Personalised circulating tumour DNA assay with large-scale mutation coverage for sensitive minimal residual disease detection in colorectal cancer.

BACKGROUND: Postoperative minimal residual disease (MRD) detection using circulating-tumour DNA (ctDNA) requires a highly sensitive analysis platform. We have developed a tumour-informed, hybrid-capture ctDNA sequencing MRD assay. METHODS: Personalised target-capture panels for ctDNA detection were designed using individual variants identified in tumour whole-exome sequencing of each patient. MRD status was determined using ultra-high-depth sequencing data of plasma cell-free DNA. The MRD positivity and its association with clinical outcome were analysed in Stage II or III colorectal cancer (CRC). RESULTS: In 98 CRC patients, personalised panels for ctDNA sequencing were built from tumour data, including a median of 185 variants per patient. In silico simulation showed that increasing the number of target variants increases MRD detection sensitivity in low fractions (<0.01%). At postoperative 3-week, 21.4% of patients were positive for MRD by ctDNA. Postoperative positive MRD was strongly associated with poor disease-free survival (DFS) (adjusted hazard ratio 8.40, 95% confidence interval 3.49-20.2). Patients with a negative conversion of MRD after adjuvant therapy showed significantly better DFS (P < 0.001). CONCLUSION: Tumour-informed, hybrid-capture-based ctDNA assay monitoring a large number of patient-specific mutations is a sensitive strategy for MRD detection to predict recurrence in CRC.

Telecom-Wavelength Bottom-up Nanobeam Lasers on Silicon-on-Insulator.

Semiconductor nanowire lasers are considered promising ultracompact and energy-efficient light sources in the field of nanophotonics. Although the integration of nanowire lasers onto silicon photonic platforms is an innovative path toward chip-scale optical communications and photonic integrated circuits, operating nanowire lasers at telecom-wavelengths remains challenging. Here, we report on InGaAs nanowire array lasers on a silicon-on-insulator platform operating up to 1440 nm at room temperature. Bottom-up photonic crystal nanobeam cavities are formed by growing nanowires as ordered arrays using selective-area epitaxy, and single-mode lasing by optical pumping is demonstrated. We also show that arrays of nanobeam lasers with individually tunable wavelengths can be integrated on a single chip by the simple adjustment of the lithographically defined growth pattern. These results exemplify a practical approach toward nanowire lasers for silicon photonics.

Monolithic InGaAs Nanowire Array Lasers on Silicon-on-Insulator Operating at Room Temperature.

Chip-scale integrated light sources are a crucial component in a broad range of photonics applications. III-V semiconductor nanowire emitters have gained attention as a fascinating approach due to their superior material properties, extremely compact size, and capability to grow directly on lattice-mismatched silicon substrates. Although there have been remarkable advances in nanowire-based emitters, their practical applications are still in the early stages due to the difficulties in integrating nanowire emitters with photonic integrated circuits. Here, we demonstrate for the first time optically pumped III-V nanowire array lasers monolithically integrated on silicon-on-insulator (SOI) platform. Selective-area growth of InGaAs/InGaP core/shell nanowires on an SOI substrate enables the nanowire array to form a photonic crystal nanobeam cavity with superior optical and structural properties, resulting in the laser to operate at room temperature. We also show that the nanowire array lasers are effectively coupled with SOI waveguides by employing nanoepitaxy on a prepatterned SOI platform. These results represent a new platform for ultracompact and energy-efficient optical links and unambiguously point the way toward practical and functional nanowire lasers.

Monolithically Integrated InGaAs Nanowires on 3D Structured Silicon-on-Insulator as a New Platform for Full Optical Links.

Monolithically integrated III-V semiconductors on a silicon-on-insulator (SOI) platform can be used as a building block for energy-efficient on-chip optical links. Epitaxial growth of III-V semiconductors on silicon, however, has been challenged by the large mismatches in lattice constants and thermal expansion coefficients between epitaxial layers and silicon substrates. Here, we demonstrate for the first time the monolithic integration of InGaAs nanowires on the SOI platform and its feasibility for photonics and optoelectronic applications. InGaAs nanowires are grown not only on a planar SOI layer but also on a 3D structured SOI layer by catalyst-free metal-organic chemical vapor deposition. The precise positioning of nanowires on 3D structures, including waveguides and gratings, reveals the versatility and practicality of the proposed platform. Photoluminescence measurements exhibit that the composition of ternary InGaAs nanowires grown on the SOI layer has wide tunability covering all telecommunication wavelengths from 1.2 to 1.8 µm. We also show that the emission from an optically pumped single nanowire is effectively coupled and transmitted through an SOI waveguide, explicitly showing that this work lays the foundation for a new platform toward energy-efficient optical links.

High Quantum Efficiency Nanopillar Photodiodes Overcoming the Diffraction Limit of Light.

InAs1-xSbx nanowires have recently attracted interest for infrared sensing applications due to the small bandgap and high thermal conductivity. However, previous reports on nanowire-based infrared sensors required low operating temperatures in order to mitigate the high dark current and have shown poor sensitivities resulting from reduced light coupling efficiency beyond the diffraction limit. Here, InAsSb nanopillar photodiodes with high quantum efficiency are achieved by partially coating the nanopillar with metal that excites localized surface plasmon resonances, leading to quantum efficiencies of ∼29% at 2390 nm. These high quantum efficiency nanopillar photodiodes, with 180 nm diameters and 1000 nm heights, allow operation at temperatures as high as 220 K and exhibit a detection wavelength up to 3000 nm, well beyond the diffraction limit. The InAsSb nanopillars are grown on low cost GaAs (111)B substrates using an InAs buffer layer, making our device architecture a promising path toward low-cost infrared focal plane arrays with high operating temperature.

High-Quality InAsSb Nanowires Grown by Catalyst-Free Selective-Area Metal-Organic Chemical Vapor Deposition.

We report on the first demonstration of InAs1-xSbx nanowires grown by catalyst-free selective-area metal-organic chemical vapor deposition (SA-MOCVD). Antimony composition as high as 15 % is achieved, with strong photoluminescence at all compositions. The quality of the material is assessed by comparing the photoluminescence (PL) peak full-width at half-max (fwhm) of the nanowires to that of epitaxially grown InAsSb thin films on InAs. We find that the fwhm of the nanowires is only a few meV broader than epitaxial films, and a similar trend of relatively constant fwhm for increasing antimony composition is observed. Furthermore, the PL peak energy shows a strong dependence on temperature, suggesting wave-vector conserving transitions are responsible for the observed PL in spite of lattice mismatched growth on InAs substrate. This study shows that high-quality InAsSb nanowires can be grown by SA-MOCVD on lattice mismatched substrate, resulting in material suitable for infrared detectors and high-performance nanoelectronic devices.

Multiband perfect absorbers using metal-dielectric films with optically dense medium for angle and polarization insensitive operation.

The cavity resonant properties of planar metal-dielectric layered structures with optically dense dielectric media are studied with the aim of realizing omnidirectional and polarization-insensitive operation. The angle-dependent coupling between free-space and cavity modes are revealed to be a key leverage factor in realizing nearly perfect absorbers well-matched to a wide range of incidence angles. We establish comprehensive analyses of the relationship between the structural and optical properties by means of theoretical modeling with numerical simulation results. The presented work is expected to provide a simple and cost-effective solution for light absorption and detection applications that exploit planar metal-dielectric optical devices.

Lasing in hybrid metal-Bragg nanocavities.

We report room-temperature lasing from an optically pumped subwavelength-scale cylindrical InGaAsP pillar surrounded by circular Bragg reflectors on a metal substrate with a dielectric spacer layer. By taking advantage of wide in-plane photonic bandgaps and proper vertical antiresonances, three dielectric Bragg pairs produce a sufficient optical feedback capable of low threshold lasing from the fundamental TE011 mode. A large spontaneous emission coupling into the lasing mode is obtained from the cavity-enhanced Purcell effects and effective suppression of nonlasing modes.

Accurate Detection of Urothelial Bladder Cancer Using Targeted Deep Sequencing of Urine DNA.

Patients with hematuria are commonly given an invasive cystoscopy test to detect bladder cancer (BC). To avoid the risks associated with cystoscopy, several urine-based methods for BC detection have been developed, the most prominent of which is the deep sequencing of urine DNA. However, the current methods for urine-based BC detection have significant levels of false-positive signals. In this study, we report on uAL100, a method to precisely detect BC tumor DNA in the urine without tumor samples. Using urine samples from 43 patients with BC and 21 healthy donors, uAL100 detected BC with 83.7% sensitivity and 100% specificity. The mutations identified in the urine DNA by uAL100 for BC detection were highly associated with BC tumorigenesis and progression. We suggest that uAL100 has improved accuracy compared to other urine-based methods for early BC detection and can reduce unnecessary cystoscopy tests for patients with hematuria.

Practical Utility of Liquid Biopsies for Evaluating Genomic Alterations in Castration-Resistant Prostate Cancer.

Traditional tissue-based assessments of genomic alterations in castration-resistant prostate cancer (CRPC) can be challenging. To evaluate the real-world clinical utility of liquid biopsies for the evaluation of genomic alterations in CRPC, we preemptively collected available plasma samples and archival tissue samples from patients that were being treated for clinically confirmed CRPC. The cell-free DNA (cfDNA) and tumor tissue DNA were analyzed using the AlphaLiquid®100-HRR panel. Plasma samples from a total of 87 patients were included in this study. Somatic mutations from cfDNA were detected in 78 (89.7%) patients, regardless of the presence of overt metastasis or concomitant treatment given at the time of plasma sample collection. Twenty-three patients were found to have known deleterious somatic or germline mutations in HRR genes from their cfDNA. Archival tissue samples from 33 (37.9%) patients were available for comparative analysis. Tissue sequencing was able to yield an NGS result in only 51.5% of the tissue samples. The general sensitivity of cfDNA for detecting somatic mutations in tissues was 71.8%, but important somatic/germline mutations in HRR genes were found to have a higher concordance (100%). Liquid biopsies can be a reasonable substitute for tissue biopsies in CRPC patients when evaluating genomic alterations.

Cancer signature ensemble integrating cfDNA methylation, copy number, and fragmentation facilitates multi-cancer early detection.

Cell-free DNA (cfDNA) sequencing has demonstrated great potential for early cancer detection. However, most large-scale studies have focused only on either targeted methylation sites or whole-genome sequencing, limiting comprehensive analysis that integrates both epigenetic and genetic signatures. In this study, we present a platform that enables simultaneous analysis of whole-genome methylation, copy number, and fragmentomic patterns of cfDNA in a single assay. Using a total of 950 plasma (361 healthy and 589 cancer) and 240 tissue samples, we demonstrate that a multifeature cancer signature ensemble (CSE) classifier integrating all features outperforms single-feature classifiers. At 95.2% specificity, the cancer detection sensitivity with methylation, copy number, and fragmentomic models was 77.2%, 61.4%, and 60.5%, respectively, but sensitivity was significantly increased to 88.9% with the CSE classifier (p value < 0.0001). For tissue of origin, the CSE classifier enhanced the accuracy beyond the methylation classifier, from 74.3% to 76.4%. Overall, this work proves the utility of a signature ensemble integrating epigenetic and genetic information for accurate cancer detection.

InAsP Quantum Dot-Embedded InP Nanowires toward Silicon Photonic Applications.

Quantum dot (QD) emitters on silicon platforms have been considered as a fascinating approach to building next-generation quantum light sources toward unbreakable secure communications. However, it has been challenging to integrate position-controlled QDs operating at the telecom band, which is a crucial requirement for practical applications. Here, we report monolithically integrated InAsP QDs embedded in InP nanowires on silicon. The positions of QD nanowires are predetermined by the lithography of gold catalysts, and the 3D geometry of nanowire heterostructures is precisely controlled. The InAsP QD forms atomically sharp interfaces with surrounding InP nanowires, which is in situ passivated by InP shells. The linewidths of the excitonic (X) and biexcitonic (XX) emissions from the QD and their power-dependent peak intensities reveal that the proposed QD-in-nanowire structure could be utilized as a non-classical light source that operates at silicon-transparent wavelengths, showing a great potential for diverse quantum optical and silicon photonic applications.

Silver superlens using antisymmetric surface plasmon modes.

Silver lenses having super-resolution are analyzed in terms of antisymmetric modes of surface plasmon which have the ability to amplify evanescent waves in UV region. Antisymmetric surface plasmon modes excited by subwavelength grating enhances the resolution and contrast of silver superlens. By using a 20 nm-thick silver superlens, the half-pitch resolution of approximately lambda(0)/8 can be achieved with good contrast at a free space wavelength of 435 nm. The resolution of silver superlens can also be improved using shorter illumination wavelength. We show that the thinner the lens, the better the imaging ability of the silver superlens due to the excitation of antisymmetric surface plasmon modes of higher propagation wave vectors. The thickness of lens is varied from 20 to 40 nm in a three layer system, SiO(2)-Ag-SiO(2). Obtained results illustrate that practical application for patterning periodic structures with good contrast and penetration depth can be achieved by using antisymmetric surface plasmon modes.

Low bending loss metal waveguide embedded in a free-standing multilayered polymer film.

Very low vertical bending loss is demonstrated in a flexible metal waveguide. The waveguide consists of an 8 nm-thick and 68 mm-long Ag strip embedded in a free-standing multilayered low-loss polymer film. The polymer film is composed of a 10 microm-thick inner cladding with a refractive index of 1.524, and a pair of 20 microm-thick outer claddings which both have a refractive index of 1.514, resulting in a total thickness of 50 microm. The measured vertical bending loss is lower than 0.3 dB/180 masculine at a wavelength of 1310 nm for the bending radii down to 2 mm.

Sub-dB/cm propagation loss in silver stripe waveguides.

We demonstrate sub-dB/cm propagation losses in polymer-based silver stripe waveguides at the wavelength of 1.31 microm. The silver stripe waveguides were fabricated in a low-loss fluorinated polymer clad. To form uniform metal stripe patterns, which are essential for reducing propagation loss, we developed a lift-off process using double layers of photoresist and SiNx. A propagation loss of less than 1.0 dB/cm was obtained with the 11- nm-thick silver stripes in the width range of 1.5 - 4.5 microm. A coupling loss of approximately 1.0 dB with a polarization maintaining single mode fiber was achieved for a width of 4.5 microm. For a width of 2.0 microm, we recorded a minimum propagation loss of 0.4 dB/cm, which is comparable with that of dielectric multi-mode waveguides.

Multiplexed single-cell RNA-seq via transient barcoding for simultaneous expression profiling of various drug perturbations.

The development of high-throughput single-cell RNA sequencing (scRNA-seq) has enabled access to information about gene expression in individual cells and insights into new biological areas. Although the interest in scRNA-seq has rapidly grown in recent years, the existing methods are plagued by many challenges when performing scRNA-seq on multiple samples. To simultaneously analyze multiple samples with scRNA-seq, we developed a universal sample barcoding method through transient transfection with short barcode oligonucleotides. By conducting a species-mixing experiment, we have validated the accuracy of our method and confirmed the ability to identify multiplets and negatives. Samples from a 48-plex drug treatment experiment were pooled and analyzed by a single run of Drop-Seq. This revealed unique transcriptome responses for each drug and target-specific gene expression signatures at the single-cell level. Our cost-effective method is widely applicable for the single-cell profiling of multiple experimental conditions, enabling the widespread adoption of scRNA-seq for various applications.

Lineage tracing using a Cas9-deaminase barcoding system targeting endogenous L1 elements.

Determining cell lineage and function is critical to understanding human physiology and pathology. Although advances in lineage tracing methods provide new insight into cell fate, defining cellular diversity at the mammalian level remains a challenge. Here, we develop a genome editing strategy using a cytidine deaminase fused with nickase Cas9 (nCas9) to specifically target endogenous interspersed repeat regions in mammalian cells. The resulting mutation patterns serve as a genetic barcode, which is induced by targeted mutagenesis with single-guide RNA (sgRNA), leveraging substitution events, and subsequent read out by a single primer pair. By analyzing interspersed mutation signatures, we show the accurate reconstruction of cell lineage using both bulk cell and single-cell data. We envision that our genetic barcode system will enable fine-resolution mapping of organismal development in healthy and diseased mammalian states.

Selective targeting of KRAS oncogenic alleles by CRISPR/Cas9 inhibits proliferation of cancer cells.

Mutations within the KRAS oncogene are associated with the proliferation of various cancers. Therapeutic approaches for treating cancers with such mutations have focused on targeting the downstream protein effectors of KRAS. However, to date, no approved treatment has targeted the mutated KRAS oncogene directly. Presently, we used the selectivity of the CRISPR/Cas9 system to directly target mutated KRAS alleles. We designed single-guide RNAs (sgRNAs) to target two specific single-nucleotide missense mutations on KRAS codon-12 located in the seed region adjacent to a protospacer adjacent motif (PAM). Lentiviral transduction of Cas9 and the sgRNAs into cancer cells with respective KRAS mutations resulted in high frequency of indels in the seed region. Indel-associated disruption of the mutant KRAS alleles correlated with reduced viability of the cancer cells. The results indicate that CRISPR-Cas9-mediated genome editing can potentially be used for the treatment of cancer patients, specifically those with oncogenic KRAS mutations.

Straightforward Delivery of Linearized Double-Stranded DNA Encoding sgRNA and Donor DNA for the Generation of Single Nucleotide Variants Based on the CRISPR/Cas9 System.

CRISPR/Cas9 for genome editing requires delivery of a guide RNA sequence and donor DNA for targeted homologous recombination. Typically, single-stranded oligodeoxynucleotide, serving as the donor template, and a plasmid encoding guide RNA are delivered as two separate components. However, in the multiplexed generation of single nucleotide variants, this two-component delivery system is limited by difficulty of delivering a matched pair of sgRNA and donor DNA to the target cell. Here, we describe a novel codelivery system called "sgR-DNA" that uses a linearized double-stranded DNA consisting of donor DNA component and a component encoding sgRNA. Our sgR-DNA-based method is simple to implement because it does not require cloning steps. We also report the potential of our delivery system to generate multiplex genomic substitutions in Escherichia coli and human cells.