Nayoung Han

Epigenetic changes in gene expression for drug-metabolizing enzymes and transporters.

Individual differences in drug response can be caused by genetic and epigenetic variability and disease determinants. Pharmacoepigenetics is a new field that studies the expression changes in pharmacogenes without changes in DNA sequences. Epigenetic control mechanisms are associated with DNA methylation, histone modification, small noncoding RNAs, and nucleosome remodeling. Researchers are actively attempting to identify epigenetic mechanisms for controlling the expression of enzymes and transporters affecting the metabolism and disposition of drugs. Current evidence suggests that epigenetic changes play a major role in cytochrome P450 enzyme expression, major transporter function, and in interactions with nuclear receptors. A thorough understanding of pharmacoepigenetics provides insight into new approaches to drug discovery and development, provides an understanding of previously observed actions of older drugs, and provides a pathway by which epigenetics can be harnessed to provide better patient‐specific therapy.

Combinational effect of intestinal and hepatic CYP3A5 genotypes on tacrolimus pharmacokinetics in recipients of living donor liver transplantation.

Background For living donor liver transplantation, the genetic association of CYP3A5 genotype of recipient’s native intestine and donor’s liver allograft with tacrolimus pharmacokinetics has not been explained completely considering liver regeneration time. The goal of the study was to investigate the longitudinal effects of recipient-donor combinational CYP3A5 genotypes on tacrolimus dose-normalized concentration (C/D ratio) in blood. Methods Tacrolimus blood concentrations were measured for 58 Korean adult living donor liver transplant recipients on tacrolimus-based immunosuppressants during 4 years of follow-up. CYP3A5 was genotyped for both recipient and donor, and the recipient-donor combinational genetic effect on tacrolimus C/D ratios were evaluated as a function of time after adjusting for covariates including demographics and clinical variables. Results CYP3A5 expresser recipients grafted from CYP3A5 expresser donors consistently had the least C/D ratio throughout the entire study period, whereas CYP3A5 expresser recipients grafted from CYP3A5 nonexpresser donors had an intermediate, and CYP3A5 nonexpresser recipients grafted from CYP3A5 nonexpresser donors had the largest C/D ratio (all P < 0.01). The CYP3A5 nonexpresser recipients grafted from CYP3A5 expresser donors showed a significant decrease from the largest to the intermediate in C/D ratio for the first month. Conclusions CYP3A5 genotypes of both recipient and donor were important factors influencing pharmacokinetic variability of tacrolimus. The recipient-donor combinational genetic effect on C/D ratio changed over time after transplantation.

Caveolin 1 Expression Correlates with Poor Prognosis and Focal Adhesion Kinase Expression in Gastric Cancer

Objective:Caveolin 1 gene is known as a tumor promoter or suppressor, depending on the tumor type and/or tumor stage. We aimed to investigate the clinical significance of caveolin 1 protein (Cav1) expression in gastric cancer (GC). Methods: Immunohistochemistry was performed on tissue array slides containing 405 GC specimens. The relationships between Cav1 expression and clinicopathological factors, prognosis, focal adhesion kinase expression, mucin phenotypes and p53 expression were analyzed. Results: In non-neoplastic gastric mucosa, Cav1 was not expressed in the epithelial compartment. In GC, positive staining of Cav1 was shown in 22 (5.4%) of 405 cases and was significantly higher in the advanced GC group than in the early GC group (p = 0.037). Also, it was significantly associated with advanced pTNM stage (p = 0.027) and lymph node metastasis (p = 0.018). Moreover, survival analysis showed that Cav1 expression was an independent prognostic factor of poor survival (p = 0.028). In addition, the expression of Cav1 was positively correlated with that of focal adhesion kinase (p = 0.034). Conclusions: These results indicate that the expression of Cav1 is significantly correlated with cancer progression and poor prognosis in GC. Thus, Cav1 could supplement its protein expression for the diagnosis and treatment of GC.

Effects of Fixation and Storage of Human Tissue Samples on Nucleic Acid Preservation

Background Because of recent advances in the molecular diagnosis of cancer patients, tissue quality has become more important in daily practice. Methods To evaluate the effects of fixative, duration of fixation, decalcification, and storage periods on nucleic acid integrity, DNA and RNA were extracted from gastrointestinal cancer tissue. The yield and purity were analyzed, and polymerase chain reaction (PCR) for glyceraldehyde 3-phosphate dehydrogenase (GAPDH; 60 bp), β-actin (148 bp), and human growth hormone (hGH; 434 bp) and real-time reverse transcription-PCR for β-actin (97 bp) were performed. Results All formalin-fixed paraffin-embedded (FFPE) and methacarn-fixed paraffin-embedded (MFPE) samples tested positive for GAPDH and β-actin by PCR. hGH was successfully detected in all MFPE samples, but in only 46.7% of the FFPE samples. Prolonged formalin fixation resulted in fewer GAPDH and β-actin PCR products, and amplification of hGH was not successful. The PCR and reverse transcription-PCR results were significantly affected by the duration of decalcification. The yield, purity, and integrity of mRNA progressively decreased with increased storage periods of paraffin blocks. Conclusions Fixation and storage should therefore be standardized in order to improve the quality of molecular pathologic diagnosis.

DNA Damage Response-Related Proteins in Gastric Cancer: ATM, Chk2 and p53 Expression and Their Prognostic Value

Objectives: The aims of this study were to assess expressions of the DNA damage response (DDR)-related proteins and to investigate their clinical significances in gastric carcinoma. Methods: Two independent cohorts, a training set (n = 524) and validation set (n = 394), of gastric cancer patients were enrolled. Ataxia telangiectasia mutated (ATM), checkpoint kinase 2 (Chk2), and p53 expressions were examined by immunohistochemistry using tissue microarray. Results: ATM loss, Chk2 loss, and p53 positivity were observed in 21.8, 14.1, and 36.1% of the training set, and in 17.3, 12.2, and 35.8% of the validation set, respectively. In the training set, the aberrant expressions of ATM, Chk2, or p53 were significantly associated with an advanced TNM stage and poor disease-specific survival. This association was verified in the validation set. Chk2 positivity and p53 negativity were significantly related to a prolonged disease-specific survival. Also, patients with nonaberrant expressional levels of all 3 DDR-related proteins had a more favorable outcome than others. Multivariate analyses showed that Chk2 loss and at least 1 aberrant DDR-related protein remained as independent prognostic factors of poor disease-specific survival. Conclusions: This study elucidated the prognostic implications of DDR-related proteins, and suggests that their aberrant expressions play critical roles in the development and progression of gastric cancer.

ABCB1 C3435T Genetic Polymorphism on Population Pharmacokinetics of Methotrexate After Hematopoietic Stem Cell Transplantation in Korean Patients: A Prospective Analysis

BACKGROUND Methotrexate (MTX) is often used to prevent graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (HSCT). However, MTX has great pharmacokinetic variability and its use can result in fatal complications and/or infections after HSCT. OBJECTIVES The purposes of this study were to build a population pharmacokinetic model of MTX treatment in Korean patients who have undergone HSCT and to identify covariates, including genetic polymorphisms, that affect the pharmacokinetic properties of MTX. METHODS Clinical characteristics and MTX concentration data for 20 post-HSCT patients were collected. For each patient, ABCB1, ABCC2, ATIC, GGH, MTHFR, and TYMS genotyping was performed. Population pharmacokinetic analysis was performed using the NONMEM program. Analysis of MTX pharmacokinetic properties was accomplished using a 2-compartment pharmacokinetic model that incorporated first-order conditional estimation methods with interaction. The effects of a variety of demographic and genetic factors on MTX disposition were investigated. RESULTS The study population consisted of 12 men (60%) and 8 women (40%). Median age and body weight were 28 years (range, 18-49 years) and 55.6 kg (range, 44.8-80.8 kg), respectively. Within the study population, the estimated mean MTX clearance (CL) was 7.08 L/h, whereas the mean central compartment volume (V(1)) of MTX distribution was 19.4 L. MTX CL was significantly affected by glomerular filtration rate (GFR), penicillin use, and the ABCB1 3435 genotype. Interindividual variabilities for CL and V(1) were 21.6% and 73.3%. A 10-mL/min GFR increase was associated with a 32% increase in mean MTX CL, whereas penicillin use was associated with a decrease in MTX CL of 61%. MTX CL was significantly greater (by ∼21%) in patients with the ABCB1 3435 CC and CT genotype than in those with the ABCB1 3435 TT genotype (P < 0.001). CONCLUSIONS There was great interindividual variation in MTX pharmacokinetic properties in patients who had undergone HSCT. GFR, concurrent penicillin use, and the presence of the ABCB1 3435 C

Prognostic implication of TSC1 and mTOR expression in gastric carcinoma

Gastric adenocarcinoma is the sixth most common and third most lethal cancer in the world. Except for HER2‐targeted therapy, targeted agents against specific molecules participating in gastric carcinogenesis, including those in the mechanistic target of rapamycin (serine/threonine kinase) (mTOR) pathway have not been proved to be effective. However, some studies have suggested that dysfunction of TSC1 may augment mTOR inhibitor activity.

Population pharmacokinetic-pharmacogenetic model of tacrolimus in the early period after kidney transplantation.

As tacrolimus has a rather narrow therapeutic range and high individual variability in its pharmacokinetics, it is important to determine the cause of the variation in tacrolimus pharmacokinetics. The purpose of this study was to establish a population pharmacokinetic–pharmacogenetic model of tacrolimus and identify covariates that affect pharmacokinetic parameters to prevent fluctuations in the tacrolimus trough concentration during the early period after transplantation. Data from 1501 trough concentrations and 417 densely collected concentrations were compiled from 122 patients who were on post‐operative days 10–20 and analysed with a nonlinear mixed‐effect model. The first‐order conditional estimation (FOCE) with interaction method was used to fit the model using the NONMEM program. Clinical/laboratory data were also collected for the same period, and CYP3A5 and ABCB1 genotypes were analysed for use in modelling from all included patients. An empirical Bayesian approach was used to estimate individual pharmacokinetic profiles. A one‐compartment model with first absorption and elimination and lag time best described the data. The estimated population mean of clearance (CL/F), volume of distribution (V/F) and absorption rate (Ka) were 21.9 L/hr, 205 L, and 3.43/hr, respectively, and the lag time was fixed at 0.25 hr. Clearance increased with days after transplantation and decreased with CYP3A5*3/*3 about 18.4% compared with CYP3A5*1 carriers (p < 0.001). A population pharmacokinetic model was developed for tacrolimus in early post‐kidney transplantation recipients to identify covariates that affect tacrolimus pharmacokinetics. Post‐operative days and CYP3A5 genotype were confirmed as critical factors of tacrolimus pharmacokinetics.

Meningeal Solitary Fibrous Tumors with Delayed Extracranial Metastasis

Background: The term solitary fibrous tumor (SFT) is preferred over meningeal hemangiopericytoma (HPC), because NAB2-STAT6 gene fusion has been observed in both intracranial and extracranial HPCs. HPCs are now considered cellular variants of SFTs. Methods: This study analyzes 19 patients with STAT6-confirmed SFTs, who were followed for over 11 years in a single institution. Ten patients (10/19, 56.2%) had extracranial metastases (metastatic group), while the remainder (9/19) did not (non-metastatic group). These two groups were compared clinicopathologically. Results: In the metastatic group, the primary metastatic sites were the lungs (n = 6), bone (n = 4), and liver (n = 3). There was a mean lag time of 14.2 years between the diagnosis of the initial meningeal tumor to that of systemic metastasis. The median age at initial tumor onset was 37.1 years in the metastatic group and 52.5 in the non-metastatic group. The 10-year survival rates of the metastatic- and non-metastatic groups were 100% and 33%, respectively. The significant prognostic factors for poor outcomes on univariate analysis included advanced age (≥45 years) and large initial tumor size (≥5 cm). In contrast, the patients with higher tumor grade, high mitotic rate (≥5/10 high-power fields), high Ki-67 index (≥5%), and the presence of necrosis or CD34 positivity showed tendency of poor prognosis but these parameters were not statistically significant poor prognostic markers. Conclusions: Among patients with SFTs, younger patients (<45 years) experienced longer survival times and paradoxically had more frequent extracranial metastases after long latent periods than did older patients. Therefore, young patients with SFTs require careful surveillance and follow-up for early detection of systemic metastases.

Copy number variability analysis of pharmacogenes in patients with lymphoma, leukemia, hepatocellular, and lung carcinoma using The Cancer Genome Atlas data.

Objective Individual differences in drug efficacy and toxicity remain an important clinical concern. We investigated copy number variation (CNV) frequencies for pharmacogenes using The Cancer Genome Atlas dataset. Materials and methods One hundred and fifty-two pharmacogenes were selected from liver hepatocellular carcinoma, lung squamous cell carcinoma (LUSC), acute myeloid leukemia, and lymphoid neoplasm diffuse large B-cell lymphoma (DLBL). The germ line and somatic CNV frequencies were analyzed. Results We found CNVs with more than 1% frequency in drug-metabolizing enzymes including CYP2A6, CYP2D6, GSTP1, CYP2E1, GSTM1, GSTT1, and SULT1A1, drug transporters such as SLC19A1 and SLC28A1, and targets such as FHIT in normal tissue or blood. GSTM1 had the highest frequency for gene gain (45.45, 39.18, 31.01, and 34.77%, respectively) and for gene loss (18.18, 29.38, 20.89, and 26.68%, respectively) in DLBL, acute myeloid leukemia, liver hepatocellular carcinoma, and LUSC. P2RY12 and P2RY1 had the highest frequency for gene gain in LUSC (26.95 and 26.68%, respectively) whereas ABCB1 and ABL2 had the highest frequency for gene gain in DLBL (27.27%) in cancer tissue or blood. Conclusion Germ line and somatic CNVs of pharmacogenes may play a role in determining individual variations in drug responses. Inclusion of CNVs in pharmacogenetic variations holds promise as biomarkers that can increase the benefits and reduce the risks of drug therapy on an individual level.