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Abstrak :
Latar belakang: Pada kanker nasofaring, tingginya angka kegagalan metastasis jauh paska terapi masih menimbulkan masalah. Sehingga, penelitian mengenai penggunaan terapi sistemik novel pada kanker nasofaring seperti imunoterapi perlu dilakukan. Terdapat beberapa biomarker yang dapat diperiksa untuk dapat memprediksi respon dari pemberian imunoterapi, salah satunya adalah microsatellite instability (MSI). Mikrosatelit merupakan area pada DNA yang memiliki banyak pengulangan kodon, sehingga rentan terjadi gangguan coding dan mengakibatkan akumulasi mutasi. Pada keadaan normal, kerusakan ini akan diperbaiki dengan sistem mismatch repair. Namun, jika terdapat gangguan atau mutasi terkait sistem ini, atau yang disebut dengan deficient mismatch repair (dMMR), akan menghasilkan fenotipe MSI. Pada kanker kolorektal dan endometrium. Namun sampai saat ini, hanya terdapat 3 penelitian yang melakukan pemeriksaan status MSI pada kanker nasofaring. Penelitian ini bertujuan untuk melihat gambara status MSI pada pasien kanker nasofaring pada pasien di RSCM, Indonesia. Metode: Penelitian ini merupakan penelitian eksploratif dengan 36 subjek penelitian. Dilakukan pemeriksaan status instabilitas mikrosatelit menggunakan pemeriksaan berbasis polymerase chain reaction (PCR) Idylla MSI. Dilakukan pula pemeriksaan MMR menggunakan pemeriksaan berbasis imunohistokimia (IHK) menggunakan 4 antibodi untuk mendapatkan keselerasan antara pemeriksaan MSI dan MMR pada pasien kanker nasofaring di RSCM. Hasil : Menggunakan pemeriksaan Idylla MSI, ditemukan MSI pada 2 dari 36 pasien (5,6%) dan dMMR menggunakan pemeriksaan IHK pada 3 dari 36 pasien (8,34%). Hasil yang konsisten ditemukan pada 2 metode pemeriksaan sebesar 96,97%. Kesimpulan: Pada kanker nasofaring ditemukan frekuensi MSI yang rendah baik menggunakan pemeriksaan IHK dan Idylla MSI. Ditemukan keselerasan yang tinggi antara pemeriksaan berbasis IHK dan pemeriksaan berbasis PCR Idylla MSI. ......Background: Despite high probability of local control after treatment, high rate of distant metastases-failure still pose as problem in the management of locally advanced nasopharyngeal carcinoma. Thus, research for novel systemic therapies for nasopharyngeal cancer, such as immunotherapy, needs to be done. There are several biomarkers that may predict the response to immunotherapy, one of which is microsatellite instability (MSI) phenotype. Microsatellites defined by areas in DNA that are prone to mutations due to repetition of 1-3 nitrogen base. However, under normal circumstances, there are repair systems that can identify and correctly repairs DNA mutations in microsatellite area, a system called mismatch repair (MMR) system. Microsatellite instability is a condition of accumulating mutations in microsatellite area due to defect in MMR system. In colorectal and endometrial cancer, MSI are known as one of prognostic and predictive markers, especially with the usage of immunotherapy immune checkpoint blockade PD-1/PD-L1. To this date, only 3 studies are available in exploring the role of MSI in nasopharyngeal cancer, and no study was done in Indonesia. We conduct this study to assess the MSI status of Indonesia's nasopharyngeal cancer patients in Ciptomangunkusumo Hospital. Methods: This is the first explorative study in exploring the role of MSI in Indonesia's nasopharyngeal cancer patients. A total of 36 subjects were recruited, and both MSI assessment using immunohistochemistry (IHC) and polymerase chain reaction (PCR) Idylla MSI was done on all study subjects. Results: MSI was found in 2 patients (5,6%) using PCR based Idylla MSI, and dMMR was found in 3 patients (8,34%). Consistent results between IHC and PCR based MSI assessment was found in 32 patients (96,97%). Conclusion: MSI was a rare event in Indonesia's nasopharyngeal cancer patients. High concordance was found between IHC and PCR MSI assessment in nasopharyngeal cancer.

Abstrak :
Metilasi DNA merupakan salah satu penyebab umum inaktivasi Mismatch Repair Gene (MMR). Gen MMR memperbaiki kesalahan penyisipan/penghapusan basa nukleotida pada proses sintesis DNA. Metilasi pada promoter gen MMR memiliki asosiasi dengan pembentukan kanker kolon, sehingga metilasi tersebut perlu diidentifikasi. Identifikasi gen MMR dapat dilakukan menggunakan teknik methylation-specific multiplex ligation-dependent probe amplification amplification (MS-MLPA). Prinsip dari teknik MS-MLPA yaitu amplifikasi probe yang menempel pada sekuens termetilasi. Tujuan dari penelitian ini yaitu untuk mengoptimasi teknik MS-MLPA dan mengidentifikasi metilasi gen MMR pada kanker kolon dengan teknik MS-MLPA. Penelitian ini menggunakan 27 sampel jaringan frozen kanker kolon yang telah tersedia di Biobank Rumah Sakit Kanker Dharmais (RSKD). Sampel tersebut dianalisis menggunakan probemix Mismatch Repair Gene [ME011-C1][C1-0518] yang telah didesain khusus untuk mendeteksi pada beberapa gen MMR yakni MLH1, PMS2, MSH6, dan MSH2. Hasil penelitian menunjukkan optimasi teknik MS-MLPA telah berhasil dilakukan, sehingga identifikasi metilasi pada gen MMR telah berhasil diperoleh pada 4 sampel pasien. Gen MMR tersebut yakni MLH1 dan MSH6, dengan persentase masing-masing 75% dan 25%. ......DNA methylation is one of the most common causes of mismatch repair gene (MMR) inactivation. The MMR gene corrects errors in the insertion/deletion of nucleotide bases in the DNA synthesis process. MMR gene promoter methylation has an association with the formation of colon cancer, so the methylation needs to be identified. Identification of the MMR gene can be done using the methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) technique. The principle of the MS-MLPA technique is the amplification of the probe attached to the methylated sequence. The purpose of this study was to optimize the MS-MLPA technique and identify MMR gene methylation in colon cancer using the MS-MLPA technique. This study used 27 samples of frozen colon cancer tissue that were available at the Dharmais Cancer Hospital Biobank (RSKD). The samples were analyzed using the Mismatch Repair Gene probemix [ME011-C1][C1-0518] which has been specially designed to detect several MMR genes, namely MLH1, PMS2, MSH6, and MSH2. The results show that the optimization of the MS-MLPA technique has been successfully carried out, so that the identification of methylation in the MMR gene has been successfully obtained in 4 patient samples. The MMR genes are MLH1 and MSH6, with a percentage of 75% and 25%, respectively. analyzed using probemix Mismatch Repair Gene [ME011-C1][C1-0518] which has been specifically designed to detect several MMR genes namely MLH1, PMS2, MSH6, and MSH2. The results showed that the optimization of the MS-MLPA technique was successful, so that identification of the methylation in the MMR gene was successfully obtained in 4 patient samples. The MMR genes are MLH1 and MSH6, with percentages of 75% and 25% respectively.

Abstrak :

Gen mismatch repair (MMR) merupakan gen yang berperan dalam mekanisme DNA repair. Gen MMR dapat memperbaiki kesalahan pasangan basa, perubahan nukleotida dan kesalahan dalam unit pengulangan pasangan basa (microsatellites) selama proses replikasi DNA. Mutasi pada MMR yang umum terjadi pada gen MLH1 dan MSH2 memiliki asosiasi dengan terjadinya Lynch Syndrome pada pasien kanker kolon. Lynch Syndrome (LS) atau Hereditary Non-Polyposis Colorectal Carcinoma (HNPCC) merupakan sindrom kanker kolon herediter yang paling umum diwariskan. LS dapat ditandai dengan mutasi gen MLH1 dan MSH2 berupa delesi dan duplikasi large genomic. Multiplex ligation-dependent probe amplification (MLPA) merupakan teknik kuantifikasi berbasis PCR yang dapat digunakan untuk mendeteksi perubahan copy number variant pada large genomic. Tujuan dari penelitian ini adalah untuk mengidentifikasi mutasi gen MLH1 dan MSH2 pada kanker kolon dengan teknik Multiplex Ligation-Dependent Probe Amplification (MLPA). Sebanyak 25 sampel darah pasien yang terdiagnosis kanker kolon yang dikumpulkan secara retrospektif di Rumah Sakit Kanker Dharmais dikoleksi dari tahun 2018-2019. Mutasi gen MLH1 dan MSH diidentifikasi pada sampel darah pasien yang terdiagnosis kanker kolon dengan teknik MLPA menggunakan Probemix P003-D1 MLH1/MSH2 (MRC-Holland). Penelitian ini berhasil melakukan optimalisasi penggunaan teknik MLPA dengan terpenuhinya kualitas fragmen kontrol MLPA, kualitas fragment analysis, dan comparative analysis pada perangkat lunak Coffalayser.Net. Hasil screening pada 25 sampel menunjukkan tidak ditemukan adanya pola mutasi gen MLH1 dan MSH2 pada sampel yang mengindikasikan tidak terjadinya Lynch Syndrome dan kanker kolon yang terjadi bersifat sporadis.

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Mismatch repair genes (MMR) plays a role in the mechanism of DNA repair. The MMR genes correct base pair errors, nucleotide changes, and errors in base pair repeating units (microsatellites) during DNA replication. Mutations in MMR are common in MLH1 and MSH2 genes that have an association with the occurrence of Lynch Syndrome in colon cancer. Lynch Syndrome or Hereditary Non-Polyposis Colorectal Carcinoma (HNPCC) is the most common inherited hereditary colon cancer syndrome. LS is characterized by MLH1 and MSH2 gene mutations in the form of large genomic deletions and duplication. Multiplex ligation-dependent probe amplification (MLPA) is a PCRbased quantification technique that can detect changes in copy number variants in large genomics. The aim of the study was to identify MLH1 and MSH2 gene mutations in colon cancer with Multiplex Ligation-Dependent Probe Amplification (MLPA) technique. Twenty-five blood samples of patients diagnosed with colon cancer were collected in Dharmais Cancer Hospital collected in 2018-2019, retrospectively. Mutations in MLH1 and MSH2 genes were identified in blood samples of patients diagnosed with colon cancer by the MLPA technique using Probemix P003-D1 MLH1/MSH2 (MRC-Holland). This study succeeded in optimizing the use of MLPA technique by fulfilling the fragments quality control of MLPA, the quality of fragment analysis, and comparative analysis in Coffalayser.Net software. The results of 25 samples screening showed no pattern of MLH1 and MSH2 gene mutations in the sample. This data indicated that there was no Lynch Syndrome and the colon cancer was sporadic.