Background and Aim:Sorafenib is now considered as a standard treatment for advanced hepatocellular carcinoma (HCC). We evaluated the effect of hepatitis B virus (HBV) DNA titers on prognosis in HCC patients treated with sorafenib. Methods: From 2008 to 2012, 78 HBV-related HCC patients who received sorafenib treatment at Severance Hospital were included in our analysis. The effect of pretreatment HBV-DNA levels on clinical outcomes for use in predicting prognosis after treatment with sorafenib was examined by univariate and multivariate analysis. Results: Median overall survival and median progression-free survival were 5.2 months (95% confidence interval: 4.0-6.4) and 3.5 months (95% confidence interval: 2.3-4.7), respectively. Multivariate analysis revealed high levels of HBV-DNA (> 2000 IU/mL) to be an independent risk factor for worse overall survival (P = 0.005; hazard ratio, 2.85) and disease progression among patients who did not receive concomitant prophylactic antiviral therapy during sorafenib treatment (P = 0.008; hazard ratio, 87.4). Moreover, viral reactivation occurred more frequently in patients who did not receive concomitant prophylactic antiviral therapy than in those who did (4/38 vs 0/40; P = 0.025). Conclusion: Higher HBV-DNA levels prior to sorafenib treatment were associated with poorer prognosis and increased viral reactivation thereafter. These results suggest the potential usefulness of prophylactic antiviral therapy when treating HBV-related HCC patients with sorafenib.

Humoral hypercalcemia of malignancy (HHM) is rarely associated with cholangiocarcinoma (CC), and represents dismal prognosis. A 63-year-old male was admitted for evaluation of an intrahepatic mass. He was diagnosed with HHM associated with locally advanced CC. As the tumor responded to the concurrent chemoradiotherapy with capecitabine and cisplatin, serum calcium level was normalized. However, according to the disease progression, he suffered recurrence of HHM and he expired approximately one year after initial diagnosis. A 68-year-old male who presented with abdominal pain was diagnosed with metastatic CC. After the eighth cycle of gemcitabine and cisplatin, progression of the disease was found with HHM. He was treated with the best supportive care, until his demise approximately one month after the diagnosis of HHM. We report on two cases of HHM associated with CC that demonstrate strong correlation between hypercalcemia and disease burden.

MHT showed considerable efficacy and tolerability in this study. Further randomized prospective study is warranted.

Introduction: Signaling pathways in acute leukemia are aberrantly activated to cause leukemogenesis and relapse after treatment. Like in many other malignancies, upregulation of WNT/beta-catenin pathway and hyperactive RAS is known to be associated with treatment resistance in leukemia. However, there are little studies about RAS singaling pathyway and leukemia, and it is a field of study that needs to be revealed. KYA1797K is a recently developed small molecule, binds directly to RGS domain of axin and enhances the beta-catenin destruction complex which activates GSK3beta and results in degradation of beta-catenin and RAS. In the current study, we tried to find the role of RAS inhibition by KYA1797K in leukemic cell lines and in patient's BM samples. Moreover, other small molecule PCK412 (Midosaturin) was also used for comparison. Materials & Methods: Leukemic cells (MOLT-4, THP-1, MOLT-4, Jurka, KG-1, MV4-11, RS4-11) were cultured in RPMI1640 media under various concentration (0.1-10µM for KYA1797K, 0.5-500nM for PKC412) for 48h and with Erlotinib (1µM) for comparison. Cell proliferation assay on each leukemic cell was done and immunoblotting for β-catenin, GSK3β, Pan-RAS, N-RAS was checked. Downstream targets of Wnt pathway (c-Myc, CD44, LEF1, Met, TCF1/TCF7) were studied by immunoblotting. MOLT-4 was stimulated with Wnt3a (200ng/mL, 4h) and changes in Wnt pathway were observed. Bone marrow samples of AML and ALL patients were evaluated for β-catenin and RAS. KYA1797K (Nat Chem Biol 2016, 12:593) was kindly provided by Prof. Kang-Yell Choi. Results: Suppression of leukemic cells by KYA1797K was evident starting from the concentration of 5 microM. (fig.1) Beta-catenin was down regulated in all cell lines by KYA1797K. Pan-RAS decreased in MOLT-4 and THP-1. All the downstream targets evaluated were down regulated by KYA1797K in MOLT-4 culture, and was evident at the concentration of 5microM. (fig.2) Stimulation of Wnt pathway by Wnt3a was inhibited by KYA1797K. (fig.3) Bone marrow samples from 7 ALL patients showed various status of β-catenin and RAS expression. Two high-risk patients showed suppression of β-cateinin and N-RAS by KYA1797K. (fig.4) In MV4-11 (FLT3 mutant) and RS4-11 (FLT3 wild type), IC50 for PKC412 was higher in RS4-11 compared to MV4-11 while KYA1797K showed same IC50 in both cell lines. β-catenin and RAS downregulation was observed by KYA1797K. Effects of KYA1797K analyzed by qRT-PCR and immunoblot for FLT3, N-RAS, MEK, ERK, ETS2 showed that KYA1797K downregulates FLT3 in MV4-11 even though FLT3 is not the main target of action. It was less effective on RS4-11. (fig.5) In bone marrow samples of ALL patient with FLT3 mutation, KYA1797K 1µM showed effect in reducing leukemia cells. (fig.6) Conclusion: This preclinical study suggests that KYA1797K may be an option for patients with acute leukemia. KYA1797K effectively destabilized β-catenin and RAS in acute leukemia even under Wnt pathway activation. FLT3, N-RAS, MEK, ERK, ETS2 were down regulated by KYA1797K, hence KYA1797K has a potential application for acute leukemia with FLT3 mutation. Extended studies including further in vivo study are needed to build up a strategy in small molecule therapy to target RAS in acute leukemia. Although with limitation, we suggest RAS inhibitor as a potential drug for leukemia. Disclosures No relevant conflicts of interest to declare.

Background: The needs for sensitive coagulation factor assays able to measure factor VIII (FVIII) and factor IX (FIX) in the range of 0.0 to 1.0 %, are continuously growing with diversification of hemophilia management. However, practical methods with sufficient analytical sensitivity available in clinical laboratory have not yet been introduced. We developed new coagulation factor assays applying various parameters derived from a turbidity based coagulometer and examined their ability to measure low-level FVIII and FIX and analytical resolution in that range. Method: We prepared 12 spiked samples with FVIII and FIX levels from 0.0 to 2.4 % and conducted conventional one-stage coagulation factor assays in repeat. We collected measured values of APTT, velocity and acceleration peaks of coagulation (peak 1 and peak 2) from each measurement. We also calculated values of peak 1 and peak 2 from the mathematical model of turbidity curves. From the measured values of these parameters we derived calibration formulae for coagulation factor assays, FVIIICT, FVIIIpeak1, FVIIIpeak2, FVIIIcalc1, FVIIIcalc2, FIXCT, FIXpeak1, FIXpeak2, FIXcalc1, and FIXcalc2. Results: The reliability interval (range of FVIII levels producing unequivocal results) of FVIIICT (the conventional FVIII assay) covered only 9 % of 0.0 to 1.0 % range. For new assays, the coverages were 54, 31, 55, and 65 % for FVIIIpeak1, FVIIIpeak2, FVIIIcalc1, and FVIIIcalc2 respectively. The resolution between immediate levels of spiked samples could be determined from modeled distributions or be checked simply by inspecting the actual assay result distributions. For FVIIIpeak1, 0.2 % and 0.6 % results stood apart from each other. For FVIIIcalc1 and FVIIIcalc2, 0.2, 0.4, and 0.6 % were distinguished from each other. When we measured recombinant human (rh) FVIII, the coverages were 7, 64, 52, 73, and 79 % for rhFVIIICT, rhFVIIIpeak1, rhFVIIIpeak2, rhFVIIIcalc1, and rhFVIIIcalc2 respectively. (rh)FVIIIpeak1, (rh)FVIIIcalc1, and (rh)FVIIIcalc2 particularly showed wide measurable ranges of guarantee. For FVIIIpeak1, 0.2 % and 0.6 % results stood apart from each other. For FVIIIcalc1 and FVIIIcalc2, 0.2, 0.4, and 0.6 % were distinguished from each other. rhFVIIIpeak1 and rhFVIIIcalc1 showed slightly better resolution than the former. rhFVIIIcalc2 was notable in that every 0.1, 0.2, 0.4, 0.6, 0.8 % result stood apart from each immediate level result. We could not determine certainty interval (the range of unequivocal values) of FIXCT and FIXpeak2 because the 0.0 % and 1.0 % ranges overlapped. Thus, the conventional FIX assay cannot measure between 0.0 and 1.0 %. FIXpeak1, FIXcalc1, and FIXcalc2 worked better and the certainty interval of unequivocal results could be determined between 0.0 and 1.0 %. The reliability interval was not available for any FIX assay. Results from rhFIX measurements were similar those of plasma FIX assays. Conclusion: We introduce new FVIII and FIX assays with superior analytical resolution in the range of 0.0 to 1.0 % in comparison to the conventional assays. Figure. Figure. Disclosures No relevant conflicts of interest to declare.