SIGNIFIKANSI KLINIKOPATOLOGI EKSPRESI PETANDA SEL PUNCA KANKER (ALDH1+/CD44+/CD24-), EKSPRESI GENA NOTCH1 DAN miRNA-200c PADA KARSINOMA PAYUDARA

ACHMAD GHOZALI, DR., SP.PA(K), Prof. Dr. dr. Teguh Aryandono Sp.BOnk(K).; dr. Totok Utoro D.M.Sc. Sp.PA (K)

2017 | Disertasi | S3 Ilmu Kedokteran

Latar belakang: Karsinoma payudara di Yogyakarta pada umumnya ditemukan pada usia muda dan stadium lanjut dengan mortalitas yang tinggi. Pertumbuhan dan progresivitas karsinoma payudara sering dihubungkan dengan peran sel punca kanker dan jalur pensinyalan Notch1. Petanda ALDH1, CD44 dan CD24 paling umum digunakan untuk mengidentifikasi sel punca kanker payudara. Sel punca kanker dengan fenotipe ALDH1+/CD44+/CD24- adalah fenotipe yang dianggap paling akurat untuk identifikasi sel punca kanker payudara. Aktivasi jalur Notch1 menentukan nasib sel sel punca embrional maupun dewasa untuk memperbarui diri, karena keterlibatannya dalam diferensiasi populasi sel punca somatik yang tidak terdiferensiasi menjadi jenis sel yang spesifik. Selain itu jalur pensinyalan notch berperan pula dalam apoptosis, proliferasi dan migrasi sel punca. Hasil-hasil penelitian tentang hubungan sel punca kanker dengan Notch1 dan dengan faktor-faktor klinikopatologis masih kontroversial. Meskipun belum diketahui dengan pasti, pada penelitian yang terbaru didapatkan indikasi bahwa Notch1 pada tingkat molekular diregulasi melalui cross talk dengan miRNA baik yang berisfat supressor tumor maupun onkomiR, yang menimbukan dugaan bahwa molekul molekul miRNA ini mempunyai peran kritikal pada tumor biologi. Tujuan penelitian: Tujuan penelitian ini adalah untuk meneliti hubungan antara keberadaan sel punca kanker dan ekspresi gena Notch1 dengan berbagai faktor klinikopatologi: Umur, grading histologis, ukuran tumor primer, keterlibatan limfonodi, subtipe molekular, dan korelasi antara ekspresi gena Notch1 dengan mirRNA-200c. Metode: Semua sampel berasal dari pasien RSUP Dr Sardjito. Sampel blok parafin untuk pemeriksaan status ekspresi ER, PR, Her2, Ki67, ALDH1, CD44 dan CD24 secara imunohistokimia, dan pemeriksaan ekspresi gena mRNA Notch1 (NCID) secara RT-PCR. Plasma darah untuk pemeriksaan ekspresi miRNA-200c secara RT-PCR juga. Selain itu dicatat pula informasi klinis dan patologi yang diperlukan, umur, ukuran tumor, status nodal. Hubungan antar variable dievaluasi secara statistic dengan uji Chi-square . Untuk menilai hubungan antara ekspresi gena Notch dan miRNA2-00c diperiksa dengan uji korelasi Spearman. Nilai signifikansi statistik sebesar p<0,05. Hasil: Sel punca positif, dengan fenotipe (ALDH1 + /CD44 + /CD24 sebanyak 54 (45%) dari 120 kasus. Berdasarkan klasifikasi menurut subtipe molekular didapatkan 47 (39,3%) luminal A, 27 (18,3%) luminalB, 13 (10,8%) Her2 dan triple negative 38 (31,7%). Tidak ada hubungan yang bermakna antara keberadaan sel punca ALDH1+/CD44+/CD24- dengan semua faktor klinikopatologis yang diteliti, kecuali dengan negativitas ER (p<0,001, PR=0,516, 95% CI:0,3467-0,7681) dan PR (p=0,007, PR=0,4636, 95% CI:0,2561-0,8393) serta indeks proliferasi (ki67) yang tinggi (p<0,001, PR=3,3611, 95% CI:1,9310- 5,8505). Sebagian besar tumor dengan subtipe triple negative dan Her2 mempunyai sel punca berfenotipe ALDH1+/CD44+/CD24-. Tidak ada hubungan yang bermakna antara ekspresi gena Notch1 dengan semua faktor kliikopatologi yang diteliti. Tidak ada korelasi antara ekspresi gena Notch1 dengan miRNA-200c, )ditemukan tetapi ada kecenderungan korelasi terbalik antara keduanya. Kesimpulan: Dari hasil penelitian ini dan pembahasannya dapat diambil kesimpulan sementara bahwa peran pensinyalan Notch1 dan miRNA-200c sebagai penyebab prevalensi yang tinggi dan proses regulasi sel punca berfenotipe (ALDH1 + /CD44 + /CD24 ) dan proses karsinogenesis kanker payudara yang diteliti belum dapat diabaikan.

Background: Carcinoma of the breast in Yogyakarta were usually found at a young age and advanced stage with high mortality. The growth and progression of breast carcinoma is often associated with the role of cancer stem cells and Notch1 signaling pathway. ALDH1 marker, CD44 and CD24 most commonly used to identify breast cancer stem cells. Cancer stem cells with the phenotype ALDH1 + / CD44 + / CD24- phenotype are considered the most accurate for the identification of breast cancer stem cells. Notch1 pathway activation determining cell fate of embryonic and adult stem cells to renew themselves, because of their involvement in the differentiation of somatic stem cell populations were not differentiated into specific cell types. Additionally notch signaling pathway plays a role also in apoptosis, proliferation and migration of stem cells. The results of studies on the association of cancer stem cells and Notch1 with clinicopathology factors still controversial. Although it is not known with certainty, the most recent studies found indications that Notch1 at the molecular level is regulated via cross talk with both miRNA tumor suppressors and onkomiR, which raises suspicion that these miRNA molecules have a critical role in tumor biology. Objective: The purpose of this study was to examine the relationship between the existence of cancer stem cells and the expression of Notch1 genes with different clinicopathologic factors: age, histological grading, primary tumor size, lymph node involvement, molecular subtypes, and the correlation between the expression of Notch1 genes with mirRNA200c Method: . All samples were from Dr Sardjito Hosptal patients. Paraffin block samples for examination of the expression of ER status, PR, Her2, Ki67, ALDH1, CD44 and CD24 by immunohistochemistry, and examination of the expression of Notch1 mRNA genes (NCID) by RT-PCR. Plasma blood for examination miRNA- 200c expression by RT-PCR as well. Besides noting the clinical and pathological information is needed, age, tumor size, nodal status. associations between variables were evaluated statistically with Chi-square test. To assess the correlation between the expression of genes Notch1 and miRNA-200c examined by Spearman correlation test. The value of statistical significance at p <0.05. Results: Positive stem cells, with (ALDH1+/CD44+/CD24-) phenotype were found as many as 54 (45%) of 120 cases. Based on the classification by molecular subtypes there were 47 (39.3%) luminal A, 27 (18.3%) luminal B, 13 (10.8%) Her2 and 38 (31.7%) triple negative.There was no significant association between the presence of stem cells ALDH1+/CD44+/CD24- with all clinicopathology factors studied, except with ER negativity (p <0.001, PR= 0.516, 95% CI: 0.3467 to 0.7681) and PR (p = 0.007, PR = 0.4636, 95% CI: 0.2561 to 0.8393) as well as the proliferation index (ki67) were higher (p <0.001, PR = 3.3611, 95% CI: 1.93105.8505). Most triple negative tumor subtype and Her2 have stem cells with phenotype ALDH1+/CD44+/CD24- There was no significant relationship between the expression of Notch1 genes with all the clinicopathology factors studied. There was no correlation between the expression of Notch1 genes with miRNA-200c, but there was a trend inverse correlation between the two. Conclusion: From the results of this research and the discussion can be concluded that the role of Notch1 signaling and miRNA-200c as the cause of high stem cell prevalence and in regulation process of (ALDH1+/CD44+/CD24) stem cell and breast cancer carcinogenesis process under study can not be ignored.

Kata Kunci : sel punca kanker payudara, petanda sel punca, pensinyalan Notch1, Breast cancer stem cells, stem cell markers, Notch1 signaling, miRNA-200c