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"SARS-CoV-2 sebagai virus penyebab COVID-19 yang berikatan dengan reseptor ACE-2 untuk masuk ke dalam sel inang melalui protein spike-1. Protein spike-1 dapat menjadi target pencegahan COVID-19 melalui pengembangan vaksin. Vaksin berbasis DNA merupakan kandidat vaksin yang menjanjikan untuk dikembangkan. Spesimen naso-oro faring pasien COVID-19 yang telah dikonfirmasi dengan RT-PCR, diekstraksi dan diamplifikasi dengan menggunakan primer kloning terhadap plasmid pUMVC4a. Hasil sekuensing dianalisis dengan SeqScape 3.0 dan MEGA 11. Analisis epitop sel B dilakukan dengan berbagai piranti lunak berbasis web. Konstruksi DNA vaksin dilakukan melalui analisis in silico menggunakan SnapGene 6.0 serta in vitro melalui teknik DNA Rekombinan. Gen spike-1 teramplifikasi dengan ukuran 2.265 bp, namun ligasi ke pUMVC4a dan transformasi ke E.coli strain DH5α belum berhasil. Berdasarkan analisis, seluruh sekuen memiliki mutasi D614G dengan isolat A dan B memiliki PNI yang dekat dengan varian Wuhan wt sementara 5 isolat (C-G) termasuk dalam varian Omicron. Berdasarkan sifat antigenisitas, toksisitas, alergenisitas, topologi dan hidrofobisitas, empat belas sekuen asam amino (pada posisi 68-678 protein S-1) diajukan sebagai epitop terpilih. Terdapat 14 sekuens asam amino pada protein spike-1 SARS-CoV-2 yang dapat diajukan sebagai domain epitop sel B dalam pengembangan vaksin COVID-19 berbasis DNA.

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SARS-CoV-2 as the virus that causes COVID-19 binds to the ACE-2 receptor to enter host cells via the spike-1 protein. Spike-1 protein can be a target for preventing COVID-19 through vaccine development. DNA-based vaccines are promising vaccine candidates to be developed. Naso-oropharyngeal specimens of COVID-19 patients confirmed by RT-PCR were extracted and amplified using clone primers against the plasmid pUMVC4a. The sequencing results were analyzed with SeqScape 3.0 and MEGA 11. B cell epitope analysis was performed with various web-based software. Vaccine DNA construction was carried out through in silico analysis using SnapGene 6.0 and in vitro using Recombinant DNA techniques. The spike-1 gene was amplified with a size of 2,265 bp, but ligation to pUMVC4a and transformation to E.coli strain DH5α were not successful. Based on the analysis, all sequences have the D614G mutation with isolate A and B having a PNI that is close to the Wuhan wt variant while 5 isolates (C-G) belong to the Omicron variant. Based on antigenicity, toxicity, allergenicity, topology and hydrophobicity, fourteen amino acid sequences (at positions 68 - 678 of protein S-1) were proposed as selected epitopes. There are 14 amino acid sequences in the SARS-CoV-2 spike-1 protein that can be proposed as B cell epitope domains in the development of a DNA-based COVID-19 vaccine."

"SARS-CoV-2 merupakan penyebab COVID-19 yang melanda dunia sejak akhir 2019. Virus ini telah menyebar secara luas di dunia akibat infektifitasnya yang tinggi. Sampai saat ini, telah muncul banyak lineage dengan karakter yang berbeda, dan beberapa diantaranya memiliki infektifitas lebih tinggi dibandingkan lineage lainnya. Perubahan nukleotida akibat mutasi menjadi penyebab munculnya lineage baru Dalam penelitian ini, telah dilakukan analisa untaian gen SARS-CoV-2 yang berasal dari beberapa lineage berbeda, yang dilanjutkan dengan analisa epitop sel B, dan sel T. Setelahnya, dilakukan desain vaksin menggunakan epitop terbaik dan dihubungkan dengan linker yang berupa asam gabungan asam amino, yang kemudian dilanjutkan dengan analisa fisikokimia dan alergenisitas kandidat vaksin. Setelahnya, kandidat vaksin dilanjutkan ke tahap simulasi molecular docking. Berdasarkan hasil yang diperoleh dari penelitian ini, terdapat 2 epitop 9-mer HLA kelas I dan 7 epitop 15-mer HLA kelas II yang dapat digunakan untuk mencakup seluruh untaian yang terpilih. Vaksin kandidat yang terpilih disusun dengan menggunakan linker tertentu dan epitop yang telah diperoleh sebelumnya. Dari simulasi molecular docking yang telah dilakukan dari vaksin kandidat terhadap 3 reseptor antigenik, diperoleh hasil simulasi berupa energi center kompleks sebesar -1161,4 kkal/mol (TLR-3), -1034,1 kkal/mol (HLA-C*14:02), -1064,3 kkal/mol (HLA-DRB1*07:01).

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SARS-CoV-2 is the cause of COVID-19 that has hit the world since late 2019. The virus has spread widely in the world due to its high infectivity. To date, many lineages have emerged with different characters, and some of them have higher infectivity than other lineages. Nucleotide changes due to mutations are the cause of the emergence of new lineages In this study, the SARS-CoV-2 gene strands from several different lineages were analysed, followed by epitope analysis of B cells and T cells. After that, vaccine design was carried out using the best epitopes and connected with a linker in the form of an amino acid combination, followed by physicochemical and allergenicity analysis of vaccine candidates. After that, the vaccine candidate is continued to the molecular docking simulation stage. Based on the results obtained from this study, there are 2 9-mer HLA class I epitopes and 7 15-mer HLA class II epitopes that can be used to cover all selected strands. The selected candidate vaccines were prepared using specific linkers and previously obtained epitopes. From the molecular docking simulation of the candidate vaccines against 3 antigenic receptors, the simulation results showed the centre complex energy of -1161.4 kcal/mol (TLR-3), -1034.1 kcal/mol (HLA-C*14:02), -1064.3 kcal/mol (HLA-DRB1*07:01)."

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Penelitian ini bertujuan untuk melakukan pengelompokan varian virus SARS-CoV-2 melalui proses clustering menggunakan metode unsupervised learning. Data yang digunakan adalah sekuens protein SARS-CoV-2 yang diekstraksi fiturnya menggunakan paket Discere dalam bahasa pemrograman Python. Sebanyak 27 fitur dihasilkan dan diseleksi dengan metode seleksi fitur Least Absolute Shrinkage and Selection Operator (LASSO). Metode Elbow digunakan untuk menentukan jumlah cluster yang optimal. Dalam penelitian ini, digunakan metode clustering K-Means dan Balanced Iterative Reducing and Clustering using Hierarchies (BIRCH). Evaluasi hasil clustering dilakukan menggunakan metrik evaluasi Silhouette Score dan Davies-Bouldin Index, serta memperhatikan waktu runtime untuk setiap simulasi. Hasil evaluasi kemudian dibandingkan untuk melihat perbedaan performa antara kedua metode clustering yang digunakan, serta pengaruh seleksi fitur terhadap performa clustering. Hasil terbaik diperoleh pada simulasi dengan metode clustering BIRCH + LASSO, dengan nilai Silhouette Score 0,74186 untuk jumlah cluster k=4 dan 0,73207 untuk k=5. Nilai Davies-Bouldin Index terbaik juga diperoleh pada simulasi tersebut, yaitu 0,42697 untuk k=4 dan 0,37949 untuk k=5. Waktu runtime terbaik tercatat pada simulasi dengan metode K-Means + LASSO, yaitu 0,21551 detik untuk k=4 dan 0,17539 detik untuk k=5. Dapat disimpulkan bahwa metode BIRCH menghasilkan cluster yang lebih baik berdasarkan metrik evaluasi, namun K-Means memberikan proses clustering yang lebih cepat. Seleksi fitur dengan metode LASSO juga membantu meningkatkan performa clustering.

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This study aims to perform clustering of SARS-CoV-2 virus variants using unsupervised learning methods. The data used consists of SARS-CoV-2 protein sequences whose features are extracted using the Discere package in the Python programming language. A total of 27 features are generated and selected using the Least Absolute Shrinkage and Selection Operator (LASSO) feature selection method. The Elbow method is employed to determine the optimal number of clusters for the clustering process. The clustering methods used in this research are K-Means clustering and Balanced Iterative Reducing and Clustering using Hierarchies (BIRCH). The clustering results are evaluated using the Silhouette Score and Davies-Bouldin Index metrics, while also considering the runtime for each simulation. The evaluation results are then compared to examine the performance differences between the two clustering methods and the impact of feature selection on clustering performance. The best Silhouette Score is obtained in the simulation using the BIRCH + LASSO clustering method, with a value of 0.74186 for k=4 and 0.73207 for k=5. The best Davies-Bouldin Index is also achieved in the same simulation, with values of 0.42697 for k=4 and 0.37949 for k=5. The fastest runtime is recorded in the simulation using the K-Means + LASSO method, with a time of 0.21551 seconds for k=4 and 0.17539 seconds for k=5. In conclusion, the BIRCH method yields better clustering results based on the evaluation metrics, while K-Means provides faster clustering processes. The LASSO feature selection method also aids in improving clustering performance.

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"[ABSTRAKPendahuluan: Infeksi dengue merupakan salah satu penyakit endemik di daerah tropis dan subtropis yang disebabkan oleh virus dengue (DENV). Hingga saat ini belum ada antiviral yang efektif untuk infeksi dengue. Penyebaran dan sirkulasi serotipe DENV berfariasi di setiap lokasi geografi, hal ini menyulitkan dalam melakukan evaluasi vaksin DENV. Oleh karena itu perlu dikembangkan kandidat vaksin DENV menggunakan strain Indonesia supaya dapat memberikan proteksi maksimal. Pada peneltian ini dikembangkan kandidat vaksin DNA tetravalen DENV berbasis gen prM-E DENV strain Indonesia.Metode: Konstruksi plasmid rekombinan kandidat vaksi dilakukan dengan cara menyisipkan gen prM-E setiap serotipe DENV ke dalam vektor pUMVC4a. Gen prM-E DENV merupakan strain Indonesia, yang diamplifikasi dari serum pasien yang terinfeksi dengan virus ini. Kemampuan plasmid rekombinan mengekspresikan protein prM-E DENV diuji di sel mamalia. Kemampuan kandidat vaksin menginduksi respon imun humoral dievaluasi secara monovalen dan tetravalen di mencit jenis ddY. Titer IgG anti dengue diperiksa menggunakan teknik ELISA, sedangkan titer antibodi netralisasi di tentukan dengan uji FRNT. Proteksi vaksin terhadap mencit yang diimunisasi dievaluasi dengan melakukan uji tantang menggunakan sel K562 yang diinfeksi DENV-2. Viremi virus di tentukan dengan menggunakan teknik foccus assay.Hasil: Konstruksi plasmid rekombinan kandidat vaksin DENV-1 dan DENV-3 sudah berhasil dilakukan. Plasmid dapat mengekspresikan protein prM-E DENV di sel mamalia, namun karakteristik dan kinetik protein masih belum dapat diketahui dengan jelas. Keempat kandidat vaksin DNA yang sedang dikembangkan dapat menginduksi respon imun, baik secara monovalen maupun tetravalen. Imunisasi secara tetravalen dapat memberikan proteksi pada mencit yang diuji tantang dengan sel K562 yang diinfeksi dengan DENV-2.;

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ABSTRACTIntroduction: Dengue infections are caused by dengue viruses (DENV) and are endemic in tropical and subtropical regions. At present, there is no effective antiviral treatment for dengue infection. Distribution and circulation of DENV serotypes varies by geographic location, it is difficult to evaluate DENV vaccine. Therefore, it is necessary to develop a vaccine candidate DENV using Indonesian strains in order to provide maximum protection. However, in this study, we constructed a recombinant plasmid-based prM-E gene from the Indonesia strain as a DENV DNA vaccine candidate.Methode: The recombinant plasmid was prepared by inserting the prM-E gene from each DENV serotypes into the plasmid backbone pUMVC4a. prM-E gene an Indonesia strain, which was amplified from patient sera infected with DENV. The ability of the recombinant plasmid expressing the prM-E DENV protein tested in mammalian cells. The ability of candidate vaccines induce humoral immune responses were evaluated monovalent and tetravalent in ddY mice. IgG titers of anti-dengue examined using ELISA technique, while neutralizing antibody titers determined with FRNT test. Vaccine protection against the immunized mice was evaluated by conducting challenge test using K562 cells infected by DENV-2. Viremia was determined by using the foccus assay.Result: Construction of recombinant plasmid vaccine candidate DENV-1 and DENV-3 was successfully performed. Plasmids can express prM-E DENV proteins in mammalian cells, but the characteristics and kinetics of protein still can not clearly known. Fourth DNA vaccine candidate that is being developed to induce an immune response, either monovalent or tetravalent. Tetravalent immunization may provide protection in mice challenged tested with K562 cells infected with DENV-2.;Introduction: Dengue infections are caused by dengue viruses (DENV) and are endemic in tropical and subtropical regions. At present, there is no effective antiviral treatment for dengue infection. Distribution and circulation of DENV serotypes varies by geographic location, it is difficult to evaluate DENV vaccine. Therefore, it is necessary to develop a vaccine candidate DENV using Indonesian strains in order to provide maximum protection. However, in this study, we constructed a recombinant plasmid-based prM-E gene from the Indonesia strain as a DENV DNA vaccine candidate.Methode: The recombinant plasmid was prepared by inserting the prM-E gene from each DENV serotypes into the plasmid backbone pUMVC4a. prM-E gene an Indonesia strain, which was amplified from patient sera infected with DENV. The ability of the recombinant plasmid expressing the prM-E DENV protein tested in mammalian cells. The ability of candidate vaccines induce humoral immune responses were evaluated monovalent and tetravalent in ddY mice. IgG titers of anti-dengue examined using ELISA technique, while neutralizing antibody titers determined with FRNT test. Vaccine protection against the immunized mice was evaluated by conducting challenge test using K562 cells infected by DENV-2. Viremia was determined by using the foccus assay.Result: Construction of recombinant plasmid vaccine candidate DENV-1 and DENV-3 was successfully performed. Plasmids can express prM-E DENV proteins in mammalian cells, but the characteristics and kinetics of protein still can not clearly known. Fourth DNA vaccine candidate that is being developed to induce an immune response, either monovalent or tetravalent. Tetravalent immunization may provide protection in mice challenged tested with K562 cells infected with DENV-2., Introduction: Dengue infections are caused by dengue viruses (DENV) and are endemic in tropical and subtropical regions. At present, there is no effective antiviral treatment for dengue infection. Distribution and circulation of DENV serotypes varies by geographic location, it is difficult to evaluate DENV vaccine. Therefore, it is necessary to develop a vaccine candidate DENV using Indonesian strains in order to provide maximum protection. However, in this study, we constructed a recombinant plasmid-based prM-E gene from the Indonesia strain as a DENV DNA vaccine candidate.Methode: The recombinant plasmid was prepared by inserting the prM-E gene from each DENV serotypes into the plasmid backbone pUMVC4a. prM-E gene an Indonesia strain, which was amplified from patient sera infected with DENV. The ability of the recombinant plasmid expressing the prM-E DENV protein tested in mammalian cells. The ability of candidate vaccines induce humoral immune responses were evaluated monovalent and tetravalent in ddY mice. IgG titers of anti-dengue examined using ELISA technique, while neutralizing antibody titers determined with FRNT test. Vaccine protection against the immunized mice was evaluated by conducting challenge test using K562 cells infected by DENV-2. Viremia was determined by using the foccus assay.Result: Construction of recombinant plasmid vaccine candidate DENV-1 and DENV-3 was successfully performed. Plasmids can express prM-E DENV proteins in mammalian cells, but the characteristics and kinetics of protein still can not clearly known. Fourth DNA vaccine candidate that is being developed to induce an immune response, either monovalent or tetravalent. Tetravalent immunization may provide protection in mice challenged tested with K562 cells infected with DENV-2.]"

"COVID-19 merupakan penyakit yang disebabkan oleh virus SARS-CoV-2 yang mengakibatkan pandemi global. Jakarta adalah salah satu kota di Indonesia dengan angka kasus dan kematian tertinggi akibat COVID-19. Salah satu cara paling efektif untuk mengurangi keparahan dan resiko penularan COVID-19 adalah dengan vaksinasi. Vaksin dapat merangsang respons imunitas humoral tubuh yang menghasilkan antibodi netralisasi. Selain vaksin, antibodi netralisasi dapat diinduksi secara natural oleh imunitas tubuh. Hybrid immunity merupakan gabungan antara antibodi netralisasi yang diinduksi secara natural dan yang diinduksi oleh vaksin. SARS-CoV-2 terus bermutasi memunculkan berbagai varian yang menyebabkan peningkatan jumlah kasus dan munculnya gelombang COVID-19 baru di Indonesia, yaitu gelombang Delta pada Juni 2021 dan gelombang Omicron pada Januari 2022. Penelitian ini bertujuan untuk mengevaluasi perubahan antibodi netralisasi 3 bulan setelah vaksinasi dosis lengkap dari beberapa jenis vaksin, yaitu vaksin virus inaktivasi (CoronaVac), vaksin viral vektor (ChAdOx1 nCoV-19), dan vaksin mRNA (BNT162b2) serta pengaruh riwayat infeksi SARS-CoV-2 pada penerima vaksin terhadap berbagai varian SARS-CoV-2 (Wuhan, Delta, Omicron B.1.1.529 dan BA.2). Penelitian dilakukan dengan menggunakan uji Surrogate Virus Neutralization Test (sVNT) yang memiliki prinsip kerja seperti enzyme-linked immunosorbent assay (ELISA) dan meniru interaksi antara receptor binding domain (RBD) dan angiotensin-converting enzyme 2 (ACE2) dalam pelat ELISA dengan RBD dan ACE2 yang telah mengalami pemurnian dengan sampel serum partisipan populasi umum (n = 76). Hasil penelitian menunjukkan adanya perbedaan signifikan antara antibodi netralisasi sebelum dan 3 bulan setelah vaksinasi dosis lengkap, tetapi tidak terdapat perbedaan signifikan pada antibodi netralisasi yang dihasilkan dari masing-masing jenis vaksin. Hal tersebut kemungkinan disebabkan oleh waktu pengambilan sampel setelah terjadi gelombang Omicron COVID-19 sehingga terjadi hybrid immunity yang menyebabkan tingginya kadar antibodi netralisasi yang merata pada setiap jenis vaksin. Partisipan dengan riwayat infeksi SARS-CoV-2 memiliki kadar antibodi netralisasi yang lebih tinggi. Terdapat perbedaan antibodi netralisasi yang signifikan terhadap berbagai varian SARS-CoV-2 dengan penurunan kadar antibodi netralisasi yang signifikan terhadap varian Omicron B.1.1.529 dan BA.2. Kesimpulan dari penelitian ini adalah vaksinasi dosis lengkap berhasil meningkatkan kadar antibodi netralisasi hingga 3 bulan pascavaksinasi yang dipengaruhi oleh riwayat infeksi SARS-CoV-2.

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COVID-19 is a disease caused by the SARS-CoV-2 virus which has resulted in a global pandemic. Jakarta is one of the cities in Indonesia with the highest number of COVID-19 cases and deaths. One of the most effective ways to reduce the severity and transmission risk of COVID-19 is by getting vaccinated. Vaccines can stimulate the body's humoral immune response to produce neutralizing antibodies. Apart from vaccines, neutralizing antibodies can be induced naturally by the body's immunity. Hybrid immunity is a combination of naturally induced neutralizing antibodies and those induced by vaccines. The continuously mutating SARS-CoV-2 has led to the emergence of various variants which have resulted in an increase in the number of cases and the emergence of new COVID-19 waves in Indonesia, namely the Delta variant which appeared in June 2021 and the Omicron variant in January 2022. This study aims to evaluate changes in neutralizing antibodies 3 months after complete doses of several types of vaccines, namely inactivated virus vaccine (CoronaVac), viral vector vaccine (ChAdOx1 nCoV-19), and mRNA vaccine (BNT162b2) and the effect of a history of SARS-CoV-2 infection in vaccine recipients against various variants SARS-CoV-2 (Wuhan, Delta, Omicron B.1.1.529 and BA.2). The study was conducted using the Surrogate Virus Neutralization Test (sVNT) test which has a working principle like the enzyme-linked immunosorbent assay (ELISA) and mimics the interaction between the receptor binding domain (RBD) and angiotensin-converting enzyme 2 (ACE2) in ELISA plates using RBD and ACE2 that had undergone purification with sera samples of general population participants (n = 76). The results showed that there were significant differences between the neutralizing antibodies before and 3 months after the full dose of vaccination, but there were no significant differences in the neutralizing antibodies produced from each type of vaccine. This was probably caused by the sampling time after the Omicron COVID-19 wave occurred, resulting in hybrid immunity which resulted in high levels of neutralizing antibodies that were evenly distributed in each type of vaccine. Participants with a history of SARS-CoV-2 infection had higher levels of neutralizing antibodies. There were significant differences in neutralizing antibodies against various variants of SARS-CoV-2 with a significant decrease in levels of neutralizing antibodies against Omicron B.1.1.529 and BA.2 variants. The conclusion of this study is that full dose vaccination has succeeded in increasing neutralizing antibody levels for up to 3 months after vaccination which are affected by a history of SARS-CoV-2 infection."

"Indonesia memiliki kekayaan alam laut yang sangat beragam, yang dapat berpotensi untuk mengandung senyawa bioaktif yang masih kurang dieksplorasi lebih dalam. Dalam penelitian ini, peneliti ingin mengeksplorasi senyawa metabolit sekunder yang berasal dari invertebrata laut yang berada di perairan Indonesia dalam kegunaannya untuk mengatasi masalah pandemi yang terjadi di dunia selama 3 tahun terakhir. Sebanyak 137 senyawa yang diperoleh dari berbagai spesies invertebrata laut seperti karang lunak, tunikata, dan spons ditapiskan dengan menggunakan penapisan virtual terhadap ACE2 dan Spike Protein Virus varian omicron. Selanjutnya setelah didapatkan 20 senyawa yang memiliki potensi dari proses penapisan, dilakukan proses studi penambatan molekuler dan prediksi ADMET untuk menemukan senyawa terbaik dari senyawa yang telah terpilih. Setelah didapatkan senyawa paling baik, dilakukan studi dinamika molekuler untuk melihat kestabilan ligan-reseptor pada senyawa yang terbaik. Dari hasil penelitian didapatkan bahwa ada 2 senyawa yaitu Acanthomanzamine E dan Cortistatin L, yang menunjukkan hasil yang cukup menjanjikan sebagai bloker terhadap ACE2 dan juga SARS-CoV-2 Omicron SPV. Acanthomanzamine E memiliki energi ikatan sebesar -12,87 kcal mol dan -8,61 kcal/mol terhadap ACE2 dan SPV, sementara Cortistatin L memiliki energi ikatan sebesar -9,96 kcal/mol dan -7,56 kcal/mol terhadap ACE2 dan SPV. Simulasi MD sebesar 50ns menunjukkan kestabilan kedua senyawa pada reseptornya masing-masing, dan juga dibandingkan terhadap senyawa pembanding yaitu XX5 untuk pembanding terhadap ACE2 dan Ceftriaxone digunakan sebagai senyawa pembanding terhadap Spike Protein Virus SARS-CoV-2 varian omicron.

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Indonesia has diversified marine creatures, which could potentially have bioactive substances still underexplored. In this research, we tried to explore these secondary metabolites from Indonesian marine invertebrates to find the lead compound in overcoming the pandemic problem that happened in the world for the last 3 years. A total of 137 compounds from different types of invertebrates, such as soft corals, tunicates, and sea sponges screened against ACE2 and Spike Protein Virus of the Omicron variant. From the virtual screening. we obtained top 20 compounds that have potential to bind with the receptors. Furthermore, molecular docking and ADMET prediction studied in the top 20 compounds to filter compound down to the best 2 compounds. We also did preliminary molecular dynamics studies to see the stability of ligand-receptor occur between the best compound and the receptor. We found that two compounds, Acanthomanzamine E and Cortistatin L, show prominent results as ACE2 and SARS-CoV-2 blockers, especially from molecular docking, which have -12,87 kcal/mol and -9,96 kcal/mol towards ACE2 and -8,61 kcal/mol and -7,56 kcal/mol towards SPV Omicron respectively. We see promising results from the 50 ns MD simulation of these compounds and their comparison which is XX5 for ACE2 receptor and Ceftriaxone in Spike Protein Virus SARS-CoV-2 omicron variant."

"Pada akhir tahun 2019, telah muncul suatu tipe virus korona baru yaitu SARS-CoV-2 yang menyebabkan pandemik global dengan tingkat kematian yang relatif sangat tinggi. Dikarenakan oleh belum adanya obat maupun vaksin yang efektif untuk mengobati virus ini maka diperlukan suatu senyawa yang bisa menginhibisi protein yang berperan dalam infeksi virus SARS-CoV-2. Virus SARS-CoV-2 terdiri dari beberapa protein penyusun. Salah satu protein yang berperan penting adalah protein nukleokapsid (NP). Protein ini berperan dalam proses transkripsi maupun replikasi dari RNA virus SARS-CoV-2. Oleh karena itu, protein ini memiliki potensi untuk dijadikan target protein yang akan diinhibisi. Pada penelitian ini akan dilakukan pemanfaatan senyawa bahan alam yang dapat menginhibisi protein target tersebut sehingga dapat mengurangi dampak dari pandemik global ini. Struktur tiga dimensi (3D) dari NP dimodel melalui sekuen yang diunduh melalui basis data GenBank. Struktur protein kemudian dioptimisasi dan dikarakterisasi untuk mengetahui keakurasian struktur hasi homologi model. Kemudian, penapisan dilakukan terhadap basis data senyawa bahan alam yang berasal dari NPASS dengan menggunakan Astex’s Rule of Three (RO3) dan sifat toksisitas untuk mendapatkan senyawa fragmen. Kemudian dilakukan simulasi penambatan molekul senyawa-senyawa fragmen ini terhadap sisi ikat dari NP menggunakan perangkat lunak MOE 2014.09. Kemudian setelah didapatkan delapan ligan terbaik, ligan-ligan tersebut ditumbuhkan dengan menggunakan proses penumbuhan fragmen yang menghasilkan 14.332 senyawa yang nantinya akan dievaluasi melalui simulasi penambatan molekul sekali lagi. Simulasi ini menghasilkan 40 ligan terbaik dengan nilai energi bebas Gibbs terendah terhadap struktur ptorein. Ligan terpilih diprediksi sifat farmakologinya secara komputasi, dan menghasilkan 2 ligan (CFG-17 dan NFG-11) yang memiliki sifat farmakologis yang baik. Kedua ligan ini divalidasi interaksinya dengan menggunakan simulasi dinamika molekul dan menunjukkan stabilitas interaksi yang baik sebagai kandidat obat untuk terapi SARS-CoV-2.

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The outbreak of COVID-19 caused by the SARS-CoV-2 virus caused a global pandemic that affects the worldwide. Because of the lacking effective prescribed drugs or vaccines as a therapeutic strategy, there is a need to identify a novel inhibitor to inhibit a protein that plays a vital role in the infection of SARS-CoV-2. The Nucleocapsid protein (NP) of the SARS-CoV-2 is a protein that is necessary for viral RNA replication and transcription. Thus, in this study, a three-dimensional (3D) NP structure was modeled using the protein sequences of NP obtained from the GenBank database. After that, the modeled protein structure was characterized and optimized. The natural compound databases obtained from the NPASS database were screened based on Astex’s Rule of Three and toxicity filter to gain lead-like fragments. The filtered fragments were docked into the binding site of the NP utilizing MOE 2014.09 software. Then potential eight potential lead-like fragments were grown to generate 14,332 new ligands by utilizing DataWarrior software. Then molecular docking simulation was performed once again with the same protocol as the first molecular docking simulation. The simulation resulted in 40 best ligands with the lowest value of Gibbs free energy binding to NP. The selected ligands were subjected to the computational pharmacological properties prediction using several tools and resulted in two compounds candidate with favorable interaction and ADME-Tox properties. Then these two compounds were further analyzed with the molecular dynamic simulation."

"Pandemi virus SARS-CoV-2 yang terjadi telah membuat kebutuhan untuk pemeriksaan massal deteksi virus menjadi meningkat. Saat ini pemeriksaan gold standard untuk mendeteksi virus SARS-CoV-2 adalah dengan metode reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) yang berbiaya mahal. Oleh karena itu dibutuhkan metode pemeriksaan alternatif yang lebih murah, cepat, dan mudah digunakan sebagai sistem deteksi virus. Metode Loop Mediated Isothermal Amplification LAMP) dapat melakukan reaksi amplifikasi nukleotida pada suhu isothermaltanpa perlu mesin thermal cycler dan dapat diamati langsung dengan penambahan zat pewarna. Metode deteksi virus berbasis LAMP yang dikembangkan menggunakan desain primer yang menarget gen S dan ORF1ab ditambah pewarna Calcein-Mangan dan senyawa tambahan Guanidine Hydrochloride (GuHCl). Sistem deteksi dilakukan optimasi konsentrasi komponen reaksi, suhu dan template yang menggunakan plasmid DNA kontrol. Optimasi didapatkan dengan konsentrasi reaksi FIP/BIP 0,4 µM, F3/B3 0,2 µM, Loop F/P 0,4 µM, Calcein-Mangan 12 µM, dan GuHCl 40 mM. Sistem LAMP yang dikembangkan dapat mendeteksi sekuens gen target pada DNA kontrol hingga 1 copy number dalam waktu sekitar 1 jam pada inkubasi suhu 55 ºC. Meski begitu, sistem LAMP yang dikembangkan belum mapu mengamplifikasi RNA virus SARS-CoV-2 hasil ekstraksi sampel swab pasien. Hal ini disebabkan oleh enzim Bst 3.0 yang dipakai dalam sistem LAMP tidak memiliki kemampuan reverse transcriptase yang diharapkan.

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The SARS-CoV-2 virus pandemic has made the need for mass screening of virus detection increased. Currently, the gold standard  test to detect SARS-CoV-2 virus is reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) method that costly. Therefore, an alternative method that is cheaper, faster, and easier to use as a virus detection system is needed. The Loop Mediated Isothermal Amplification (LAMP) method can perform nucleotide amplification reactions at isothermal temperatures  without the need of thermal cycler  machine and can be observed directly with the addition of coloring agents. The LAMP-based virus detection method development use primers design targeting the S and ORF1ab genes with Calcein-Manganese dyes and the additive compound Guanidine Hydrochloride (GuHCl). The detection system performed optimization of the concentration of reaction components, temperature and template using plasmid DNA control. Optimization was obtained with reaction concentration of FIP/BIP 0.4 μM, F3/B3 0.2 μM, Loop F/P 0.4 μM, Calcein-Manganese 12 μM, and GuHCl 40 mM. The developed LAMP system can detect target gene sequences in control DNA up to  1 copy number in about 1 hour at 55 ºC incubation temperature. Even so, the LAMP system developed still unable to amplify the RNA of the SARS-CoV-2 virus from the extraction of patient swab samples. This issue occurred due to the Bst 3.0 enzyme that used in the reaction did not have reverse transcriptase activity as expected."

"Pandemi yang disebabkan oleh SARS-CoV-2 telah memicu situasi darurat kesehatan di seluruh dunia. Varian Omicron yang menyebar dengan cepat semakin mendesak pencarian terapi yang tepat untuk menghindari infeksi yang lebih berat. TMPRSS2 manusia dan protein spike SARS-CoV-2 varian Omicron diidentifikasi sebagai protein target melalui penapisan secara komputasi. Metode yang digunakan adalah penapisan virtual berbasis struktural; analisis prediksi absorption, distribution, metabolism, excretion, dan toxicity (ADMET); dan simulasi dinamika molekuler. Ligan uji yang digunakan adalah senyawa metabolit sekunder invertebrata laut Indonesia. Camostat dan nafamostat (ko-kristal) digunakan sebagai ligan pembanding terhadap penghambatan TMPRSS2 sedangkan mefloquine ligan pembanding terhadap Protein Spike. Berdasarkan hasil penambatan molekul, acanthomanzamine C (-9,75 kkal/mol) dan cortistatin G (-9,39 kkal/mol) memiliki aktivitas yang lebih baik terhadap penghambatan TMPRSS2 dibandingkan dengan camostat (-8,25 kkal/mol) dan nafamostat (-6,52 kkal/mol). Sebagai inhibitor protein spike SARS-CoV-2 varian Omicron, acanthomanzamine C (-9,19 kkal/mol) dan cortistatin J (-8,89 kkal/mol) juga menunjukkan penghambatan yang lebih baik dibandingkan dengan mefloquine (-6,34 kkal/mol). Ligan uji tersebut juga telah memenuhi seluruh kriteria ADMET yang ditetapkan. Dari hasil analisis simulasi dinamika molekuler menunjukkan pengikatan yang stabil senyawa ligan uji terhadap protein target setelah simulasi berjalan 60 nanodetik dan memiliki energi ikatan bebas MMGBSA dan MMPBSA yang lebih baik dibandingkan ligan pembanding diantaranya TMPRSS2–acanthomanzamine C (-28,2067; -24,6639 kkal/mol), TMPRSS2–cortistatin G (-29,9908; -24,8869 kkal/mol), protein spike–acanthomanzamine C (-45,1414; -27,8749 kkal/mol), dan protein spike–cortistatin J (-37,8537; -35,6439 kkal/mol). Hasil penelitian ini menunjukkan bahwa acanthomanzamine C, cortistatin G, dan cortistatin J merupakan senyawa hits sebagai kandidat terapi untuk infeksi SARS-CoV-2.

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The pandemic caused by SARS-CoV-2 has triggered a global health emergency. The rapid spread of the Omicron variant has further intensified the urgency to search for appropriate therapies to prevent severe infections. The human TMPRSS2 and spike protein of the SARS-CoV-2 Omicron variant were identified as the target proteins through computational screening. The methods used are structure-based virtual screening; absorption, distribution, metabolism, excretion, and toxicity (ADMET) analysis; and molecular dynamics simulation. Bioactive marine invertebrates from Indonesia were employed as test ligands. Camostat and nafamostat (co-crystal) were utilized as reference ligands against TMPRSS2, whereas mefloquine was used as a reference ligand against spike protein. Following a molecular docking, acanthomanzamine C (-9,75 kcal/mol) and cortistatin G (-9,39 kcal/mol) had better activity against TMPRSS2 inhibition compared to camostat (-8,25 kcal/mol) and nafamostat (-6,52 kcal/mol). As inhibitors of spike protein of SARS-CoV-2 Omicron variant, acanthomanzamine C (-9,19 kcal/mol) and cortistatin J (-8,89 kcal/mol) also showed better inhibition compared to mefloquine (-6,34 kcal/mol). The test ligands have also met all the established ADMET criteria. The results of the molecular dynamics analysis showed stable binding of the test ligands to the target proteins after the initial 60 nanoseconds and had free binding energies of MMGBSA/MMPBSA that were better than the comparison ligands, including TMPRSS2–acanthomanzamine C (-28,2067; -24,6639 kcal/mol), TMPRSS2–cortistatin G (-29,9908; -24,8869 kcal/mol), spike protein–acanthomanzamine C (-45,1414; -27,8749 kcal/mol), and spike protein–cortistatin J (-37,8537; -35,6439 kcal/mol).  These results indicate that acanthomanzamine C, cortistatin G, and cortistatin J are hits compounds as candidate therapies for SARS-CoV-2 infection."