Biology subjects

Gao, A.

Publications and source records attributed to Gao, A..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Dental topography and diet in marsupials and comparisons with primates

Dental topography is a powerful tool for investigating the relationship between diet and tooth shape in mammals. Here, we test whether dental topographic metrics (DTMs) can accurately predict diet in marsupials based on 81 lower molars from 43 extant species in 12 families (representing six of the seven extant orders), and compare them to DTMs of primates with similar diets. Specific DTMs tested included Dirichlet normal energy (DNE) and variants (ariaDNE, convex DNE, and ariaDNE CV), orientation patch count rotated (OPCR), and Relief Index (RFI). We also investigate the use of the ratio of trigonid to talonid height (TriTaHI) as an additional DTM. Highest dietary classification accuracy using a leave-one-out quadratic discriminant function analysis for marsupials was reached when ariaDNE, RFI, molar size, and TriTaHI of lower second and third molars were combined, resulting in 69.2% dietary accuracy. In contrast, the primate sample of lower second molars reached the highest classification accuracy of 80.4% when only ariaDNE, RFI, and molar size were used. Notably, 3D meshes from epoxy casts showed consistently higher OPCR and ariaDNE CV values than meshes from original specimens. When using a cross-validated approach, using the primate sample to predict marsupial diets and vice versa, only ariaDNE, RFI, and molar size were needed to obtain the maximum classification accuracy (69.7% when classifying primates using the marsupial training set; and 61.8% vice versa). The comparative dataset of this study will be very useful for studies aiming to reconstruct the diets of fossil mammals that lack phylogenetically close extant analogues.

evolutionary biology↗

bioRxiv2025

Structural basis of QueC-family protein function in qatABCD anti-phage defense

QueC proteins are nucleoside biosynthesis enzymes required for production of the 7-deazaguanine derivative queuosine. Recently, QueC-family proteins were also shown to catalyze a deazaguanylation protein-nucleobase conjugation reaction in type IV CBASS bacterial anti-phage defense. Here we determine the structural basis of QueC-family protein function in a distinct bacterial immunity system named qatABCD. We demonstrate that the QueC-family protein QatC forms a specific complex with the immunity protein QatB and that this complex is minimally required for qatABCD defense. Crystal structures of the QatBC complex enable direct comparison of qatABCD and type IV CBASS defense and support a shared role for QueC-family proteins in targeting protein substrates for N-terminal modification. We show that the QatB unstructured N-terminus and N-terminal glycine motif are essential for qatABCD defense in vivo, suggesting a modification occurs analogous to CBASS deazaguanylation. These findings highlight broad roles of QueC proteins beyond nucleoside biosynthesis and suggest that adaptation of QueC-like proteins for specialized biochemical functions is a common strategy in bacterial anti-phage immunity.

biochemistry↗

bioRxiv2025

Inflammatory cytokines promote interferon regulatory factor (IRF) transcriptional activity in human pulmonary epithelial cells through the induction of IRF1 by nuclear factor-κB

Transcription factors of the interferon regulatory factor (IRF) family play key roles during viral and bacterial infections. However, IRF regulation by inflammatory cytokines, including interleukin (IL)-1{beta} and tumor necrosis factor (TNF) , remains underexplored. Since airway epithelial cells (AECs) modulate lung inflammation, IRF expression was characterized in pulmonary A549 and bronchial BEAS-2B epithelial cells, plus primary human AECs grown in submersion or highly differentiated air-liquid interface culture. Whereas most IRF mRNAs were expressed across models, IRF4 and IRF8 showed consistently low expression, and IRF6 was only expressed highly in primary cells. IRF3 and IRF9 were highly expressed in each model, and while protein expression was confirmed in A549 cells, this remained primarily cytoplasmic post-IL-1{beta} treatment. IRF2 was constitutively nuclear in A549 cells and showed moderate/high mRNA expression in each model. IRF1 mRNA was induced by IL-1{beta}-in primary and A549 cells, where IL-1{beta} and TNF dramatically increased IRF1 protein expression. TNF also increased IRF1 protein in BEAS-2B cells. In A549 cells, an IRF reporter system combined with RNA silencing did not support transcriptional roles for IRF2 or IRF3 but suggested a modest role for IRF9 following IL-1{beta} stimulation. IL-1{beta}-induced IRF1 localized to the nucleus of A549 cells revealing a major transcriptional role. IRF1 expression induced by IL-1{beta}/TNF required NF-{kappa}B and four novel 5' enhancer regions to IRF1 bound the NF-{kappa}B subunit, p65. These regions drove IL-1{beta}/TNF-inducible reporter activity via consensus NF-{kappa}B motifs. Three such regions recruited RNA polymerase-2 and were flanked by the active chromatin mark, histone 3 lysine 27 acetylation which, given the high levels of genetic conservation, support involvement in IRF1 transcription. Finally, IRF1 expression, transcription rate, and enhancer activity induced by IL-1{beta}/TNF were relatively unaffected by glucocorticoid co-treatment. IRF1-dependent gene expression could therefore be insensitive to glucocorticoid and may contribute to glucocorticoid-resistant diseases such as severe asthma.

molecular biology↗

bioRxiv2025

3D-Printed Magnetic Levitation Device with Deep Learning-Assisted Particle Tracking and Analysis for High-Throughput Sorting

Intrinsic density-based particle separation is fundamental to biomedical research and materials science. Magnetic levitation offers an accessible and label-free approach; however, current platforms are limited by throughput, complex fabrication requirements, and manual analysis methods. Here, we demonstrate a high-throughput magnetic levitation-based microfluidic device fabricated using commercial 3D printing, integrated with dual automated analysis systems. The device features optimized magnet configurations and wide channel design (1 mm x 1 mm) that enables gentle separation (<1 PSI) at throughputs of 66 mL/hr-- a ten-fold improvement over existing levitation platforms. We developed two complementary analysis tools: "Phase" for static levitation height measurements, and a deep learning pipeline combining CNN-based particle classification (>95% accuracy) with SORT (Simple Online and Realtime Tracking) algorithm for dynamic analysis. The automated system showed excellent correlation with manual counting (Pearson coefficients: 0.91-0.99, p<0.001). Through systematic optimization of magnet spacing and paramagnetic medium concentration (150 mM Gd), the platform achieved robust continuous-flow sorting while maintaining exceptional purity (>90%) and resolving density differences as small as 0.03 g/mL. This work establishes a versatile platform for particle sorting, enabling sophisticated analysis without specialized facilities or extensive operator training, with broad applications in biomedical research and diagnostics.

bioengineering↗

bioRxiv2024

Naturally ornate RNA-only complexes revealed by cryo-EM

Myriad families of natural RNAs have been proposed, but not yet experimentally shown, to form biologically important structures. Here we report three-dimensional structures of three large ornate bacterial RNAs using cryogenic electron microscopy at resolutions of 2.9-3.1 [A]. Without precedent among previously characterized natural RNA molecules, Giant, Ornate, Lake- and Lactobacillales-Derived (GOLLD), Rumen-Originating, Ornate, Large (ROOL), and Ornate Large Extremophilic (OLE) RNAs form homo-oligomeric complexes whose stoichiometries are retained at concentrations lower than expected in the cell. OLE RNA forms a dimeric complex with long co-axial pipes spanning two monomers. Both GOLLD and ROOL form distinct RNA-only multimeric nanocages with diameters larger than the ribosome. Extensive intra- and intermolecular A-minor interactions, kissing loops, an unusual A-A helix, and other interactions stabilize the three complexes. Sequence covariation analysis of these large RNAs reveals evolutionary conservation of intermolecular interactions, supporting the biological importance of large, ornate RNA quaternary structures that can assemble without any involvement of proteins.

biophysics↗

bioRxiv2024

Microbial Metabolite Trimethylamine N-Oxide Promotes Campylobacter jejuni Infection by Escalating Intestinal Inflammation, Epithelial Damage, and Barrier Disruption

The interactions between Campylobacter jejuni, a critical foodborne cause of gastroenteritis, and the intestinal microbiota during infection are not completely understood. The crosstalk between C. jejuni and its host is impacted by the gut microbiota through mechanisms of competitive exclusion, microbial metabolites, or immune response. To investigate the role of gut microbiota on C. jejuni pathogenesis, we examined campylobacteriosis in the IL10KO mouse model, which was characterized by an increase in the relative abundance of intestinal proteobacteria, E. coli, and inflammatory cytokines during C. jejuni infection. We also found a significantly increased abundance of microbial metabolite Trimethylamine N-Oxide (TMAO) in the colonic lumens of IL10KO mice. We further investigated the effects of TMAO on C. jejuni pathogenesis. We determined that C. jejuni senses TMAO as a chemoattractant and the administration of TMAO promotes C. jejuni invasion into Caco-2 monolayers. TMAO also increased the transmigration of C. jejuni across polarized monolayers of Caco-2 cells, decreased TEER, and increased C. jejuni-mediated intestinal barrier damage. Interestingly, TMAO treatment and presence during C. jejuni infection of Caco-2 cells synergistically caused an increased inflammatory cytokine expression, specifically IL-1{beta} and IL-8. These results establish that C. jejuni utilizes microbial metabolite TMAO for increased virulence during infection.

microbiology↗

bioRxiv2024

Single-cell transcriptional landscape of temporal neutrophil response to burn wound in larval zebrafish

Neutrophils accumulate early in tissue injury. However, the cellular and functional heterogeneity of neutrophils during homeostasis and in response to tissue damage remains unclear. Here, we use larval zebrafish to understand neutrophil responses to thermal injury. Single-cell transcriptional mapping of myeloid cells during a 3-day time course in burn and control larvae revealed distinct neutrophil subsets and their cell-cell interactions with macrophages across time and conditions. The trajectory formed by three zebrafish neutrophil subsets resembles human neutrophil maturation, with varying transition patterns between conditions. Through ligand-receptor cell-cell interaction analysis, we found neutrophils communicate more in burns in a pathway and temporal manner. Finally, we identified the correlation between zebrafish myeloid signatures and human burn severity, establishing GPR84+ neutrophils as a potential marker of early innate immune response in burns. This work builds the molecular foundation and a comparative single-cell genomic framework to identify neutrophil markers of tissue damage using model organisms.

immunology↗

bioRxiv2024

General Capillary Endothelial Cells Undergo Reprogramming into Arterial Endothelial Cells in Pulmonary Hypertension through HIF-2a/Notch4 Pathway

Pulmonary arterial hypertension (PAH) is characterized by a progressive increase of pulmonary vascular resistance and obliterative pulmonary vascular remodeling that result in right heart hypertrophy, failure, and premature death. The underlying mechanisms of loss of distal capillary endothelial cells (ECs) and obliterative vascular lesion formation remain unclear. Our recent single-cell RNA sequencing, spatial transcriptomics analysis, RNASCOPE, and immunostaining analysis showed that arterial ECs accumulation and loss of capillary ECs were evident in human PAH patients and pulmonary hypertension (PH) rodents. Pseudotime trajectory analysis of the single-cell RNA sequencing data suggest that lung capillary ECs transit to arterial ECs during the development of PH. Our study also identified CXCL12 as the marker for arterial ECs in PH. Capillary EC lineage tracing approach using capillary specific-Dre;Tdtomato reporter mice demonstrated that capillary ECs gave rise to arterial ECs during PH development. Genetic deletion of HIF-2a or pharmacological inhibition of Notch4 normalized the arterial programming in PH. In conclusion, our study demonstrates that capillary endothelium transits to arterial endothelium through the HIF-2a-Notch4 pathway during the development of PAH. Thus, targeting arterial EC transition might be a novel approach for treating PAH patients.

physiology↗

bioRxiv2023

Colonic spermidine promotes the proliferation and migration of intestinal epithelial cells.

The gut microbiome, comprising trillions of diverse microorganisms, profoundly influences the efficient development and maintenance of the intestinal barrier. While shifts in microbial composition are observed in Inflammatory Bowel Disease (IBD) and Colorectal Cancer (CRC), the causal relationship between these changes and resolution of inflammation remains elusive. Notably, IBD is not only marked by shifts in microbial composition but also by changes in microbial metabolites. Polyamines, produced by both gut bacteria and human cells, have emerged as potential regulators of gut pathology, cancer and mucosal repair. Investigating how elevated polyamine levels influence intestinal epithelial cells (IEC) can provide insights into their role in the regeneration of mucosal epithelia and restoration of gut barrier functions. To dissect the complex interplay between the gut microbiome, polyamines, and IEC, we focused on the overrepresented bacterium B. uniformis and its primary metabolite, spermidine. Here, we show that B. uniformis, a dominant member of gut microbiota, expands during the repair & resolution phase of the chemically induced chronic murine colitis. Furthermore, we found that the abundance of colonic polyamines was also altered, with spermidine being the abundant polyamine. Our RNA sequencing and transcriptomic analysis of cultured colonic epithelial cells demonstrate that spermidine regulates the expression of genes and pathways involved in different cellular functions, including proliferation, differentiation, adhesion, lipid metabolism, migration, chemotaxis, and receptor expression. We also found that spermidine stimulates the proliferation of cultured colonic epithelial cells in vitro. Additionally, our findings indicate that spermidine enhances the migrations of enterocytes. Our study emphasizes the crucial functions of the gut microbiome and polyamines in governing the functions of intestinal epithelial cells. Thus, these microorganisms and their metabolic byproducts hold promise as prospective therapeutic agents.

microbiology↗

bioRxiv2023

Development of Multiomics in situ Pairwise Sequencing (MiP-Seq) for Single-cell Resolution Multidimensional Spatial Omics

Delineating the spatial multiomics landscape will pave the way to understanding the molecular basis of physiology and pathology. However, current spatial omics technology development is still in its infancy. Here, we developed a high-throughput multiomics in situ pairwise sequencing (MiP-Seq) strategy to efficiently decipher multiplexed DNAs, RNAs, proteins, and small biomolecules at subcellular resolution. We delineated dynamic spatial gene profiles in the hypothalamus using MiP-Seq. Moreover, MiP-Seq was unitized to detect tumor gene mutations and allele-specific expression of parental genes and to differentiate sites with and without the m6A RNA modification at specific sites. MiP-Seq was combined with in vivo Ca2+ imaging and Raman imaging to obtain a spatial multiomics atlas correlated to neuronal activity and cellular biochemical fingerprints. Importantly, we proposed a "signal dilution strategy" to resolve the crowded signals that challenge the applicability of in situ sequencing. Together, our method improves spatial multiomics and precision diagnostics and facilitates analyses of cell function in connection with gene profiles.

molecular biology↗

bioRxiv2022

Increased mRNA expression of CDKN2A is a transcriptomic marker of clinically aggressive meningiomas

BackgroundHomozygous loss of CDKN2A/B is a genetic alteration found in many cancer types including meningiomas, where it is associated with poor clinical outcome. It is now also a diagnostic criterion for grade 3 meningiomas in the 2021 WHO classification for central nervous system tumors. However, as in other cancers, the relationship between copy number loss of CDKN2A/B and expression of its gene product is unclear and may be either commensurate or paradoxical in nature. Therefore, we aimed to investigate the association of CDKN2A mRNA expression with clinical prognosis, WHO grade, and other molecular biomarkers in meningiomas such as DNA methylation, molecular group, and proteomics. MethodsWe used multidimensional molecular data of 490 meningioma samples from 4 independent cohorts to examine the relationship between mRNA expression of CDKN2A and copy number status, its correlation to clinical outcome, the transcriptomic pathways altered in differential CDKN2A expression, and its relationship with DNA methylation, and proteomics using an integrated molecular approach. ResultsMeningiomas without any copy number loss were dichotomized into high (CDKN2Ahigh) and low (CDKN2Alow) CDKN2A mRNA expression groups. Patients with CDKN2Ahigh meningiomas had poorer progression free survival (PFS) compared to those with CDKN2Alow meningiomas. CDKN2A mRNA expression was increased in more aggressive molecular groups, and in higher WHO grade meningiomas across all cohorts. CDKN2Ahigh meningiomas and meningiomas with CDKN2A copy number loss shared common up-regulated cell cycling pathways. CDK4 mRNA expression was increased in CDKN2Ahigh meningiomas and both p16 and CDK4 protein were more abundant in CDKN2Ahigh meningiomas. CDKN2Ahigh meningiomas were frequently hypermethylated at the gene body and UTR compared to CDKN2Alow meningiomas and found be more commonly Rb-deficient. ConclusionsAn intermediate level of CDKN2A mRNA expression appears to be optimal as significantly low (CDKN2A deleted) or high expression (CDKN2Ahigh) are associated with poorer outcomes clinically. Though CDK4 is elevated in CDKN2Ahigh meningiomas, Rb-deficiency may be more common in this group, leading to lack of response to CDK inhibitors.

cancer biology↗

bioRxiv2022

Serotyping Salmonella Enteritidis and Typhimurium Using Whole Cell Matrix Assisted Laser Desorption Ionization - Time of Flight Mass Spectrometry (MALDI-TOF MS) through Multivariate Analysis and Artificial Intelligence

Salmonella is one of the most frequent food-borne zoonoses, while Salmonella Typhimurium and Enteritidis are the major serovars of concern in public health. 113 Salmonella strains including 38 S. Enteritidis (SE), 38 S. Typhimurium (ST) and 37 strains of 32 other Salmonella serovars (SG) were tested in quadruplicate by whole-cell MALDI-TOF MS. Ions were studied and aligned from the raw data of mzXML files using Mass-Up (http://www.sing-group.org/mass-up), resulting in 1,741 aligned peaks. Datasets of ions (presence/absence) selected using a home-developed criteria on their specificity and detectability were subjected to multivariate analyses and artificial intelligence tools. Principle Component Analysis based on 88 selected ions separated SE, ST and SG without overlap on the first three principle components. The network and forest based deep learning tools were more sophisticated than the decision tree-based models. Neural Network carried out consistently well in training model, but no advantage was gained over the other models in validation results. HP (high performance) Neural, Support Vector Machine, HP Forest and Gradient Boasting were able to identify SE, ST and SG up to 100% correctly in both training and validation when 88 selected ions were used in analysis. Among them, HP Neural seemed to perform slightly better and relatively stable. Selection of serovar specific ions helps develop serotyping by increasing signal to noise. MALDI-TOF MS used with appropriate data processing strategies and classification tools could be applied to quickly alert when Salmonella serotypes of concern are suspected among routinely processed samples.

microbiology↗

bioRxiv2022

Comparative analysis about the structure of complete chloroplast genomes in genus Mangifera and accuracy verification about phylogenetic analysis based on gene ycf2 in genus level

Mango is an evergreen plant belonging to the genus Mangifera of the Anacardiaceae family. Genus Mangifera has 69 species of Mangifera around the world that are mainly distributed in tropical and subtropical countries, including India, Indonesia, the Malay Peninsula, Thailand, and South China. It is a popular tropical fruit known as the "King of Tropical Fruits". However, the study of the structure information of the complete chloroplast genome of Mangifera was microscopic, there was no report about the comparison of SSR, Ka/Ks, codons analysis and RNA editing, so in this study, we sequenced the 6 Mangifera samples and used three different ways to analyze the relationship of 6 species of Mangifera. Then we got some results, through the RNA editing and Ka/Ks calculating, we found the species could be divided into two groups, and the difference between the two groups was protein-coding gene ccsA. Moreover, all RNA editing occurred conversion of C to T and the gene ndhB had the most RNA editing sites in all species. In Ka/Ks analysis, the gene atpB, cemA, clpP, ndhD, petD, petB and ycf15 would be suffered from the positive selection after divergence. We also find the IR regions in these seven samples were very conservation through IR contraction and expansion and Sequence Divergence Analysis. Finally, we tried to confirm the relationship between 7 samples of Mangifera in Angiosperms in 3 different ways. Then we got that ML210 and MP090 had a closer relationship than others, MS796 had a closer relationship with ML210 and MP090 than others. At the same time, the method of phylogenetic analysis based on the gene ycf2 was not more accurate at the genus level than the method based on complete cp genome and proteincoding genes.

molecular biology↗

bioRxiv2021

The rs1421085 variant within FTO promotes but not inhibits thermogenesis and is potentially associated with human migration

Disease-associated GWAS loci are predominantly scattered among noncoding regions of the human genome, which impedes causality estimation. One lead risk signal of obesity-rs1421085 T>C within the FTO gene-is reported to functional in vitro but lack of organismal evidence. Here, we established global and the brown-adipocyte specific locus-knock-in mice to recapitulate this homologous variant in humans, and discovered the minor allele (C-allele) as one candidate thermogenic locus. Mice carrying the C-alleles showed increased thermogenic capacity and a resistance to high-fat diet-induced adiposity. In terms of mechanism, the knock-in models showed enhanced FTO expression, while FTO knockdown or inhibition effectively eliminated the increased thermogenic ability of brown adipocytes. In humans, the C-allele was associated with lower birthweight, and its allele frequency increases following the environmental temperature decreases. Cumulatively, these findings demonstrated rs1421085 T>C as a functional variant regulating whole-body thermogenesis, and this variation was possibly related to early human migration from hot to cold environments.

genetics↗

bioRxiv2021

A novel approach to evaluate alpha-synuclein seeding shows a wide heterogeneity in multiple system atrophy

Several in vitro and in vivo findings have consistently shown that -synuclein derived from multiple system atrophy (MSA) subjects has more seeding capacity than Parkinsons disease-derived -synuclein. However, reliable detection of -synuclein derived from MSA using seeded amplification assays, such as the Real-Time Quaking-induced Conversion, has remained challenging. Here we demonstrate that the interaction of the Thioflavin T dye with -synuclein from MSA and Parkinsons disease patients can be modulated by the type of salt, pH, and ionic strength used to generate strain-specific reaction buffers. Employing this novel approach, we have generated a streamlined Real-Time Quaking-induced Conversion assay capable of categorizing MSA brains according to their -synuclein seeding behavior, and to unravel a previously unrecognized heterogeneity in seeding activity between different brain regions of a given individual that goes beyond immunohistochemical observations and provide a framework for future molecular subtyping of MSA.

neuroscience↗

bioRxiv2020

Structure and inhibition of the SARS-CoV-2 main protease reveals strategy for developing dual inhibitors against Mpro and cathepsin L

The main protease (Mpro) of SARS-CoV-2, the pathogen responsible for the COVID-19 pandemic, is a key antiviral drug target. While most SARS-CoV-2 Mpro inhibitors have a {gamma}-lactam glutamine surrogate at the P1 position, we recently discovered several Mpro inhibitors have hydrophobic moieties at the P1 site, including calpain inhibitors II/XII, which are also active against human cathepsin L, a host-protease that is important for viral entry. To determine the binding mode of these calpain inhibitors and establish a structure-activity relationship, we solved X-ray crystal structures of Mpro in complex with calpain inhibitors II and XII, and three analogues of GC-376, one of the most potent Mpro inhibitors in vitro. The structure of Mpro with calpain inhibitor II confirmed the S1 pocket of Mpro can accommodate a hydrophobic methionine side chain, challenging the idea that a hydrophilic residue is necessary at this position. Interestingly, the structure of calpain inhibitor XII revealed an unexpected, inverted binding pose where the P1 pyridine inserts in the S1 pocket and the P1 norvaline is positioned in the S1 pocket. The overall conformation is semi-helical, wrapping around the catalytic core, in contrast to the extended conformation of other peptidomimetic inhibitors. Additionally, the structures of three GC-376 analogues UAWJ246, UAWJ247, and UAWJ248 provide insight to the sidechain preference of the S1, S2, S3 and S4 pockets, and the superior cell-based activity of the aldehyde warhead compared with the -ketoamide. Taken together, the biochemical, computational, structural, and cellular data presented herein provide new directions for the development of Mpro inhibitors as SARS-CoV-2 antivirals.

biophysics↗

bioRxiv2020

Predicting the Immunogenicity of T cell epitopes: From HIV to SARS-CoV-2

We describe a physics-based learning model for predicting the immunogenicity of Cytotoxic T Lymphocyte (CTL) epitopes derived from diverse pathogens, given a Human Leukocyte Antigen (HLA) genotype. The model was trained and tested on experimental data on the relative immunodominance of CTL epitopes in Human Immunodeficiency Virus infection. The method is more accurate than publicly available models. Our model predicts that only a fraction of SARS-CoV-2 epitopes that have been predicted to bind to HLA molecules is immunogenic. The immunogenic CTL epitopes across all SARS-CoV-2 proteins are predicted to provide broad population coverage, but the immunogenic epitopes in the SARS-CoV-2 spike protein alone are unlikely to do so. Our model predicts that several immunogenic SARS-CoV-2 CTL epitopes are identical to those contained in low-pathogenicity coronaviruses circulating in the population. Thus, we suggest that some level of CTL immunity against COVID-19 may be present in some individuals prior to SARS-CoV-2 infection.

immunology↗

bioRxiv2019

Microscaled Proteogenomic Methods for Precision Oncology

Cancer proteogenomics integrates genomics, transcriptomics and mass spectrometry (MS)-based proteomics to gain insights into cancer biology and treatment efficacy. A proteogenomics approach was therefore developed for frozen core biopsies using tissue-sparing specimen processing with a \"microscaled\" proteomics workflow. For technical proof-of-principle, biopsies from ERBB2 positive breast cancers before and 48-72 hours after the first dose of neoadjuvant trastuzumab-based chemotherapy were analyzed. ERBB2 protein and phosphosite levels, as well as mTOR target phosphosites, were significantly more suppressed upon treatment in cases associated with pathological complete response, suggesting MS-based pharmacodynamics is achievable. Furthermore, integrated analyses indicated potential causes of treatment resistance including the absence of ERBB2 amplification (false-ERBB2 positive) and insufficient ERBB2 activity for therapeutic sensitivity despite ERBB2 amplification (pseudo-ERBB2 positive). Candidate resistance features in true-ERBB2+ cases, including androgen receptor signaling, mucin expression and an inactive immune microenvironment were observed. Thus, proteogenomic analysis of needle core biopsies is feasible and clinical utility should be investigated.

cancer biology↗