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Background A cisplatin plus irinotecan (CPT‐11) regimen is used for patients with extensive disease small cell lung cancer (ED‐SCLC). Amrubicin (AMR) is primarily used for relapsed SCLC. The HOT1401/NJLCG1401 trial, an open‐label randomized phase II trial, was designed to assess the benefit of maintenance therapy in patients with ED‐SCLC who responded to induction therapy. Methods Patients with histologically‐ or cytologically‐confirmed ED‐SCLC were included and were treated with an induction therapy of four cycles of cisplatin (60 mg/m2 on day 1) plus CPT‐11 (60 mg/m2 on days 1, 8, and 15) every four weeks. After induction therapy, patients who had nonprogressive disease were randomized to receive either maintenance CPT‐11 (60 mg/m2 on days 1 and 8) every three weeks, or AMR (35 mg/m2 on days 1–3) every three weeks. Results A total of 34 patients were enrolled; 20 patients had progressive disease or received incomplete induction chemotherapy. Finally, 14 patients were randomly assigned to receive CPT‐11 (n = 7) or AMR (n = 7). This study was terminated prematurely because of low patient accrual. The overall objective response rate was 73%, the median PFS was 5.7 months (95% confidence interval [CI]: 3.6–11.8), and the median overall survival was 20.1 months (95% CI: 13.7–not reached). No statistically significant difference in progression‐free survival (PFS) were noted between patients treated with CPT‐11 and those treated with AMR. There were no treatment‐related deaths in this study. Conclusions Maintenance therapy with CPT‐11 or AMR after induction therapy might be effective in some patients., There were some patients who achieved long disease control. Maintenance therapy with CPT‐11 or AMR after induction therapy could have potential for treating ED‐SCLC.

Triamcinolone acetonide (TA) has been shown to improve morphological and functional outcome in diabetic macular edema (DME) patients. However, the functional mechanism of TA has not been elucidated yet. In this study we investigated the detailed functional mechanism of TA using culture cells and retinopathy mouse models in which retinal inflammation and abnormal angiogenesis were induced by pericyte depletion. TA significantly prevented retinal hemorrhage, edema and partially improved abnormal angiogenesis. TA decreased retinal vascular endothelial growth factor (VEGF) concentration, presumably by preventing recruitment of macrophages into retina and TA also inhibited expression of inflammatory cytokines in retina. TA inhibited proliferation/migration of vascular endothelial cells and vessel sprouting. No direct inhibition of VEGF receptor 2 (VEGFR2) autophosphorylation was observed by TA. These results suggested that TA improved inflammatory retinal events which were induced in pericyte-deleted mice by mainly decreasing macrophage-derived VEGF and expression of inflammatory cytokines followed by attenuation of vascular permeability and proliferation/migration of endothelial cells. Furthermore, in these processes, translocation of glucocorticoid receptor (GR) was partially involved.

Progranulin is a secreted growth factor associated with multiple physiological functions in ischemic pathophysiology. However, it is still not fully understood how progranulin is involved in ischemic lesion and cardiac remodeling after myocardial infarction (MI). In this study, we investigated the effects of progranulin on myocardial ischemia and reperfusion injury. We investigated progranulin expression using Western blotting and immunostaining after permanent left coronary artery (LCA) occlusion in mice. Infarct size and the number of infiltrating neutrophils were measured 24 h after permanent LCA occlusion. Recombinant mouse progranulin was administered before LCA occlusion. In addition, we evaluated cardiac function using cardiac catheterization and echocardiography, and fibrosis size by Masson’s trichrome staining after myocardial ischemia/reperfusion in rabbits. Recombinant human progranulin was administered immediately after induction of reperfusion. Progranulin expression increased in the myocardial ischemic area 1, 3, and 5 days after permanent LCA occlusion in mice. The administration of recombinant mouse progranulin significantly attenuated infarct size and infiltrating neutrophils 24 h after permanent LCA occlusion in mice. We also found that administration of recombinant human progranulin ameliorated the deterioration of cardiac dysfunction and fibrosis after myocardial ischemia/reperfusion in rabbits. These findings suggest that progranulin may protect myocardial ischemia/reperfusion injury.

Recent studies have indicated that afatinib is beneficial for patients with non‐small cell lung cancer (NSCLC) harboring uncommon epidermal growth factor receptor (EGFR) mutations, while the effectiveness of afatinib rechallenge has not been fully defined. Here, we report a long‐term survival case of NSCLC harboring concomitant EGFR G719C and S768I mutations who received afatinib rechallenge followed by chemotherapy. The present case suggests that combined therapeutic strategies such as afatinib plus sequential chemotherapy would be beneficial based on appropriately timed rebiopsies from recurrent lesions. Key points Significant findings of the study and what this study adds A NSCLC patient carrying EGFR G719X/S768I mutations survived for a long period of time with afatinib rechallenge followed by chemotherapy. Combined therapeutic strategies should be considered based on rebiopsies in appropriate timing in NSCLC with uncommon EGFR mutations.

Background/aim The efficacy of the combination of amrubicin and bevacizumab against advanced non small-cell lung cancer (NSCLC), as a second or third-line treatment, was evaluated. Patients and methods Amrubicin was administered for 3 days to patients with previously treated advanced NSCLC, whereas bevacizumab was administered on day 1 of each cycle; this regimen was repeated every 3 weeks. Results Among the 16 patients, an overall response rate of 12.5% (for two patients) was achieved, and the overall disease control rate was 93.7%. Progression free survival and overall survival were 8.5 and 16.6 months, respectively. Grade 3 or 4 haematological toxicities were leukopenia, neutropenia, and febrile neutropenia. Grade 3 proteinuria and infection were the non haematological adverse events. Conclusion The combination of amrubicin and bevacizumab is a promising regimen in the second or third-line treatment for advanced non-squamous NSCLC; however, physicians must recognise the risk of proteinuria related with this regimen.

Anti-vascular endothelial growth factor (VEGF) therapies are now the first-line treatment for many ocular diseases, but some patients are non-responders to these therapies. The purpose of this study was to determine whether the level of adiponectin increased the pathogenesis of retinal edema and neovascularization in the retina of progressive ocular vascular diseases. We examined the role played by adiponectin in two types of cells and animal models which are retinal vein occlusion (RVO) and oxygen-induced retinopathy (OIR) mice. Our results showed that an injection of anti-adiponectin antibody ameliorated the retinal edema and ischemia through the depression of the expression level of VEGF-related factors and tight junction-related proteins in the retina of RVO mice. The intravitreal injection of anti-adiponectin antibody also decreased the degree of retinal neovascularization in an OIR mice. In addition, exposure of human retinal microvascular endothelial cells and human brain microvascular pericytes in culture to adiponectin increased both the vascular permeability and neovascularization through the increase of inflammatory factor and the dropout of the pericytes. These findings indicate that adiponectin plays a critical role in retinal edema and neovascularization, and adiponectin is a potential therapeutic target for the treatment of diabetic macular edema, proliferative diabetic retinopathy, and RVO.

Purpose In patients with epidermal growth factor receptor (EGFR)-mutated, advanced, non-small cell lung cancer (NSCLC), common gefitinib-sensitive EGFR mutations that predict a greater response to therapy include the exon 19 deletion and L858R point mutation. The objective of this study was to evaluate whether body surface area (BSA), body weight (BW), and body mass index (BMI) affect gefitinib efficacy in such patients. Methods The medical charts of 138 consecutive patients with advanced NSCLC harboring sensitive EGFR mutations, who underwent gefitinib treatment, were reviewed. The median BSA and BW were used as cutoff values to evaluate their impact on gefitinib efficacy. BMI was categorized as underweight (

Although docetaxel plus ramucirumab has shown superior treatment efficacy over docetaxel monotherapy for patients with non-small cell lung cancer (NSCLC), the high rate of febrile neutropenia (FN) presents a clinical problem. This study aimed to validate the primary prophylactic use of pegfilgrastim with docetaxel and ramucirumab treatment in Japanese patients with NSCLC. Patients with NSCLC with progression after at least one round of chemotherapy were enrolled and administered docetaxel (60 mg/m2) plus ramucirumab (10 mg/kg) intravenously on day 1, followed by pegylated-granulocyte colony-stimulating factor (3.6 mg) on day 2 of a 21-day treatment cycle. The primary study endpoint was the percentage of patients who developed FN. Secondary endpoints included overall survival, progression-free survival, overall response rate, and safety. Overall, 20 patients (15 men and 5 women) were enrolled, of whom one developed FN, resulting in an overall FN rate of 5%. The response and disease control rates were 40% and 85%, respectively. The median progression-free survival was 6.6 (95% confidence interval [CI], 0.5-NR) months. The median overall survival was 18.4 (95% CI, 2.2–11.0) months. Six patients aged over 75 years were included in this study, and although most adverse events were durable, ramucirumab-associated adverse events occurred more frequently in these patients. We observed a 5% FN rate using primary prophylactic pegylated-granulocyte colony-stimulating factor with docetaxel plus ramucirumab in Japanese patients with NSCLC. While most adverse events were durable, elderly patients should be closely monitored.

Objective: This study aimed to examine whether DC101 (anti-VEGFR2 antibody)- modified micelles have applications as novel drug delivery devices, which allow small molecule antiangiogenic agents to deliver to angiogenic sites on a murine laser-induced choroidal neovascularization (CNV) model. Materials and Method: CNV was induced by photocoagulation on the unilateral eye of each mouse under anesthesia. Immediately after laser coagulation, E7974-loaded DC101-modified micelles and motesanib-loaded DC101-modified micelles were intravitreally administrated. Two weeks after photocoagulation, CNV was visualized using fluorescein-conjugated dextran (MW=2,000 kDa), and the CNV area was measured in retinal pigment epithelium (RPE)-choroidal flat mounts. Results: Intravitreal administration of both DC101-modified micelles loaded with E7974 at 2 µM and motesanib at 2 µM significantly reduced CNV area in the murine laser-induced CNV model at a clearly lower concentration than the effective dose of each agent. Conclusion: These results suggest that DC101-modified micelle might be effective drug carrier system for treating CNV and other ocular angiogenic diseases.

Purpose Ocular angiogenesis, including retinopathy of prematurity, diabetic retinopathy, and exudative age-related macular degeneration, are closely related to oxidative stress. Many reports have shown that the cellular protective mechanism against oxidative stress and inflammatory response has nuclear factor-erythroid 2-related factor-2 (Nrf2) activity. The aim of this study was to investigate the effectiveness and mechanism of Nrf2 activation in treating the ocular diseases with abnormal vessels. Methods The effects of Nrf2 activators, bardoxolone methyl (BARD) and RS9, were evaluated against vascular endothelial growth factor (VEGF)-induced cell migration in human retinal microvascular endothelial cells (HRMECs). We measured the expression of the Nrf2 target genes, Ho-1 and Nqo-1 mRNA, in mouse retinas after a single injection of BARD and RS9. The effects and mechanisms of RS9 against retinal angiogenesis were evaluated using an oxygen-induced retinopathy (OIR) model in mice. Moreover, the effect of RS9 against choroidal neovascularization (CNV) was evaluated in a laser-induced CNV monkey model. Results Both BARD and RS9 decreased VEGF-induced cell migration, and significantly increased Ho-1 mRNA expression; however, only RS9 significantly increased Nqo-1 mRNA. RS9 decreased retinal neovascularization through suppressing VEGF expression and increasing Nrf2, HO-1, platelet-derived growth factor receptor (PDGFR)-β, and tight junction proteins in OIR murine retinas. Furthermore, RS9 showed a tendency toward decreasing CNV lesions, and improved vascular leakage in a CNV monkey model. Conclusions These data indicate that a Nrf2 activator might be a candidate for treatment of ocular diseases characterized by pathophysiological angiogenesis and hyperpermeability.

Despite the high frequency of KRAS mutations in non-small cell lung cancer (NSCLC), therapeutic modalities targeting KRAS-mutated NSCLC have not been established. Based on our previous findings that mutant KRAS knockdown sensitized NSCLC cells to a p38 inhibitor, the growth-inhibitory effect of dual MEK and p38 inhibition on tumor growth in NSCLC cells harboring KRAS mutations was investigated. In KRAS-mutated NSCLC cells, the MEK inhibitor, selumetinib, inhibited cell growth in a dose-dependent manner, and its growth-inhibitory effect was enhanced by combined treatment with the p38 inhibitor LY2228820. Similarly, another pair of MEK and p38 inhibitors also exhibited antitumor activity. Small interfering RNAs (siRNAs) against MAPK14, which encodes p38α MAPK, enhanced the growth-inhibitory effect of the MEK inhibitors in NSCLC cells with KRAS mutations. Notably, MEK inhibitors reduced p38 expression levels but increased p38 phosphorylation levels, resulting in sensitization to p38 inhibitors in KRAS-mutated NSCLC cells. These results provide evidence that dual MEK and p38 inhibition could be a potent therapeutic strategy against oncogenic KRAS-driven NSCLC.

Despite the high frequency of KRAS mutations in non-small cell lung cancer (NSCLC), therapeutic modalities targeting KRAS-mutated NSCLC have not been established. Based on our previous findings that mutant KRAS knockdown sensitized NSCLC cells to a p38 inhibitor, the growth-inhibitory effect of dual MEK and p38 inhibition on tumor growth in NSCLC cells harboring KRAS mutations was investigated. In KRAS-mutated NSCLC cells, the MEK inhibitor, selumetinib, inhibited cell growth in a dose-dependent manner, and its growth-inhibitory effect was enhanced by combined treatment with the p38 inhibitor LY2228820. Similarly, another pair of MEK and p38 inhibitors also exhibited antitumor activity. Small interfering RNAs (siRNAs) against MAPK14, which encodes p38α MAPK, enhanced the growth-inhibitory effect of the MEK inhibitors in NSCLC cells with KRAS mutations. Notably, MEK inhibitors reduced p38 expression levels but increased p38 phosphorylation levels, resulting in sensitization to p38 inhibitors in KRAS-mutated NSCLC cells. These results provide evidence that dual MEK and p38 inhibition could be a potent therapeutic strategy against oncogenic KRAS-driven NSCLC.

Background Cisplatin plus irinotecan (PI) regimen is used for patients with extensive disease (ED)-SCLC. Amrubicin is mainly used for relapsed SCLC. HOT1401/NJLCG1401 trial, open-label randomized phase II trials was designed to assess the benefit of maintenance therapy in patients with ED-SCLC who responded induction therapy. Methods Eligible patients were aged 20-74 years; had histologically or cytologically confirmed, ED-SCLC. After induction therapy PI (P: 60mg/m2 on day 1, I: 60mg/m2 on days 1, 8, 15, every 4 weeks for 4 cycles), patients who attained CR, PR and SD were randomized to either maintenance irinotecan (I: 60mg/m2 on days 1, 8, every 3 weeks) or amrubicin (A: 35 mg/m2 on days 1-3 every 3 weeks). The primary endpoint is 6-months PFS rate. Results A total of 34 patients were enrolled from 2014 to 2017. Twenty patients had disease progression or incomplete of induction chemotherapy, finally 14 (41.1%) patients were randomly assigned to receive irinotecan (n = 7) and amrubicin (n = 7). This study was terminated prematurely because of low patient accrual. Of the evaluable patients (n = 34) the overall objective response rate was 73%, median progression free survival was 5.7 months (95% CI: 3.6 -11.8), and median overall survival was 20.1 months (95% CI: 13.7-NR). Six months PFS rate was 47 % (95%CI: 31.4-63.2). Sub group analysis showed the patients receive maintenance group showed longer PFS and OS than those without (15.8 months and 3.5 months, respectively p Conclusions Maintenance therapy of irinotecan or amrubicin after induction therapy was well tolerated. Some patients might be effective for maintenance therapy. Clinical trial identification UMIN 000013882. Legal entity responsible for the study HOT/NJLCG. Funding Has not received any funding. Disclosure All authors have declared no conflicts of interest.

The efficacy of gefitinib [an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor] in elderly patients with non-small cell lung cancer (NSCLC) and EGFR mutation has not been elucidated. Therefore, the objective of this study was to investigate the efficacy and feasibility of gefitinib in elderly chemotherapy-naive patients with NSCLC harboring sensitive EGFR mutations. We retrospectively evaluated the clinical effects of gefitinib as a first-line treatment for elderly (≥75 years) NSCLC patients with EGFR mutations (exon 19 deletion or exon 21 L858R mutation). All patients were initially treated with gefitinib (250 mg/day) at seven institutions. Between January 2006 and December 2012, 62 patients (17 men, 45 women) with a median age of 80 years (range, 75–89 years) were included in our analysis. The overall response and disease control rates were 61.2 and 83.8 %, respectively, and the median progression-free survival and overall survival were 13.2 and 19.0 months, respectively. Common adverse events included rash, diarrhea, and liver dysfunction. Major grade 3 or 4 toxicities included skin rash (3.2 %) and increased levels of aspartate aminotransferase or alanine aminotransferase (21.0 %). Gefitinib treatment was discontinued owing to adverse events of liver dysfunction in 3 patients, drug-induced pneumonitis in 2, and diarrhea in 1. First-line gefitinib could be a preferable standard treatment in elderly patients with advanced NSCLC harboring sensitive EGFR mutations.

This study aimed to establish an experimental exudative age-related macular degeneration (AMD) model in the common marmoset (Callithrix jacchus), which is a small New World monkey. Choroidal neovascularization (CNV) was induced by laser irradiation on the left eye of each animal under anesthesia. Eight laser spots were applied around the macular area using the image-guided laser system (532 nm) attached with Micron III at 650 mW-2,000 mW power. Laser pulse duration and spot size were fixed at 100 ms and 50 µm, respectively. At 14 days after laser irradiation, fluorescein angiograms were observed. At 21 days after laser irradiation, the fluorescein angiograms were transcardially perfused to the bilateral common carotid arteries with 4% paraformaldehyde for the transverse section or with fluorescein-conjugated dextran (MW = 2,000 kDa) for the retinal pigment epithelia (RPE)-choroidal flatmount. At 14 days after laser irradiation, late hyperfluorescence and leakage within or beyond the lesion borders were observed in a laser power-dependent manner. In the RPE-choroidal flatmount, the mean size of the CNV lesions at 1,500 mW was 1.34 ± 0.49 × 10(5) µm(2) (Mean ± S.D., n = 29), and the coefficient of variation for each CNV area was 36.5% (n = 29). In conclusion, we succeeded in producing an experimental exudative type of AMD model in the common marmoset. This model may be useful in elucidating the pathophysiological mechanism and screening of new candidates for exudative AMD.

We describe the case of an 85-year-old man diagnosed with chronic myelomonocytic leukemia whose disease was treated with hydroxyurea for 3 months. He developed respiratory symptoms that were extensively investigated. Despite the intensive treatment, he died of respiratory failure eleven days later. An autopsy revealed diffuse interstitial inflammation of both lungs consistent with drug-induced inflammation. A drug lymphocyte stimulation test was positive for hydroxyurea. Taken together these findings demonstrated that severe interstitial pneumonitis was induced by this drug. Physicians using hydroxyurea must be aware of its potentially life-threatening pulmonary toxicity.

Objective— The incidence of blindness is increasing because of the increase in abnormal ocular neovascularization. Anti-VEGF (vascular endothelial growth factor) therapies have led to good results, although they are not a cure for the blindness. The purpose of this study was to determine what role HB-EGF (heparin-binding epidermal growth factor–like growth factor) plays in ocular angiogenesis. Approach and Results— We examined the role played by HB-EGF in ocular neovascularization in 2 animal models of neovascularization: laser-induced choroidal neovascularization (CNV) and oxygen-induced retinopathy. We also studied human retinal microvascular endothelial cells in culture. Our results showed that the neovascularization was decreased in both the CNV and oxygen-induced retinopathy models in HB-EGF conditional knockout mice compared with that in wild-type mice. Moreover, the expressions of HB-EGF and VEGF were increased after laser-induced CNV and oxygen-induced retinopathy, and their expression sites were located around the neovascular areas. Exposure of human retinal microvascular endothelial cells to HB-EGF and VEGF increased their proliferation and migration, and CRM-197 (cross-reactive material-197), an HB-EGF inhibitor, decreased the HB-EGF–induced and VEGF-induced cell proliferation and migration. VEGF increased the expression of HB-EGF mRNA. VEGF-dependent activation of EGFR (epidermal growth factor receptor)/ERK1/2 (extracellular signal-regulated kinase 1/2) signaling and cell proliferation of endothelial cells required stimulation of the ADAM17 (a disintegrin and metalloprotease) and ADAM12. CRM-197 decreased the grades of the fluorescein angiograms and size of the CNV areas in marmoset monkeys. Conclusions— These findings suggest that HB-EGF plays an important role in the development of CNV. Therefore, further investigations of HB-EGF are needed as a potential therapeutic target in the treatment of exudative age-related macular degeneration.

Oxidative stress plays a pivotal role in developing and accelerating retinal diseases including age-related macular degeneration (AMD), glaucoma, diabetic retinopathy (DR), and retinal vein occlusion (RVO). An excess amount of reactive oxygen species (ROS) can lead to functional and morphological impairments in retinal pigment epithelium (RPE), endothelial cells, and retinal ganglion cells (RGCs). Here we demonstrate that edaravone, a free radical scavenger, decreased apoptotic cell death, oxidative damage to DNA and lipids, and angiogenesis through inhibiting JNK and p38 MAPK pathways in AMD, glaucoma, DR, and RVO animal models. These data suggest that the therapeutic strategy for targeting oxidative stress may be important for the treatment of these ocular diseases, and edaravone may be useful for treating retinal diseases associated with oxidative stress.

Kallikrein is a serine protease involved in the kallikrein–kinnin system. Kallikrein is derived from the blood plasma or tissue, and is correlated with aggravation and improvement in eye diseases, such as, glaucoma, diabetic retinopathy, age-related macular degeneration, and ocular ischemic syndrome. The plasma kallikrein stimulates retinal vascular permeability and intraocular hemorrhage. On the other hand, we had reported that the tissue kallikrein normalizes retinal vasopermeability and inhibited retinal neovascularization and retinal ischemic injury. The protective mechanisms of the tissue-derived kallikrein include the cleavage of vascular endothelial growth factor (VEGF), which suggests that the tissue kallikrein could be potentially-effective against any disease involving the VEGF production.

This study aimed to evaluate the effect of liposomes loaded with diclofenac, a potent cyclooxygenase (COX)-1 and COX-2 inhibitor, on laser-induced choroidal neovascularization (CNV) in mice and non-human primates (common marmosets). CNV was induced by laser irradiation on the unilateral or bilateral eye of each mouse or common marmoset, respectively, under anesthesia. The CNV was visualized using fluorescence labeling with intravenous injection of fluoresceinconjugated dextran (molecular weight = 2,000 kDa), and quantified in the retinal pigment epithelia (RPE)-choroidal flatmounts. Diclofenac-loaded liposome or diclofenac ophthalmic solution was instillated to the eye surface daily for 14 days and 21 days in mice and common marmosets, respectively. In the mouse CNV model, 0.1% diclofenac-loaded liposome eye drops administered four times a day (q.i.d.) significantly reduced CNV formation in the RPE-choroidal flatmounts compared with those in empty liposome eye drops. Diclofenac-loaded liposome (0.1%) eye drops, administered once a day (s.i.d.), twice a day (b.i.d.), and three times a day (t.i.d.), also reduced CNV formation in a frequency-dependent manner. Furthermore, diclofenac-loaded liposome (0.03% and 0.1%) eye drops administered t.i.d. reduced CNV formation in a dose-dependent manner, significantly so at 0.1%. In the common marmoset CNV model, late hyperfluorescence and leakage by fluorescein angiograms was observed within or beyond the lesion borders at 17 days after laser irradiation, and diclofenac-loaded liposome eye drops (0.1% t.i.d.) tended to attenuate the late hyperfluorescence and leakage. Diclofenac-loaded liposomes had significantly reduced CNV formation in the RPE- choroidal flatmounts at 21 days after laser irradiation. In conclusion, diclofenac-loaded liposome eye drops enhance penetration to the RPE-choroid, and reduce the CNV formation. These results suggest that a drug-loaded liposome is a useful tool for drug delivery into the posterior segment of the eye.

Hepatocellular carcinomas (HCC) are the fifth most common tumor type and the third most common cause of cancer-related death worldwide. Some tumor markers for HCC, such as α-fetoprotein and des-γ-carboxyprothrombin, are used clinically. Recent advances in proteomics and glyco-proteomics might provide various types of novel tumor markers for HCC. While the clinical availability of these tumor markers is important, the molecular mechanisms underlying the production of tumor markers requires further clarification. Our group has investigated the glycobiology of tumor markers. In this review, we describe the impact of novel HCC markers and their possible implications for clinical use.

Fucosylated alpha-fetoprotein (AFP) is a specific tumor marker for hepatocellular carcinomas (HCC). However, the mechanisms underlying the increase in fucosylated AFP in serum of patients with HCC are largely unknown. Recently, we reported that fucosylation is a possible signal for the secretion of glycoproteins into bile in the liver. This finding might lead to the selective secretion of fucosylated AFP into bile and the selective secretion might be disrupted in hepatocarcinogenesis. In this study, therefore, we analyzed the oligosaccharide structures of glycoproteins in bile and serum of LEC rats, which are a rat model of spontaneous hepatocarcinogenesis. Lectin microarraying showed enhanced binding of 13 lectins to bile, compared with in serum from normal LEC rats, and the binding of these lectins to serum of LEC rats bearing HCC was higher than in normal rats. Structural analyses involving HPLC and mass spectrometry showed that the fucosylation levels of serum glycoproteins were not increased in CH rats but were in HCC rats, although the fucosylation levels of biliary glycoproteins were increased in both CH and HCC rats. These results suggested that the sorting machinery through fucosylation might be disrupted in the liver with HCC.

Mutant KRAS and BRAF are important oncogenic drivers in non-small cell lung cancer (NSCLC), however, effective treatment strategies for most of these mutants have not been established. Immuno-oncology clinical trials have shown promising results of anti-programmed death 1 (PD-1) antibodies for a subset of NSCLC patients and tumor expression of programmed-death ligand 1 (PD-L1) is regarded as a predictive marker for anti-PD-1 therapy. We found that CD274 mRNA and PD-L1 protein expression were reduced in NSCLC lines by small interfering RNAs (siRNAs) targeting mutant KRAS, but not wild-type KRAS, and also by pharmacologic inhibitors of MEK and ERK. In addition, there was a positive correlation between CD274 expression and KRAS expression in NSCLC lines, and analyses of The Cancer Genome Atlas (TCGA) database revealed that NSCLCs with high KRAS expression exhibited higher levels of CD274 compared to the tumors with low KRAS expression among KRAS-mutant NSCLCs. Similarly, siRNA-mediated BRAF knockdown and inhibitors of BRAF and MEK reduced PD-L1 expression in BRAF-mutant NSCLC cells, suggesting that PD-L1 expression was also upregulated by oncogenic BRAF. Mechanistically, these results suggest that oncogenic KRAS and BRAF induce PD-L1 overexpression in NSCLCs by activation of the MEK-ERK pathway. For clinical translation, our results suggest that: 1. in NSCLCs the combination of KRAS and PD-L1 overexpression could be a surrogate predictive marker for anti-PD-1 treatment; and 2. that since the goal of checkpoint inhibitor immuno-oncology therapy is to inhibit the PD-L1 axis, that a combination of anti-PD-1 and targeted therapy to inhibit the MEK-ERK pathway could be beneficial. Citation Format: Yosuke Miura, Noriaki Sunaga, Pinjie Bao, Luc Girard, Yusuke Tsukagoshi, Tomomi Masuda, Norimitsu Kasahara, Reiko Sakurai, Kyoichi Kaira, Takeshi Hisada, John Minna, Masanobu Yamada. PD-L1 overexpression induced by oncogenic KRAS and BRAF mutations through MAPK pathway activation in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5740.

Hypoxic stress is a risk factor of ocular neovascularization. Hypoxia visualization may provide clues regarding the underlying cause of angiogenesis. Recently, we developed a hypoxia-specific probe, protein transduction domain-oxygen-dependent degradation domain-HaloTag-Rhodamine (POH-Rhodamine). In this study, we observed the localization of HIF-1α proteins by immunohistochemistry and the fluorescence of POH-Rhodamine on RPE-choroid flat mounts. Moreover, we compared the localization of POH-Rhodamine with pimonidazole which is a standard reagent for detecting hypoxia. Next, we investigated the effects of triamcinolone acetonide (TAAC) against visual function that was evaluated by recording electroretinogram (ERG) and choroidal neovascularization (CNV) development. Mice were given laser-induced CNV using a diode laser and treated with intravitreal injection of TAAC. Finally, we investigated POH-Rhodamine on CNV treated with TAAC. In this study, the fluorescence of POH-Rhodamine and HIF-1α were co-localized in laser-irradiated sites and both the POH-Rhodamine and pimonidazole fluorescent areas were almost the same. Intravitreal injection of TAAC restored the reduced ERG b-wave but not the a-wave and decreased the mean CNV area. Furthermore, the area of the POH-Rhodamine-positive cells decreased. These findings indicate that POH-Rhodamine is useful for evaluating tissue hypoxia in a laser-induced CNV model, suggesting that TAAC suppressed CNV through tissue hypoxia improvement.

We conducted a phase I dose escalation study to determine the maximum tolerated dose (MTD), the recommended dose (RD) and the safety profile of amrubicin (AMR) plus paclitaxel (PTX) combination regimen for patients with previously treated non-small cell lung cancer (NSCLC).PTX was administered at a fixed dose of 150 mg/m(2)/day on day 1 and AMR was intravenously administered at a starting dose of 25 mg/m(2)/day on days 1-3, and this was repeated every 4 weeks. Doses of each drug were planned as follows-level 0, 20/150; level 1, 25/150; level 2, 30/150; level 3, 30/180 AMR mg/m(2) per day/PTX mg/m(2) per day.Twelve patients were enrolled in this study. The dose-limiting toxicity (DLT) of the regimen was assessed during the first cycle. At level 1, all three patients developed a DLT due to grade 4 neutropenia lasting4 days, grade 4 thrombocytopenia and grade 3 febrile neutropenia. Therefore, level 1 was considered the MTD and level 0 was selected as the RD. Objective responses were seen in two patients (response rate 16.7 %). Overall disease control rate was 91.7 %.The combination of AMR and PTX is a feasible and well-tolerated regimen for the treatment of patients with previously treated advanced NSCLC. Although our study included a small number of patients, encouraging disease control and progression-free survival were achieved at the recommended doses. Further clinical trials are warranted.

Gefitinib is an effective first-line chemotherapy for advanced non-small cell lung cancer (NSCLC) patients harboring sensitive EGFR mutations. However, whether second-line platinum combination chemotherapy after first-line gefitinib treatment shows similar effects to first-line platinum combination chemotherapy in these patients remains unclear. Therefore, we here aimed to investigate the efficacy of platinum combination chemotherapy after first-line gefitinib treatment in NSCLC patients harboring sensitive EGFR mutations.We retrospectively evaluated the clinical effects of second-line platinum combination chemotherapy after first-line gefitinib treatment in NSCLC patients harboring sensitive EGFR mutations (exon 19 deletion or exon 21 L858R mutation) at five institutions. All patients were initially treated with gefitinib (250 mg/day) followed by platinum combination chemotherapy as second-line chemotherapy.Between January 2006 and December 2012, 42 patients [8 men, 34 women; median age, 63 years (range 39-75 years)] were enrolled. The overall response rate, disease control rate, and median progression-free survival (PFS) were 26.2, 61.9%, and 5.1 months, respectively, after the second-line treatment. The corresponding values for first-line gefitinib treatment were 69.0, 95.2%, and 11.1 months, respectively. Moreover, second-line platinum combination chemotherapy with pemetrexed or bevacizumab-containing regimens was independently associated with improved PFS.Second-line platinum combination chemotherapy after first-line gefitinib treatment in NSCLC patients harboring sensitive EGFR mutations was effective and showed equivalent outcomes to first-line platinum combination chemotherapy. After failure of first-line gefitinib therapy, second-line platinum combination chemotherapy with pemetrexed or bevacizumab might result in improved PFS.

The ring expansion of zerumbone to a 12-membered ring was studied via a ring opening system or a ring closure system of zerumbone. We succeeded in the synthesis of a zerumbone derivative with 12-membered ring, an allene type zerumbone. For the first time, a Doering–LaFlamme allene synthesis method was adopted and the structure was confirmed by monocrystal X-ray diffraction. It was obtained in total 27.7% yield from zerumbone. We believe that this compound is not only an important building block in synthesizing the BC ring of paclitaxel, but also plays an important role in a novel structure formation and a reactive discovery.

The Sharpless asymmetric epoxidation of (+)-zerumbol, itself obtained by the reduction of zerumbone 1 gave only all-erythro bisepoxide 3 which has five stereogenic centers. Chiral monoepoxide 4, which decomposed easily under acidic or high-temperature conditions, could also be obtained. The reaction mechanism was revealed to be diastereoselective but non-enantioselective monoepoxidation at the 2,3-position, which proceeded quantitatively followed by a second epoxidation at the 10,11-position, and also proceeded extremely enantioselectively giving an efficient kinetic resolution. Novel asymmetric diepoxydihydroxyzerumbol 6 and triepoxyzerumbol 7 each with seven stereogenic centers were formed with complete diastereoselectivity by dihydroxylation and epoxidation of bisepoxyzerumbol, respectively.

The achiral sesquiterpene zerumbone ( 1 ), readily available from a wild ginger, has unique functionality and reactivity which make it a potential starting material for conversion to useful compounds such as paclitaxel, provided that it can easily be transformed to chiral derivatives. Optically active zerumbol 2 and its acetate 3 were synthesized by lipase-catalyzed stereoselective transesterification of racemic 2 . In the best conditions found, a lipase from Pseudomonas fluorescens (Amano AK) and isopropenyl acetate in THF at 30 °C afforded ( R )- 2 and ( S )- 3 with an E value of 56. The absolute configuration of ( R )- 2 was determined by Sharpless epoxidation with l -diethyltartrate ( l -DET) to a bisepoxide of known configuration.

Purpose To determine the relationship between the different isoforms of apolipoprotein E (ApoE) and retinal neovascularization. Methods The concentrations of ApoE and VEGF in vitreous humor samples with either a macular hole (MH), or diabetic macular edema (DME), or proliferative diabetic retinopathy (PDR) with or without intravitreal injection of bevacizumab (IVB) were measured by ELISA. The effects of each isoform of ApoE on human retinal microvascular endothelial cells (HRMECs) in culture or on the retina of oxygen-induced retinopathy (OIR) mice were investigated. Results The concentrations of ApoE and VEGF were significantly higher in the vitreous humor of patients with PDR and DME than in patients with an MH. There was a significant positive correlation between the concentrations of ApoE and VEGF in vitreous humor of patients. In vitro assays showed that ApoE2 and ApoE3, but not ApoE4, promoted the VEGF-induced cell proliferation and migration. In vivo assays showed that intravitreal injections of ApoE2 and ApoE3 increased the number and area of nodes in the retina of OIR mice. Moreover, ApoE was expressed in the vascular endothelial cell in both normal and OIR retinas, but their expression levels were different at postnatal day (P) 12 and P17. Conclusions These results demonstrate that ApoE2 and ApoE3, but not ApoE4, have proangiogenic effects, and the increased expression of ApoE in the vitreous humor of patients with PDR and DME indicates that ApoE2 and ApoE3 are involved in the development of retinal neovascularization in eyes.

The Sharpless asymmetric epoxidation was applied to zerumbol 2 in order to introduce chirality into the readily available achiral sesquiterpene zerumbone 1. Single bisepoxides (+)-3 and (−)-3 were obtained in nearly 100% enantiomeric purity, characterized as the unexpected all erythro configuration by X-ray analysis.

A method for the synthesis of novel disaccharides was developed. It involved the following two steps. The first step consisted of two continuous reactions: the conversion of maltose to β- d -glucose-1-phosphate and d -glucose by the phosphorolytic activity of maltose phosphorylase and the specific consumption of only d -glucose by incubation with glucose-consuming yeast cells. The second step involved the addition of an appropriate carbohydrate and its condensation with the remaining β- d -glucose-1-phosphate by the synthetic activity of maltose phosphorylase or trehalose phosphorylase. Several factors affecting the yields of disaccharides were optimized. Using this method, five maltose-like derivatives and two trehalose-like derivatives were synthesized from maltose and the corresponding carbohydrates. Among these, 4- O -α- d -glucopyranosyl- l -fucopyranose (Glc(α1–4)Fuc) and α- d -glucopyranosyl α- d -fucopyranoside (Glc(α1-1α)Fuc) were purified, and identified by 1 H NMR and 13 C NMR.

1,5-Anhydro-D-glucitol (1,5-AG) was found to inhibit trehalase and trehalose phosphorylase activities competitively, because of its structural similarity with D-glucose. Trehalase from Nocardia sp., one of the most 1,5-AG-sensitive enzymes, was used in the determination of 1,5-AG concentration, which is a useful marker for the diagnosis of diabetes. A good linear relationship was observed between 1,5-AG concentration in the range of 0.02 to 1.0 mM and the extent of trehalase inhibition by 1,5-AG. The 1,5-AG concentration range could be determined by estimating enzymatically the amount of the reaction product, D-glucose, produced by the trehalase.

Maltose phosphorylase (EC 2.4.1.8) from Propionibacterium freudenreichii was purified and characterized. The enzyme ctalyzed both the phosphorolysis and the synthesis of maltose. In particular, in the synthetic reaction, the enzyme could use any of nine sugars other than d -glucose as a sugar acceptor, which resulted in the formation of new disaccharides, in which the first carbon of d -glucose and the fourth carbon of the other sugar were connected by an α-glycosidic linkage.

Objective The purpose of this study was to investigate the relationship between craniofacial and nasopharyngeal morphology and velopharyngeal function in submucous cleft palate. Design and Patients Fifty-two lateral cephalometric radiographs of 46 sub-mucous cleft palate (SMCP) patients with velopharyngeal competence (24 patients) and incompetence (22 patients) at 4 and 7 years of age were studied. The patients had not received any surgical or orthodontic treatment prior to cephalography being performed. Results Significant differences were found between cephalometric variables (N-Ba, N-S-Ba angle) in children with velopharyngeal competence and incompetence. However, the results of our study showed that cephalometric data alone are not useful for predicting velopharyngeal function and can not serve as an absolute prognostic indicator. Conclusion There are many factors that can influence velopharyngeal function in SMCP patients. Cephalometric data did not demonstrate a strong relationship to velopharyngeal function.

Trehalose phosphorylase was purified from the cell extracts of Catellatospora ferruginea. The enzyme had an apparent molecular weight of 400,000 by gel filtration and 98,000 by SDS-PAGE, suggesting that the enzyme was a tetramer. The enzyme was specific for trehalose in phosphorolysis and specific for beta-D-glucose 1-phosphate in synthesis. In addition to D-glucose, D-xylose and D-fucose were also possible sugar acceptors during synthesis. Phosphate ions were a key to the activity and stability of the enzyme, controlling the equilibrium of the reversible reaction and the heat stability of the enzyme. The enzyme was strongly inhibited by p-chloromercuribenzoate and pyridoxal phosphate. The enzyme was inactivated by heat or by storage frozen with ammonium chloride and lithium chloride.

A range of microorganisms was screened for new and high producer strains of trehalose phosphorylase (EC 2.4.1.64). Trehalose phosphorylase activity was found in cells of actinomyctes of the genera Actinomadura, Amycolata, Catellatospora, Kineosporia, and Nocardia. Among them, Catellatospora ferruginea showed the highest enzyme activity. Trehalose phosphorylase from C. ferruginea was able to catalyse both the phosphorolysis of trehalose into beta-glucose 1-phosphate and D-glucose and the synthesis of trehalose from beta-glucose 1-phosphate and D-glucose.