balli, Robert Petit, Michael Princiotta. Targeting shared hotspot cancer mutations with a Listeria monocytogenes immunotherapy induce potent anti-tumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-149.

Modeling The Effects of Shock Pressure and Pore Morphology on Hot Spot Mechanisms in HMX

AbstractWe investigate the effects of shock pressure and pore morphology on the formation and growth of hot spots in HMX (octahydro‐1,3,5,7‐tetranitro‐1,3,5,7‐tetrazocine). Both non‐reactive and reactive ALE3D simulations are used in these studies. Our non‐reactive simulations show a viscous‐dominated pore collapse mode at lower shock pressures (2–10 GPa) with shear band formation and a hydrodynamic‐dominated mode at higher shock pressures (20‐40 GPa) due to bulk melting. When normalized by bulk shock heating, viscous‐dominated pore collapse modes are more efficient at generating hot spots. Pore morphology influences the post‐collapse temperature distributions and reaction rate for a fixed pore area and shock pressure. We find that multiple surface pores at the binder‐grain interface tend to react the fastest. Due to their upstream location in the HMX grain, the surface pores collapse sooner than interior pores; thus, the extent of reaction will generally favor these morphologies because they have more time to grow. In general, multiple smaller hot spots tend to react faster than a single larger hot spot because they accelerate one another's burning. The rank order of morphology effects, however, is not the same for non‐reactive and reactive simulations. For example, while multiple surface pores produce the highest reaction rates they do not produce the highest (non‐reactive) hot spot temperatures. Our numerical studies provide insights on hot spot mechanisms in lieu of direct measurements and can be used to develop advanced shock initiation models.

Unexpected large nanoparticle size of single dimer hotspot systems for broadband SERS enhancement.

We have numerically demonstrated the feasibility and possibility to achieve broadband surface-enhanced Raman scattering (SERS) enhancement in the visible and near-infrared wavelength range using single nanoparticle (NP) dimer hotspot systems. Instead of the conventionally reported sub-100nm, we find that the optimal NP size is as large as 200nm in diameter for both Ag and Au. The key lies in the continuous arising of the bonding dipole plasmon mode and higher-order resonances at shorter wavelengths. Further, it is revealed that the near- and far-field optical responses of these hotspot systems correlate well with each other, despite the intrinsic enormous near- to far-field redshift for individual large NPs. The physical principles demonstrated here benefit significantly the fundamental understanding and engineering optimization of broadband SERS substrates.

Plasmonic Heterodimers with Binding Site-Dependent Hot Spot for Surface-Enhanced Raman Scattering.

A novel plasmonic heterodimer nanostructure with a controllable self-assembled hot spot is fabricated by the conjugation of individual Au@Ag core-shell nanocubes (Au@Ag NCs) and varisized gold nanospheres (GNSs) via the biotin-streptavidin interaction from the ensemble to the single-assembly level. Due to their featured configurations, three types of heterogeneous nanostructures referred to as Vertice, Vicinity, and Middle are proposed and a single hot spot forms between the nanocube and nanosphere, which exhibits distinct diversity in surface plasmon resonance effect. Herein, the calculated surface-enhanced Raman scattering enhancement factors of the three types of heterodimers show a narrow distribution and can be tuned in orders of magnitude by controlling the size of GNSs onto individual Au@Ag NCs. Particularly, the Vertice heterodimer with unique configuration can provide extraordinary enhancement of the electric field for the single hot spot region due to the collaborative interaction of lightning rod effect and interparticle plasmon coupling effect. This established relationship between the architecture and the corresponding optical properties of the heterodimers provides the basis for creating controllable platforms which can be exploited in the applications of plasmonic devices, electronics, and biodetection.

Simultaneous identification of clinically relevant single nucleotide variants, copy number alterations and gene fusions in solid tumors by targeted next-generation sequencing.

In this study, we have set-up a routine pipeline to evaluate the clinical application of Oncomine™ Focus Assay, a panel that allows the simultaneous detection of single nucleotide hotspot mutations in 35 genes, copy number alterations (CNAs) in 19 genes and gene fusions involving 23 genes in cancer samples. For this study we retrospectively selected 106 patients that were submitted to surgical resection for lung, gastric, colon or rectal cancer.We found that 56 patients out of 106 showed at least one alteration (53%), with 47 patients carrying at least one relevant nucleotide variant, 10 patients carrying at least one CNA and 3 patients carrying one gene fusion. On the basis of the mutational profiles obtained, we have identified 22 patients (20.7%) that were potentially eligible for targeted therapy.The most frequently mutated genes across all tumor types included KRAS (30 patients), PIK3CA (16 patients), BRAF (6 patients), EGFR (5 patients), NRAS (4 patients) and ERBB2 (3 patients) whereas CCND1, ERBB2, EGFR and MYC were the genes most frequently subjected to copy number gain. Finally, gene fusions were identified only in lung cancer patients and involved MET [MET(13)–MET(15) fusion] and FGFR3 [FGFR3(chr 17)–TACC3(chr 11)].In conclusion, we demonstrate that the analysis with a multi-biomarker panel of cancer patients after surgery, may present several potential advantages in clinical daily practice, including the simultaneous detection of different potentially druggable alterations, reasonable costs, short time of testing and automated interpretation of results.

Hotspot Analysis of Double-Layer Microchannel Heat Sinks

ABSTRACTThe performance of double-layer microchannel heat sinks are evaluated comparatively for the parallel flow, counter flow, and transverse flow configurations with and without hotspots as heating condition. Conjugate heat transfer analysis is performed by solving three-dimensional Navier–Stokes and energy equations using a finite volume solver. The flow is considered to be steady, incompressible, and laminar. Functional relations between the thermophysical properties of water and temperature are developed and used for numerical calculations with variable fluid properties. The thermal resistances, maximum temperature increase at the hotspots, temperature variation among the hotspots, and pressure drops are evaluated for the three heat-sink designs with two hotspot schemes (single hotspot at the center of the heat sink and multiple hotspots distributed uniformly at six peripheral locations). For the single-hotspot case, the parallel flow heat sink exhibited the lowest thermal resistance and temperature rise at the hotspot. For all the six multiple hotspot cases, the transverse flow heat sink exhibited the lowest thermal resistance and temperature variation among the hotspots.

Studying the Dynamics of Hybrid X-Pinches

Since the first experiments with hybrid X-pinches (HXP), it has been shown that a short wire between two conical tungsten electrodes produces the same radiation as a standard X-pinch. Since HXP has a simple construction, it is commonly used in many laboratories instead of standard pinches. Although details of the HXP dynamics remain poorly studied, the main factors governing HXP formation are investigated experimentally in this work. It is shown that the process of the electrode plasma formation is one of the main factors that lead to single hot spot bursts. Experimental results allow the fitting of HXP parameters and determination of the size of the source of X-ray radiation. It is also shown the radiation of HXP can be used as a source for point projection radiography in the wavelength range of soft to hard X-rays. Such sources can be also used for absorption spectroscopy and even electron beams.

Numerical and Experimental Study on Multi-Focal Metallic Fresnel Zone Plates Designed by the Phase Selection Rule via Virtual Point Sources

We propose a novel design method for multi-focal metallic Fresnel zone plates (MFZPs), which exploits the phase selection rule by putting virtual point sources (VPSs) at the desired focal points distant to the MFZP plane. The phase distribution at the MFZP plane reciprocally formed by the VPSs was quantized in a binary manner based on the phase selection rule, thereby leading to a corresponding on-off amplitude pattern for the targeted MFZP. The resultant phase distribution was dependent on the complex amplitudes of the VPSs, so that they could be determined from the perspective of both multi-focal functionality and fabrication feasibility. As a typical example, we utilized the particle swarm optimization algorithm to determine them. Based on the proposed method, we designed and numerically analyzed two types of novel MFZPs—one for a monochromatic multi-focal application and the other for a multi-chromatic mono-focal application—verifying the effectiveness and validity of the proposed method. We also fabricated them onto Au-deposited glass substrates, using electron beam evaporation and a focused ion beam milling process. We experimentally characterized them and also verified that they successfully demonstrated their feasibilities. The former produced distinct hot spots at three different focal distances of 10, 15, and 20 μ m for monochromatic incidence at 650 nm, and the latter produced a single hot spot at a focal distance of 15 μ m for multi-chromatic incidence at 660, 532, and 473 nm. The experimental results were also in good agreement with their corresponding numerical results. We expect that both MFZPs will have various applications, such as laser micromachining, optical trapping, biomedical sensing, confocal collimation, achromatic optics, etc.

Hotspot Thermal Management via Thin-Film Evaporation—Part I: Experimental Characterization

The presence of hotspots with extreme heat fluxes and temperatures in high-performance electronics has led to severe thermal management challenges in the semiconductor industry. Our work experimentally investigates the potential of capillary-fed thin-film evaporation as a thermal management solution for devices where hotspots are superposed with mild background heating. The front side of our test device incorporates well-defined silicon micropillar arrays for passive fluidic transport via capillary wicking, and the backside is patterned with thin-film resistive heaters to emulate $640 \times 620~\mu \text{m}^{2}$ hotspots and $1 \times 1$ cm2 uniform background heating. Our micropillar wick design facilitates efficient fluidic and thermal transport and dissipated ${\approx} 6$ kW/cm2 from a single hotspot without background heating beyond which viscous losses exceeded the capillary pressure and dryout occurred due to insufficient liquid supply. While the capillary-limited hotspot dryout heat flux decreased by creating concurrent hotspots as well as by superposing a hotspot with mild uniform background heating, it was unaltered (within the measurement error) when the location of the hotspot was varied within the $1 \times 1$ cm2 microstructured area. The ultrahigh heat fluxes in our experiments were achieved by suppressing boiling by judiciously designing the fluidic transport where the microstructured surface maintains a thin liquid film and syphons only the required amount of liquid to sustain evaporation. Our experimental results show that thin-film evaporation is a viable thermal management solution for the next-generation high-performance power electronics, among others, where hotspots pose a significant challenge.