Efficient long-term amplification of hepatitis B virus isolates after infection of slow proliferating HepG2-NTCP cells

Kö,nig, Alexander,Yang, Jaewon,Jo, Eunji,Park, Kyu Ho Paul,Kim, Hyun,Than, Thoa Thi,Song, Xiyong,Qi, Xiaoxuan,Dai, Xinghong,Park, Soonju,Shum, David,Ryu, Wang-Shick,Kim, Jung-Hee,Yoon, Seung Kew,P Elsevier 2019 Journal of hepatology Vol.71 No.2

<P><B>Background & Aims</B></P> <P>As hepatitis B virus (HBV) spreads through the infected liver it is simultaneously secreted into the blood. HBV-susceptible <I>in vitro</I> infection models do not efficiently amplify viral progeny or support cell-to-cell spread. We sought to establish a cell culture system for the amplification of infectious HBV from clinical specimens.</P> <P><B>Methods</B></P> <P>An HBV-susceptible sodium-taurocholate cotransporting polypeptide-overexpressing HepG2 cell clone (HepG2-NTCPsec+) producing high titers of infectious progeny was selected. Secreted HBV progeny were characterized by native gel electrophoresis and electron microscopy. Comparative RNA-seq transcriptomics was performed to quantify the expression of host proviral and restriction factors. Viral spread routes were evaluated using HBV entry- or replication inhibitors, visualization of viral cell-to-cell spread in reporter cells, and nearest neighbor infection determination. Amplification kinetics of HBV genotypes B-D were analyzed.</P> <P><B>Results</B></P> <P>Infected HepG2-NTCPsec+ secreted high levels of large HBV surface protein-enveloped infectious HBV progeny with typical appearance under electron microscopy. RNA-seq transcriptomics revealed that HBV does not induce significant gene expression changes in HepG2-NTCPsec+, however, transcription factors favoring HBV amplification were more strongly expressed than in less permissive HepG2-NTCPsec−. Upon inoculation with HBV-containing patient sera, rates of infected cells increased from 10% initially to 70% by viral spread to adjacent cells, and viral progeny and antigens were efficiently secreted. HepG2-NTCPsec+ supported up to 1,300-fold net amplification of HBV genomes depending on the source of virus. Viral spread and amplification were abolished by entry and replication inhibitors; viral rebound was observed after inhibitor discontinuation.</P> <P><B>Conclusions</B></P> <P>The novel HepG2-NTCPsec+ cells efficiently support the complete HBV life cycle, long-term viral spread and amplification of HBV derived from patients or cell culture, resembling relevant features of HBV-infected patients.</P> <P><B>Lay summary</B></P> <P>Currently available laboratory systems are unable to reproduce the dynamics of hepatitis B virus (HBV) spread through the infected liver and release into the blood. We developed a slowly dividing liver-derived cell line which multiplies infectious viral particles upon inoculation with patient- or cell culture-derived HBV. This new infection model can improve therapy by measuring, in advance, the sensitivity of a patient’s HBV strain to specific antiviral drugs.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Cell culture system that mimicks complete HBV life cycle from entry to egress. </LI> <LI> Efficient <I>in vitro</I> infection with crude HBV patient sera. </LI> <LI> Up to 50- and 1,300-fold net amplification of patient- and cell culture-derived input HBV in the supernatant. </LI> <LI> Polyethylene glycol-independent HBV spread to adjacent cells, forming infected cell clusters. </LI> <LI> Evaluation of patient- and cell culture-derived HBV amplification w/wo antivirals over 8 weeks. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

Less Volatile Value-at-Risk Estimation Under a Semi-parametric Approach

David K. Wang,Shih-Feng Huang 한국증권학회 2023 Asia-Pacific Journal of Financial Studies Vol.52 No.3

In this study, we propose a two-step, less-volatile value-at-risk (LVaR) estimation using a generalized nearly isotonic regression (GNIR) model. In the proposed approach, a VaR sequence is first produced under the generalized autoregressive conditional heteroskedasticity (GARCH) framework. Then, the VaR sequence is adjusted by GNIR, and the generated estimate is denoted as LVaR. The results of an empirical investigation show that LVaR outperformed other VaR estimates under the classic equally weighted and exponentially weighted moving-average frameworks. Furthermore, we show not only that LVaR is less volatile, but also that it performed reasonably well in various backtests.

A feature descriptor based on the local patch clustering distribution for illumination-robust image matching

Wang, Han,Yoon, Sang Min,Han, David K.,Ko, Hanseok Elsevier 2017 Pattern recognition letters Vol.94 No.-

<P><B>Abstract</B></P> <P>This paper proposes a feature descriptor based on the local patch clustering distribution (LPCD), which preserves the salient features of a given image following changes in illumination. To mitigate the effects of illumination change, the proposed LPCD methodology consists of two steps. First, a local patch clustering assignment map is constructed by pairing the source image with a reference image. To resolve the quantization problem caused by an illumination change, a dual-codebook clustering method is employed so that an effective local patch clustering feature space can be constructed. Second, in the feature encoding process, the impact of the informative local patches that contain textural information is enhanced when using a saliency detection response as a method of weighting every local patch when the histogram feature is extracted. Experimental results show that the proposed local patch clustering space is more robust than the conventional intensity order-based space in response to changes in illumination.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Local patch clustering distribution is proposed for use in illumination-robust image matching. </LI> <LI> A dual codebook is proposed to generate local patch clustering distributions. </LI> <LI> A saliency detection response increases the effectiveness of the proposed LPCD. </LI> </UL> </P>

Development of A Chimeric Antigen Receptor Targeting C-Type Lectin-Like Molecule-1 for Human Acute Myeloid Leukemia

Laborda, Eduardo,Mazagova, Magdalena,Shao, Sida,Wang, Xinxin,Quirino, Herlinda,Woods, Ashley K.,Hampton, Eric N.,Rodgers, David T.,Kim, Chan Hyuk,Schultz, Peter G.,Young, Travis S. MDPI 2017 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.18 No.11

<P>The treatment of patients with acute myeloid leukemia (AML) with targeted immunotherapy is challenged by the heterogeneity of the disease and a lack of tumor-exclusive antigens. Conventional immunotherapy targets for AML such as CD33 and CD123 have been proposed as targets for chimeric antigen receptor (CAR)-engineered T-cells (CAR-T-cells), a therapy that has been highly successful in the treatment of B-cell leukemia and lymphoma. However, CD33 and CD123 are present on hematopoietic stem cells, and targeting with CAR-T-cells has the potential to elicit long-term myelosuppression. C-type lectin-like molecule-1 (CLL1 or CLEC12A) is a myeloid lineage antigen that is expressed by malignant cells in more than 90% of AML patients. CLL1 is not expressed by healthy Hematopoietic Stem Cells (HSCs), and is therefore a promising target for CAR-T-cell therapy. Here, we describe the development and optimization of an anti-CLL1 CAR-T-cell with potent activity on both AML cell lines and primary patient-derived AML blasts in vitro while sparing healthy HSCs. Furthermore, in a disseminated mouse xenograft model using the CLL1-positive HL60 cell line, these CAR-T-cells completely eradicated tumor, thus supporting CLL1 as a promising target for CAR-T-cells to treat AML while limiting myelosuppressive toxicity.</P>

Radio frequency-mediated local thermotherapy for destruction of pancreatic tumors using Ni–Au core–shell nanowires

Hopkins, Xiaoping,Gill, Waqas Amin,Kringel, Rosemarie,Wang, Guankui,Hass, Jamie,Acharya, Suresh,Park, Jungrae,Jeon, In Tak,An, Boo Hyun,Lee, Ji Sung,Ryu, Jong Eun,Hill, Rod,McIlroy, David,Kim, Young K IOP 2017 Nanotechnology Vol.28 No.3

<P>We present a novel method of radio frequency (RF)-mediated thermotherapy in tumors by remotely heating nickel (Ni)–gold (Au) core–shell nanowires (CSNWs). Ectopic pancreatic tumors were developed in nude mice to evaluate the thermotherapeutic effects on tumor progression. Tumor ablation was produced by RF-mediated thermotherapy via activation of the paramagnetic properties of the Ni–Au CSNWs. Histopathology demonstrated that heat generated by RF irradiation caused significant cellular death with pyknotic nuclei and nuclear fragmentation dispersed throughout the tumors. These preliminary results suggest that thermotherapy ablation induced via RF activation of nanowires provides a potential alternative therapy for cancer treatment.</P>