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Accurate Diagnosis of SARS-CoV-2 JN.1 by Sanger Sequencing of Receptor-Binding Domain Is Needed for Clinical Evaluation of Its Immune Evasion

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Journal of Biosciences and Medicines 2024年第4期12卷 69-78页

作者： sin hang lee Milford Molecular Diagnostics Laboratory Milford CT USA

Background: Omicron JN.1 has become the dominant SARS-CoV-2 variant in recent months. JN.1 has the highest number of amino acid mutations in its receptor binding domain (RBD) and has acquired a hallmark L455S mutation. The immune evasion capability of JN.1 is a subject of scientific investigation. The US CDC used SGTF of TaqPath COVID-19 Combo Kit RT-qPCR as proxy indicator of JN.1 infections for evaluation of the effectiveness of updated monovalent XBB.1.5 COVID-19 vaccines against JN.1 and recommended that all persons aged ≥ 6 months should receive an updated COVID-19 vaccine dose. Objective: Recommend Sanger sequencing instead of proxy indicator to diagnose JN.1 infections to generate the data based on which guidelines are made to direct vaccination policies. Methods: The RNA in nasopharyngeal swab specimens from patients with clinical respiratory infection was subjected to nested RT-PCR, targeting a 398-base segment of the N-gene and a 445-base segment of the RBD of SARS-CoV-2 for amplification. The nested PCR amplicons were sequenced. The DNA sequences were analyzed for amino acid mutations. Results: The N-gene sequence showed R203K, G204R and Q229K, the 3 mutations associated with Omicron BA.2.86 (+JN.1). The RBD sequence showed 24 of the 26 known amino acid mutations, including the hallmark L455S mutation for JN.1 and the V483del for BA.2.86 lineage. Conclusions: Sanger sequencing of a 445-base segment of the SARS-CoV-2 RBD is useful for accurate determination of emerging variants. The CDC may consider using Sanger sequencing of the RBD to diagnose JN.1 infections for statistical analysis in making vaccination policies.

关键词： Omicron JN.1 SARS-CoV-2 Sanger Sequencing RBD L455S Mutation Immune Evasion Vaccination Policies CDC

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Sanger Sequencing for Molecular Diagnosis of SARS-CoV-2 Omicron Subvariants and Its Challenges

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Journal of Biosciences and Medicines 2022年第9期10卷 182-223页

作者： sin hang lee Milford Molecular Diagnostics Laboratory Milford USA

Large population passages of the SARS-CoV-2 in the past two and a half years have allowed the circulating virus to accumulate an increasing number of mutations in its genome. The most recently emerging Omicron subvariants have the highest number of mutations in the Spike (S) protein gene and these mutations mainly occur in the receptor-binding domain (RBD) and the N-terminal domain (NTD) of the S gene. The European Centre for Disease Prevention and Control (eCDC) and the World Health Organization (WHO) recommend partial Sanger sequencing of the SARS-CoV-2 S gene RBD and NTD on the polymerase chain reaction (PCR)-positive samples in diagnostic laboratories as a practical means of determining the variants of concern to monitor possible increased transmissibility, increased virulence, or reduced effectiveness of vaccines against them. The author’s diagnostic laboratory has implemented the eCDC/WHO recommendation by sequencing a 398-base segment of the N gene for the definitive detection of SARS-CoV-2 in clinical samples, and sequencing a 445-base segment of the RBD and a 490 - 509-base segment of the NTD for variant determination. This paper presents 5 selective cases to illustrate the challenges of using Sanger sequencing to diagnose Omicron subvariants when the samples harbor a high level of co-existing minor subvariant sequences with multi-allelic single nucleotide polymorphisms (SNPs) or possible recombinant Omicron subvariants containing a BA.2 RBD and an atypical BA.1 NTD, which can only be detected by using specially designed PCR primers. In addition, Sanger sequencing may reveal unclassified subvariants, such as BA.4/BA.5 with L84I mutation in the S gene NTD. The current large-scale surveillance programs using next-generation sequencing (NGS) do not face similar problems because NGS focuses on deriving consensus sequence.

关键词： eCDC WHO Sanger Sequencing Omicron Variant Minor Subvariants BA.4/BA.5 L84I BA.2 Multi-Allelic Recombinant

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Detection of human papillomavirus L1 gene DNA fragments in postmortem blood and spleen after Gardasil^(■) vaccination—A case report

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Advances in Bioscience and Biotechnology 2012年第8期3卷 1214-1224页

作者： sin hang lee Milford Hospital and Milford Molecular Laboratory MilfordConnecticutUSA

A same-nested PCR was used to re-amplify the amplicon of a hypervariable region of the HPV-16 L1 gene DNA in the postmortem blood and splenic tissue obtained at autopsy of a formerly healthy teenage girl who suffered a sudden unexpected death in sleep 6 months after 3 intramuscular injections of a quadrivalent HPV vaccine, Gardasil?. A full autopsy analysis revealed no cause of death. The HPV-16 gene DNA detected in the postmortem materials was similar to the HPV-16 gene DNA fragments in Gardasil? in that both were in non-B-conformation, requiring nondegenerate GP6 and MY11 primers to re-amplify the PCR amplicon for detection and to generate a template useful for direct DNA sequencing. A sequence excised from the base-calling DNA sequencing electropherogram was analyzed by Basic Local Alignment Search Tool (BLAST) alignment and a 45 - 60 base sequence fully matched with a standard hypervariable region of the HPV-16 L1 gene retrieved from the National Center for Biotechnology Information database validated the correct genotyping for HPV- 16 L1 gene DNA. These naked non-proliferating HPV- 16 L1 gene DNA fragments appeared to be in the macrophages of the postmortem blood and spleen, and were protected from degradation by binding firmly to the particulate aluminum adjuvant used in vaccine formulation. The significance of these HPV DNA fragments of a vaccine origin found in post-mortem materials is not clear and warrants further investigation.

关键词： Gardasil^(■) Postmortem HPV DNA Aluminum Vaccine

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Topological conformational changes of human papillomavirus(HPV)DNA bound to an insoluble aluminum salt—A study by low temperature PCR

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Advances in Biological Chemistry 2013年第1期3卷 76-85页

作者： sin hang lee Milford Hospital and Milford Molecular Laboratory MilfordConnecticutUSA

A low temperature (LoTemp?) polymerase chain reaction (PCR), conducted at cycling temperatures not to exceed 85℃and catalyzed by a novel highly processive HiFi? DNA polymerase with proofreading function, was used to study the topological conformational changes of the human papillomavirus (HPV) L1 gene DNA fragments bound to the insoluble amorphous aluminum hydroxyphosphate sulfate (AAHS) adjuvant in the quadrivalent HPV vaccine, Gardasil?. L1 gene DNA fragments of HPV-11, HPV-18 and HPV-16 were detected in the AAHS particles by nested PCR, but all were lacking a region that was amplifiable by an MY09 degenerate primer. In addition, a pair of degenerate consensus GP6/MY11 primers was able to amplify a target segment of the HPV-11 L1 gene DNA and the HPV-18 L1 gene DNA bound to the AAHS particles as expected for any HPV DNA in the B-conformation. However, there was no co-amplification of the HPV-16 L1 gene DNA known to coexist in the same samples. The lack of co-amplification was verified by direct DNA sequencing of the PCR amplicons. The companion HPV-16 L1 gene DNA in the same sample required repeated PCRs with a pair of modified non-degenerate GP6/ MY11 primers for detection. This melting profile of the HPV-16 L1 gene DNA was similar to that of the HPV-16 L1 gene DNA recently discovered in the postmortem blood of a young woman who suffered a sudden unexpected death 6 months after Gardasil? vaccination. The findings suggest that the topological conformational changes in the HPV L1 gene DNA residues bound to the AAHS adjuvant may be genotype-related. The special non-B-conformation may prevent the HPV-16 L1 gene DNA from being degraded in the body of the vaccine recipients after in- tramuscular injection.

关键词： Low Temperature PCR HPV DNA Conformation Gardasil^■ Postmortem AAHS Adjuvant Aluminum

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Small intestine angioleiomyoma as a rare cause of perforation:A case report

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World Journal of Clinical Cases 2024年第12期12卷 2116-2121页

作者： Teng-Yuan Hou Wei-Juo Tzeng Pei-hang lee Department of General Surgery Kaohsiung Chang Gung Memorial HospitalKaohsiung 833Taiwan Department of Anatomical Pathology Kaohsiung Chang Gung Memorial HospitalKaohsiung 833Taiwan

BACKGROUND Angioleiomyoma is a rare and benign stromal tumor typically found in subcutaneous *** rarely occurs in the gastrointestinal *** the reported cases,the most common complication was gastrointestinal *** has only been reported as a complication in the last few *** SUMMARY This case report detailed the discovery of intestinal angioleiomyoma in a 47-yearold male presenting with abdominal pain that had persisted for 3 *** suspecting hollow organ perforation,surgical intervention involving intestinal resection and anastomosis was *** The report underscores the significance of early surgical intervention in effectively treating angioleiomyoma while emphasizing the pivotal role of timely and appropriate measures for favorable outcomes.

关键词： Angioleiomyoma Intestinal perforation Abdomen Acute Diarrhea Case report

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Inferior vena cava reconstruction in extended right hepatectomy

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Hepatobiliary & Pancreatic Diseases International 2024年第5期23卷 533-534页

作者： Karin KY Ho Sui Ling sin Kin Pan Au Henry HY lee Daniel TL Chan Albert Chan Department of Surgery The University of Hong Kong102 Pok Fu Lam RoadHong KongChina Department of Surgery Yan Chai Hospital7 Yan Chai StreetHong KongChina State Key Laboratory of Liver Research(Li Ka Shing Faculty of Medicine The University of Hong Kong)102 Pok Fu Lam RoadHong KongChina

To the Editor : Barcelona Clinic Liver Cancer(BCLC) Stage C hepatocellular carcinoma(HCC) with vascular invasion has been considered unresectable. However, the curative rate from systemic treatment is rather dismal. Even with the use of immunotherapy, complete remission is less than 10% [ 1 ]. On the contrary, recent advances in transplant oncology, especially in the application of liver transplant-related surgical techniques in complex liver resections, make the operation possible for patients with locally advanced HCC who were previously considered to be unresectable [ 2 ]. We hereby present the surgical management of a patient with giant HCC in right liver lobe with inferior vena cava(IVC) invasion.

关键词： invasion vena Liver

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Highly efficient and stable organic solar cells with SnO_(2)electron transport layer enabled by UV-curing acrylate oligomers

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Journal of Energy Chemistry 2024年第5期92卷 124-131页

作者： Mwende Mbilo Du Hyeon Ryu Seungjin lee Muhammad Haris Julius Mwakondo Mwabora Robinson Juma Musembi hang Ken lee Sang Kyu lee Chang Eun Song Won Suk Shin Department of Physics Faculty of Science and TechnologyUniversity of Nairobi(UoN)P.O.Box 30197-00100NairobiKenya Advanced Energy Materials Research Center Korea Research Institute of Chemical Technology(KRICT)Daejeon 34114Republic of Korea Advanced Materials and Chemical Engineering University of Science and Technology(UST)Daejeon 34113Republic of Korea

The interfaces between the inorganic metal oxide and organic photoactive layer are of outmost importance for efficiency and stability in organic solar cells(OSCs).Tin oxide(SnO_(2))is one of the promising candidates for the electron transport layer(ETL)in high-performance inverted *** a solution-processed SnO_(2)ETL is employed,however,the presence of interfacial defects and suboptimal interfacial contact can lower the power conversion efficiency(PCE)and operational stability of ***,highly efficient and stable inverted OSCs by modification of the SnO_(2)surface with ultraviolet(UV)-curable acrylate oligomers(SAR and OCS)are *** highest PCEs of 16.6%and 17.0%are achieved in PM6:Y6-BO OSCs with the SAR and OCS,respectively,outperforming a device with a bare SnO_(2)ETL(PCE 13.8%).The remarkable enhancement of PCEs is attributed to the optimized interfacial contact,leading to mitigated surface *** strikingly,improved light-soaking and thermal stability strongly correlated with the interfacial defects are demonstrated for OSCs based on SnO_(2)/UV cross-linked resins compared to OSCs utilizing bare SnO_(2).We believe that UV cross-linking oligomers will play a key role as interfacial modifiers in the future fabrication of large-area and flexible OSCs with high efficiency and stability.

关键词： Organic solar cells SnO_(2) Surface defects Ultraviolet resins Stability Cross-linking oligomers Non-halogenated solvent

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Development and application of photocatalytic coating for roadside NO_(x) mitigation in Hong Kong

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Chinese Chemical Letters 2024年第3期35卷 153-157页

作者： Xinwei Li Pengge Wang Shuwen Han Yu Huang Wingkei Ho Steven Sai hang Ho Shun-cheng lee Meng Wang Department of Civil and Environmental Engineering The Hong Kong Polytechnic UniversityHong KongChina State Key Laboratory of Loess and Quaternary Geology(SKLLQG)and Key Laboratory of Aerosol Chemistry and Physics Institute of Earth EnvironmentChinese Academy of SciencesXi'an 710061China Department of Science and Environmental Studies The Education University of Hong KongHong KongChina Division of Atmospheric Sciences Desert Research InstituteRenoNV 89512United States

A facile chemical method for the development of photocatalytic coating products was proposed based on practical application perspective for the Hong Kong roadside nitrogen oxides(NO_(x))***_(2)-based photocatalytic coating PC-C film with crystallized size of around 5–6 nm was synthesized with the peptization of H_(2)O_(2).The PC-C coating possesses a super-hydrophilicity surface and is proven to have a NO_(x)degradation rate of 46.8%with an optimum pH level of *** addition,the PC-C coating presents a promising photocatalytic NO_(x)degradation compared with other commercially available coating products and P25 when applied on two building materials of poly-methyl methacrylate(PMMA)and concrete surface.A weather resistance simulation and a 180-day on-site field trial were carried out the attenuation effects of photocatalytic coating applied in outdoor *** on epidemiological estimation and field investigation,hospital admissions for respiratory diseases(HARD)and mortality cases(MC)could be reduced with the application of PC-C coating along the street *** work demonstrates the feasibility of air pollution control measures for the local roadside NO_(x)using photocatalytic technology,offering promising health benefits with environmental remediation.

关键词： Photocatalysis Coating NO_(x) mitigation Hong Kong Roadside

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Efficacy of fexuprazan compared with rebamipide in Korean patients with gastritis:A matching-adjusted indirect comparison

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World Journal of Clinical Cases 2024年第19期12卷 3890-3897页

作者： Gwang Ha Kim hang Lak lee Department of Internal Medicine Pusan National University School of Medicine and Biomedical Research InstitutePusan National University HospitalBusan 49241South Korea Department of Internal Medicine Hanyang University College of MedicineSeoul 04763South Korea

BACKGROUND Gastritis is one of the most frequently diagnosed diseases requiring medical treatment in South ***,a novel potassium-competitive acid blocker,has been approved for treating gastritis and erosive ***,rebamipide is the most commonly used mucoprotective agent for acute and chronic gastritis in real-world settings in South ***,there have been no studies comparing the efficacy of these two drugs *** To compare the efficacy of fexuprazan with that of rebamipide for acute and chronic *** This was a matching-adjusted indirect *** patient data from a phase III study of fexuprazan(10 mg BID)were compared with cumulative data from two matching studies of rebamipide(100 mg TID).Erosion improvement and healing rates were compared between two weeks of fexurapan,two weeks of rebamipide,and four weeks of *** two main outcome variables were presented as percentages,and the risk differences(RD)and 95%confidence intervals(CI)were calculated for the relative treatment *** In the primary analysis,the erosion improvement and healing rates after a twoweek treatment with fexuprazan were 64.5%and 53.2%,respectively,while a twoweek treatment with rebamipide resulted in erosion improvement and healing rates of 43.6%(RD:21.0%;95%CI:9.6-32.3;P<0.01)and 35.6%(RD:17.6%;95%CI:6.1-29.2;P=0.003),*** the additional analysis,the erosion improvement and healing rates for the two-week fexuprazan treatment(64.2%and 51.2%,respectively)were similar to those obtained during a four-week treatment with rebamipide(60.6%;RD:3.6%;95%CI:-9.8,17.0;P=0.600 and 53.5%;RD:-2.3%;95%CI:-16.1,11.5;P=0.744,respectively).CONCLUSION The two-week fexuprazan treatment was superior to the two-week rebamipide treatment and similar to the fourweek rebamipide treatment for patients with gastritis.

关键词： Gastritis Erosive gastritis Fexuprazan Rebamipide Matching-adjusted indirect comparison Indirect treatment comparison

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Immunoregulatory and Anti-cancer Activities of Combination Treatment of Novel Four-Herb Formula and Doxorubicin in 4T1-Breast Cancer Bearing Mice

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Chinese Journal of Integrative Medicine 2024年第4期30卷 311-321页

作者： Lea Ling-Yu Kan Ben Chung-Lap Chan Grace Gar-lee Yue LI Pei-ting Sharon Sze-Man Hon HUANG Dan-qi Miranda sin-Man Tsang Clara Bik-San Lau Ping-Chung Leung Chun-Kwok Wong Institute of Chinese Medicine The Chinese University of Hong KongHong Kong SARChina State Key Laboratory of Research on Bioactivities and Clinical Applications of Medicinal Plants The Chinese University of Hong KongHong KongChina Department of Chemical Pathology The Chinese University of Hong KongHong Kong SARChina China-Australia International Research Centre for Chinese Medicine School of Health and Biomedical SciencesSTEM CollegeRMIT UniversityBundooraVictoriaAustralia Li Dak Sum Yip Yio Chin R&D Centre for Chinese Medicine The Chinese University of Hong Kong SARHong KongChina

Objective To investigate the in vivo immunomodulatory and anti-tumor mechanisms of the combined treatment of novel Four-Herb formula(4HF)and doxorubicin in triple-negative breast cancer(TNBC).Methods Murine-derived triple-negative mammary carcinoma cell line,4T1 cells,was cultured and inoculated into mouse mammary ***-six mice were randomly assigned into 6 groups(n=11 in ench):naive,control,LD 4HF(low dose 4HF),HD 4HF(high dose 4HF),LD 4HF+D(low dose and doxorubicin),and D(doxorubicin).Apart from the naive group,each mouse received subcutaneous inoculation with 5×10^(5)4T1 cells resuspended in 100µL of normal saline in the mammary fat *** from the day of tumor cell inoculation,tumors were grown for 6 *** LD and HD groups received daily oral gavage of 658 and 2,630 mg/kg 4HF,*** LD 4HF+D group received daily oral gavage of 658 mg/kg 4HF and weekly intraperitoneal injection of doxorubicin(5 mg/kg).The D group received weekly intraperitoneal injections of doxorubicin(5 mg/kg).The treatment naive mice received daily oral gavage of 0.2 mL double distilled water and 0.1 mL normal saline via intraperitoneal injection once a *** control group received daily oral gavage of 0.2 mL double-distilled *** treatment period was 30 *** the end of treatment,mice organs were harvested to analyze immunological activities via immunophenotyping,gene and multiplex analysis,histological staining,and gut microbiota *** Mice treated with the combination of 4HF and doxorubicin resulted in significantly reduced tumor and spleen burdens(P<0.05),altered the hypoxia and overall immune lymphocyte landscape,and manipulated gut microbiota to favor the anti-tumor immunological ***,immunosuppressive genes,cytokines,and chemokines such as C-C motif chemokine 2 and interleukin-10 of tumors were significantly downregulated(P<0.05).4HF-doxorubicin combination treatment demonstrated synergetic activities and was most effective

关键词： adjunctivetherapy breast cancer cytokines Chinese medicine tumor microenvironment

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