Microfluidics guided by deep learning for cancer immunotherapy screening.

Immunocyte infiltration and cytotoxicity play critical roles in both inflammation and immunotherapy. However, current cancer immunotherapy screening methods overlook the capacity of the T cells to penetrate the tumor stroma, thereby significantly limiting the development of effective treatments for solid tumors. Here, we present an automated high-throughput microfluidic platform for simultaneous tracking of the dynamics of T cell infiltration and cytotoxicity within the 3D tumor cultures with a tunable stromal makeup. By recourse to a clinical tumor-infiltrating lymphocyte (TIL) score analyzer, which is based on a clinical data-driven deep learning method, our platform can evaluate the efficacy of each treatment based on the scoring of T cell infiltration patterns. By screening a drug library using this technology, we identified an epigenetic drug (lysine-specific histone demethylase 1 inhibitor, LSD1i) that effectively promoted T cell tumor infiltration and enhanced treatment efficacy in combination with an immune checkpoint inhibitor (anti-PD1) in vivo. We demonstrated an automated system and strategy for screening immunocyte-solid tumor interactions, enabling the discovery of immuno- and combination therapies.

Adsorption Properties of Dissolved Gas Molecules in Transformer Oil on the ReSe2 Monolayer: A DFT Study.

Based on density functional theory, the adsorption behavior of seven typical dissolved gas molecules (CO, CO2, H2, CH4, C2H2, C2H4, and C2H6) and H2O molecule on the ReSe2 monolayer was systematically investigated. The interactions between the ReSe2 monolayer and eight gas molecules were investigated by calculating the adsorption energies, charge transfer, density of states (DOS), and deformation charge density (DCD) for eight different adsorption systems. The gas sensitivity of the ReSe2 monolayer toward these gases was studied using frontier molecular orbital theory and work function analysis. The results demonstrate that compared to other gas molecules, the ReSe2 monolayer exhibits a stronger interaction with CO with an adsorption energy of -1.49 eV. It also displays excellent sensitivity and selectivity toward CO making it a promising candidate for CO gas sensing applications. We aspire that this research will offer theoretical guidance for the development of ReSe2-based gas sensors and contribute to state monitoring technology in oil-immersed power equipment.

Stability of Protein Pharmaceuticals: Recent Advances.

There have been significant advances in the formulation and stabilization of proteins in the liquid state over the past years since our previous review. Our mechanistic understanding of protein-excipient interactions has increased, allowing one to develop formulations in a more rational fashion. The field has moved towards more complex and challenging formulations, such as high concentration formulations to allow for subcutaneous administration and co-formulation. While much of the published work has focused on mAbs, the principles appear to apply to any therapeutic protein, although mAbs clearly have some distinctive features. In this review, we first discuss chemical degradation reactions. This is followed by a section on physical instability issues. Then, more specific topics are addressed: instability induced by interactions with interfaces, predictive methods for physical stability and interplay between chemical and physical instability. The final parts are devoted to discussions how all the above impacts (co-)formulation strategies, in particular for high protein concentration solutions.'

Colistin monotherapy or combination for the treatment of bloodstream infection caused by Klebsiella pneumoniae: a systematic review and meta-analysis.

BACKGROUND: Bloodstream infection of Klebsiella pneumoniae (BSI-KP) were associated with increased mortality. Klebsiella pneumoniae was tested to susceptible to colistin by E-test and broth microdilution method in clinical laboratory. This study aimed to assess the efficacy of colistin versus tigecycline, carbapenem monotherapy and combination in the treatment of BSI-KP. METHODS: Electronic databases such as PubMed, Web of Science and Embase were searched. The last search was in November 24th, 2022, addressing the colistin, carbapenems and tigecycline monotherapy and combination treatments in patients with BSI-KP. The primary outcomes were 30-day or 28-day mortality. OR where available with 95% CI were pooled in random-effects meta-analysis. RESULTS: Following the outlined search strategy, a total of 658 articles were identified from the initial database searching. Six studies, 17 comparisons were included. However, they all were observational design, lacking high-quality randomized controlled trials (RCTs). Moderate or low-quality evidences suggested that colistin monotherapy was associated with an OR = 1.35 (95% CI = 0.62-2.97, P = 0.45, Tau2 = 0.00, I2 = 0%) compared with tigecycline monotherapy, OR = 0.81 (95% CI = 0.27-2.45, P = 0.71, Tau2 = 0.00, I2 = 0%) compared with carbapenem monotherapy. Compared with combination with tigecycline or carbapenem, Colistin monotherapy resulted in OR of 3.07 (95% CI = 1.34-7.04, P = 0.008, Tau2 = 0.00, I2 = 0%) and 0.98 (95%CI = 0.29-3.31, P = 0.98, Tau2 = 0.00, I2 = 0% ), respectively. CONCLUSIONS: Colistin, carbapenem and tigecycline monotherapy showed similar treatment effects in patients who suffered from BSI-KP. Compared with colistin monotherapy, colistin combined tigecycline therapy might play the synergism effects. TRIAL REGISTRATION: retrospectively registered.

Traceability Research on Geographic Erigeron breviscapus Based on High-Resolution Mass Spectrometry and Chemometric Analysis.

A method was developed to identify and trace the geographic sources of Erigeron breviscapus using high-resolution mass spectrometry and chemometrics. The representative samples were collected from the geographic area of Honghe Dengzhanhua and other areas in Yunnan province and Guizhou province. The data points could be determined well using the PCA and PLS-DA diagram. A total of 46 characteristic compounds were identified from Honghe Dengzhanhua and within Guizhou province, but 37 compounds were different from Honghe Dengzhanhua and other counties in Yunnan province. Two biomarkers were found from three regions. Their structures were inferred as 8-amino-7-oxononanoic acid and 8-hydroxyquinoline, and they had the same molecular composition. This may suggest that a possible synthesis pathway can be proven in the future.

Three-dimensional force detection and decoupling of a fiber grating sensor for a humanoid prosthetic hand.

A fiber Bragg grating (FBG) based three-dimensional (3D) force sensor for a humanoid prosthetic hand is designed, which can precisely detect 3D force and compensate for ambient temperature. FBG was encapsulated in polydimethylsiloxane (PDMS) for force sensitization and immobilization, and the structural parameters of the sensor were optimized by using finite element simulation, so that its sensitivity to 3D force is enhanced. In the meantime, the calibration experiments for normal force fZ, shear force fX/fY, and temperature were conducted, and the 3D force data were decoupled using the least square (LS) and backpropagation (BP) neural networks decoupling methods, so that an overall decoupling error is 0.038. The results show that the sensor has a simple structure, high sensitivity, high linearity, good creep resistance, and rapid decoupling, providing a successful design for the 3D force detection of a humanoid prosthetic hand.

FBG-based three-dimensional micro-force sensor with axial force sensitivity-enhancing and temperature compensation for micro-forceps.

During retinal microsurgery, excessive interaction force between surgical instruments and intraocular tissue can cause serious accidents such as tissue injury, irreversible retinal damage, and even vision loss. It is essential to accurately sense the micro tool-tissue interaction force, especially for the Ophthalmic Microsurgery Robot. In this study, a fiber Bragg grating (FBG) three-dimensional (3-D) micro-force sensor for micro-forceps is proposed, which is integrated with the drive module as an end-effector and can be conveniently mounted onto the ophthalmic surgical robot. An innovative axial force sensitivity-enhancing structure is proposed based on the principles of flexure-hinge and flexible levers to overcome the low sensitivity of axial force measurement. A dual-grating temperature compensation method is adopted for axial force measurement, which considers the differential temperature sensitivity of the two FBGs. Three FBGs are arranged along the circumference of the guide tube in this study to measure transverse forces and compensate for effects caused by changes in temperature. The experimental results demonstrate that the micro-forceps designed in this study achieved a resolution of 0.13 mN for transverse force and 0.30 mN for axial force. The temperature compensation experiments show that the 3-D micro-force sensor can simultaneously compensate for temperature effects in axial and transverse force measurement.

Safety and effectiveness of neoadjuvant PD-1 inhibitor (toripalimab) plus chemotherapy in stage II-III NSCLC (LungMate 002): an open-label, single-arm, phase 2 trial.

BACKGROUND: This trial aimed to analyse the safety, effectiveness and transcriptomic characteristics of neoadjuvant toripalimab plus chemotherapy in II-III non-small-cell lung cancer (NSCLC). METHODS: Patient eligibility mainly involved treatment-naive, clinical stage II-III and wild-type EGFR/ALK NSCLC. The patients received 2-4 cycles of toripalimab (240 mg q3w) plus carboplatin-based chemotherapy. After the second treatment cycle, all patients were re-evaluated by a multidisciplinary team. Candidates eligible for surgery underwent surgery; otherwise, patients received the remaining treatment cycles. The primary endpoints were safety and major pathological response (MPR). Secondary endpoints were R0 resection rate, progression-free survival (PFS) and overall survival (OS). RNA sequencing of baseline and post-treatment samples was conducted to explore the transcriptomic characteristics of the therapeutic response. RESULTS: In total, 50 eligible patients were enrolled, including 12 (24.0%) with resectable disease (RD) and 38 (76.0%) with potentially resectable disease (PRD). Treatment-related adverse events (TRAEs) were recorded in 48 cases (96.0%). Severe TRAEs occurred in 3 (6.0%) cases, including myelosuppression, drug-induced liver injury and death related to haemoptysis. The objective response rate (ORR) was 76.0%, with 8 (16.0%) patients having a complete response (CR), 30 (60.0%) partial response (PR), 10 (20.0%) stable disease (SD) and 2 (4.0%) progressive disease (PD). Surgery could be achieved in 12 (100%) patients with RD and 25 (65.8%) with PRD; 1 (2.0%) with PRD refused surgery. Therefore, R0 resection was performed for all 36 (100%) patients who underwent surgery; 20 (55.6%) achieved MPR, including 10 (27.8%) with a complete pathological response (pCR). The CHI3L1 (chitinase-3-like protein 1) immunohistochemistry (IHC) expression of baseline tumour samples could predict the therapeutic response (AUC=0.732), OS (P=0.017) and PFS (P=0.001). Increased PD-1 expression, T cell abundance and immune-related pathway enrichment were observed in post-treatment samples compared to baseline in the response group (CR+PR) but not in the non-response group (SD+PD). CONCLUSIONS: Neoadjuvant toripalimab plus chemotherapy was safe and effective, with a high MPR and manageable TRAEs for II-III NSCLC, even converting initially PRD to RD. Disparate transcriptomic characteristics of therapeutic efficiency were observed, and CHI3L1 expression predicted therapeutic response and survival. TRIAL REGISTRATION: ChiCTR1900024014, June 22, 2019.

Simultaneous Determination and Health Risk Assessment of Four High Detection Rate Pesticide Residues in Pu'er Tea from Yunnan, China.

Four pesticides with a high detection rate in Pu'er tea have been determined by a QuEChERS (quick, easy, cheap, effective, rugged, safe) method with multiwalled carbon nanotubes (MWCNTs), and combined ultrahigh-performance liquid chromatography-triple quadrupole linear ion trap-tandem mass spectrometry (UHPLC-QTRAP-MS/MS). MWCNs have been compared with other common purification materials, and found to be superior. The matrix effect was systematically studied, and the results show that the MWCNs can quickly and effectively reduce matrix interference values, which were in the range from -17.8 to 13.8. The coefficients (R2) were greater than 0.99, with the limit of quantification ranging from 0.1 to 0.5 µg/kg, and the recovery rate ranging from 74.8% to 105.0%, while the relative standard deviation (RSD) ranged from 3.9% to 6.6%. A total of 300 samples, taken from three areas in which Yunnan Pu'er tea was most commonly produced, tested for four pesticides. The results show that the detection rate of tolfenpyrad in Pu'er tea was 35.7%, which is higher than other pesticides, and the lowest was indoxacarb, with 5.2%. The residual concentrations of chlorpyrifos, triazophos, tolfenpyrad and indoxacarb ranged from 1.10 to 5.28, 0.014 to 0.103, 1.02 to 51.8, and 1.07 to 4.89 mg/kg, respectively. By comparing with China's pesticide residue limits in tea (GB 2763-2021), the over standard rates of chlorpyrifos, tolfenpyrad, and indoxacarb were 4.35%, 0.87% and 0%, respectively. The risk assessment result obtained with the hazard quotient (HQ) method shows that the HQ of the four pesticides was far less than one, indicating that the risk is considered acceptable for the four pesticides in Pu'er tea. The largest HQ was found for tolfenpyrad, 0.0135, and the smallest was found for indoxacarb, 0.000757, but more attention should be paid to tolfenpyrad in daily diets in the future, because its detection rate, and residual and residual median were all relatively high.

The development and controversy of competitive endogenous RNA hypothesis in non-coding genes.

As a momentous post-transcriptional regulator, microRNAs (miRNAs) are attracting more and more attention. The classical miRNAs regulated mechanism shows it binds to the targets' 3'UTR thus play the role in post-transcription. Meanwhile, single miRNA can target multiple genes, so those should compete to bind that miRNA. Vice versa, single gene can sponge mass of miRNAs as well. Thus the competitive endogenous RNAs (ceRNAs) hypothesis was put forward in 2011. The ceRNA hypothesis has made huge achievements, in particular in non-coding genes, which including long non-coding RNAs (lncRNAs), circle RNAs (circRNAs) and pseudogenes, even viral transcripts. It also contributed greatly to epigenetics development. However, an increasing number of controversies have occurred with applause. Based on this situation, this review introduces something in detail about the ceRNAs hypothesis achieved in lncRNAs, circRNAs, pseudogenes and viral transcripts, respectively. Meanwhile, it also covers controversy of the ceRNAs hypothesis.

Immune infiltration landscape and immune-marker molecular typing of pulmonary fibrosis with pulmonary hypertension.

BACKGROUND: Pulmonary arterial hypertension (PH) secondary to pulmonary fibrosis (PF) is one of the most common complications in PF patients, it causes severe disease and usually have a poor prognosis. Whether the combination of PH and PF is a unique disease phenotype is unclear. We aimed to screen the key modules associated with PH-PF immune infiltration based on WGCNA and identify the hub genes for molecular typing. METHOD: Using the gene expression profile GSE24988 of PF patients with or without PH from the Gene Expression Omnibus (GEO) database, we evaluated immune cell infiltration using Cibersortx and immune cell gene signature files. Different immune cell types were screened using the Wilcoxon test; differentially expressed genes were screened using samr. The molecular pathways implicated in these differential responses were identified using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes functional enrichment analyses. A weighted co-expression network of the differential genes was constructed, relevant co-expression modules were identified, and relationships between modules and differential immune cell infiltration were calculated. The modules most relevant to this disease were identified using weighted correlation network analysis. From these, we constructed a co-expression network; using the STRING database, we integrated the values into the human protein-protein interaction network before constructing a co-expression interaction subnet, screening genes associated with immunity and unsupervised molecular typing, and analyzing the immune cell infiltration and expression of key genes in each disease type. RESULTS: Of the 22 immune cell types from the PF GEO data, 20 different immune cell types were identified. There were 1622 differentially expressed genes (295 upregulated and 1327 downregulated). The resulting weighted co-expression network identified six co-expression modules. These were screened to identify the modules most relevant to the disease phenotype (the green module). By calculating the correlations between modules and the differentially infiltrated immune cells, extracting the green module co-expression network (46 genes), extracting 25 key genes using gene significance and module-membership thresholds, and combining these with the 10 key genes in the human protein-protein interaction network, we identified five immune cell-related marker genes that might be applied as biomarkers. Using these marker genes, we evaluated these disease samples using unsupervised clustering molecular typing. CONCLUSION: Our results demonstrated that all PF combined with PH samples belonged to four categories. Studies on the five key genes are required to validate their diagnostic and prognostic value.

Beam flatness modulation for a flattening filter free photon beam utilizing a novel direct leaf trajectory optimization model.

Flattening filter free (FFF) linear accelerators produce a fluence distribution that is forward peaked. Various dosimetric benefits, such as increased dose rate, reduced leakage and out of field dose has led to the growth of FFF technology in the clinic. The literature has suggested the idea of vendors offering dedicated FFF units where the flattening filter (FF) is removed completely and manipulating the beam to deliver conventional flat radiotherapy treatments. This work aims to develop an effective way to deliver modulated flat beam treatments, rather than utilizing a physical FF. This novel optimization model is an extension of the direct leaf trajectory optimization (DLTO) previously developed for volumetric modulated radiation therapy (VMAT) and is capable of accounting for all machine and multileaf collimator (MLC) dynamic delivery constraints, using a combination of linear constraints and a convex objective function. Furthermore, the tongue and groove (T&G) effect was also incorporated directly into our model without introducing nonlinearity to the constraints, nor nonconvexity to the objective function. The overall beam flatness, machine deliverability, and treatment time efficiency were assessed. Regular square fields, including field sizes of 10 × 10 cm2 to 40 × 40 cm2 were analyzed, as well as three clinical fields, and three arbitrary contours with "concave" features. Quantitative flatness was measured for all modulated FFF fields, and the results were comparable or better than their open FF counterparts, with the majority having a quantitative flatness of less than 3.0%. The modulated FFF beams, due to the included efficiency constraint, were able to achieve acceptable delivery time compared to their open FF counterpart. The results indicated that the dose uniformity and flatness for the modulated FFF beams optimized with the DLTO model can successfully match the uniformity and flatness of their conventional FF counterparts, and may even provide further benefit by taking advantage of the unique FFF beam characteristics.

A Digital Acoustofluidic Pump Powered by Localized Fluid-Substrate Interactions.

The precise transportation of small-volume liquids in microfluidic and nanofluidic systems remains a challenge for many applications, such as clinical fluidical analysis. Here, we present a reliable digital pump that utilizes acoustic streaming induced by localized fluid-substrate interactions. By locally generating streaming via a C-shaped interdigital transducer (IDT) within a triangle-edged microchannel, our acoustofluidic pump can generate a stable unidirectional flow (∼nanoliter per second flow rate) with a precise digital regulation (∼second response time), and it is capable of handling aqueous solutions (e.g., PBS buffer) as well as high viscosity liquids (e.g., human blood) with a nanoliter-scale volume. Along with our acoustofluidic pump's low cost, programmability, and capacity to control small-volumes at high precision, it could be widely used for point-of-care diagnostics, precise drug delivery, and fundamental biomedical research.

Optical-fiber-based dynamic measurement system for 3D tip clearance of rotating blades.

This paper proposes a dynamic model for measuring the 3D tip clearance of rotating blades using an isosceles-right-triangle sensing structure consisting of three identical two-circle coaxial optical fiber bundles and a demodulation approach using the ratios of difference signals between any two bundles based on the Taylor expansion principle. The dynamic system comprising the optical fiber probe, hardware circuits, and measurement software is designed and established according to the characteristics of dynamic sensing signals. Finally, the feasibility of the system for the dynamic measurement of 3D tip clearance is experimentally verified using the simulated blades of a rotor test bench.

Demodulation technique for 3-D tip clearance measurements based on output signals from optical fiber probe with three two-circle coaxial optical fiber bundles.

In this work, we propose a new demodulation technique for three-dimensional (3-D) tip clearance measurements using the output signals acquired from three two-circle coaxial optical fiber bundles. This technique is based on the ratio of the difference in the signal intensities between any two sensing units of the optical fiber probe, and we derived the demodulation equations using the second-order Taylor expansion for a three-variable function. We verified the feasibility of the demodulation technique by experiments and demodulation error curves, which indicates that the method is viable for 3-D tip clearance measurements.

Modifications of recombinant monoclonal antibodies in vivo.

Therapeutic recombinant monoclonal antibodies are subject to various modifications during cell culture, and to a lesser degree, during purification. These modifications are expected to remain relatively constant during storage with the protection of appropriate formulations. However, after administration to patients, the levels of modifications may vary over time in circulation, where the recombinant monoclonal antibodies are exposed to the physiological conditions. Scientific understanding of those in vivo modifications can help drug candidate selection to choose the most stable molecules and set appropriate specifications for product release, which ultimately ensures safety and efficacy.

Sample Average Approximation with Sparsity-Inducing Penalty for High-Dimensional Stochastic Programming.

The theory on the traditional sample average approximation (SAA) scheme for stochastic programming (SP) dictates that the number of samples should be polynomial in the number of problem dimensions in order to ensure proper optimization accuracy. In this paper, we study a modification to the SAA in the scenario where the global minimizer is either sparse or can be approximated by a sparse solution. By making use of a regularization penalty referred to as the folded concave penalty (FCP), we show that, if an FCP-regularized SAA formulation is solved locally, then the required number of samples can be significantly reduced in approximating the global solution of a convex SP: the sample size is only required to be poly-logarithmic in the number of dimensions. The efficacy of the FCP regularizer for nonconvex SPs is also discussed. As an immediate implication of our result, a flexible class of folded concave penalized sparse M-estimators in high-dimensional statistical learning may yield a sound performance even when the problem dimension cannot be upper-bounded by any polynomial function of the sample size.

Asparagine Deamidation in a Complementarity Determining Region of a Recombinant Monoclonal Antibody in Complex with Antigen.

Asparagine deamidation in the complementarity determining regions of recombinant monoclonal antibodies has been extensively studied and shown to have a negative impact on antigen binding. Those asparagine residues are typically exposed and thus have a higher tendency toward deamidation, depending also on local structure and environmental factors such as temperature and pH. Deamidation rates and products of a susceptible asparagine residue in the complementarity determining regions of a recombinant monoclonal antibody free in solution or in the antibody-antigen complex were studied. The results demonstrated that incubation of the antibody or its antigen complex generated a similar amount of aspartate. The expected amount of isoaspartate product was detected in free antibody, but it was completely lacking in the antibody-antigen complex.

Characterization of succinimide stability during trypsin digestion for LC-MS analysis.

LC-MS peptide mapping is the most commonly used method to analyze protein modifications. The proteins are generally digested using trypsin at a slightly basic pH at 37 °C from several hours to overnight. Assay-induced artifacts can be generated during this procedure, potentially causing false-positive or false-negative results for a given modification. Unfortunately, for the analysis of succinimide, both false-negative and false-positive results can be generated within the same procedure. This study evaluates the stability of succinimide during the peptide mapping procedure and has demonstrated that up to 13% of pre-existing succinimide was lost during a 4 h trypsin digestion at pH 5.0 which was previously determined to be optimal for the detection of succinimide. The same procedure was able to simultaneously generate approximately 3% succinimide. Using the optimized procedure, it was also found that two aspartate residues that are followed by glycine residues in the conserved Fc region of a recombinant monoclonal antibody were not prone to isomerization. On the other hand, an aspartate residue followed by a glycine in the heavy chain variable domain was highly susceptible to isomerization. Interestingly, the antibody containing the succinimide eluted from an SEC column after the monomer peak.

Characterization of charge variants of a monoclonal antibody using weak anion exchange chromatography at subunit levels.

An efficient strategy to characterize recombinant monoclonal antibody charge variants was established using weak anion exchange chromatography, LC-MS and IdeS digestion to allow subunit level characterization. Significantly higher resolution was achieved at subunit levels by weak anion exchange chromatography and LC-MS. In addition, subunit analysis localized potential modifications to either F(ab')2 or Fc fragments to facilitate further characterization. Peptide mapping of fractions from various charge variants after IdeS digestion identified aspartate isomerization, asparagine deamidation and glycation as the modifications. Although, aspartate isomerization does not generate net charge difference directly, it does generate antibody basic species. Antibodies with either isoaspartate or aspartate from deamidation showed different retention times by chromatography. Even more interestingly, the antibody contained succinimide as the isomerization intermediate, which though more basic compared to aspartate, eluted off the weak anion exchange column as an acidic species. The results demonstrated not only the utility of subunit level characterization but also the unpredictable chromatographic behavior of antibody charge variants.