The Association between Estradiol Level, Remission, and Survival in Breast Cancer Patients at Dr. Moewardi Hospital, Surakarta, Central Java

Background: The mortality rate of breast cancer patients was relatively high 80% – 60% while the survival rate was only <40%. High estrogen levels and long-term exposure were associated with an increased risk of breast cancer. Recent studies have reported that estradiol can act independently and contribute to the risk of distant metastases in breast cancer patients with negative Estrogen Receptor (ER) tumors. The aim of this study was to estimate the estradiol effect on remission and survival in breast cancer patients in Dr. Moewardi Hospital, Surakarta, Central Java. Subjects and Method: This was a cohort study conducted at the Department of Surgical Oncology, Dr. Moewardi Hospital, Surakarta, Central Java, from March to May 2015. A sample of 303 breast cancer patients was selected for this study and followed up until 3 years. The independent variable was estradiol level. The dependent variables were remission and survival. The data were collected by clinical examination and analyzed by Cox Proportional Hazard Model and Kapplan-Meier. Results: High estradiol level decreased the possibility of remission (HR= 0.31; 95% CI= 0.09 to 1.11, p= 0.003) in breast cancer patients. Normal estradiol level increased survival (HR= 4.03; 95% CI= 1.94 to 8.37; p<0.001). Conclusion: High estradiol level decreases the possibility of remission, while normal estradiol level increases survival in breast cancer patients. Keywords: remission, survival, estradiol, breast cance

Histopathological Features of Early Onset Indonesian Breast Cancer Pointing to Brca1/2 Germline Mutations

Background: Breast cancer under 40 years concerns a relatively small subgroup of cases that tend to display a more aggressive phenotype. Compatible with this, early age of onset has been known as one of clinical characteristic of hereditary breast cancers associated with germline BRCA1 or BRCA1 mutations. As early onset breast cancer is frequent in Indonesia, we investigated the histopathological and immunohistochemical characteristics of early onset (< 40 years) Indonesian breast cancer patients, as such features can be used to distinguish between BRCA and non-BRCA germline mutation carriers among these young women.Method: Thirty-five formalin-fixed and paraffin-embedded tissue sections of young women (mean 36 years, range 22-40 years) who underwent surgical resection at the Department of Surgery of the Sardjito Hospital Yogyakarta were examined for pathological features, estrogen and progesterone receptor status, proliferation as determined by Ki67 labeling, EGFR and CK5/6 and the presence of HER-2/neu and p53 protein. Additionally, mutation analysis for BRCA1 and BRCA2 was performed in 30 young women. The control group consisted ofcarcinomas from women above 50 years (mean 59.02, range 50-80 years).Result: Carcinomas occurring in women aged below 40 years were more often advanced stage and higher proliferating (p=0.006). Among the early onset breast cancer patients, ductal type, grade 3, ER and HER-2/neu negativity, high Ki67 index and CK5/6 and EGFR positivity were typical for BRCA1 patients. Unfortunately, there were no typical phenotypical features for BRCA2 carriers. However, grade I and lobular cases were never BRCA1/2 germline mutated.Conclusion: Early onset Indonesian breast cancer shows increased proliferation compared to late onset patients. Within the early onset group, the strongest features pointing to a sporadic cancer seem to be grade I and lobular differentiation. Features increasing the chance of a germline BRCA1/2 mutation are CK5/6 and EGFR expression, p53 accumulation and high proliferation as measured by Ki67 labeling. This is potentially useful to optimize selection of early onset breast cancer patients for BRCA1/2 mutation testing

Histopathological Features of Early Onset Indonesian Breast Cancer Pointing to Brca1/2 Germline Mutations

Background: Breast cancer under 40 years concerns a relatively small subgroup of cases that tend to display a more aggressive phenotype. Compatible with this, early age of onset has been known as one of clinical characteristic of hereditary breast cancers associated with germline BRCA1 or BRCA1 mutations. As early onset breast cancer is frequent in Indonesia, we investigated the histopathological and immunohistochemical characteristics of early onset (< 40 years) Indonesian breast cancer patients, as such features can be used to distinguish between BRCA and non-BRCA germline mutation carriers among these young women.Method: Thirty-five formalin-fixed and paraffin-embedded tissue sections of young women (mean 36 years, range 22-40 years) who underwent surgical resection at the Department of Surgery of the Sardjito Hospital Yogyakarta were examined for pathological features, estrogen and progesterone receptor status, proliferation as determined by Ki67 labeling, EGFR and CK5/6 and the presence of HER-2/neu and p53 protein. Additionally, mutation analysis for BRCA1 and BRCA2 was performed in 30 young women. The control group consisted ofcarcinomas from women above 50 years (mean 59.02, range 50-80 years).Result: Carcinomas occurring in women aged below 40 years were more often advanced stage and higher proliferating (p=0.006). Among the early onset breast cancer patients, ductal type, grade 3, ER and HER-2/neu negativity, high Ki67 index and CK5/6 and EGFR positivity were typical for BRCA1 patients. Unfortunately, there were no typical phenotypical features for BRCA2 carriers. However, grade I and lobular cases were never BRCA1/2 germline mutated.Conclusion: Early onset Indonesian breast cancer shows increased proliferation compared to late onset patients. Within the early onset group, the strongest features pointing to a sporadic cancer seem to be grade I and lobular differentiation. Features increasing the chance of a germline BRCA1/2 mutation are CK5/6 and EGFR expression, p53 accumulation and high proliferation as measured by Ki67 labeling. This is potentially useful to optimize selection of early onset breast cancer patients for BRCA1/2 mutation testing.Keywords: breast cancer, early onset, histopathology, immunohistochemistry, BRCA1, BRCA

Resistance to doxorubicin correlated with dysregulation of microRNA-451 and P-glyoprotein, caspase 3, estrogen Receptor on Breast Cancer cell line

Doxorubicin (Dox)has beenused widely in breast cancer therapy. One of the problems in chemotherapy is the development of resistance to chemotherapy that lead to metastasis and relapse aggressiveness of cancer. MicroRNAs (miRNAs) are small non-coding RNA that regulate protein expression and play role in carcinogenesis, as well as cancer chemotherapy resistance. MiR-451 is classified as tumour suppressor miRNA, that binds to messenger RNA (mRNA) of MDR1, and leads disruption of  P-glycoprotein (Pgp) expression. Thestudy aimed to investigate the association between miR-451 and Pgp related with Dox resistance mechanism. In silico analysis was conducted to predict the binding affinity between miR-451 and mRNA of MDR1. The MCF-7 cell line was used as wild type model, while MCF-7/Dox was used as a model of resistance. qPCR was conducted to calculated miR-451 expression and immunocytochemistry was used to observe Pgp expression. miRNA was down-regulated in both on MCF-7 and MCF-7/Dox. On the other hand, Pgp expression was detectable in the cytoplasmic and cytoplasmic membrane in MCF-7/Dox. The Pgp expression was higher in the MCF-7/Dox compared to MCF-7. In conlusion, the over expression of Pgp is associated with the resistance to MCF-7/Dox

The Expression of miR-141 and mRNA PTEN with Cisplatin Therapy on NPC

The incidence of NPC in Yogyakarta province is 6.2/100 000 population and the tendency is increasing in the younger population. It has been known that drugs resistance also issues in NPC therapy. Cisplatin is drugs of choice that used in NPC therapy, besides miR-141 and mRNA PTEN are also had a role in chemotherapyresistance in some of cancer. We conducted the examination of the expression of miR-141 from blood plasma pre and post-therapy patient and blood plasma in healthy control. The sample collected from whole blood and then plasma separation. RNA extracted using RNA Isolation Kit miRCURY-Biofluid, synthesis cDNA with cDNASynthesis kit. mRNA analysis with KAPA SYBR® FAST One-Step qRT-PCR Kit then detected by qRT-PCR. Data analysis with comparative analysis and statistical analysis. MiR-141 showed upregulation by 1.49 (p-value: 0.075) and mRNA PTEN showed downregulation by 0.65 (p-value: 0.323) in patients NPC compared to healthy control and there is a relationship between miR-141 and mRNA PTEN (p-value:0.001). The expression levels of miR-141 in patients pre- and post-therapy experience down-regulated (fold change:0.61, two-tailed t-test: 0.09) and mRNA expression levels of PTEN experience down-regulated (fold change:0.5, t-test two-tailed:0.09). The level expression of mRNA PTEN is still down-regulated through the expression of miR-141 is up-regulated indicated that the possibility of drugs resistance in therapy with cisplatin. Then, miR-141 is one of the important factors in cisplatin drugs resistance with inhibition of mRNA PTEN. Keywords: miR-141, mRNA PTEN, Cisplatin, Drugs resistance, NPC

miR-21, miR-29c, and miR-155 as Biomarker to Develop Minimal Invasive Diagnostic in Hepatocellular Carcinoma Patient

Background: Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies and the third leading cause of cancer-related deaths worldwide. Incidence and mortality of HCC  increase annually because of its poor prognosis. The most common cause of this condition is its characteristics with high metastasis and high recurrence after surgical treatment. So, it is become important to find new serological biomarkers for early stage detection of HCC. Plasma microRNA are being actively investigated as minimal invasive biomarkers in human cancers, including HCC.Objective: The aim of this study is to investigate the level expression of miR-21, miR-29c, and miR-155 as novel serological biomarkers for hepatocellular carcinoma.Methods: This study is quasi experimental and remain as preliminary study. Collecting for more samples is currently underway. HCC patient blood samples obtained from RSUP dr. Sardjito Yogyakarta based on specific inclusion and exclusion criteria. Blood samples were collected from 8 patients and 8 healthy controls. The collected blood samples were treated as follows: plasma isolation, RNA total isolation, cDNA synthesis, quantification by qRT-PCR, data analysis with Biorad CFX ManagerTM Softwere to determine Cq, followed by the calculation of expression levels using Livax Methods.Result: The result showed that miR-21 and miR-155 were upregulated 1.68 fold and 2.38 fold respectively compared with healthy control, while miR-29c was downregulated 3,45 fold compared with healthy control.Conclusion: Based on the result of preliminary study, it can be concluded that miR-21 act as oncomiR, while miR-29c and miR-155 act as tumor supressor miR in HCC. The three microRNAs can be detected in HCC and can be used as minimal invasive biomarkers to detect HCC.Keywords: HCC, miR-21, miR-29c, miR-155, minimal invasive, biomarke

THE EXPRESSION OF Hsa-miR-21-5p AS MINIMAL INVASIVE MARKER TO ADJUVANT CHEMOTHERAPY IN BREAST CANCER PATIENTS

AbstractBreast cancer remains the leading cause of death among women, and there is a need to develop minimally invasive marker. In our previous study based on clinicopathologic in pre-chemotherapy patients showed miR-21 was upregulated 1.32 times higher at advanced stage compared with early stage. Therefore the matched patients for post-chemotherapy samples were used. The aim of this research is to examine the expression of miR-21 as potential marker to adjuvant chemotherapy in breast cancer patients. The samples were taken by using cross sectional method with total 39 blood plasma samples from breast cancer patients in adjuvant chemotherapy and 12 healthy control samples. Plasma was obtained from blood samples and then RNA isolated were performed. Total RNA was reverse transcribed using cDNA synthesis. The expression of miR-21 was then analyzed using specific primer for miR-21 and miR-16 as the reference gene. Livak Method was used to calculate the expression level in each group. The result showed that there is significant downregulated expression of miR-21 in postchemotherapy 2.61 fold compared with pre-chemotherapy (p<0.05). The expression of miR-21 upregulated 2.2 folds (p<0.05) in pre-chemotherapy compared with healthy control, while in post-chemotherapy compared with healthy control, the expression of miR-21 was 0.8 fold (p<0.05). In conclusion, Hsa-miR-21-5p can be used as marker for adjuvant chemotherapy response in breast cancer because there is significant different expression between prechemotherapy, post-chemotherapy and healthy control. The continuation research in the near future for detecting the expression of tumor suppressor protein regulated by miR-21 is needed.Keywords: breast cancer, adjuvant chemotherapy, miR-21, minimal invasive marke

Expression of Circulating microRNA-141 and mRNA of PTEN (Phosphatase and Tensin Homolog) in Blood Plasma of Ovarian Tumor and Epithelial Ovarian Cancer Patient

Epithelial Ovarian Cancer (EOC) is the most lethal gynecological malignancies among woman. The majority of this disease is diagnosed at the advanced stage due to lack of specific symptoms and effective screening methods. Therefore, an adequate biomarker for early detection is needed and may improve patient survival. microRNA is a small non-coding RNA that regulates gene expression in post-transcriptional level. Several studies have shown the ability to detect microRNA in blood circulation so microRNA may be used as a minimally invasive biomarker for EOC. microRNA-141 (miR-141) plays a major role in EOC by regulating expression of several tumor suppressor gene. Previous study have  confirmed that miR-141 regulated  PTEN gene directly by interacting with 3’UTR sequence of PTEN mRNA, and upregulation of miR-141 caused downregulation of PTEN expression in vivo. PTEN is important tumor suppressor gene that its inactivation found in various human cancer. PTEN is protein with lipid phosphatase activity that negatively regulate PI3K-AKT signaling pathway, thus playing a important role in various cellular process such as proliferation, growth, cell survival, EMT, cell motility, and angiogenesis. When various studies found that PTEN mRNA and protein expression is significantly downregulated in EOC tissue, little is known about the expression of PTEN mRNA in blood circulation of EOC patient, especially in Yogyakarta population.The aims of this study is to measure and to investigate the correlation of miR-141 and mRNA PTEN expression in plasma of ovarian tumor patient and EOC patient.This study used cross-sectional design. 25 blood plasma of ovarian tumor and 25 blood plasma of EOC were collected. Total RNA was isolated and reverse transcribed to obtain cDNA. The expression of miR-141 and mRNA PTEN were measured by quantitative real-time polymerase chain reaction assay (qPCR). The 2^(-∆∆cq) method was used to calculate relative quantification of miR-141 and mRNA PTEN using miR-16 as reference gene for microRNA and beta-actin mRNA as reference genes for PTEN mRNA. Expression of miR-141 is significantly elevated in blood plasma of epithelial ovarian cancer patient compared to the ovarian tumor (p=0,001, fold change=7,59). Expression of PTEN mRNA significantly downregulated in blood plasma of epithelial ovarian cancer patient compared to the ovarian tumor (p=0,001, fold change=11,63). There was a significant negative correlation between miR-141 expression and mRNA PTEN expression in blood plasma of epithelial ovarian cancer patient (p=0,033; r=-0,428).miR-141 and mRNA PTEN differentially expressed in blood plasma of ovarian tumor and epithelial ovarian cancer patient. There was a negative correlation between miR-141 and mRNA PTEN expression in blood plasma of epithelial ovarian cancer patient.  Keywords: Epithelial ovarian cancer, circulating microRNA, miR-141, PTEN mRN

The Expression of Homo Sapiens microRNA-21 (Hsa-miR-21-5p) and mRNA Reversion Inducing Cysteine Rich Protein with Kazal Motifs (RECK) in Plasma of Epithelial Ovarian Cancer

ABSTRACT Epithelial Ovarian Cancer (EOC) is malignant cancer that caused death for most women in Indonesia. The emergence of EOC showed no specific symptoms in its early stages; that makes the screening mostly occur when patients are in advanced stage. Treatment of EOC at an advanced stage will be more challenging with poor prognosis. Therefore, minimally invasive biomarkers are needed to diagnose at the early stage. microRNA is one of the potential biomarkers which not only expressed inside the cell but also secreted outside the cell with exosome protection. This protection makes microRNA stable. Moreover, several studies have shown the ability to detect microRNA in the blood sample. microRNA-21 (miR-21) is oncomiR which targeted tumor suppressor mRNA RECK based on in silico analysis.The first aim is to determine the expression of miR-21 in plasma samples of EOC patients compared with healthy controls. The second aim is to investigate the expression correlation between miR-21 and RECK mRNA.Blood samples were collected from 30 patients and 30 healthy controls. Plasma was then obtained from centrifugated blood samples. The total RNA was isolated and reverse transcribed to produce cDNAs. cDNAs were then quantified using qPCR using specific primer for miR-21 and RECK mRNA. The expression analysis was done relative expression method by Livak. The expression of miR-21 was calculated using the miR-16 expression as the reference gene. Also, Beta-actin was used as reference gene for RECK mRNA calculation. The correlation between the expression of miR-21 and mRNA RECK was analyzed using the Spearman rho correlation analysis.In this study showed the expression of miR-21 in patients with EOC increased 4.7579-fold compared with healthy controls (p <0.05). On the other hand, the miR-21 target, RECK mRNA, decreased 4,2 times with fold change 0.237665 on plasma EOC patients compared with healthy controls (p <0.05). The statistical calculation of the expression of miR-21 and mRNA RECK was inversely proportional to mRNA RECK with a strong correlationThis study has been able to prove that the expression of miR-21 is up-regulated in EOC patients and confirmed by the down-regulation of RECK expression. The next research challenge is to make anti-miR-21 to suppress the expression of miR-21 in EOC, which can be analyzed the effect of miR-21 in the development of EOC