Isolating Neural Signatures of Conscious Speech Perception with a No-Report Sine-Wave Speech Paradigm.

Identifying neural correlates of conscious perception is a fundamental endeavor of cognitive neuroscience. Most studies so far have focused on visual awareness along with trial-by-trial reports of task-relevant stimuli, which can confound neural measures of perceptual awareness with postperceptual processing. Here, we used a three-phase sine-wave speech paradigm that dissociated between conscious speech perception and task relevance while recording EEG in humans of both sexes. Compared with tokens perceived as noise, physically identical sine-wave speech tokens that were perceived as speech elicited a left-lateralized, near-vertex negativity, which we interpret as a phonological version of a perceptual awareness negativity. This response appeared between 200 and 300 ms after token onset and was not present for frequency-flipped control tokens that were never perceived as speech. In contrast, the P3b elicited by task-irrelevant tokens did not significantly differ when the tokens were perceived as speech versus noise and was only enhanced for tokens that were both perceived as speech and relevant to the task. Our results extend the findings from previous studies on visual awareness and speech perception and suggest that correlates of conscious perception, across types of conscious content, are most likely to be found in midlatency negative-going brain responses in content-specific sensory areas.

Colocalization of Chikungunya Virus with Its Receptor MXRA8 during Cell Attachment, Internalization, and Membrane Fusion.

Arthritogenic alphaviruses, including chikungunya virus (CHIKV), preferentially target joint tissues and cause chronic rheumatic disease that adversely impacts the quality of life of patients. Viruses enter target cells via interaction with cell surface receptor(s), which determine the viral tissue tropism and pathogenesis. Although MXRA8 is a recently identified receptor for several clinically relevant arthritogenic alphaviruses, its detailed role in the cell entry process has not been fully explored. We found that in addition to its localization on the plasma membrane, MXRA8 is present in acidic organelles, endosomes, and lysosomes. Moreover, MXRA8 is internalized into cells without a requirement for its transmembrane and cytoplasmic domains. Confocal microscopy and live cell imaging revealed that MXRA8 interacts with CHIKV at the cell surface and then enters cells along with CHIKV particles. At the moment of membrane fusion in the endosomes, many viral particles are still colocalized with MXRA8. These findings provide insight as to how MXRA8 functions in alphavirus internalization and suggest possible targets for antiviral development. IMPORTANCE The globally distributed arthritogenic alphaviruses have infected millions of humans and induce rheumatic disease, such as severe polyarthralgia/polyarthritis, for weeks to years. Alphaviruses infect target cells through receptor(s) followed by clathrin-mediated endocytosis. MXRA8 was recently identified as an entry receptor that shapes the tropism and pathogenesis for multiple arthritogenic alphaviruses, including chikungunya virus (CHIKV). Nonetheless, the exact functions of MXRA8 during the process of viral cell entry remain undetermined. Here, we have provided compelling evidence for MXRA8 as a bona fide entry receptor that mediates the uptake of alphavirus virions. Small molecules that disrupt MXRA8-dependent binding of alphaviruses or internalization steps could serve as a platform for unique classes of antiviral drugs.

Functional and genetic analysis of viral receptor ACE2 orthologs reveals a broad potential host range of SARS-CoV-2.

The pandemic of COVID-19, caused by SARS-CoV-2, is a major global health threat. Epidemiological studies suggest that bats (Rhinolophus affinis) are the natural zoonotic reservoir for SARS-CoV-2. However, the host range of SARS-CoV-2 and intermediate hosts that facilitate its transmission to humans remain unknown. The interaction of coronavirus with its host receptor is a key genetic determinant of host range and cross-species transmission. SARS-CoV-2 uses angiotensin-converting enzyme 2 (ACE2) as the receptor to enter host cells in a species-dependent manner. In this study, we characterized the ability of ACE2 from diverse species to support viral entry. By analyzing the conservation of five residues in two virus-binding hotspots of ACE2 (hotspot 31Lys and hotspot 353Lys), we predicted 80 ACE2 proteins from mammals that could potentially mediate SARS-CoV-2 entry. We chose 48 ACE2 orthologs among them for functional analysis, and showed that 44 of these orthologs-including domestic animals, pets, livestock, and animals commonly found in zoos and aquaria-could bind the SARS-CoV-2 spike protein and support viral entry. In contrast, New World monkey ACE2 orthologs could not bind the SARS-CoV-2 spike protein and support viral entry. We further identified the genetic determinant of New World monkey ACE2 that restricts viral entry using genetic and functional analyses. These findings highlight a potentially broad host tropism of SARS-CoV-2 and suggest that SARS-CoV-2 might be distributed much more widely than previously recognized, underscoring the necessity to monitor susceptible hosts to prevent future outbreaks.

Susceptibilities of Human ACE2 Genetic Variants in Coronavirus Infection.

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in more than 235 million cases worldwide and 4.8 million deaths (October 2021), with various incidences and mortalities among regions/ethnicities. The coronaviruses SARS-CoV, SARS-CoV-2, and HCoV-NL63 utilize the angiotensin-converting enzyme 2 (ACE2) as the receptor to enter cells. We hypothesized that the genetic variability in ACE2 may contribute to the variable clinical outcomes of COVID-19. To test this hypothesis, we first conducted an in silico investigation of single-nucleotide polymorphisms (SNPs) in the coding region of ACE2. We then applied an integrated approach of genetics, biochemistry, and virology to explore the capacity of select ACE2 variants to bind coronavirus spike proteins and mediate viral entry. We identified the ACE2 D355N variant that restricts the spike protein-ACE2 interaction and consequently limits infection both in vitro and in vivo. In conclusion, ACE2 polymorphisms could modulate susceptibility to SARS-CoV-2, which may lead to variable disease severity. IMPORTANCE There is considerable variation in disease severity among patients infected with SARS-CoV-2, the virus that causes COVID-19. Human genetic variation can affect disease outcome, and the coronaviruses SARS-CoV, SARS-CoV-2, and HCoV-NL63 utilize human ACE2 as the receptor to enter cells. We found that several missense ACE2 single-nucleotide variants (SNVs) that showed significantly altered binding with the spike proteins of SARS-CoV, SARS-CoV-2, and NL63-HCoV. We identified an ACE2 SNP, D355N, that restricts the spike protein-ACE2 interaction and consequently has the potential to protect individuals against SARS-CoV-2 infection. Our study highlights that ACE2 polymorphisms could impact human susceptibility to SARS-CoV-2, which may contribute to ethnic and geographical differences in SARS-CoV-2 spread and pathogenicity.

A novel cell culture system modeling the SARS-CoV-2 life cycle.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the global pandemic of COVID-19. SARS-CoV-2 is classified as a biosafety level-3 (BSL-3) agent, impeding the basic research into its biology and the development of effective antivirals. Here, we developed a biosafety level-2 (BSL-2) cell culture system for production of transcription and replication-competent SARS-CoV-2 virus-like-particles (trVLP). This trVLP expresses a reporter gene (GFP) replacing viral nucleocapsid gene (N), which is required for viral genome packaging and virion assembly (SARS-CoV-2 GFP/ΔN trVLP). The complete viral life cycle can be achieved and exclusively confined in the cells ectopically expressing SARS-CoV or SARS-CoV-2 N proteins, but not MERS-CoV N. Genetic recombination of N supplied in trans into viral genome was not detected, as evidenced by sequence analysis after one-month serial passages in the N-expressing cells. Moreover, intein-mediated protein trans-splicing approach was utilized to split the viral N gene into two independent vectors, and the ligated viral N protein could function in trans to recapitulate entire viral life cycle, further securing the biosafety of this cell culture model. Based on this BSL-2 SARS-CoV-2 cell culture model, we developed a 96-well format high throughput screening for antivirals discovery. We identified salinomycin, tubeimoside I, monensin sodium, lycorine chloride and nigericin sodium as potent antivirals against SARS-CoV-2 infection. Collectively, we developed a convenient and efficient SARS-CoV-2 reverse genetics tool to dissect the virus life cycle under a BSL-2 condition. This powerful tool should accelerate our understanding of SARS-CoV-2 biology and its antiviral development.

Comparative analysis reveals the species-specific genetic determinants of ACE2 required for SARS-CoV-2 entry.

Coronavirus interaction with its viral receptor is a primary genetic determinant of host range and tissue tropism. SARS-CoV-2 utilizes ACE2 as the receptor to enter host cell in a species-specific manner. We and others have previously shown that ACE2 orthologs from New World monkey, koala and mouse cannot interact with SARS-CoV-2 to mediate viral entry, and this defect can be restored by humanization of the restrictive residues in New World monkey ACE2. To better understand the genetic determinants behind the ability of ACE2 orthologs to support viral entry, we compared koala and mouse ACE2 sequences with that of human and identified the key residues in koala and mouse ACE2 that restrict viral receptor activity. Humanization of these critical residues rendered both koala and mouse ACE2 capable of binding the spike protein and facilitating viral entry. Our study shed more lights into the genetic determinants of ACE2 as the functional receptor of SARS-CoV-2, which facilitates our understanding of viral entry.

Establishment of an orthotopic prostate cancer xenograft mouse model using microscope-guided orthotopic injection of LNCaP cells into the dorsal lobe of the mouse prostate.

BACKGROUND: Orthotopic LNCaP xenograft mouse models closely mimic the progression of androgen-dependent prostate cancer in humans; however, orthotopic injection of LNCaP cells into the mouse prostate remains a challenge. METHODS: Under the guidance of a stereoscopic microscope, the anatomy of the individual prostate lobes in male Balb/c athymic nude mice was investigated, and LNCaP cells were inoculated into the mouse dorsal prostate (DP) to generate orthotopic tumors that mimicked the pathophysiological process of prostate cancer in humans. Real-time ultrasound imaging was used to monitor orthotopic prostate tumorigenesis, contrast-enhanced ultrasonography (CEUS) was used to characterize tumor angiogenesis, and macroscopic and microscopic characteristics of tumors were described. RESULTS: The DP had a trigonal bipyramid-shape and were located at the base of the seminal vesicles. After orthotopic inoculation, gray scale ultrasound imaging showed progressive changes in tumor echotexture, shape and location, and tumors tended to protrude into the bladder. After 8 weeks, the tumor take rate was 65% (n = 13/20 mice). On CEUS, signal intensity increased rapidly, peaked, and decreased gradually. Observations of gross specimens showed orthotopic prostate tumors were well circumscribed, round, dark brown, and soft, with a smooth outer surface and a glossy appearance. Microscopically, tumor cells were arranged in acini encircled by fibrous septa with variably thickened walls, mimicking human adenocarcinoma. CONCLUSIONS: This study describes a successful approach to establishing an orthotopic LNCaP xenograft Balb/c athymic nude mouse model. The model requires a thorough understanding of mouse prostate anatomy and proper technique. The model represents a valuable tool for the in vivo study of the biological processes involved in angiogenesis in prostate cancer and preclinical evaluations of novel anti-angiogenic therapies.

Diagnosis of chronic prostatitis by noninvasive methods in elderly patients with benign prostatic hyperplasia in China.

Chronic prostatitis is hard to be identified in BPH patients in clinical works. This study aimed to diagnose chronic prostatitis in BPH patients by noninvasive methods. BPH patients who received transurethral resection of prostate from January 2014 to July 2015 were enrolled in current study. Patients were received examinations of PSA, sex hormones, inflammatory cytokines, metabolic panel and transrectal ultrasonography. According to histological results, patients were divided into two group of BPH with/without prostatitis. Logistic regression was used to find risk factors of chronic prostatitis. As a result, 181 men with an average age of 72.15 ± 8.41 years were enrolled in this study, including 116 patients with prostatitis and 65 patients without prostatitis. The storage sub-score, PSA and IL-2R were significantly higher in patients with prostatitis than those without prostatitis. Based on logistic regression analysis, the above three parameters were also the risk factors of BPH with prostatitis. The diagnostic model was calculated as: 0.317 × storage sub-score + 0.092 × PSA + 0.003 × IL-2R - 4.296. The AUC was 0.725. Histological prostatitis in BPH patients can be diagnosed by the combination of serum IL-2R, PSA and storage sub-score. Identification of chronic prostatitis in BPH patients could more efficiently alleviate urinary symptoms and reduce the risk of disease progression.

The new insight of prostate-specific antigen reduction during finasteride therapy in aging men.

OBJECTIVE: To evaluate the effect of finasteride on prostate-specific antigen (PSA) in Chinese population. MATERIALS AND METHODS: From Feb 2011 to Jan 2012, 83 benign prostatic hyperplasia (BPH) patients with prostate volume (PV) >30 mL were enrolled in our study. All the patients were older than 50 years and all of them received combined therapy (finasteride + doxazosin). All the patients were required for 1-year follow-up. PSA level and PV was measured at the start, 6 and 12 months, respectively. RESULTS: 79 patients completed the follow up. PSA level reduced by approximately 40 % during finasteride therapy. We defined baseline PSA as PSA1, PSA at 6 months as PSA2, PSA at 12 months as PSA3. PSA1 was significantly correlated with PSA2/PSA1 and PSA3/PSA1. However, prostate volume was not correlated with PSA1. We divided the patients into three groups according to PSA level. Groups 1, 2, 3 represented the patients with PSA less than 2 ng/mL, between 2 and 4 ng/mL and greater than 4 ng/mL, respectively. Both the PSA2/PSA1 and the PSA3/PSA1 had significant difference among three groups. Furthermore, group 1 and group 2 both showed the fairly large data variance. CONCLUSIONS: When baseline PSA level was greater than 4 ng/mL, the doubling rule could be used for screening. When baseline PSA level was less than 4 ng/Ml, the doubling rule might not be an accurate predictor. We can use the PSA rise from nadir or proPSA to predict prostate cancer.

Efficacy of 5α-reductase inhibitors for patients with large benign prostatic hyperplasia (>80 mL) after transurethral resection of the prostate.

PURPOSE: To prospectively evaluate 5α-reductase inhibitors (5αRIs) for benign prostatic hyperplasia (BPH) patients with a large prostate (>80 mL) after transurethral resection of the prostate (TURP). MATERIALS AND METHODS: Eighty-seven patients were recruited from January 2007 to October 2014. Patients were randomized into a trial and a control group. The trial group was treated with 5αRIs for 3 years after TURP, while the control group received a placebo. We evaluated the indicators before, peri and after TURP. RESULTS: There were no significant differences in the indicators before and peri-TURP. Six months later, there were significant differences in PSA and hematuria (HU). Three years after TURP, there were significant differences in prostate volume (PV), level of prostate-specific antigen (PSA), the maximum flow rate (Qm), and HU between the trial and control groups. Additionally, there were significant differences in the PV, PSA, international prostate symptom score (IPSS), patient quality of life (QoL) in the trial group alone between those treated with finasteride and those treated with dutasteride. CONCLUSIONS: After TURP for large BPH, administration of 5αRIs for 3 years improved PV, PSA, Qm and HU. Additionally, dutasteride produced superior improvements in PV, PSA, IPSS and QoL compared with finasteride.

In Vivo Molecular MRI Imaging of Prostate Cancer by Targeting PSMA with Polypeptide-Labeled Superparamagnetic Iron Oxide Nanoparticles.

The prostate specific membrane antigen (PSMA) is broadly overexpressed on prostate cancer (PCa) cell surfaces. In this study, we report the synthesis, characterization, in vitro binding assay, and in vivo magnetic resonance imaging (MRI) evaluation of PSMA targeting superparamagnetic iron oxide nanoparticles (SPIONs). PSMA-targeting polypeptide CQKHHNYLC was conjugated to SPIONs to form PSMA-targeting molecular MRI contrast agents. In vitro studies demonstrated specific uptake of polypeptide-SPIONs by PSMA expressing cells. In vivo MRI studies found that MRI signals in PSMA-expressing tumors could be specifically enhanced with polypeptide-SPION, and further Prussian blue staining showed heterogeneous deposition of SPIONs in the tumor tissues. Taken altogether, we have developed PSMA-targeting polypeptide-SPIONs that could specifically enhance MRI signal in tumor-bearing mice, which might provide a new strategy for the molecular imaging of PCa.

Prostate cancer detection with real-time elastography using a bi-plane transducer: comparison with step section radical prostatectomy pathology.

PURPOSE: To evaluate elastography using a bi-plane transducer for localizing prostate cancer (PCa) in patients scheduled for radical prostatectomy (RP), in comparison with step section pathological analysis. METHODS: Fifty-six consecutive PCa patients underwent real-time elastography examination with a bi-plane transducer before RP. Transverse elastographic images were obtained from the apex to the base by slightly compressing and releasing the prostate tissue using the probe. The diagnostic performance of elastography was evaluated in correlation with step section RP histopathology. RESULTS: In 56 PCa patients, gray-scale ultrasonography detected at least one lesion in 36 patients, whereas elastography detected at least one lesion in 53 patients (P = 0.001). The overall sensitivity, specificity and accuracy of elastography in depicting tumor lesions were 67.6, 89.5 and 82.7 %, respectively. The detection rate of a PCa lesion with elastography was best in the left posterior region, followed by the right posterior region. Elastography was more sensitive in detecting PCa lesions with higher Gleason scores, diameter >5 mm and extracapsular extension. CONCLUSIONS: The additional use of elastography with the bi-plane transducer can improve PCa detection rate by providing more information about tissue stiffness within the prostate gland.

Correlation between three-dimensional ultrasound features and pathological prognostic factors in breast cancer.

OBJECTIVES: To investigate the correlation of three-dimensional (3D) ultrasound features with prognostic factors in invasive ductal carcinoma. METHODS: Surgical resection specimens of 85 invasive ductal carcinomas of 85 women who had undergone 3D ultrasound were included. Morphology features and vascularization perfusion on 3D ultrasound were evaluated. Pathologic prognostic factors, including tumour size, histological grade, lymph node status, oestrogen and progesterone receptor status (ER, PR), c-erbB-2 and p53 expression, and microvessel density (MVD) were determined. Correlations of 3D ultrasound features and prognostic factors were analysed. RESULTS: The retraction pattern in the coronal plane had a significant value as an independent predictor of a small tumour size (P = 0.014), a lower histological grade (P = 0.009) and positive ER or PR expression status (P = 0.001, 0.044). The retraction pattern with a hyperechoic ring only existed in low-grade and ER-positive tumours. The presence of the hyperechoic ring strengthened the ability of the retraction pattern to predict a good prognosis of breast cancer. The increased intra-tumour vascularization index (VI, the mean tumour vascularity) reflected a higher histological grade (P = 0.025) and had a positive correlation with MVD (r = 0.530, P = 0.001). CONCLUSIONS: The retraction pattern and histogram indices of VI provided by 3D ultrasound may be useful in predicting prognostic information about breast cancer. KEY POINTS: Three-dimensional ultrasound can potentially provide prognostic evaluation of breast cancer. The retraction pattern and hyperechoic ring in the coronal plane suggest good prognosis. The increased intra-tumour vascularization index reflects a higher histological grade. The intra-tumour vascularization index is positively correlated with microvessel density.

DF-GAM: Cross-Domain Ultrasound Image High-Quality Reconstruction Using a Dual Frequency-Domain Guided Adaptation Model.

OBJECTIVE: To enhance the quality of low-resolution (LR) ultrasound images and mitigate artifacts and speckle noise, which can impede accurate medical diagnosis, a novel method called the dual frequency-domain guided adaptation model (DF-GAM) is proposed. The method aims to achieve high-quality image reconstruction across diverse domains, including different ultrasound machines, diseases and phantom images. METHODS: DF-GAM utilizes a dual-branch network architecture combined with frequency-domain self-adaptation and self-supervised edge regression. This approach enables cross-domain enhancement by focusing on the reconstruction of clear tissue structures and speckle patterns. The model is designed to adapt to various ultrasound imaging (USI) scenarios, ensuring its applicability in real-world clinical settings. RESULTS: Experimental evaluations of DF-GAM were conducted using five different datasets. The results demonstrated the method's effectiveness, with DF-GAM outperforming existing enhancement techniques. The average peak signal-to-noise ratio (PSNR) achieved was 34.62, and the structural similarity index (SSIM) was 0.91, indicating a significant improvement in image quality compared to other methods. CONCLUSION: DF-GAM shows great potential in improving medical image diagnosis and interpretation. Its ability to enhance LR ultrasound images across various domains without the need for extensive training data makes it a valuable tool for clinical use. The high PSNR and SSIM scores validate the method's effectiveness, suggesting that DF-GAM could significantly contribute to the field of USI diagnostics.

B-mode ultrasound to elastography synthesis using multiscale learning.

Elastography is a promising diagnostic tool that measures the hardness of tissues, and it has been used in clinics for detecting lesion progress, such as benign and malignant tumors. However, due to the high cost of examination and limited availability of elastic ultrasound devices, elastography is not widely used in primary medical facilities in rural areas. To address this issue, a deep learning approach called the multiscale elastic image synthesis network (MEIS-Net) was proposed, which utilized the multiscale learning to synthesize elastic images from ultrasound data instead of traditional ultrasound elastography in virtue of elastic deformation. The method integrates multi-scale features of the prostate in an innovative way and enhances the elastic synthesis effect through a fusion module. The module obtains B-mode ultrasound and elastography feature maps, which are used to generate local and global elastic ultrasound images through their correspondence. Finally, the two-channel images are synthesized into output elastic images. To evaluate the approach, quantitative assessments and diagnostic tests were conducted, comparing the results of MEIS-Net with several deep learning-based methods. The experiments showed that MEIS-Net was effective in synthesizing elastic images from B-mode ultrasound data acquired from two different devices, with a structural similarity index of 0.74 ± 0.04. This outperformed other methods such as Pix2Pix (0.69 ± 0.09), CycleGAN (0.11 ± 0.27), and StarGANv2 (0.02 ± 0.01). Furthermore, the diagnostic tests demonstrated that the classification performance of the synthetic elastic image was comparable to that of real elastic images, with only a 3 % decrease in the area under the curve (AUC), indicating the clinical effectiveness of the proposed method.

A novel hACE2 knock-in mouse model recapitulates pulmonary and intestinal SARS-CoV-2 infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission is responsible for the coronavirus disease 2019 (COVID-19) pandemic. SARS-CoV-2 uses the angiotensin-converting enzyme 2 (ACE2) receptor to enter the host, and the gastrointestinal tract is a potential infection site as this receptor is expressed on it. Multiple studies have indicated that an increasing number of COVID-19 patients presented with gastrointestinal symptoms that are highly associated with disease severity. Moreover, emerging evidence has demonstrated that alterations in the gut immune microenvironment induced by intestinal SARS-CoV-2 infection can regulate respiratory symptoms. Therefore, targeting the intestines may be a candidate therapeutic strategy in patients with COVID-19; however, no mouse model can serve as an appropriate infection model for the development of fatal pneumonia while mimicking intestinal infection. In this study, a novel human ACE2 knock-in (KI) mouse model (or hACE2-KI) was systemically compared with the popular K18-hACE2 mice; it showed differences in the distribution of lung and intestinal infections and pathophysiological characteristics. These newly generated hACE2-KI mice were susceptible to intranasal infection with SARS-CoV-2, and not only developed mild to severe lung injury, but also acquired intestinal infection. Consequently, this model can be a useful tool for studying intestinal SARS-CoV-2 infection and developing effective therapeutic strategies.

The CH24H metabolite, 24HC, blocks viral entry by disrupting intracellular cholesterol homeostasis.

Cholesterol-24-hydroxylase (CH24H or Cyp46a1) is a reticulum-associated membrane protein that plays an irreplaceable role in cholesterol metabolism in the brain and has been well-studied in several neuro-associated diseases in recent years. In the present study, we found that CH24H expression can be induced by several neuroinvasive viruses, including vesicular stomatitis virus (VSV), rabies virus (RABV), Semliki Forest virus (SFV) and murine hepatitis virus (MHV). The CH24H metabolite, 24-hydroxycholesterol (24HC), also shows competence in inhibiting the replication of multiple viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). 24HC can increase the cholesterol concentration in multivesicular body (MVB)/late endosome (LE) by disrupting the interaction between OSBP and VAPA, resulting in viral particles being trapped in MVB/LE, ultimately compromising VSV and RABV entry into host cells. These findings provide the first evidence that brain cholesterol oxidation products may play a critical role in viral infection.

Development of Highly Potent Noncovalent Inhibitors of SARS-CoV-2 3CLpro.

The 3C-like protease (3CLpro) is an essential enzyme for the replication of SARS-CoV-2 and other coronaviruses and thus is a target for coronavirus drug discovery. Nearly all inhibitors of coronavirus 3CLpro reported so far are covalent inhibitors. Here, we report the development of specific, noncovalent inhibitors of 3CLpro. The most potent one, WU-04, effectively blocks SARS-CoV-2 replications in human cells with EC50 values in the 10-nM range. WU-04 also inhibits the 3CLpro of SARS-CoV and MERS-CoV with high potency, indicating that it is a pan-inhibitor of coronavirus 3CLpro. WU-04 showed anti-SARS-CoV-2 activity similar to that of PF-07321332 (Nirmatrelvir) in K18-hACE2 mice when the same dose was administered orally. Thus, WU-04 is a promising drug candidate for coronavirus treatment.

CDCP1-targeted nanoparticles encapsulating phase-shift perfluorohexan for molecular US imaging in vitro.

BACKGROUND: Molecular targeted contrast-enhanced ultrasound (CEUS) imaging is a potential imaging strategy to improve the diagnostic accuracy of conventional ultrasound (US) imaging. US contrast agents are usually micrometer-sized and non-target gas bubbles while nano-sized and targeted agents containing phase-shift materials absorb more attractions for their size and the liquid core and excellent molecular imaging effect. METHODS: PLGA12k-mPEG2k-NH2, DSPE-mPEG2k and perfluorohexan (PFH) were used to construct a new targeted ultrasound contrast agent with CUB domain-containing protein 1 (CDCP1) receptor for the detection and diagnosis of prostate cancer. The potential of tumor-targeted nanoparticles (CDCP1-targeted perfluorohexan-loaded phase-transitional nanoparticles, anti-CDCP1 NPs) as contrast agents for ultrasound (US) imaging was assessed in vitro. Moreover, studies on the cytotoxicity and the targeting ability of anti-CDCP1 NPs assisted by US were carried out. RESULTS: The results showed that anti-CDCP1 NPs had low cytotoxicity, and with the increasing of polymer concentration in anti-CDCP1 NPs, the CEUS imaging of agent gradually enhanced, and enhanced imaging associated with the length of observing time. Furthermore, it was testified that anti-CDCP1 assisted the agent to target cells expressing CDCP1, which demonstrated the active targeting of anti-CDCP1 NPs in vitro. CONCLUSION: All in all, the feasibility of using targeted anti-CDCP1 NPs to enhance ultrasound imaging has been demonstrated in vitro, which laid a solid foundation for molecular US imaging in vivo, and anti-CDCP1 NPs might have a great clinical application prospect.