CAR-Modified Vγ9Vδ2 T Cells Propagated Using a Novel Bisphosphonate Prodrug for Allogeneic Adoptive Immunotherapy.

The benefits of CAR-T therapy could be expanded to the treatment of solid tumors through the use of derived autologous αß T cell, but clinical trials of CAR-T therapy for patients with solid tumors have so far been disappointing. CAR-T therapy also faces hurdles due to the time and cost intensive preparation of CAR-T cell products derived from patients as such CAR-T cells are often poor in quality and low in quantity. These inadequacies may be mitigated through the use of third-party donor derived CAR-T cell products which have a potent anti-tumor function but a constrained GVHD property. Vγ9Vδ2 TCR have been shown to exhibit potent antitumor activity but not alloreactivity. Therefore, in this study, CAR-T cells were prepared from Vγ9Vδ2 T (CAR-γδ T) cells which were expanded by using a novel prodrug PTA. CAR-γδ T cells suppressed tumor growth in an antigen specific manner but only during a limited time window. Provision of GITR co-stimulation enhanced anti-tumor function of CAR-γδ T cells. Our present results indicate that, while further optimization of CAR-γδ T cells is necessary, the present results demonstrate that Vγ9Vδ2 T cells are potential source of 'off-the-shelf' CAR-T cell products for successful allogeneic adoptive immunotherapy.

Efficacy and feasibility of cryoballoon ablation for atrial fibrillation in patients with heart failure: A large-scale multicenter study.

INTRODUCTION: Data are limited regarding outcomes of cryoballoon ablation for atrial fibrillation (AF) in patients with heart failure (HF). This large-scale multicenter study aimed to evaluate the prognosis of patients with HF after cryoballoon ablation for AF. METHODS: Among 3655 patients undergoing cryoballoon ablation at 17 institutions, 549 patients (15%) (391 with paroxysmal AF and 158 with persistent AF) diagnosed with HF preoperatively were analyzed. Clinical endpoints were recurrence, mortality, and HF hospitalization after ablation. RESULTS: Most patients had a preserved left ventricular ejection fraction (LVEF) ≥ 50%. During a mean follow-up period of 25.7 months, recurrence, all-cause death, and HF hospitalization occurred in 29%, 4.0%, and 4.8%, respectively. Cardiac function on echocardiography and B-type natriuretic peptide (BNP) levels significantly improved postoperatively, and the effect was more pronounced in the nonrecurrence group. Major complications occurred in 33 patients (6.0%), but most complications were phrenic nerve palsy (3.6%). Although death and HF hospitalization occurred more frequently in patients with LVEF ≤ 40% (n = 73) and New York Heart Association (NYHA) class III-IV (n = 19) than other subgroups, the BNP levels, and LVEF significantly improved after ablation in all LVEF and NYHA class subgroups. High BNP levels, NHYA class, CHADS2 score, and structural heart disease, but not postablation recurrence, independently predicted death, and HF hospitalization on multivariate analysis. The patients with tachycardia-induced cardiomyopathy had better recovery of BNP levels and LVEF after ablation than those with structural heart disease. CONCLUSIONS: Cryoballoon ablation for AF in HF patients is feasible and leads to significantly improved cardiac function.

Impact of heart failure severity on bone mineral density among older patients with heart failure.

The study aimed to identify factors related to bone mineral density (BMD) among older patients with heart failure (HF). A total of 70 consecutive patients with HF aged 65 years or older who were admitted to an acute hospital due to worsening condition were enrolled before discharge. BMD of the femoral neck was evaluated using the DEXA method. Physical function, as well as echocardiographic and laboratory findings including biomarker of HF severity were collected. Bivariate and multiple regression analyses were employed to determine the association between BMD and the clinical variables. Bivariate analysis determined that age, grip strength, walking speed, serum albumin, and N-terminal pro B-type natriuretic peptide (NT-proBNP) were significantly correlated with BMD (P < 0.01), whereas other clinical parameters were not. The multiple regression analysis identified NT-proBNP as an independent related factor for BMD after adjusting with confounding clinical variables. NT-proBNP was independently related to BMD among older patients with HF. Our results suggest the inclusion of bone fracture prevention strategies in disease management programs, especially for older patients with HF.

Antitumor activity of CAR-T cells targeting the intracellular oncoprotein WT1 can be enhanced by vaccination.

The recent success of chimeric antigen receptor (CAR)-T cell therapy for treatment of hematologic malignancies supports further development of treatments for both liquid and solid tumors. However, expansion of CAR-T cell therapy is limited by the availability of surface antigens specific for the tumor while sparing normal cells. There is a rich diversity of tumor antigens from intracellularly expressed proteins that current and conventional CAR-T cells are unable to target. Furthermore, adoptively transferred T cells often suffer from exhaustion and insufficient expansion, in part, because of the immunosuppressive mechanisms operating in tumor-bearing hosts. Therefore, it is necessary to develop means to further activate and expand those CAR-T cells in vivo. The Wilms tumor 1 (WT1) is an intracellular oncogenic transcription factor that is an attractive target for cancer immunotherapy because of its overexpression in a wide range of leukemias and solid tumors, and a low level of expression in normal adult tissues. In the present study, we developed CAR-T cells consisting of a single chain variable fragment (scFv) specific to the WT1235-243/HLA-A*2402 complex. The therapeutic efficacy of our CAR-T cells was demonstrated in a xenograft model, which was further enhanced by vaccination with dendritic cells (DCs) loaded with the corresponding antigen. This enhanced efficacy was mediated, at least partly, by the expansion and activation of CAR-T cells. CAR-T cells shown in the present study not only demonstrate the potential to expand the range of targets available to CAR-T cells, but also provide a proof of concept that efficacy of CAR-T cells targeting peptide/major histocompatibility complex can be boosted by vaccination.

Predictive impact of early mobilization on rehospitalization for elderly Japanese heart failure patients.

The aim of this study was to determine whether early mobilization was associated with rehospitalization among elderly heart failure patients. We measured the time from admission to mobilization and other clinical characteristics for 190 heart failure patients (mean age, 80.7 years). The primary outcome was heart failure rehospitalization. Kaplan-Meier survival curves were plotted and the hazard ratios for rehospitalization were determined using Cox proportional hazards regression models. During a median follow-up period of 750 days, 58 patients underwent rehospitalization. The time from admission to mobilization was significantly longer for these patients than for those who were not rehospitalized. Univariate and multivariate Cox proportional hazards analyses showed that the time from admission to mobilization was an independent predictor of rehospitalization, and receiver-operating characteristic analysis determined an optimal cutoff value of 3 days for differentiating the patients more likely to experience a subsequent cardiac event (sensitivity, 76%; specificity, 69%; area under the curve, 0.667). Kaplan-Meier survival curve analysis showed a significantly lower event rate in the ≤ 3-day group (p = 0.001, log-rank test). In conclusion, the time from admission to mobilization may be one of the strongest predictors of rehospitalization in elderly heart failure patients. Early mobilization within 3 days may be an initial target for the acute phase treatment of heart failure.

Usefulness of Exchanged Protein Directly Activated by cAMP (Epac)1-Inhibiting Therapy for Prevention of Atrial and Ventricular Arrhythmias in Mice.

BACKGROUND: It has been suggested that protein directly activated by cAMP (Epac), one of the downstream signaling molecules of ß-adrenergic receptor (ß-AR), may be an effective target for the treatment of arrhythmia. However, there have been no reports on the anti-arrhythmic effects or cardiac side-effects of Epac1 inhibitors in vivo. Methods and Results: In this study, the roles of Epac1 in the development of atrial and ventricular arrhythmias are examined. In addition, we examined the usefulness of CE3F4, an Epac1-selective inhibitor, in the treatment of the arrhythmias in mice. In Epac1 knockout (Epac1-KO) mice, the duration of atrial fibrillation (AF) was shorter than in wild-type mice. In calsequestrin2 knockout mice, Epac1 deficiency resulted in a reduction of ventricular arrhythmia. In both atrial and ventricular myocytes, sarcoplasmic reticulum (SR) Ca2+ leak, a major trigger of arrhythmias, and spontaneous SR Ca2+ release (SCR) were attenuated in Epac1-KO mice. Consistently, CE3F4 treatment significantly prevented AF and ventricular arrhythmia in mice. In addition, the SR Ca2+ leak and SCR were significantly inhibited by CE3F4 treatment in both atrial and ventricular myocytes. Importantly, cardiac function was not significantly affected by a dosage of CE3F4 sufficient to exert anti-arrhythmic effects. CONCLUSIONS: These findings indicated that Epac1 is involved in the development of atrial and ventricular arrhythmias. CE3F4, an Epac1-selective inhibitor, prevented atrial and ventricular arrhythmias in mice.

Vidarabine, an anti-herpesvirus agent, prevents catecholamine-induced arrhythmias without adverse effect on heart function in mice.

Sympathetic activation causes clinically important arrhythmias including atrial fibrillation (AF) and ventricular tachyarrhythmia. Although the usefulness of ß-adrenergic receptor blockade therapy is widely accepted, its multiple critical side effects often prevent its initiation or continuation. The aim of this study is to determine the advantages of vidarabine, an adenylyl cyclase (AC)-targeted anti-sympathetic agent, as an alternative treatment for arrhythmia. We found that vidarabine, which we identified as a cardiac AC inhibitor, consistently shortens AF duration and reduces the incidence of sympathetic activation-induced ventricular arrhythmias. In atrial and ventricular myocytes, vidarabine inhibits adrenergic receptor stimulation-induced RyR2 phosphorylation, sarcoplasmic reticulum (SR) Ca2+ leakage, and spontaneous Ca2+ release from SR, the last of which has been considered as a potential arrhythmogenic trigger. Moreover, vidarabine also inhibits sympathetic activation-induced reactive oxygen species (ROS) production in cardiac myocytes. The pivotal role of vidarabine's inhibitory effect on ROS production with regard to its anti-arrhythmic property has also been implied in animal studies. In addition, as expected, vidarabine exerts an inhibitory effect on AC function, which is more potent in the heart than elsewhere. Indexes of cardiac function including ejection fraction and heart rate were not affected by a dosage of vidarabine sufficient to exert an anti-arrhythmic effect. These findings suggest that vidarabine inhibits catecholamine-induced AF or ventricular arrhythmia without deteriorating cardiac function in mice.

Role of G protein-regulated inducer of neurite outgrowth 3 (GRIN3) in ß-arrestin 2-Akt signaling and dopaminergic behaviors.

The G protein-regulated inducer of neurite growth (GRIN) family has three isoforms (GRIN1-3), which bind to the Gαi/o subfamily of G protein that mediate signal processing via G protein-coupled receptors (GPCRs). Here, we show that GRIN3 is involved in regulation of dopamine-dependent behaviors and is essential for activation of the dopamine receptors (DAR)-ß-arrestin signaling cascade. Analysis of functional regions of GRIN3 showed that a di-cysteine motif (Cys751/752) is required for plasma membrane localization. GRIN3 was co-immunoprecipitated with GPCR kinases 2/6 and ß-arrestins 1/2. Among GRINs, only GRIN3, which is highly expressed in striatum, strongly interacted with ß-arrestin 2. We also generated GRIN3-knockout mice (GRIN3KO). GRIN3KO exhibited reduced locomotor activity and increased anxiety-like behavior in the elevated maze test, as well as a reduced locomoter response to dopamine stimulation. We also examined the phosphorylation of Akt at threonine 308 (phospho308-Akt), which is dephosphorylated via a ß-arrestin 2-mediated pathway. Dephosphorylation of phospho308-Akt via the D2R-ß-arrestin 2 signaling pathway was completely abolished in striatum of GRIN3KO. Our results suggest that GRIN3 has a role in recruitment and assembly of proteins involved in ß-arrestin-dependent, G protein-independent signaling.

The role of Epac in the heart.

As one of the most important second messengers, 3',5'-cyclic adenosine monophosphate (cAMP) mediates various extracellular signals including hormones and neurotransmitters, and induces appropriate responses in diverse types of cells. Since cAMP was formerly believed to transmit signals through only two direct target molecules, protein kinase A and the cyclic nucleotide-gated channel, the sensational discovery in 1998 of another novel direct effecter of cAMP [exchange proteins directly activated by cAMP (Epac)] attracted a great deal of scientific interest in cAMP signaling. Numerous studies on Epac have since disclosed its important functions in various tissues in the body. Recently, observations of genetically manipulated mice in various pathogenic models have begun to reveal the in vivo significance of previous in vitro or cellular-level findings. Here, we focused on the function of Epac in the heart. Accumulating evidence has revealed that both Epac1 and Epac2 play important roles in the structure and function of the heart under physiological and pathological conditions. Accordingly, developing the ability to regulate cAMP-mediated signaling through Epac may lead to remarkable new therapies for the treatment of cardiac diseases.

[Diffuse Malignant Peritoneal Mesothelioma with Secondary Liver Invasion Diagnosed Using Laparoscopy - A Case Report].

A 69-year-old man with right upper quadrant abdominal pain and fever was referred to our hospital. He had a history of asbestosis exposure. Computed tomography(CT)revealed a mass at the right subhepatic space, and an antibiotic was administered after a diagnosis of an abdominal abscess. However, the patient did not respond to the treatment, and finally, exploratory laparoscopy was performed. A sheet of combined white nodules surrounding the right lobe of the liver was found, and the mass was continuous with the covering particles. Biopsy of the mass and immunohistochemical examination was performed. The resulting diagnosis was diffuse epithelial malignant peritoneal mesothelioma(MPM). Postoperative systematic chemotherapy of pemetrexed and cisplatin was administered. Laparoscopy was useful to evaluate the distribution of the MPM, which led to adequate therapeutic determination.

[A Case of Metastatic Carcinoma of Anal Fistula].

A 58-year-old man underwent rectal resection(D2 dissection)for rectal cancer and liver/lung metastases in August 2009. Histopathological findings were Ra, type 2, 70×80mm, tub1>tub2, int, pSI(peritoneum), INF b, ly1, v1, pN1(2/13), pPM0, pDM0, M1a(H1, PUL1), fStage IV . The lung metastasis had disappeared on chest CT after postoperative chemotherapy and we were able to perform radical resection of the liver metastasis by performing hepatectomy twice. In October 2013, anal pain appeared and a painful tumor approximately 2 cm in size was found in the 5 o'clock direction of the anus. Biopsy revealed a well-differentiated tubular adenocarcinoma similar to rectal cancer, and it was diagnosed as a fistula metastasis of rectal cancer.We performed chemoradiotherapy(S-1 120mg/day plus RT 60 Gy/30 Fr)as surgery was recommended but refused. As a result, the tumor reduced markedly in size. In December 2015, the tumor enlarged in size and the patient and family requested surgery. We, therefore, performed abdominoperineal resection. Currently, the patient is alive at 18 months after surgery with no recurrence.

[Laparoscopic Distal Gastrectomy for Elderly Patients with Advanced Gastric Cancer and Gastric Outlet Obstruction - A Case Report].

A 90-year-old female patient was admitted to our hospital with a chief complaint of vomiting.Gastroscopy revealed type 3 gastric cancer and gastric outlet obstruction(GOO).Abdominal computed tomography revealed thickening of the antral wall and suggested the presence of 3 perigastric lymph node metastases, but there was no ascitic fluid or distant metastasis.The clinical diagnosis was T4a(SE)N2H0CYXP0M0, Stage III B, according to the Japanese Classification of Gastric Carcinoma.Her general conditions including kidney and cardiac function were good, we considered that she was able to tolerate radical distal gastrectomy.We planned laparoscopic distal gastrectomy(LDG)and D2 lymphadenectomy after getting sufficient informed consent.The patient experienced an uneventful post-operative recovery, and was discharged in good health 11 days after surgery.

[Early Liver Metastases after Curative Surgery for Stageâ Gastric Cancer with Neuroendocrine Differentiation-A Case Report].

An 87-year-old male patient was admitted to our hospital with a chief complaint of vomiting. Gastroscopy revealed Type 0-Ⅱc+Ⅱa tumor at the posterior wall in the middle third of the stomach. A biopsy indicated moderately differentiated adenocarcinoma. Abdominal CT revealed no lymph node or distant metastases. The clinical diagnosis was cT2(MP), N0, M0, cStage Ⅰ. Laparoscopic distal gastrectomy with D2 lymphadenectomy was performed. The pathological findings revealed moderately differentiated adenocarcinoma containing synaptophysin, chromogranin A, and CD56-positive tumor cells. He was then diagnosed with adenocarcinoma with neuroendocrine differentiation. The pathological diagnosis was pT2(MP), pN0, M0, pStage ⅠB. MRI revealed multiple liver metastases 5 months postoperatively. S-1 alone chemotherapy was started, and the patient showed partial response(PR)after 3 courses, according to the Response Evaluation Criteria in Solid Tumor (RECIST).

[Invasive Micropapillary Carcinoma of the Colon That Was Difficult to Diagnose Preoperatively].

A 73-year-old man was admitted with sigmoid colon diverticulitis. Although a biopsy did not indicate malignancy, the sigmoid colon was completely obstructed following conservative treatment. After sigmoidectomy, the histopathological findings revealed a well-differentiated adenocarcinoma localized to the mucosal surface; invasive micropapillary carcinoma (IMPC)accounting for>95% of the tumor volume spread extensively below the submucosal layer. IMPC is highly malignant and difficult to diagnose preoperatively, possibly due to the presence of poorly differentiated histological sub-types in the deepest portions of the tumor.

[Sigmoid colon cancer with intussusception prolapsing through the anus treated by elective laparoscopic radical surgery].

An 86-year-old man was brought in ambulance to our hospital because of sudden hematochezia and abdominal pain during defecation. Intestinal prolapse approximately 80mm from the anus and a type 1 tumor 50mm in size on the mucosal surface were detected. The intestinal prolapse was manually repositioned, and the reduction of the intussusception was confirmed by computed tomography (CT). Following colonoscopy and abdominal-enhanced CT, a sigmoid colon cancer without distant metastases was detected. Elective laparoscopic radical surgery was performed. The present study described a rare case of sigmoid colon cancer with an intussusception prolapsing through the anus and highlighted the treatment strategy by reviewing 48 previous cases. The treatment strategy employed was as follows:first, manual repositioning of the intestinal prolapse was attempted;and second, the presence of intussusception was confirmed by CT. In cases when repositioning of the intussusception was not possible, even with the use of an endoscope or contrast enema, emergency surgery was required.

Cardiac overexpression of Epac1 in transgenic mice rescues lipopolysaccharide-induced cardiac dysfunction and inhibits Jak-STAT pathway.

Pro-inflammatory cytokines are released in septic shock and impair cardiac function via the Jak-STAT pathway. It is well known that sympathetic stimulation leads to coupling of the ß-adrenergic receptor/Gs/adenylyl cyclase, a membrane-bound enzyme that catalyzes the conversion of ATP to cAMP, thereby stimulating protein kinase A (PKA) and ultimately compensating for cardiac dysfunction. The mechanism of such compensation by catecholamine has been traditionally understood as PKA-mediated enforcement of cardiac contractility. We hypothesized that exchange protein activated by cyclic AMP (Epac), a new target of cAMP signaling that functions independently of protein kinase A, also plays a key role in protection against acute stresses or changes in hemodynamic overload. Lipopolysaccharide injection induced cytokine release and severe cardiac dysfunction in mouse. In mouse overexpressing Epac1 in the heart, however, the magnitude of such dysfunction was significantly smaller. Epac1 overexpression inhibited the Jak-STAT pathway, as indicated by decreased phosphorylation of STAT3 and increased SOCS3 expression, with subsequent inhibition of iNOS expression. In cultured cardiomyocytes treated with isoproterenol or forskolin, the increase of SOCS3 expression was blunted when Epac1 or PKCα was silenced with siRNA. Activation of the cAMP/Epac/PKCα pathway protected the heart against cytokine-induced cardiac dysfunction, suggesting a new role of catecholamine signaling in compensating for cardiac dysfunction in heart failure. Epac1 and its downstream pathways may be novel targets for treating cardiac dysfunction in endotoxemia.

Prediction of difficult laparoscopic cholecystectomy for acute cholecystitis.

BACKGROUND: No report has described the predictive factor of surgical difficulty for laparoscopic cholecystectomy (LC) by preoperative computed tomography (CT) findings. This study aimed to investigate whether dynamic CT findings can predict the difficulty of LC for acute cholecystitis. MATERIALS AND METHODS: Fifty-seven patients who underwent emergency LC and dynamic CT preoperatively were enrolled. Difficult LC (DLC) was defined as any patient with an operative time ≥3 h, bleeding volume ≥300 mL, common bile duct injury, partial cholecystectomy, the need for a second surgeon, and/or conversion to open surgery. Patients were assigned to either the DLC (+) or DLC (-) group. We determined the CT attenuation ratio of the arterial phase (ARAP) to represent the degree of transient focal enhancement of the liver adjacent to the gallbladder. The ARAP cutoff value for a DLC predictor was determined using receiver operating characteristic curve analysis. Patients' characteristics and CT findings, including the ARAP, were compared between the groups. The Fisher exact test for categorical variables and the Mann-Whitney U test for continuous variables with Bonferroni correction were used to evaluate the significance of differences. RESULTS: Fifteen patients were assigned to the DLC (+) group. The ARAP was significantly higher in the DLC (+) group than in the DLC (-) group (P = 0.006). The ARAP cutoff value was 1.55. Regarding the CT findings, an ARAP ≥1.55 (P = 0.005) was significantly correlated with DLC. CONCLUSIONS: Among dynamic CT findings, an increased ARAP is a predictive factor for DLC.

The iron chelating agent, deferoxamine detoxifies Fe(Salen)-induced cytotoxicity.

Iron-salen, i.e., µ-oxo-N,N'-bis(salicylidene)ethylenediamine iron (Fe(Salen)) was a recently identified as a new anti-cancer compound with intrinsic magnetic properties. Chelation therapy has been widely used in management of metallic poisoning, because an administration of agents that bind metals can prevent potential lethal effects of particular metal. In this study, we confirmed the therapeutic effect of deferoxamine mesylate (DFO) chelation against Fe(Salen) as part of the chelator antidote efficacy. DFO administration resulted in reduced cytotoxicity and ROS generation by Fe(Salen) in cancer cells. DFO (25 mg/kg) reduced the onset of Fe(Salen) (25 mg/kg)-induced acute liver and renal dysfunction. DFO (300 mg/kg) improves survival rate after systematic injection of a fatal dose of Fe(Salen) (200 mg/kg) in mice. DFO enables the use of higher Fe(Salen) doses to treat progressive states of cancer, and it also appears to decrease the acute side effects of Fe(Salen). This makes DFO a potential antidote candidate for Fe(Salen)-based cancer treatments, and this novel strategy could be widely used in minimally-invasive clinical settings.

[A Case of Rectal Cancer Presenting with Schistosomiasis Japonica Eggs].

A 87-year-old man had used to live in Shizuoka Prefecture. He got colonoscopy due to melena, and a type 1 tumor about 3 cm was detected in the rectum. He was diagnosed with rectal cancer. We performed a laparoscopic low anterior resection with lymphadenectomy. Histopathological findings shows tub1, pSM(7mm), med, INF a, ly1, v0, pPM0, pDM0, pN0(0/7), T1bN0M0, Stage I . Schistosoma japonica eggs were seen in submucosal of the rectum not around the tumor but also normal tissue. The eggs embolized microvessels. It has been 5 year since the operation, the patient has survived without recurrence.

[Esophagogastric Junction Cancer Resection with Bilateral Adrenal Metastasis - A Case Report].

The patient was a 64-year-old man with esophagogastric junction cancer. We performed right thoracotomy-laparotomy for lower esophageal and cardiac gastric resection, D2 lymphadenectomy, and reconstruction of a gastric tube in October 2011. Histopathology confirmed T4aN1M1(LYM), Stage IV cancer(Japanese Classification of Gastric Carcinoma, 14th edition) with R0 resection. Because of preexisting alcoholic cirrhosis, postoperative chemotherapy was not an option. In March 2014, we performed left adrenalectomy for left adrenal metastasis, and in December 2014, we performed right adrenalectomy for metastasis to the right adrenal gland. The patient was prescribed 20mg/day of hydrocortisone postoperatively. Survival from the right adrenalectomy was 2 years and 2 months, and survival from the first operation was 5 years and 4 months, without recurrence. This case of esophagogastric junction cancer resection with bilateral adrenal metastasis is rare, with only one previously reported case in Japan.