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The administration of an accurate and effective POCUS course is a crucial tool in improving health education and thus the health care system in low- to middle-income countries. The development of the ultrasound curriculum in these countries during the pandemic era is a major challenge for medical educators. Therefore, this study aims to survey the learner experience after implementing the POCUS curriculum for first-year emergency medicine residents. All learners responded to the survey. Our results demonstrated that the ultrasound rotation and our ultra-sound learning materials were useful tools which showed a positive impact on POCUS knowledge for our learners. However, some obstacles of POCUS learning were identified to assist in closing faculty development gaps, including the availability of handheld devices, as well as the re-modeling of the ultrasound rotation course, which should be managed according to the feedback we received. This study demonstrated a clear need for constant updates in higher education, medical program development, accuracy of local learning materials, and the explosion of virtual and online learning platforms during this decade.

BACKGROUND: Patients with uncontrolled blood loss often require immediate blood transfusion after the bleeding is stopped. If it is an emergency situation, blood that has not been tested for compatibility (uncrossmatched red blood cell [URBC] products) can be used. However, no studies have been conducted to evaluate the effectiveness of this protocol. AIM: The aim of the study is to evaluate the effectiveness of URBC transfusion in Srinagarind Hospital’s emergency department (ED). METHODS: This was a cross-sectional study that reviewed the medical records of ninty Thai patients over 18 years of age who received at least one unit of blood through URBC transfusion in the Srinagarind Hospital ED from September 2016 to August 2018. RESULTS: The average age of the patients was 47.23 ± 18.2 years, and 73.3% were male. A total of 149 units of URBC were provided, with 54.44% of recipients being trauma patients and 27.78% being gastrointestinal bleeding patients. The 24-h and in-hospital mortality rates were 58.89 and 72.22%, respectively. There were no cases of acute blood transfusion complications or inappropriate URBC transfusion. CONCLUSIONS: The transfusion of URBC necessary in patients with uncontrolled bleeding. No complications were found due to acute blood transfusion.

BACKGROUND: Coronavirus disease 2019 has caused a major worldwide disruption to nearly all aspects of the medical education process, which includes the teaching of point-of-care ultrasound (POCUS). However, the impacts of the pandemic on POCUS training during this time have not been well studied. AIM: This was a retrospective observational study of the 1st year of emergency medicine (EM) residents at the Department of EM who had received POCUS training at a tertiary university hospital in Thailand. METHODS: The study was conducted throughout the 2019–2020 Academic year. The primary aim of this study was to evaluate the new POCUS training methodology, which, due to health and safety protocols, was carried out through virtual learning. RESULTS: A total of 18 1st-year EM residents were evaluated. The average pre-training scores in the 2019–2020 academic years were 5.25 and 5, respectively. The average post-training scores in the 2019 and 2020 academic years were 8.5 and 8.67, respectively. In terms of the ultrasound (US) skills test, the total average scores in the 2019–2020 academic year were 17 out of 20 (85%) and 14.875 out of 20 (74.38%), respectively (p < 0.001). CONCLUSIONS: Virtual learning is effective for teaching theoretical US knowledge. However, it was not suitable for teaching practical skills, including US skills.

BACKGROUND: Point of care ultrasound (POCUS) plays an important role as a crucial tool for the diagnosis of various conditions in emergency and critical ill patients. However, POCUS examination in elderly populations has not been well studied. AIM: This was a retrospective observational study of elderly patients at the Department of Emergency Medicine who had received POCUS examination at a tertiary university hospital, Thailand. METHODS: The study was conducted throughout January 2020–December 2020. Patients’ characteristics and ultrasonography findings were recorded. RESULTS: A total of 191 elderly patients were evaluated. Median patient age was 75.65 years; 56.02% of patients were female. Chief complaints where POCUS scans were applied were respiratory (36.65%) and cardiovascular system-related (21.99%). The most frequent procedures performed were cardiac, lung, and inferior vena cava examinations. Abnormal ultrasound findings were discovered in 133 patients (68.91%). The 66.17% of abnormal ultrasound findings were associated with final diagnosis. The admission rate of elderly patients (56.82%) was highest among patients with positive ultrasound findings associated with final diagnosis. CONCLUSIONS: POCUS utilization in elderly patients was useful in aiding emergency physicians amid diagnosis of various diseases, especially life-threatening ones.

BACKGROUND: Point-of-care ultrasound (POCUS) is the core competency in the Emergency Medicine (EM) residency training. However, there are many methods that can be used to evaluate this competency, and the best practices for teaching ultrasonography to residents have yet to be determined. AIM: The researchers aimed at evaluating the POCUS knowledge and skills of the EM residents after having participated in the POCUS training during their first ultrasound rotation in the Emergency Department. METHODS: A curriculum was developed in the form of a 2-week rotation in the EM residency program at the Department of EM at Khon Kaen University’s Srinagarind Hospital. It consisted of didactic lectures, bedside ultrasound trainings, the journal club, and the process of reviewing the images. Tools were developed, which included a knowledge exam. For each resident, the assessments were administered before and after the rotation. Furthermore, an ultrasound skills test was developed to be used at the end of the 1st year EM residency program. RESULTS: Nine EM residents completed their rotations and the tests. The average pre-training score and post-training scores were 5.25 ± 1.03 and 8.50 ± 1.20, respectively. The mean difference score between pre- and post-test was 3.25 ± 1.28. (95% CI −4.321, −2.178). In terms of the ultrasound skills test, the average total score was 26.13 out of 30 (87.1%). Moreover, the residents had higher scores in the aspects of image acquisition (87.5%) and image interpretation (87.5%). However, for the aspect of clinical decision-making, the average score was 75%. The survey questions indicated that with respect to all of the academic activities, the “Bedside ultrasound,” which had encouraged the residents to learn POCUS, was given the highest score (4.75 of 5). CONCLUSIONS: The 2-week ED ultrasound rotation had improved the residents’ EM ultrasound knowledge and skills.

© 2021 Korakot Apiratwarakul, Takaaki Suzuki, Ismet Celebi, Kamonwon Ienghong, Thanat Tangpaisarn, Somsak Tiamkao, Vajarabhongsa Bhudhisawasdi.BACKGROUND: Concerning the COVID-19 outbreak in Thailand, the number of patients has been increasing. Emergency medical services (EMS) operating duration differs from normal services due to equipment preparation, number of personnel, and on-board ambulance procedures. Notably, there have been no studies examining EMS duration regarding COVID-19 patients. AIM: The aim of this study was to compare the EMS time for COVID-19 patients and routine services. METHODS: This cross-sectional study was conducted in a tertiary university hospital in Khon Kaen, Thailand. Information gathering was carried out by employing the Srinagarind Hospital EMS database throughout January 1, 2020, and February 10, 2021. RESULTS: A total of 2420 EMS operations were examined, of which five tested positive for COVID-19 (0.21%). The mean age of the COVID-19 patients was 35.6 ± 7.2 years, with the activation interval for COVID-19 and routine services at 64.20 ± 10.14 and 1.42 ± 0.42 min, respectively (p < 0.001). The on-scene time for COVID-19 and routine services was 3.20 ± 0.44 and 5.20 ± 2.20 min, respectively. CONCLUSIONS: EMS operating time amid the activation interval for COVID-19 patients was significantly longer than in the normal group. However, on-scene time for COVID-19 patients was less time than in normal operations.

BACKGROUND: Motorcycles (motorlance) are often deployed as ambulances to the scene of an emergency to reduce response time. The COVID-19 pandemic has affected emergency medical services (EMS) in Thailand in many respects, and this study was conducted to examine its effect on motorlance operation time. AIM: The aim of the study was to examine motorlance operation time during the COVID-19 pandemic in comparison to normal periods. METHODS: This cross-sectional study examined all EMS motorlance operations dispatched from Srinagarind Hospital (Thailand). Data were collected from the Srinagarind Hospital EMS operation database and hospital information database system. Data from June 1, 2018, to December 31, 2019 (normal period) were compared with those from January 13 to April 21, 2020 (COVID-19). RESULTS: Eight hundred seventy-one EMS operations were examined over two periods. Mean patient age during the COVID-19 pandemic was 41.5 ± 6.2 years, and 54.6% (n = 59) were male. Average response time was 6.20 ± 1.35 min during the normal period and 3.48 ± 1.01 min during the pandemic (p = 0.021). Transport time was also significantly shorter during the latter period (2.35 vs. 5.20 min). CONCLUSIONS: Motorlance response and transport time during the COVID-19 pandemic were significantly shorter than usual.

Aim of the study Intraductal papillary neoplasm of the bile duct (IPNB) can present at various stages of the disease. Each stage needs different treatment. The neutrophil-to-lymphocyte ratio (NLR) and the platelet-to-lymphocyte ratio (PLR) have been described as predictive markers for several tumors. There has been no investigation on the role of NLR and PLR in IPNB. Material and methods We retrospectively reviewed the medical records of 112 patients who underwent curative-intent hepatic resection for IPNB between January 2007 and December 2011. All clinical parameters and survival were analyzed for their association with NLR and PLR. Results For prediction of malignancy, the best respective cut-off for NLR and PLR was 2.74 and 130, with area under the ROC curve being 0.662 and 0.763. For micro-papillary IPNB, both markers well predict malignancy and lymph node involvement. The respective area under the ROC curve of NLR and PLR for prediction of malignancy was 0.78 and 0.88. Both markers had an area under the ROC curve for prediction of lymph node involvement of 1.0. The median overall survival of those with PLR < 130 was 86.4 months compared with 45.0 months for those with PLR > 130 (p = 0.02). Conclusions NLR and PLR seem likely candidates for predicting malignancy, lymph node involvement, and survival of the patients. PLR performed better than NLR for all predictions. The markers worked very well for micro-papillary IPNB; however, we recommend using these markers in conjunction with the radiologic appearance of tumors.

Background Cholangiocarcinoma (CCA) is a leading cause of cancer death in northeastern Thailand. We reported on the incidence of CCA using only one method. In the current study, we used three different statistical methods to forecast future trends and estimate relative survival. Methods We reviewed the CCA cases diagnosed between 1989 and 2018 recorded in the population-based Khon Kaen Cancer Registry (KKCR). Annual percent change (APC) was calculated to quantify the incidence rate trends using Joinpoint regression. Age-period-cohort models (APC model) were used to examine the temporal trends of CCA by age, calendar year, and birth cohort. We projected the incidence of CCA up to 2028 using three independent approaches: the Joinpoint, Age-period-cohort, and Nordpred models. Survival assessments were based on relative survival (RS). Results The respective APC in males and females decreased significantly (-3.1%; 95%CI: -4.0 to -2.1 and -2.4%; 95%CI: -3.6 to -1.2). The APC model—AC-P for male CCA—decreased according to a birth-cohort. The CCA incidence for males born in 1998 was 0.09 times higher than for those born in 1966 (Incidence rate ratios, IRR = 0.09; 95%CI: 0.07 to 0.12). The relative incidence for female CCA similarly decreased according to a birth-cohort (IRR = 0.11; 95%CI: 0.07 to 0.17). The respective projection for the age-standardized rate for males and females for 2028 will be 7.6 per 100,000 (102 patients) and 3.6 per 100,000 (140 patients). The five-year RS for CCA was 10.9% (95%CI: 10.3 to 11.6). Conclusion The incidence rate of CCA has decreased. The projection for 2028 is that the incidence will continue to decline. Nevertheless, the survival of patients with CCA remains poor.

BACKGROUND: The use of lights and sirens (L&S) alerts other drivers of the presence of an ambulance and that they are required yield, increasing the speed and safety of emergency medical services (EMS) operations. However, there have been no studies examining the effect of L&S on pre-hospital time conducted in Thai EMS agencies. AIM: The aim of the study was to compare the operation times of ambulances with and without the use of L&S. METHODS: This was a cross-sectional study consisting of patients over 18 years of age assessed and treated through the Srinagarind Hospital EMS between April 2019 and March 2020. Data were collected from the Srinagarind Hospital EMS operation database and hospital information database system. RESULTS: A total of 1764 patients were enrolled, 1426 (80.8%) of whom were transported in an ambulance that used L&S. The mean age of patients in the L&S group was 45.2 ± 6.2 years and 742 (52.0%) were male. The average response times in the L&S and non-L&S groups were 10.2 min and 18.2 min, respectively (p < 0.001). Average L&S transport time was 11.1 min and non-L&S transport time was 17.1 min (p = 0.008). CONCLUSIONS: The use L&S reduced the response and transport times of EMS operations but not affect on-scene time.

Background. Major hepatectomy is the mainstay of the treatment for cholangiocarcinoma. Infrahepatic inferior vena cava (IVC) clamping is an effective maneuver for reducing blood loss during liver transection. The impact of this procedure on major hepatectomy for cholangiocarcinoma is unknown. This study evaluated the effect of infrahepatic IVC clamping on blood loss during liver transection. Methods. Clinical and pathological data were collected retrospectively for 116 cholangiocarcinoma patients who underwent major hepatectomy between January 2015 and December 2016, to investigate the benefit of infrahepatic IVC clamping. Two of five surgeons adapted the policy performing infrahepatic IVC clamping during liver transection in all cases. Patients, therefore, were divided into those ( n = 39 ; 33.6%) who received infrahepatic IVC clamping during liver transection (C1) and those ( n = 77 ; 66.4%) who did not (C0). Results. The patients’ backgrounds, operative parameters, and extent of hepatectomy did not differ significantly between the 2 groups, except for gender. A significantly lower blood loss ( p = 0.028 ), blood transfusion ( p = 0.011 ), and rate of vascular inflow occlusion requirement ( p < 0.001 ) were observed in the C1 group. The respective blood losses in the C1 group and the C0 group were 498.9 (95% CI: 375.8-622.1) and 685.6 (95% CI: 571-800.2) millilitres. Conclusions. The current study found infrahepatic IVC clamping during liver transection for cholangiocarcinoma reduces blood loss, blood transfusion, and rate of vascular inflow occlusion requirement.

BACKGROUND: COVID-19 outbreaks occurring in many countries around the world have had a significant impact on emergency medical services (EMS) in terms of the number of operations and procedures performed, including those on ambulances. However, the number of EMS amid the rising number of COVID-19 cases in Khon Kaen, Thailand has not been well studied. AIM: The aim of this study was to identify and analyze the relationship between the use of EMS and the outbreak of COVID-19. METHODS: A cross-sectional study was done in Srinagarind Hospital, Khon Kaen, Thailand with a single EMS centered. Data was collected between March 27, 2021 and April 9, 2021 and designated as (Day-7 to Day 7 of pandemic services) as well as normal services which collected data between March 27 and April 9, 2019 (Day-7 to Day 7 of normal services). RESULTS: Three hundred eight EMS operations were examined. A total of 77.9% (n = 95) of pandemic services were for males and the mean age of the patients was 32.1 ± 5.4 years. During the normal services in 2019, the average number of EMS operations was 13.3 ± 2.4 times per day over the two weeks of the study. In 2021, before the outbreak (Day-7 to Day 1), the average number of EMS operations was 13.2 ± 1.8 times/day. After Day 1, there was a drop in the number of EMS operations which was associated with an increase in COVID-19 infections (p < 0.001). CONCLUSIONS: The number of EMS users during the COVID-19 outbreak decreased considerably compared to the pre-epidemic levels and normal service intervals, including the severity of the users, were more severe than normal.

© 2021 Kamonwon Ienghong, Takaaki Suzuki, Ismet Celebi, Vajarabhongsa Bhudhisawasdi, Somsak Tiamkao, Dhanu Gaysonsiri, Korakot Apiratwarakul.BACKGROUND: B-line artifacts (BLAs) play an important role in identifying lung pathology. They may indicate different diseases. However, the diagnostic study of BLA as applied to emergency patients has not been well studied. AIM: The aim of this study was to determine the diagnostic accuracy of BLA in various conditions. METHODS: This was a retrospective observational study of emergency patients who had received lung ultrasound at Srinagarind Hospital’s Emergency Department throughout January 2020–December 2020. Ultrasound artifacts were recorded. Ultrasonography findings were correlated with final diagnosis. Sensitivity and specificity were also calculated. RESULTS: A total of 105 patients were evaluated. The most prevalent condition which BLA found in this study was pulmonary edema (44.12%) with 88.24% sensitivity and 46.48% specificity. BLA also indicated pneumonia with 66.67% sensitivity and 35.71% specificity. Diffuse BLA indicated pulmonary edema with 70% sensitivity and 70.42% specificity. Focal BLA indicated pneumonia with 28.57% sensitivity and 76.19% specificity. CONCLUSIONS: The sensitivity of BLA for pulmonary edema and pneumonia diagnosis in this study was of moderate to good sensitivity, but low specificity. BLA may become crucial in the diagnosis of lung pathology in the emergency department.

BACKGROUND: Sepsis is a major problem with a high mortality rate, and emergency rooms are the first service point at which sepsis patients are treated. Blood presepsin levels are used as a tool to identify patients who may suffer from sepsis caused by bacterial infection, necessitating immediate antibiotic treatment.AIM: The aim of the study was to evaluate presepsin levels in patients with bacterial and viral infections in the emergency department. METHODS: This was a prospective study consisting of 88 patients over 18 years of age who visited the Srinagarind Hospital emergency room and fulfilled one or more criteria for systemic inflammatory response syndrome. Presepsin levels in the leftover blood specimens were tested. RESULTS: In the bacterial infection group, 63.6% of the patients were male, and the mean age was 43.7 ± 10.5 years. Mean presepsin levels were 2904.73, pg/ml in the bacterial infection group and 204.88 pg/ml in the viral infection group. CONCLUSION: Presepsin levels in the bacterial infection group were significantly higher than in the viral infection group, in which they were below the standard cutoff point.

An effective serum biomarker may improve cholangiocarcinoma (CCA) management. Periostin (PN) has been demonstrated to be associated with aggressive CCA. The current study evaluated PN in blood serum for its diagnostic and prognostic potential in patients with CCA. Sera of 68 patients with CCA were collected prior to treatment, and PN levels were measured using an ELISA. Sera from 50 normal controls, 6 patients with benign liver diseases, 2 with hepatocellular carcinoma and 21 with breast cancer were analyzed. Immunohistochemistry of PN in CCA tissues was also investigated. The data were analyzed using the Mann-Whitney U test, Kaplan-Meier log rank tests, Cox proportional hazard regression models and Fisher's exact tests. The median serum PN level in patients with CCA was significantly increased compared with that in healthy controls, patients with benign liver diseases and patients with breast cancer (all P94 ng/ml) accordingly. High serum and tissue PN levels were significantly associated with reduced survival rate (P

Background & Aims Intraductal papillary neoplasm of the bile duct (IPNB) is a specific type of bile duct tumor. Studies about the surgical outcomes for IPNB are few; therefore, we investigated the survival of patients who underwent curative surgical resection of IPNB. Methods We retrospectively reviewed the medical and pathological records of 148 IPNB patients who underwent curative-intent hepatic resection between January 2005 and December 2011, to examine the prognosis of IPNB. All demographic and operative parameters were analyzed the effect on survival of patients. Results: The median survival of IPNB patients was 1,326 days with a respective 1, 3, and 5 year overall survival of 83.6% (95%CI:76.5-88.7), 64.4% (95%CI:56.0-71.6), and 47% (95%CI:38.4-55.7). The level of invasiveness of IPNB predicted survival very well. For malignant IPNB, univariate analysis showed that serum CA19-9 level, lymph node metastasis, and completeness of resection were significant prognostic factors. Lymph node metastasis and completeness of resection were found in multivariate analysis to be significantly related to survival of the patients. Conclusions The level of invasiveness and lymph node status were found to be associated with patient survival, as was adequacy of surgery. We recommend R0 resection be attempted for patients with IPNB.

Background: Liver cancer is the second leading cause of cancer-related deaths worldwide. With a predicted 2.4-fold rise in liver cancer incidence by 2020, there is an urgent need for early, inexpensive diagnostic biomarkers to deploy in the clinic. Methods: We employed ultraperformance liquid chromatography tandem mass-spectrometry (UPLC/MS-MS) for the quantitation of four metabolites, creatine riboside (CR), N-acetylneuraminic acid (NANA), cortisol sulfate, and a lipid molecule designated as 561+, in urine samples from the NCI-MD cohort comprising 98 hepatocellular carcinoma (HCC) cases, 101 high-risk subjects, and 95 controls. Validation was carried out in the TIGER-LC cohort [n = 370 HCC and intrahepatic cholangiocarcinoma (ICC) cases, 471 high-risk subjects, 251 controls], where ICC, the second most common primary hepatic malignancy, is highly prevalent. Metabolite quantitation was also conducted in TIGER-LC tissue samples (n = 48 ICC; n = 51 HCC). Results: All profiled metabolites were significantly increased in liver cancer when compared with high-risk subjects and controls in the NCI-MD study. In the TIGER-LC cohort, the four-metabolite profile was superior at classifying ICC than a clinically utilized marker, CA19-9, and their combination led to a significantly improved model (AUC = 0.88, P = 4E-8). Metabolites CR and NANA were significantly elevated in ICC when compared with HCC cases in both urine and tissue samples. High levels of CR were associated with poorer prognosis in ICC. Conclusions: Four metabolites are significantly increased in HCC and ICC and are robust at classifying ICC in combination with the clinically utilized marker CA19-9. Impact: Noninvasive urinary metabolite biomarkers hold promise for diagnostic and prognostic evaluation of ICC.

Introduction and aim. The carcinogenesis of tubular and papillary cholangiocarcinoma (CCA) differ. The available epidemiologic studies about risk factors for CCA do not differentiate between the tubular and papillary type. The current study investigated the relationship between the number of repeated use of Praziquantel (PZQ) treatments and each type of CCA. Material and methods. This was a hospital-based, matched, case-control study of patients admitted to Srinagarind Hospital, Khon Kaen University. The patients were 210 pathologically-confirmed cases of CCA, while the controls were 840 subjects diagnosed with other diseases. The 4 controls were individually matched with each case by sex, age, and date of admission. The cases were classified according to location (intrahepatic vs. extrahepatic) and cell type (papillary vs. tubular). Multivariable conditional logistic regression was used for the analysis. Results. After adjusting for confounders, there were statistically significant associations between intrahepatic and papillary CCA and repeated use of PZQ treatment. The respective odds of developing intrahepatic CCA for those who used PZQ once, twice, or more was 1.54 (95%CI:0.92-2.55 ), 2.28 (95%CI:0.91-5.73), and 4.21 (95%CI:1.61-11.05). The respective odds of developing papillary CCA for those who used PZQ once, twice, or more was 1.45 (95%CI:0.80-2.63), 2.96 (95%CI:1.06-8.24), and 3.24 (95%CI:1.09-9.66). There was no association between number of uses of PZQ treatment and developing extrahepatic or tubular CCA. Conclusion. The current study found an association between papillary and intrahepatic CCA and repeated use of PZQ treatment. We suggest further study on the risk factors for papillary and tubular CCA should be performed separately.

Intrahepatic cholangiocarcinoma (ICC) is an aggressive cancer, arising in the biliary ducts that extend into the liver. The highest incidence of ICC occurs in Southeast Asia, particularly in the Mekong River Basin countries of Thailand, Laos, Cambodia, and Vietnam, where it is strongly associated with chronic infection by the food-borne liver fluke Opisthorchis viverrini (OV), one of only three eukaryote pathogens considered Group one carcinogens. Intrahepatic cholangiocarcinoma is usually diagnosed at an advanced stage, with a poor prognosis and survival often less than 24months. Hence, biomarkers that enable the early detection of ICC would be desirable and have a potentially important impact on the public health in the resource-poor regions where this cancer is most prevalent. As microRNAs (miRNAs) remain well preserved after formalin fixation, there is much interest in developing them as biomarkers that can be investigated using tumor biopsy samples preserved in formalin fixed paraffin embedded (FFPE) tumor blocks. Recently, we reported the first comprehensive profiling of tissue-based miRNA expression using FFPE from the three most common subtypes of OV-induced ICC tumors: moderately differentiated ICC, papillary ICC, and well-differentiated ICC. We observed that each subtype of OV-induced ICC exhibited a distinct miRNA profile, which suggested the involvement of specific sets of miRNAs in the progression of this cancer. In addition, non-tumor tissue adjacent to ICC tumor tissue on the same FFPE block shared a similar miRNA dysregulation profile with the tumor tissue than with normal (non-tumor) liver tissue (individuals without ICC or OV infection). Herein, we provide a detailed description of the microarray analysis procedures used to derive these findings.

Background & Aims Intrahepatic cholangiocarcinoma (ICC) is a significant public health problem in East Asia, where it is strongly associated with chronic infection by the food-borne parasite Opisthorchis viverrini (OV). We report the first comprehensive miRNA expression profiling by microarray of the most common histologic grades and subtypes of ICC: well differentiated, moderately differentiated, and papillary ICC. Methods MicroRNA expression profiles from FFPE were compared among the following: ICC tumour tissue (n=16), non-tumour tissue distally macrodissected from the same ICC tumour block (n=15), and normal tissue (n=13) from individuals undergoing gastric bypass surgery. A panel of deregulated miRNAs was validated by qPCR. Results Each histologic grade and subtype of ICC displayed a distinct miRNA profile, with no cohort of miRNAs emerging as commonly deregulated. Moderately differentiated ICC showed the greatest miRNA deregulation in quantity and magnitude, followed by the papillary subtype, and then well differentiated ICC. Moreover, when ICC tumour tissues were compared to adjacent non-tumour tissue, similar miRNA dysregulation profiles were observed. Conclusions We show that common histologic grades and subtypes of ICC have distinct miRNA profiles. As histological grade and subtypes are associated with ICC aggressiveness, these profiles could be used to enhance the early detection and improve the personalised treatment for ICC. These findings also suggest the involvement of specific miRNAs during ICC tumour progression and differentiation. We plan to use these insights to (a) detect these profiles in circulation and (b) conduct functional analyses to decipher the roles of miRNAs in ICC tumour differentiation.

Cholangiocarcinoma is the most common cancer in the northeast of Thailand. Most of the patients present when the disease is in an advanced stage. Improvement of preoperative diagnoses and surgical techniques provide more satisfactory results. Herein we reviewed our 30-year experience in management of perihilar cholangiocarcinoma in Khon Kaen northeast Thailand. Between 1982 and 2012 we reviewed four specific studies of perihilar cholangiocarcinoma in Srinagarind Hospital, Khon Kaen, Thailand. The first study focused on advanced surgical pathology and palliative surgery, which were used to treat obstructive jaundice cholangiocarcinoma patients. Long-term survival in this study was rare with a one-year survival of just 15%. The second study was conducted on 30 consecutive cases of perihilar cholangiocarcinoma who presented with obstructive jaundice without preoperative biliary drainage. All the patients underwent major liver resection with bilio-enteric reconstruction. Perioperative mortality was 6.7% without a 5-year survival. The third study aimed to analyze the survival rates and factors affecting survival in extrahepatic CCA patients following surgical treatment at Srinagarind Hospital and concluded that resection margins are an important prognostic factor. The last study objective was the analysis of curative surgical attempt in 99 consecutive perihilar cholangiocarcinoma and results showed that R0 resection could improve long-term survival. We evaluated four studies of perihilar cholangiocarcinoma in Srinagarind Hospital, Khon Kaen, Thailand from 1982-2012. Viewed chronologically there has been a progressive improvement of diagnosis and surgical treatment during the past 30 years. Despite these advances the 5-year survival rate remains unsatisfactorily low. Future improvement of patient selection and surgical techniques can lead to a greater survival rate for patients.

Purpose To determine whether FOLFOX4 (infusional fluorouracil, leucovorin, and oxaliplatin) administered as palliative chemotherapy to patients with advanced hepatocellular carcinoma (HCC) provides a survival benefit and efficacy versus doxorubicin. Patients and Methods This multicenter, open-label, randomized, phase III study in mainland China, Taiwan, Korea, and Thailand involved 371 patients age 18 to 75 years who had locally advanced or metastatic HCC and were ineligible for curative resection or local treatment. They were randomly assigned at a ratio of one to one to receive either FOLFOX4 (n = 184) or doxorubicin (n = 187). The primary end point was overall survival (OS); secondary end points included progression-free survival (PFS), response rate (RR) by RECIST (version 1.0), and safety. Results At the prespecified final analysis, median OS was 6.40 months with FOLFOX4 (95% CI, 5.30 to 7.03) and 4.97 months with doxorubicin (95% CI, 4.23 to 6.03; P = .07; hazard ratio [HR], 0.80; 95% CI, 0.63 to 1.02). Median PFS was 2.93 months with FOLFOX4 (95% CI, 2.43 to 3.53), and 1.77 months with doxorubicin (95% CI, 1.63 to 2.30; P < .001; HR, 0.62; 95% CI, 0.49 to 0.79). RR was 8.15% with FOLFOX4 and 2.67% with doxorubicin (P = .02). On continued follow-up, the trend toward increased OS with FOLFOX4 was maintained (P = .04; HR, 0.79; 95% CI, 0.63 to 0.99). Toxicity was consistent with previous experiences with FOLFOX4; proportions of grade 3 to 4 adverse events were similar between treatments. Conclusion Although the study did not meet its primary end point, the trend toward improved OS with FOLFOX4, along with increased PFS and RR, suggests that this regimen may confer some benefit to Asian patients, but an OS benefit cannot be concluded from these data.

Background: Cholangiocarcinoma is relatively rare worldwide. Most previous reports collected only patients with pathological diagnosis. In fact, however, many patients coming to hospital are diagnosed by clinical suspicion with radiologic imaging and receive treatment without histological confirmation. Real survival data and outcome of each treatment, especially for patients that do not have histologic confirmation, are lacking. In this study, therefore, we aimed to analyze the survival rates of CCA patients and the proportions of patients receiving different treatments. Materials and Methods: A total of 270 patients clinically suspected of CCA and visiting Srinagarind Hospital in May-July 2010, were prospectively followed until December 2014. After checking their clinical records, 163 of 270 patients were finally diagnosed as having CCA, and the data of this group were analyzed for survival rate and received treatments. Results: Of the 163 patients, 96 (58.9%) had intrahepatic, 56 (34.4%) had perihilar and 11 (6.7%) had distal CCA. The majority [107 (65.6%, 95%CI, 57.8-73.0)] received only supportive care. Overall median survival was 4 months (95%CI, 3.3-4.7), and 2-years survival was only 8.1% (95%CI,4.5-12.9). However, the 4 year survival of the R0 resection group was 100%. Conclusions: The present results show that the prognosis of CCA is very poor in North-east Thailand. Most CCA patients receive only treatment to alleviate symptoms due to their advanced stage of disease. Complete surgical resection at the early stage is the only treatment that significantly improves patient survival.

Intraductal papillary neoplasm of the bile duct (IPNB) is a specific type of bile duct tumor. It has been proposed that it could be the biliary counterpart of the intraductal papillary neoplasm of the pancreas (IPMN-P). This hypothesis is supported by the presence of simultaneous intraductal tumors of both the bile duct and pancreas. There have been five reports of patients with simultaneous IPNB and IPMN-P. In all of these cases, biliary involvement was limited to the intrahepatic and perihilar bile duct, which had characteristics similar to IPMN-P and usually had slow progression in nature. Herein, we present the first case of extensive intraductal neoplasm involving the extrahepatic bile duct, intrahepatic bile duct, and entire length of the pancreas with a poor outcome, even after being treated aggressively with radical surgery and adjuvant chemotherapy. Additionally, we summarize previous case reports of simultaneous intraductal lesions of the bile duct and pancreas.

Objectives: To investigate the association of expression status of alpha-enolase (ENO1) and clinicopathological outcomes of CCA patients. Design and methods: Two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) were used to compare differential expressed protein profiles of four human CCA cell lines and H69, a non-malignant biliary cell line, as a control. Immunohistochemical analysis was carried out in tissue-microarray of human CCA tissues (n=301). Results: We identified ENO1 in all CCA cell lines but not H69 by proteomics based. About 75% of patients with CCA showed over-expression of ENO1 in hyperplastic bile duct and the tumors compared with that in tumor-adjacent normal tissue counterparts. Moreover, over-expression of ENO1 is significantly associated with poor prognosis and tumor invasion of CCA patients. Conclusions: ENO1 may serve as a prognostic marker to monitor the disease progression of these patients.

We performed a comparative proteomic analysis of protein expression profiles in 4 cholangiocarcinoma cell lines: K100, M156, M213, and M139. The H69 biliary cell line was used as a control. Peroxiredoxin 1 and ezrin-radixin-moesin-binding phosphoprotein 50 were selected for further validation by immunohistochemistry using a cholangiocarcinoma tissue microarray (n = 301) to assess their prognostic value in this cancer. Both peroxiredoxin 1 and ezrin-radixin-moesin-binding phosphoprotein 50 were overexpressed in cholangiocarcinoma tissues compared with normal liver tissues. Of the 301 cholangiocarcinoma cases, overexpression of peroxiredoxin 1 in 103 (34.3%) was associated with an age-related effect in young patients (P = .011) and the absence of cholangiocarcinoma in lymphatic vessels and perineural tissues (P = .004 and P = .037, respectively). Expression of radixin-moesin-binding phosphoprotein 50 correlated with histopathologic type, with 180 (59.8%) of moderately or poorly differentiated tumors (P = .039) being higher, and was associated with the presence of cholangiocarcinoma in lymphatic and vascular vessels (P < .001 and P < .001, respectively). The high expression of radixin-moesin-binding phosphoprotein 50 and the low expression of peroxiredoxin 1 correlated with reduced survival by univariate analysis (P = .017 and P = .048, respectively). Moreover, the impact of peroxiredoxin 1 and radixin-moesin-binding phosphoprotein 50 expression on patient survival was an independent predictor in multivariate analyses (P = .004 and P = .025, respectively). Therefore, altered expression of peroxiredoxin 1 and radixin-moesin-binding phosphoprotein 50 may be used as prognostic markers in cholangiocarcinoma.

Cholangiocarcinoma is a crucial health problem in northeast Thailand. Although rare, it is a highly fatal disease and the prognosis of CCA patients is very poor. To determine if expression of specific genes is useful for diagnosis and prognosis for CCA. We examined the expression of HSP70, HSP90, RB1, cyclin D1, and HDAC6 in 50 resections of human CCA tissues by quantitative real-time PCR. The expression of HSP70, RB1, and HDAC6 was "dominant down-regulation," while the expression of cyclin D1 and HSP90 was "dominant up-regulation." There were no correlations between RB1, cyclin D1, HSP90, and clinicopathological parameters such as status, histology type, histological grading, stage of CCA, and metastasis. A significant association was found between HDAC6 and CCA staging (p = 0.000), CCA gross type and HSP70 (p = 0.046) as well as RB1 expression (p = 0.046). Patients with down-regulation of HSP70 had significantly poorer prognosis than those in the up-regulation group (p = 0.002). Expression of HSP70 may be useful as a new prognostic marker for CCA.

The retinoblastoma interacting zinc finger (RIZ1) gene is adjacent to D1S228 where microsatellite instability has been associated with poor patient survival in liver fluke- associated cholangiocarcinoma (CCA). An understanding of the molecular mechanisms underlying the carcinogenesis and pathogenesis of CCA is necessary to improve patient survival. Therefore, we determined the genetic and epigenetic alterations of RIZ1 in 81 CCA samples and 69 matched non-tumor tissues. Methylation was found in 31 of 81 (38%) tumor samples and in 5 of 69 (7%) matched non-tumor tissues. Frameshift mutations (2 of 81) and loss of heterozygosity (LOH) (14 of 81) were not common. Statistical analysis found no significant correlation between RIZ1 alterations and clinicopathological features, but RIZPro704 LOH was associated with patient survival in the multivariate analysis. RIZ1 hypermethylation may be one of the crucial molecular events contributing to cholangiocarcinogenesis, and RIZPro704 LOH may adversely impact patient survival. The biological function of RIZ1 in CCA should be further investigated in order to verify its potential role in regulating this cancer.

Background Infection by the liver fluke, Opisthorchis viverrini, remains an important public health problem in Thailand and has resulted in the highest prevalence of infection and incidence of subsequent cholangiocarcinoma (CCA) in the world. Praziquantel (PZQ) is the antihelminthic drug of choice for treatment. Previous studies in hamsters showed that repeated infection and PZQ treatment could increase the risk of CCA. However, the few available epidemiology studies in humans have shown unclear evidence of an increased risk of CCA with frequency of PZQ intake. The present study investigated the relationship between the number of repeated PZQ treatments and CCA. Methods A hospital-based matched case–control study was conducted. All cases and controls were inpatients of a tertiary hospital in Northeast Thailand. During 2012–2014 a total of 210 incident cases of pathologically diagnosed CCA and 840 control subjects were selected from a hospital inpatient database (four controls per case). The four recruited controls were individually matched with CCA cases by gender, age and date of admission. Data were collected in face-to-face interviews using a standardised pre-tested questionnaire. Multivariable conditional logistic regression was used in the analysis of the data. Results The frequencies of PZQ usage among the 210 cases and 840 controls were 48.6 vs. 66.0 for never, 32.9 vs. 24.4 for once, 8.6 vs. 4.9 for twice, and 10.0 % vs. 4.8 % for more than twice, respectively. There was a statistically significant dose–response relationship (p

BACKGROUND The authors recently showed that MUC5AC mucin, which is expressed aberrantly in tumor tissue, is present in significant concentrations in serum from patients with cholangiocarcinoma. Subsequently, determination of serum MUC5AC had high sensitivity and specificity for cholangiocarcinoma. In this study, the possible association between serum MUC5AC mucin and the clinical findings of the patients and their prognostic value were explored. METHODS The expression of MUC5AC mucin in serum samples from 179 patients with histologically confirmed cholangiocarcinoma were determined using immunoblotting. RESULTS Detection of serum MUC5AC was associated with patients with blood group Type A, larger-sized tumors (> 5 cm), and advanced-stage disease. Patients who had positive serum MUC5AC status had a significantly poorer prognosis (median survival, 127 days; 95% confidence interval [95% CI], 107–180 days) compared with patients who had negative serum MUC5AC status (median survival, 329 days; 95% CI, 199–458 days; P < 0.001). Multivariate analysis with adjustment for all covariates showed that patients who had positive serum MUC5AC status had a 2.5-fold higher risk of death compared with patients who had negative serum MUC5AC status (P < 0.001). CONCLUSIONS Serum MUC5AC was associated with tumor burden. The determination of serum MUC5AC may be predictive of poor patient outcome and may be useful in selecting possible treatment options for patients with cholangiocarcinoma. Cancer 2003;98:1438–43. © 2003 American Cancer Society. DOI 10.1002/cncr.11652

Background: High levels of serum total sialic acid (TSA) have been reported in cholangiocarcinoma (CCA) patients. In this study, the clinical value and possible cause of increased total sialic acid in the serum in cholangiocarcinoma patients were examined. Methods: Total sialic acid was determined in 172 serum and 25 tumor tissue samples taken from cholangiocarcinoma patients using the periodate thiobarbituric acid method. Results: The total sialic acid content of the tumor tissue was significantly greater than that of the serum and not related to the concentration found in the serum. The serum total sialic acid was not correlated with age, sex, body mass index, blood group, tumor location, tumor stage, metastatic condition, histological types and survival of the patients. The increased total sialic acid in the serum had a significant correlation with serum MUC5AC mucin, alkaline phosphatase and the CA19-9, and the numbers of white blood cell and neutrophils. Conclusions: The concentration of serum sialic acid was not associated with clinicopathologic features or the tumor burden. The glycoproteins secreted from the tumor and inflammatory cells might be responsible for the increased total sialic acid in the serum in these patients.

Hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC) are two distinct histological liver cancers. They are clinically and biologically heterogeneous and highly resistant to treatment, making liver cancer the second most lethal malignancy in the world. In Thailand, liver cancer represents the primary cause of cancer-related death and is a major health problem. While HBV and HCV are major etiological factors for HCC globally, liver fluke infection (O. viverrini) is a major etiological factor for ICC in Thailand, especially in north-eastern Thailand where O. viverrini is endemic and approximately 70% of liver cancers are ICC. These unique risk factor patterns provide an opportunity to study cancer heterogeneity and unique liver tumor biology. The Thailand Initiative in Genomics and Expression Research for Liver Cancer (TIGER-LC) consortium was established to identify genomic and expression factors that may modify HCC and ICC susceptibility and progression. Here, we determined molecular subtypes and features of HCC and ICC through systems integration of genomic, transcriptomic and metabolic profiles. We performed genome wide profiling of 398 surgical specimens derived from 199 Thai liver cancer patients. We employed the Affymetrix Human Transcriptome Array 2.0, the Affymetrix Genome-Wide Human SNP Array 6.0, Metabolon's DiscoveryHD4 platform and Exome Sequencing to examine transcriptome profiles, somatic copy number alterations (SCNA), cancer metabolic profiles and mutation patterns, respectively. The results were validated in 847 independent Asian or Caucasian HCC or ICC cases. Transcriptomic analyses revealed that Thai HCC consisted of 3 stable subgroups (C1-C3), while Thai ICC contained 4 stable subgroups (C1-C4). Interestingly, HCC-C1 and ICC-C1 subtypes shared a similar gene expression matrix, as did HCC-C2 and ICC-C2, which correlated with patient survival. These prognostic subtypes were validated in independent Asian HCC and ICC cohorts, but not in Caucasian patients, and were associated with tumor biology rather than etiology. GSEA revealed that the C1 subtype is enriched for mitotic checkpoint anomalies, while the C2 subtype is related to cytokine and chemokine signaling. We found that the C1 subtype encompassed a higher degree of SCNA when compared to the C2 subtype, suggesting an association with a genomic instability phenotype. Further analysis showed that the C2 subtype is linked to an increased body mass index, inflammatory responses and unique tumor metabolic activities. HCC and ICC from Asian populations, while clinically treated as separate entities, share common subtypes with similar actionable drivers which can be targeted to improve precision therapy. Citation Format: Anuradha Budhu, Jittiporn Chaisaingmongkol, Hien Dang, Siritida Rabibhadana, Benjarath Pupacdi, So Mee Kwon, Marshonna Forgues, Yotsawat Pomyen, Vajarabhongsa Bhudhisawasdi, Nirush Lertprasertsuke, Anon Chotirosniramit, Chawalit Pairojkul, Chirayu U. Auewarakul, Thaniya Sricharunrat, Kannika Phornphutkul, Suleeporn Sangrajrang, Maggie Cam, Ping He, Stephen M. Hewitt, Xiaolin Wu, Snorri S. Thorgeirsson, Joshua J. Waterfall, Yuelin J. Zhu, Jennifer Walling, Holly S. Stevenson, Daniel Edelman, Paul S. Meltzer, Christopher A. Loffredo, Robert H. Wiltrout, Curtis C. Harris, Chulabhorn Mahidol, Mathuros Ruchirawat, Xin W. Wang. The Thailand initiative in genomics and expression research in liver cancer: Race related common molecular subtypes among Asian hepatocellular carcinoma and cholangiocarcinoma identified by integrated genomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4390. doi:10.1158/1538-7445.AM2017-4390

Background Intrahepatic cholangiocarcinoma (ICC) is a highly aggressive tumor of the bile duct, and a significant public health problem in East Asia, where it is associated with infection by the parasite Opisthorchis viverrini. ICC is often detected at an advanced stage and with a poor prognosis, making a biomarker for early detection a priority. Methods We have comprehensively profiled miRNA expression levels in ICC tumor tissue using small RNA-Seq and validated these profiles using quantitative PCR on matched plasma samples. Results Distinct miRNA profiles were associated with increasing histological differentiation of ICC tumor tissue. We also observed that histologically normal tissue adjacent to ICC tumor displayed miRNA expression profiles more similar to tumor than liver tissue from healthy donors. In plasma samples, an eight-miRNA signature associated with ICC, regardless of the degree of histological differentiation of its matched tissue, forming the basis of a circulating miRNA-based biomarker for ICC. Conclusions The association of unique miRNA profiles with different ICC subtypes suggests the involvement of specific miRNAs during ICC tumor progression. In plasma, an eight-miRNA signature associated with ICC could form the foundation of an accessible (plasma-based) miRNA-based biomarker for the early detection of ICC. Electronic supplementary material The online version of this article (doi:10.1186/s12885-015-1270-5) contains supplementary material, which is available to authorized users.

Objectives: This study was undertaken to establish the diagnostic utility of serum total sialic acid (TSA) for patients with cholangiocarcinoma (CCA). Design and methods: Serum TSA was determined in 89 histologically confirmed CCA patients, 38 with benign hepatobiliary diseases (BHD) and 43 healthy persons. To check whether the test could adequately discriminate between these groups, complete statistical Receiver Operating Characteristic (ROC) curves were analyzed. Results: The mean value of serum TSA in CCA patients (2.75 ± 0.67 mmol/L) was significantly higher than that in the BHD (2.33 ± 0.69 mmol/L p < 0.002) and healthy persons (1.89 ± 0.46 mmol/L p < 0.001) groups. The areas under the ROC curves were 0.6699 and 0.8558, respectively. A cut-off value of 2.33 mmol/L discriminated between the CCA, BHD and healthy groups with a sensitivity of 71.9% and a positive predictive value range of 80 to 89%. Conclusion: Determination of TSA yielded high diagnostic values for differentiating between CCA, BHD and healthy persons. The determination of serum TSA would be most useful as an adjunct diagnosis rather than an early detection and screening tool because of the apparent nonspecificity of SA to a given disease.

Intrahepatic cholangiocarcinoma is defined as adenocarcinoma originating from bile ductules and segmental and lobar intrahepatic ducts. Four types of surgical pathology have been identified in the Khon Kaen endemic area in Thailand: peripheral, type I; intermediate, type II; central, type III; and diffuse, type IV. We report our experience with intrahepatic cholangiocarcinoma with emphasis on the surgical pathology, operative procedure, and associated survival time. We reviewed the records of patients treated for cholangiocarcinoma at Srinagarind Hospital from January 1, 1992 to February 28, 1997. There was a total of 411 patients, and 138 were intrahepatic and non-jaundiced. Tumors in the proximity of the gray zone i.e., portal, periportal with jaundice, were excluded. Patient profiles, surgical pathology, operative procedure, postoperative morbidity, and mortality were recorded. The data were analyzed using Kaplan-Meier survival curves. Of the 138 patients with intrahepatic disease who were non-jaundiced, 116 had type I, 10 had type II-III, and 12 had type IV. The wear ages of the patients were: 53.0, SE 9.2 years in type I; 57.1, SE 4.6 years in type II-III, and 50.2, SE 9.2 years in type IV. The male-to-female ratios in the three groups were 1.4 : 1, 1.5 : 1, and 5 : 1, respectively. The mean survival times in the three groups were 556, SE 63 days 374, SE 149 days and 97, SE 35 days. Most of the surgical procedures were tumor excisions (108/138). Right hepatectomy was performed in 63 patients, extended right hepatectomy in 8, left hepatectomy in 18, and extended left hepatectomy in 1. Palliative procedures were performed in the other patients because tumors were in both lobes. The mean survival time was 582 days (SE, 75), for right lobe surgery; 458 days (SE, 89) for left lobe surgery; and 127 days (SE, 58) for the other procedures. Mean survival time was 1039 days (SE, 201) in tumor stage III, 773 days (SE, 123) in stage IVa, and 382 days (SE, 60) in stage IVb. There were no significant differences in survival time according to age or sex. The results of surgery in type I and type II-III were better than the results in type IV. Survival time after right hepatectomy was better than that after left hepatectomy, although without statistical significance, but survival time was significantly better after both operations than after palliative procedures. The results of surgery according to pathological staging showed that survival time in stage III was better than that in either stage IVa or IVb, but only the difference from stage IVb was significant.

Objectives: To investigate the association of expression status of alpha-enolase (ENO1) and clinicopathological outcomes of CCA patients. Design and methods: Two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) were used to compare differential expressed protein profiles of four human CCA cell lines and H69, a non-malignant biliary cell line, as a control. Immunohistochemical analysis was carried out in tissue-microarray of human CCA tissues (n=301). Results: We identified ENO1 in all CCA cell lines but not H69 by proteomics based. About 75% of patients with CCA showed over-expression of ENO1 in hyperplastic bile duct and the tumors compared with that in tumor-adjacent normal tissue counterparts. Moreover, over-expression of ENO1 is significantly associated with poor prognosis and tumor invasion of CCA patients. Conclusions: ENO1 may serve as a prognostic marker to monitor the disease progression of these patients.

ObjectivesTo investigate whether Helicobacter spp. infection and the cagA of H. pylori are associated with hepatobiliary pathology, specifically biliary inflammation, cell proliferation and cholangiocarcinoma (CCA).MethodsHelicobacter species including H. pylori, H. bilis and H. hepaticus were detected in the specimens using the polymerase chain reaction (PCR). Biliary inflammation of the liver and gallbladders was semi-quantitatively graded on hematoxylin and eosin (H&E)-stained slides. Biliary proliferation was evaluated by immunohistochemistry using the Ki-67-labelling index.ResultsHelicobacter pylori was found in 66.7%, 41.5% and 25.0% of the patients in the CCA, cholelithiasis and control groups (P 0.05), and was absent in the control group. The cagA gene of H. pylori was detected in 36.2% and 9.1% of the patients with CCA and cholelithiasis, respectively (P

Background: Although 5-fluorouracil (5-FU) is the drug of choice for the palliative treatment of cholangiocarcinoma (CCA), resistance to the drug is a therapeutic obstacle. The aim of this study was to explore the mechanisms underlying 5-FU resistance of CCA using cell lines derived from CCA associated with liver fluke infection. Methods: A stepwise exposure was used for inducing 5-FU-resistant CCA cell lines, and the expression of nine genes associated with 5-FU resistance was analyzed using real-time (RT)-PCR. Results: Altered expression of several genes involved in 5-FU resistance in CCA cell lines was observed. The expression levels of almost all target genes investigated including TP, DPD, ENT1, UNG1, TOP2A, BIRC5, TP73 and DeltaNp73 appeared to be significantly altered in these resistant strains. The expression of the TS gene tended to be increased but the fold change was not significantly different from their parental cell lines. UNG1 (a DNA repairing enzyme) and BIRC5 (an apoptotic inhibitor) expressions were increased whereas TP73 (a proapoptotic factor) expression levels decreased concomitantly. Conclusion: Our study showed that increases in UNG1 and BIRC5 expression and concomitant decreases in TP73 expression may be associated with development of acquired 5-FU resistance in CCA lines and their phenotypes.

Hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) represent two major histological cancer subtypes confined within the liver. They are clinically and biologically heterogeneous, and are highly resistant to treatment, which makes them the second most lethal cancer for men in the world. In Thailand, liver cancer represents the primary cause of cancer-related death and is a major health problem, especially in north-eastern area of Thailand where liver fluke (O. viverrini) is endemic and approximately 70% of liver cancers are CCA. While HBV and HCV are major etiological factors for HCC globally, liver fluke infection is a major etiological factor for CCA in Thailand. These unique risk factor patterns provide an opportunity to study cancer heterogeneity and its unique tumor biology. The Thailand Initiative in Genomics and Expression Research for Liver Cancer (TIGER-LC) consortium was established to identify genomic and expression factors that may modify HCC and CCA susceptibility and progression. In a Phase I study, we determined molecular subtypes of HCC and CCA. We performed genomic profiling of 398 surgical specimens derived from 199 liver cancer patients. We employed the Affymetrix Human Transcriptome Array 2.0 to examine transcriptome profiles. Unsupervised Consensus Clustering (cCluster), Subclass Mapping (SM) and Gene Set Enrichment Analysis (GSEA) algorithms were used to analyze transcriptome data. The results were validated in 247 Asian HCC cases and 104 Caucasian CCA cases. We found that the Thai HCC cases consisted of 3 stable subgroups (C1-C3), while the Thai CCA cases contained 4 stable subgroups (C1-C4) based on gene expression patterns determined by cCluster. SM analysis revealed that CCA-C1 and HCC-C1 subtypes shared a similar gene expression matrix, as did CCA-C2 and HCC-C2 for a separate pattern. Interestingly, patients in both CCA-C1 and HCC-C1 had a poor prognosis, while those in CCA-C2 and HCC-C2 had a good prognosis. These prognostic subtypes were validated in an independent Asian HCC cohort but not in a Caucasian CCA cohort. GSEA revealed that among 17 significantly altered canonical pathways in the C1 subtype, 8 are related to mitotic checkpoint signaling. In contrast, the main signaling pathways associated with the C2 subtype were related to cytokine and chemokine signaling. We found that certain mitotic checkpoint genes are highly activated only in C1, but not in the C2 subtype. These results are consistent with the hypothesis that CCA and HCC from Asian populations consist of molecularly-similar tumor subgroups with similar prognostic impacts and unique tumor biology and that the C1 subtype may be sensitive to mitotic checkpoint blockage. Our ability to rigorously classify and validate both HCC and CCA using these tools may represent a new avenue for the development of targeted therapeutic interventions. Citation Format: The TIGER-LC Consortium, Jittiporn Chaisaingmongkol, Anuradha Budhu, Hien Dang, Siritida Rabibhadana, Benjarath Pupacdi, Marshonna Forgues, Vajarabhongsa Bhudhisawasdi, Nirush Lertprasertsuke, Anon Chotirosniramit, Chawalit Pairojkul, Chirayu U. Auewarakul, Thaniya Sricharunrat, Kannika Phornphutkul, Suleeporn Sangrajrang, Maggie Cam, Ping He, Stephen M. Hewitt, Xiaolin Wu, Snorri S. Thorgeirsson, Paul S. Meltzer, Christopher A. Loffredo, Robert H. Wiltrout, Curtis C. Harris, Chulabhorn Mahidol, Mathuros Ruchirawat, Xin W. Wang. The Thailand initiative in genomics and expression research for liver cancer (TIGER-LC): Defining novel subtypes of hepatocellular carcinoma and cholangiocarcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-173. doi:10.1158/1538-7445.AM2015-LB-173

To analyze the DNA copy number of target genes NF2, TIMP3, ST13, TOB2, BIK, and TP and the reference microsatellite markers D22S283, D22S423, and D22S274 mapped on 22q12-qter in liver fluke related cholangiocarcinoma (CCA) and define its correlation with clinical parameters.Quantitative real time PCR (qPCR) was used for determining allelic imbalances in 65 liver fluke related CCA tissues. Statistical correlations between allelic imbalances and clinicopathological parameters, i.e. age, sex, tumor stage, histological type, blood vessel invasion, nerve invasion and lymphatic invasion were evaluated by means of the chi2 test. Cox regression analysis was used for determining patient's survival.Amplifications of the TP (22q13.33), TOB2 (22q13.2-13.31), D22S283 (22q12.3), TIMP3 (22q12.3) and NF2 (22q12.2) were found in 35 (53.8%), 28 (43.1%), 27 (41.5%), 24 (36.9%), and 24 (36.9%), respectively. Losses at the D22S423 (22q13.1-13.2) and BIK (22q13.31) were detected in 26 (40%) and 23 (35.4%), respectively. Significant correlations were observed between lymphatic invasion and allelic losses of BIK (P = 0.025) and D22S283 (P = 0.041). Univariate and multivariate Cox regression analysis revealed D22S283 amplification as an independent predictor of good prognosis (P = 0.006, death hazard ratio = 0.411, 95% CI = 0.217-0.779) and blood vessel invasion as an independent poor prognostic factor (P = 0.042, death hazard ratio = 1.911, 95% CI = 1.022-3.571) in CCA patients.This study provides evidence for the involvement of gene amplification and deletion on chromosome 22q in liver fluke related CCA. This is the first report of D22S283 amplification as an independent indicator of favorable prognosis in liver fluke related CCA.

AIM: To investigate loss of heterozygosity (LOH) and microsatellite instability (MSI) on the chromosomal region 1p36-pter in cholangiocarcinoma (CCA) patients and determine the association between microsatellite alterations and clinicopathological parameters. METHODS: Ten polymorphic microsatellite markers were determined for LOH and MSI using GS-3000 gel scan fragment autoanalyzer. RESULTS: Sixty-eight out of 90 cases (75.6%) showed LOH in one or more loci. LOH was found most frequently at D1S199 (40.0%), D1S507 (34.6%), D1S2845 (30.5%), and D1S2734 (30.1%). MSI was found in 34 of 90 cases (37.8%) at one or more loci. Fine mapping at 1p36 showed two distinctive regions of common loss, which were D1S2845 and the 25.5-cM region between D1S507 and D1S2734, indicating the existence of putative tumor suppressor genes that is likely to play important roles in the development of CCA. Patients with LOH at D1S234 showed less lymphatic invasion (P = 0.017), whereas patients with LOH at D1S2676 exhibited more lymphatic invasion than those without (P = 0.031). LOH at D1S2845 showed a significant correlation with nerve invasion (P = 0.029). Moreover, patients who demonstrated MSI at D1S228 showed a poor prognosis (P = 0.0026). CONCLUSION: Allelic loss plays a major role in microsatellite alterations at chromosome 1p36, which may contribute to carcinogenesis and pathogenesis of liver fluke related CCA and these alterations can be used as molecular prognostic indicators for CCA patients.

AIM: To establish and characterize a new cholangiocarcinoma cell line from a patient living in the Opisthorchis viverrini (O. viverrini) endemic area of Northeast Thailand. METHODS: Fresh liver biopsy and bile specimens were obtained from a 65-year-old Thai woman with cholangiocarcinoma of the porta hepatis. After digestion, the cells were cultured in Ham’s F12 media. The established cell line was then characterized for growth kinetics, cell morphology, imm-unocytochemistry and cytogenetics. Tumorigenicity of the cell line was determined by heterotransplanting in nude mice. RESULTS: The primary tumor was a poorly differentiated tubular adenocarcinoma. Examination of the bile revealed malignant cells with O. viverrini eggs. The cholangioc-arcinoma cell line KKU-100 was established 4 mo after the primary culture-population doubling time was 72 h. KKU-100 possesses compact and polygonal-shaped epithelial cells. Immunocytochemically, this cell line exhibited cytokeratin, EMA, CEA, and CA125, but not α-fetoprotein (AFP), CA19-9, desmin, c-met, or p53. Such protein expressions parallel those of the primary tumor. Cytogenetic analysis identified aneuploidy karyotypes with a modal chromosome number of 78 and marked chromosomal structural changes. Inoculation of KKU-100 cells into nude mice produced a transplantable, poorly differentiated aden-ocarcinoma, similar to the original tumor. CONCLUSION: KKU-100 is the first egg-proven, Opisthorchis-associated cholangiocarcinoma cell line, which should prove useful for further investigations of the tumor biology of this cancer.

Measurement of the activities of alkaline phosphatase isoenzyme has been used for the identification and monitoring of diseases associated with the isoenzyme. Biliary alkaline phosphatase (BALP), an isoform of liver-ALP, has been found in the serum of patients with biliary obstruction and metastatic liver cancer. This study compared the BALP isoform in the serum of patients with cholangiocarcinoma (CCA) with that of non-jaundiced benign hepatobiliary disease, other cancers, and healthy persons. ALP isoforms were separated using cellulose acetate electrophoresis and the activity was demonstrated using indolyl blue reagent. The BALP isoform was demonstrated in 65% of CCA patients independently of jaundice condition or histological grading of the tumor. The level of serum BALP in non-jaundiced CCA was significantly lower than that of jaundiced CCA, and not correlated with serum bilirubin. No BALP was detected in healthy persons. In the patients with high serum ALP (>147U/l), BALP can differentiate non-jaundiced CCA patients from other non-jaundiced carcinoma patients with 85% sensitivity, 79% specificity, and positive and negative predictive values of 81% and 83%, respectively. The demonstration of serum BALP, in particular in non-jaundiced patients with high serum ALP, may indicate the presence of tumor in the bile duct.

Aberrant expression of MUC5AC mucin is obvious in cholangiocarcinoma tissues, however, this mucin has never been detected in the serum. Using immunoblotting marked with antibody vs. MUC5AC core protein, we could detect MUC5AC mucin in the serum of 112 from 179 cholangiocarcinoma patients (62.6% sensitivity), two of the 62 with benign hepatobiliary diseases, six of the 60 with hepato-gastrointestinal cancer, and none in either the 60 active opisthorchiasis or 74 healthy persons. Detection of serum mucin in the serum of cholangiocarcinoma patients corresponded well to the MUC5AC expressed in individual tissues. Serum MUC5AC may be used to enhance the diagnostic accuracy of cholangiocarcinoma.

206 Background: Cholangiocarcinoma in Thailand and Laos is associated with the liver fluke Opisthorchis viverrini (OV). Molecular differences between OV-CCA and Non-OV CCA remain to be defined. MiRNAs are regulators of mRNA that result in transcriptional repression or gene silencing. MiRNA profiles of OV vs. non-OV CCA have not yet been studied. Methods: 50 OV-CCA and 16 non-OV CCA pathological samples were obtained from Liver Fluke and Cholangiocarcinoma Center, Khonkaen University, Thailand and University of Texas, MD Anderson Cancer Center (MDACC), USA, respectively. Macrodissected common bile duct samples from MDACC were used as control groups. Global miRNA expression profiling was performed using the miRCURY LNA microRNA Array (Exiqon).qRT-PCR and in situ hybridization (ISH) were used to validate the miRNAs noted to be significant on the microRNA array. Results: Ten samples of OV and non-OV CCA were analyzed for global miRNA expression; 46 were used for validation. MiRNA microarray identified 71 miRNAs that were differentially expressed between OV-CCA and non-OV CCA (p

Opisthorchis viverrini is considered among the most important of the food-borne trematodes due to its strong association with advanced periductal fibrosis and bile duct cancer (cholangiocarcinoma). We investigated the relationship between plasma levels of Interleukin (IL)-6 and the risk of developing advanced fibrosis and bile duct cancer from chronic Opisthorchis infection. We show that IL-6 circulates in plasma at concentrations 58 times higher in individuals with advanced fibrosis than age, sex, and nearest-neighbor matched controls and 221 times higher in individuals with bile duct cancer than controls. We also observed a dose-response relationship between increasing levels of plasma IL-6 and increasing risk of advanced fibrosis and bile duct cancer; for example, in age and sex adjusted analyses, individuals with the highest quartiles of plasma IL-6 had a 19 times greater risk of developing advanced periductal fibrosis and a 150 times greater risk of developing of bile duct cancer than individuals with no detectable level of plasma IL-6. Finally, we show that a single plasma IL-6 measurement has excellent positive predictive value for the detection of both advanced bile duct fibrosis and bile duct cancer in regions with high O. viverrini transmission. These data support our hypothesis that common mechanisms drive bile duct fibrosis and bile duct tumorogenesis from chronic O. viverrini infection. Our study also adds a unique aspect to the literature on circulating levels of IL-6 as an immune marker of hepatobiliary pathology by showing that high levels of circulating IL-6 in plasma are not related to infection with O. viverrini, but to the development of the advanced and often lethal pathologies resulting from chronic O. viverrini infection., Author Summary O. viverrini is among the few parasites considered a Class 1 carcinogen because of its strong association with bile duct cancer (cholangiocarcinoma). Currently, more than 40 million people are infected with O. viverrini worldwide. Thailand has the highest prevalence of O. viverrini at 10 million people infected and also the highest incidence of Opisthorchis-associated bile duct cancer (cholangiocarcinoma) in the world. In the current study, we also show that levels of IL-6 in plasma are associated in a dose-dependent fashion with Opisthorchis-induced bile duct fibrosis (periductal fibrosis) and cholangiocarcinoma. More importantly, we show that O. viverrini infection alone does not elevate IL-6 levels in the plasma. It is only in the presence of these advanced pathologies (advanced fibrosis or cholangiocarcinoma) from chronic O. viverrini infection that significantly elevates plasma levels of IL-6 are observed. Moreover, we show that plasma IL-6 is an easily accessible biomarker for the detection of advanced periductal fibrosis and cholangiocarcinoma, which would be a critical advance for this region of Thailand and other countries in Southeast Asia, where the prevalence of O. viverrini infection can reach as high as 80% and the incidence of bile duct cancer is the highest in the world.

To determine whether expression of certain enzymes related to 5-fluorouracil (5-FU) metabolism predicts 5-FU chemosensitivity in cholangiocarcinoma (CCA).The histoculture drug response assay (HDRA) was performed using surgically resected CCA tissues. Tumor cell viability was determined morphologically with hematoxylin and eosin- and terminal deoxynucleotide transferase-mediated dUTP nick-end labeling-stained tissues. The mRNA expression of thymidine phosphorylase (TP), orotate phosphoribosyl transferase (OPRT), thymidylate synthase (TS), and dihydropyrimidine dehydrogenase (DPD) was determined with real-time reverse transcriptase-polymerase chain reaction. The levels of gene expression and the sensitivity to 5-FU were evaluated.Twenty-three CCA tissues were obtained from patients who had been diagnosed with intrahepatic CCA and who underwent surgical resection at Srinagarind Hospital, Khon Kaen University from 2007 to 2009. HDRA was used to determine the response of these CCA tissues to 5-FU. Based on the dose-response curve, 200 μg/mL 5-FU was selected as the test concentration. The percentage of inhibition index at the median point was selected as the cut-off point to differentiate the responding and non-responding tumors to 5-FU. When the relationship between TP, OPRT, TS and DPD mRNA expression levels and the sensitivity of CCA tissues to 5-FU was examined, only OPRT mRNA expression was significantly correlated with the response to 5-FU. The mean expression level of OPRT was significantly higher in the responder group compared to the non-responder group (0.41 ± 0.25 vs 0.22 ± 0.12, P0.05).OPRT mRNA expression may be a useful predictor of 5-FU chemosensitivity of CCA. Whether OPRT mRNA could be used to predict the success of 5-FU chemotherapy in CCA patients requires confirmation in patients.