Purpose/Objective(s) Although immune checkpoint inhibitors (ICI) targeting for PD-1 axis is a promising approach for advanced gastric cancer (GC) patients, the response rate is still limited. Induction of synergistic effect of irradiation with ICI targeting for the PD-1 axis can be an attractive strategy. We have conducted and reported a single-arm, phase 1/2 trial against advanced GC treated with combination of palliative radiotherapy and nivolumab, an anti-PD-1 antibody (CIRCUIT trial: NCT03453164). The 3-years follow-up results are presented herein. Materials/Methods The eligible patients were unresectable advanced or recurrent GC who developed progression after primary and secondary chemotherapy with more than 1 lesions assessable (also, 1 lesion must be ≥ 2 cm) in diagnostic imaging. The number of enrolled patients was 41. Thirty-four patients were male and 7 were female. Median age was 70 years old (range = 36–86 years old). Thirty-three patients (81%) had 5 or more cancer lesions and 34 patients (83%) had 2 or more organs with cancer. The biggest or symptomatic tumors were irradiated in 22.5 Gy/5 fractions/5 days (EQD2 = 34.8Gy). Three patients had multiple sites irradiated. Ten patients had additional newly appeared tumors irradiated after first irradiation and 5 patients were re-irradiated after local recurrence (same site). The total number of irradiated field was 65 (primary tumor site (stomach): 11, metastatic lymph node site: 29, metastatic liver tumor site: 14, and other site: 11). Toxicities were graded based on the Common Terminology Criteria for Adverse Events version 4.0. Results The median survival time and progression-free survival time were 218 days (range = 21-1842 days) and 92 days (range = 21-1514 days), respectively. The overall survival rate and progression-free survival rate were 28% and 7% at 1-year, 15% and 7% at 2-year, and 13% and 7% at 3-year, respectively. Three patients have lived without disease progression for more than 3 years. The local control rate (LCR) at 1-, 2- and 3-years for the irradiated tumor were 80%, 67% and 67%, respectively. The LCR at 3-year according to sites were 100% at primary tumor (stomach), 48% at lymph node metastasis, 67% at liver metastasis, and 100% at other site. Severe adverse effect (≥ G3) was occurred in 18 patients (44%). However, severe adverse effect (≥ G3) caused by radiotherapy was not seen. Conclusion Addition of radiotherapy to nivolumab therapy was safe and seemed to be beneficial for survival. Furthermore, good LCR was observed even though palliative irradiation dose.

BACKGROUND:The guideline-recommended low-density lipoprotein cholesterol target level of <70 mg/dL may not be achieved with statin administration in some patients with acute coronary syndrome (ACS). Therefore, the proprotein convertase subtilisin-kexin type 9 (PCSK9) antibody can be added to high-risk patients with ACS. Nevertheless, the optimal duration of PCSK9 antibody administration remains unclear.METHODS AND RESULTS:Patients were randomized to receive either 3 months of lipid lowering therapy (LLT) with the PCSK9 antibody followed by conventional LLT (with-PCSK9-antibody group) or 12 months of conventional LLT alone (without-PCSK9-antibody group). The primary endpoint was the composite of all-cause death, myocardial infarction, stroke, unstable angina, and ischemia-driven revascularization. A total of 124 patients treated with percutaneous coronary intervention (PCI) were randomly assigned to the two groups (n = 62 in each). The primary composite outcome occurred in 9.7% and 14.5% of the patients in the with- and without-PCSK9-antibody groups, respectively (hazard ratio: 0.70; 95% confidence interval: 0.25 to 1.97; p = 0.498). The two groups showed no significant differences in hospitalization for worsening heart failure and adverse events.CONCLUSIONS:In ACS patients who underwent PCI, short-term PCSK9 antibody therapy with conventional LLT was feasible in this pilot clinical trial. Long-term follow-up in a larger scale clinical trial is warranted.

Although basic, translational and clinical research suggest a possibility of synergistic effect of radiation-induced immunogenic cell death with immune checkpoint inhibitors, the effectiveness of concurrent therapy with radiotherapy and immunotherapy is not fully established. Phase I/II, open single arm, prospective clinical trial was conducted and eligible patients were unresectable advanced or recurrent gastric cancer patients (n = 40) who developed progression after primary and secondary chemotherapy with multiple metastasis assessable in imaging (one lesion must be ≥2cm). Radiotherapy of total 22.5 Gy/5 fractions/5 days was given to the largest or symptomatic lesion and nivolumab was administered day 15-22 at 3 mg/kg or 240mg/body every 2 weeks to a total of 6 administrations. The primary endpoint is disease control rate of non-irradiated lesions as an abscopal effect. The secondary endpoints are MST, safety and proportion of local control rate for irradiated lesion. As an ancillary analysis, immunologic parameters including high-dimensional MHC multimer analysis and TCR repertoire analysis, and ctDNA analysis are designed. Total 41 patients were enrolled and 40 patients were evaluated as full analysis set, since 1 patient was judged as an unmatched case to inclusion criteria. The clinical evaluation as an abscopal effect were CR=1 (2.5%), PR=5 (12.5%), SD=3 (7.5%), PD=15 (37.5%) and NE=16 (40%). The disease control rate and the response rate for non-irradiated target lesions was 22.5% and 15%, respectively. The MST was 230 days (157-330, 95%CI) and 1-year survival rate was 28.6%. Any adverse event more than Grade 3 was observed in 16 cases in 41 patients (39%). The most frequent AE more than grade 3 were anemia (19%) and appetite loss (12%). The local control for irradiated lesions were seen in CR=5 (12.5%), PR=6 (15%), SD=5 (12.5%), PD=4 (10%) and NE=20 (50%). The ancillary analysis for immunological monitoring is under investigation. The combination of nivolumab with radiotherapy demonstrated promising anti-tumor activity with marked prolonged survival time in gastric cancer. No new safety issues were detected in the combination.

Although beneficial effects of non-secreting intracellular renin (ns-renin) against ischemia have been reported, the precise mechanism remains unclear. In this study, we investigated the roles of ns-renin and mitochondrial extracellular signal-related kinase (ERK) 1/2 on mitochondrial permeability transition pore (mPTP) opening during ischemia in diabetes mellitus (DM) hearts. When isolated hearts from Wistar rats (non-DM hearts) and Goto-Kakizaki rats (DM hearts) were subjected to ischemia for 70 min by left anterior descending coronary artery ligation, DM hearts exhibited higher left ventricular (LV) developed pressure and lower LV end-diastolic pressure than non-DM hearts, suggesting ischemic resistance. In addition, DM hearts showed increased intracellular renin (int-renin, including secreting and non-secreting renin) in the ischemic area, and a direct renin inhibitor (DRI; aliskiren) attenuated ischemic resistance in DM hearts. ERK1/2 was significantly phosphorylated after ischemia in both whole cell and mitochondrial fractions in DM hearts. In isolated mitochondria from DM hearts, rat recombinant renin (r-renin) significantly phosphorylated mitochondrial ERK1/2, and hyperpolarized mitochondrial membrane potential (ΔΨm) in a U0126 (an inhibitor of mitogen-activated protein kinases/ERK kinases)-sensitive manner. R-renin also attenuated atractyloside (Atr, an mPTP opener)-induced ΔΨm depolarization and Atr-induced mitochondrial swelling in an U0126-sensitive manner in isolated mitochondria from DM hearts. Furthermore, U0126 attenuated ischemic resistance in DM hearts, whereas it did not alter the hemodynamics in non-DM hearts. Our results suggest that the increased int-renin during ischemia may inhibit mPTP opening through activation of mitochondrial ERK1/2, which may be involved in ischemic resistance in DM hearts.

Background: Conventional hands-on chest compression, in cardiopulmonary resuscitation, is often inadequate, especially when the rescuers are weak or have a small physique. Objectives: This study aimed to investigate the potential of leg-foot chest compression, with and without a footstool, during cardiopulmonary resuscitation. Methods and Results: We prospectively enrolled 21 medical workers competent in basic life support. They performed cardiopulmonary resuscitation on a manikin for 2 min using conventional hands-on compression (HO), leg-foot compression (LF), and leg-foot compression with a footstool (LF + FS). We analyzed the compression depths, changes in the rescuers’ vital signs, and the modified Borg scale scores after the trials. The compression depth did not differ between the cases using HO and LF. In the case of LF + FS, compression depths ⩾5 cm were more frequently observed (median, inter-quartile range: 93%, 81%–100%) than in HO (9%, 0%–57%, p < 0.01) and LF (28%, 11%–47%, p < 0.01). The increase in the heart rate or modified Borg scale scores, after the trials, did not differ between the HO and LF group; however, the values were the lowest in the case of LF + FS (49 ± 18 beats/min and 5 (4–7) in HO, 46 ± 18 and 6 (5–7) in LF, and 32 ± 11 and 2 (1–3) in LF + FS, respectively, p < 0.01). However, the increase in blood pressure, SpO2, and respiratory rate were not different among each group. The increases in the heart rate and modified Borg scale scores negatively were correlated with the rescuers’ body size, in the case of HO and LF, but not LF + FS. Conclusion: LF can be used as an alternative to HO, when adequate HO is difficult. LF + FS could be used when rescuers are weak or have a small physique and when the victims are bigger than the rescuers.

Local recurrent tumors after radiotherapy are radio-resistant tumors, and carbon-ion is expected to overcome the refractory disease. We performed prospective study to evaluate the efficacy and safety of re-irradiation using carbon-ion radiotherapy for isolated recurrent tumor. The inclusion criteria were as follows: clinically proven recurrent tumor; measurable tumor by CT or MRI; Age >=16, performance status 0-2; isolated tumor at previously irradiated site; and life expectancy more than 6 months. The exclusion criteria were as follows: tumor invasion to gastrointestinal tract or major blood vessel; uncontrolled infection; early recurrent tumor less than 3 months; and severe concomitant diseases. The primary end-point was the 1-year local control rate, and the secondary end-points included the overall survival rate and adverse events. Local control and overall survival rates were statistically calculated by the Kaplan-Meier method and log-rank tests. A P value < 0.05 was defined as a statistical significance. Acute and late adverse events were evaluated according to the Common Terminology Criteria for Adverse Events, version 4.0. This study was reviewed and approved by our Institutional Review Board (No. 1108). Between December 2013 and March 2016, 22 patients were enrolled in this prospective study. All patients were re-irradiated carbon-ion radiotherapy with radical intent. The median age was 67 years old (range: 17-89), and number of male patients was 13 (59%). There were 7 patients with rectal cancer, 4 with sarcoma, 4 with lung cancer, and 3 with hepatic cell carcinoma, and 4 with other tumors. The median follow-up time was 26 months. Eight patients developed local recurrence, and the 1- and 2-year local control rates were 71% and 60%, respectively. Eight patients died of the cancers and 2 died of other disease. The 1- and 2 year overall survival rates were 76% and 67%, respectively. There were no acute adverse events grade ≧3. There were 4 patients with late adverse events grade ≧3, including 3 patients with grade 3 urinary tract obstruction requiring ureteral stent and 1 with skin ulcer and upper limb disability. There were no late adverse events grade ≧4. Re-irradiation of carbon-ion radiotherapy with radical intent was safely performed with tolerable toxicities for the selected patients. The re-irradiation has the potential to improve the treatment for isolated local recurrent tumor, and the further investigations are required to confirm the therapeutic efficacy. UMIN000014513.

Accumulating evidence has revealed pivotal roles of glycogen synthase kinase-3β (GSK3β) inactivation on cardiac protection. Because the precise mechanisms of cardiac protection against ischemia/reperfusion (I/R) injury by GSK3β-inactivation remain elusive, we investigated the relationship between GSK3β-mediated mitochondrial hexokinase II (mitoHK-II; a downstream target of GSK3β) dissociation and mitochondrial permeability transition pore (mPTP) opening. In Langendorff-perfused hearts, GSK3β inactivation by SB216763 improved the left ventricular-developed pressure and retained mitoHK-II binding after I/R. In permeabilized myocytes, GSK3β depolarized mitochondrial membrane potential with accelerated mitochondrial calcein release (suggesting GSK3β-mediated mPTP opening) and decreased mitoHK-II bindings. GSK3β-mediated mPTP opening depended on mitoHK-II binding, i.e., it was accelerated by dissociation of mitoHK-II (dicyclohexylcarbodiimide) and attenuated by enhancement of mitoHK-II binding (dextran). However, inactivation of mitoHK-II by glucose-depletion or glucose-6-phosphate inhibited the GSK3β-mediated mPTP opening. We conclude that GSK3β-mediated mPTP opening may be involved in I/R injury and regulated by mitoHK-II binding and activity.

Experimental testicular teratomas (ETTs) can be induced in 129/Sv mouse by E12.5 fetal testes transplant into adult testes. Previously, we conducted linkage analysis to explore candidate genes possibly involved in ETT development using F2 intercross fetuses derived from F1[LTXBJ × 129/Sv- + /Ter (+ /+)] hybrids. By linkage analysis on Chr 18 and Chr 19, we identified the genomic locus for experimental testicular teratoma 1 (ett1) on Chr 18. In the present study, we conducted additional mapping and linkage analysis on teratoma susceptibility and genome composition on Chr 1–17. The results revealed two new candidate loci, experimental testicular teratoma 2 (ett2) and experimental testicular teratoma 3 (ett3), on Chr 3 and 7. Interestingly, the rates of ETT generation were increased in the case of ett2 and ett3 regions replaced with LTXBJ strain. To determine whether a polymorphic gene was present, we performed exome analysis of 129/Sv- + /Ter (+ /+) and LTXBJ. This revealed the presence of SNPs in all three loci, ett1 to ett3. ett1 contains polymorphic Mc4r; ett2 contains polymorphic Polr3c, Cd160, and Pdzk1; and ett3 contains polymorphic Prmt3. We found additional loci responsible for ETT formation, namely, ett2 and ett3, and identified candidate genes in these regions by exome analysis.

Purpose: Although beneficial effects of intracellular renin (int-renin) against ischemia were reported in diabetic hearts, the precise mechanisms have remained elusive.We here investigated the roles of int-renin and mitochondrial ERK1/2 on the mitochondrial permeability transition pore (mPTP) opening during ischemia in diabetic hearts.Methods and results: (1) When isolated hearts from Goto-Kakizaki (DM-hearts) and Wistar rats (non-DM-hearts) were subjected to ischemia for 70 min, DMhearts exhibited higher LV developed pressure (113±3 mmHg vs. 90±7 mmHg of non-DM-hearts, P<0.05) and lower LV end-diastolic pressure (10.3±0.7 mmHg vs. 15.9±0.8mmHg of non-DM-hearts, P<0.05) than non-DM-hearts.Immunohistochemistory revealed increased int-renin expression, especially in ischemic area, in DM-hearts.Aliskiren (1 mM; a direct renin inhibitor) abolished the ischemic resistance in DM-hearts.These results suggested that an increased int-renin is involved in the ischemic resistance in DM-hearts.(2) DM-hearts exhibited significant increase in phosphorylated ERK1/2 after ischemia both in whole cell and mitochondrial fraction.In addition, U0126 (0.5 μM; an ERK1/2 inhibitor) abolished the ischemic resistance in DM-hearts, whereas did not altered hemodynamics in the non-DM-hearts.(3) In the isolated mitochondria, both DM-and non-DMhearts exhibited the increased phosphorylation in mitochondrial ERK1/2 after a rat recombinant renin (renin: 100 ng/ml) exposure.Similarly, both DM-and non-DM-hearts showed hyperpolarized mitochondrial membrane potential (DYm) after renin (JC-1 ratio; non-DM: 3.062 of renin, vs. 2.961 of control, P<0.05, DM: 2.812 of renin, vs. 2.629 of control, P<0.05), and U0126 inhibited the renin-induced DYm hyperpolarization (non-DM: 2.912 of renin+U0126 vs. renin, P<0.05, DM: 2.639 of renin+U0126, vs. renin, P<0.05).( 4) Furthermore, renin attenuated the atractyloside (Atr; 50 μM, an mPTP opener)-induced DYm depolarization (2.091 of Atr, vs. 2.246 of Atr+renin, P<0.05) in an U0126-sensitive manner (2.099 of Atr+renin+U0126, vs. Atr+renin, P<0.05) in DM-hearts, suggesting that renin inhibited mPTP opening through the activation of mitochondrial ERK1/2.Conclusions: Our results suggest that the increased int-renin expression after ischemia inhibits mPTP opening through the activated mitochondrial ERK1/2, which may result in the ischemic resistance in DM-hearts.

Amyloid deposits are frequently found in degenerated cardiac valves of elderly patients. However, their precursor proteins have not detected to date. The aim of this study is to analyze the contents to reveal their original proteins. Materials and Methods Amyloid fibrils (AFs) were collected from fresh cardiac valves of aortic stenosis patients. AFs were separated by saline precipitation, collagenase or pepsin digestion and water extraction. AFs were dissolved in 6 M urea and isolated by SDS‐PAGE. Main bands in the gel were analyzed by MS spectrometry. Several candidate proteins were picked up and their location on the histological sections were detected by immunohistochemistry. Fibronectins, fibrinogen alpha chains (FGA), vitronectins, lysozymes and complement 3 were stained whether they were co‐localized with amyloid. Results Only FGA were revealed to be present with amyloid deposits. FGA were scattered in the valves with random orientation and some areas showed co‐localization with amyloid deposits. Conclusion FGA is the precursor protein of localized cardiac valve amyloidosis. Support or Funding Information This work was supported by the Japan Society for the Promotion of Science KAKENHI, Scientific Research Grant Number (c) 15K08375.

Purpose: Diabetes mellitus (DM) is one of most critical risks of heart failure. Although the ischemic resistance in DM hearts have been reported, the precise mechanism remains elusive and involvement of intracellular renin has not been studied. Methods & Results: (1) When isolated hearts from Wistar rats (non-DM) and Goto-Kakizaki rats (DM) were subjected to ischemia for 70 min, DM hearts exhibited higher LV developed pressure (113 ± 3 mmHg vs. 90 ± 7 mmHg of non-DM, P < .05) and lower LV end-diastolic pressure (10.3 ± 0.7 mmHg vs. 15.9 ± 0.8 mmHg of non-DM, P < .05) than those of non-DM hearts, indicating the ischemic resistance in DM hearts. (2) The ischemic resistance in DM hearts was abolished by aliskiren (a direct renin inhibitor) or U0126 (an ERK1/2 inhibitor). (3) DM hearts exhibited the increased intracellular renin and activated mitochondrial ERK1/2. (4) In DM hearts, renin hyperpolarized mitochondrial membrane potential (JC-1; 2.81 ± 0.01 of renin, vs. 2.63 ± 0.01 of control, P < .05) in an U0126-sensitive manner (2.64 ± 0.02 of renin + U0126, vs. renin, P < .05). (5) Renin attenuated the atractyloside (Atr, an mPTP opener)-induced membrane potential depolarization (JC-1; 2.09 ± 0.01 of Atr, vs. 2.25 ± 0.02 of Atr + renin, P < .05) in an U0126-sensitive manner (2.10 ± 0.01 of Atr + renin + U0126, vs. Atr + renin, P < .05) in DM hearts. Conclusions: Intracellular renin may inhibit mPTP opening during ischemia through the activated mitochondrial ERK1/2, which may result in the ischemic resistance in DM hearts.

BACKGROUND:Emerging evidence suggested the preferable effects of eicosapentaenoic acid (EPA; n-3 polyunsaturated fatty acid) against cardiac lipotoxicity, which worsens cardiac function by means of excessive serum free fatty acids due to chronic adrenergic stimulation under heart failure. Nonetheless, the precise molecular mechanisms remain elusive. In this study, we focused on dynamin-related protein-1 (Drp1) as a possible modulator of the EPA-mediated cardiac protection against cardiac lipotoxicity, and investigated the causal relation between AMP-activated protein kinase (AMPK) and Drp1. METHODS AND RESULTS:When differentiated H9c2 myocytes were exposed to palmitate (PAL; saturated fatty acid, 400µM) for 24h, these myocytes showed activation of caspases 3 and 7, enhanced caspase 3 cleavage, depolarized mitochondrial membrane potential, depleted intracellular ATP, and enhanced production of intracellular reactive oxygen species. These changes suggested lipotoxicity due to excessive PAL. PAL enhanced mitochondrial fragmentation with increased Drp1 expression, as well. EPA (50µM) restored the PAL-induced apoptosis, mitochondrial dysfunction, and mitochondrial fragmentation with increased Drp1 expression by PAL. EPA activated phosphorylation of AMPK, and pharmacological activation of AMPK by 5-aminoimidazole-4-carboxamide ribonucleotide ameliorated the PAL-induced apoptosis, mitochondrial dysfunction, and downregulated Drp1. An AMPK knockdown via RNA interference enhanced Drp1 expression and attenuated the protective effects of EPA against the PAL-induced lipotoxicity. CONCLUSION:EPA ameliorates the PAL-induced lipotoxicity via AMPK activation, which subsequently suppresses mitochondrial fragmentation and Drp1 expression. Our findings may provide new insights into the molecular mechanisms of EPA-mediated myocardial protection in heart failure.

A 78-year-old male was admitted to our hospital due to frequent palpitation. His electrocardiogram (ECG) presented regular narrow QRS tachycardia with 170 bpm, and catheter ablation was planned. During electroanatomical mapping of the right atrium (RA) with a multiloop mapping catheter, the catheter head was entrapped nearby the ostium of inferior vena cava. Rotation and traction of the catheter failed to detach the catheter head from the RA wall. Exfoliation of connective tissue twined around catheter tip by forceps, which were designed for endomyocardial biopsy, succeeded to retract and remove the catheter. Postprocedural echocardiography and pathologic examination proved the existence of Chiari’s network. The handling of complex catheters in the RA has a potential risk of entrapment with Chiari’s network.

PurposeTo examine how left ventricular (LV) volume and function affect flow dynamics by analyzing 3D intra-LV vortex features using 4D-Flow.Materials and MethodsTwenty-one patients with preserved (LVEF > 60%) and 14 with impaired LV function (LVEF < 40%) underwent 4D-Flow (at 3T).ResultsIn patients with preserved LV function, the intra-LV vortices developed in both the early and late diastolic phases. The shift of inflow vectors at the basal LV toward the posterior-lateral side of the LV and the mid-ventricular turn of inflow vectors toward the LV outflow could explain clearer vortex formation in the late diastolic phase. In patients with impaired LV function, the intra-LV vortices during the diastolic phase located at the more apical LV were larger and more spherically shaped. Both the distance to the vortex core and the vortex area correlated significantly with LV end-diastolic volume (r = 0.66 and 0.73), LVEF (r = -0.74 and -0.68), LV sphericity index (r = -0.60 and -0.65), and peak filling rate (r = -0.61 and -0.64), respectively (P < 0.01). The intra-LV vortices developed during the systolic phase in 10 cases. In those, some of the particles at the apical LV rotated within the LV, whereas in patients with preserved LV function, all of the particles were directed straight to the ascending aorta with accelerated flow velocity (256.8 120.2 cm/s vs. 414.3 +/- 88.2 cm/s, P < 0.01).ConclusionVortex formation during the diastolic phase may be critical for both LV filling and ejection. 4D-Flow showed the 3D alterations of intra-LV flow dynamics by LV dilatation and dysfunction in a noninvasive and comprehensive manner. J. Magn. Reson. Imaging 2016;44:1493-1503.

The aim of this study was to reveal the pathogenesis of aortic stenosis (AS) and regurgitation (AR) by comparing differences in mechanical and biochemical alterations. We applied scanning acoustic microscopy (SAM) to measure the speed of sound (SOS) through valves to estimate the elasticity and monitor sensitivity to protease treatment, as the SOS is correlated with the stiffness of materials, which is reduced after digestion by proteases. The fibrosa of both the AS and AR groups were stiffer than the fibrosa of the normal group. The AR group displayed significantly stiffer fibrosa than the AS group, with the exception of calcified areas. The AS group showed significantly decreased SOS values following protease digestion, whereas the AR showed little reduction. The AS group presented type III collagen in the fibrosa and the ventricularis. In the AR group, both type I collagen and type III collagen coexisted in the fibrosa and the ventricularis. Upon immunostaining for advanced glycation end-products, the AS group showed sparse, weak staining, whereas the AR group presented a strong, band-like positive reaction in the fibrosa. In conclusion, tissue remodelling associated with damage and repair is associated with AS pathogenesis, whereas static chemical alterations with slow collagen turnover induce AR.