Genomic analysis of multiple myeloma using targeted capture sequencing in the Japanese cohort

Previous genomic studies have revealed the genomic landscape of myeloma cells. Although some of the genomic abnormalities shown are believed to be correlated to the molecular pathogenesis of multiple myeloma and/or clinical outcome, these correlations are not fully understood. The aim of this study is to elucidate the correlation between genomic abnormalities and clinical characteristics by targeted capture sequencing in the Japanese multiple myeloma cohort. We analysed 154 patients with newly diagnosed multiple myeloma. The analysis revealed that the study cohort consisted of a less frequent hyperdiploid subtype (37·0%) with relatively high frequencies of KRAS mutation (36·4%) and IGH-CCND1 translocation (26·6%) compared with previous reports. Moreover, our targeted capture sequencing strategy was able to detect rare IGH-associated chromosomal translocations, such as IGH-CCND2 and IGH-MAFA. Interestingly, all 10 patients harboured MAX mutations accompanied by 14q23 deletion. The patients with del(17p) exhibited an unfavourable clinical outcome, and the presence of KRAS mutation was associated with shorter survival in patients with multiple myeloma, harbouring IGH-CCND1. Thus, our study provides a detailed landscape of genomic abnormalities, which may have potential clinical application for patients with multiple myeloma.     

Nonagonistic Dectin-1 ligand transforms CpG into a multitask nanoparticulate TLR9 agonist

CpG DNA, a ligand for Toll-like receptor 9 (TLR9), has been one of the most promising immunotherapeutic agents. Although there are several types of potent humanized CpG oligodeoxynucleotide (ODN), developing “all-in-one” CpG ODNs activating both B cells and plasmacytoid dendritic cells forming a stable nanoparticle without aggregation has not been successful. In this study, we generated a novel nanoparticulate K CpG ODN (K3) wrapped by the nonagonistic Dectin-1 ligand schizophyllan (SPG), K3-SPG. In sharp contrast to K3 alone, K3-SPG stimulates human peripheral blood mononuclear cells to produce a large amount of both type I and type II IFN, targeting the same endosome where IFN-inducing D CpG ODN resides without losing its K-type activity. K3-SPG thus became a potent adjuvant for induction of both humoral and cellular immune responses, particularly CTL induction, to coadministered protein antigens without conjugation. Such potent adjuvant activity of K3-SPG is attributed to its nature of being a nanoparticle rather than targeting Dectin-1 by SPG, accumulating and activating antigen-bearing macrophages and dendritic cells in the draining lymph node. K3-SPG acting as an influenza vaccine adjuvant was demonstrated in vivo in both murine and nonhuman primate models. Taken together, K3-SPG may be useful for immunotherapeutic applications that require type I and type II IFN as well as CTL induction.CpG oligodeoxynucleotide (CpG ODN) is a short (∼20 bases), single-stranded synthetic DNA fragment containing the immunostimulatory CpG motif, a potent agonist for Toll-like receptor 9 (TLR9), which activates dendritic cells (DCs) and B cells to produce type I interferons (IFNs) and inflammatory cytokines (1, 2) and acts as an adjuvant toward both Th1-type humoral and cellular immune responses, including cytotoxic T-lymphocyte (CTL) responses (3, 4). Therefore, CpG ODN has been postulated as a possible immunotherapeutic agent against infectious diseases, cancer, asthma, and pollinosis (2, 5).There are at least four types of CpG ODN, each of which has a different backbone, sequence, and immunostimulatory properties (6). D-type (also called A) CpG ODNs typically comprise one palindromic CpG motif with a phosphodiester (PO) backbone and phosphorothioate (PS) poly(G) tail, and activates plasmacytoid DCs (pDCs) to produce a large amount of IFN-α but fails to induce pDC maturation and B-cell activation (7, 8). The three other types of ODN consist of a PS backbone. K-type (also called B) CpG ODN contains nonpalindromic multiple CpG motifs, and strongly activates B cells to produce IL-6 and pDCs to maturation but barely produces IFN-α (8, 9). Recently, C and P CpG ODNs have been developed; these contain one and two palindromic CpG sequences, respectively, both of which can activate B cells like K-type and pDC like D-type, although C CpG ODN induces weaker IFN-α production compared with P CpG ODN (10–12).D and P CpG ODNs have been shown to form higher-order structures, Hoogsteen base pairing to form parallel quadruplex structures called G tetrads, and Watson–Crick base pairing between cis- and trans-palindromic portions, respectively, that are required for robust IFN-α production by pDCs (12–14). Although such higher-order structures appear necessary for localization to early endosomes and signaling via TLR9, they suffer from product polymorphisms, aggregation, and precipitation, thereby hampering their clinical application (15). Therefore, only K and C CpG ODNs are generally available as immunotherapeutic agents and vaccine adjuvants for human use (16, 17). Although K CpG ODN enhances the immunogenicity of vaccines targeting infectious diseases and cancers in human clinical trials (6, 17), chemical or physical conjugation between antigen and K CpG ODN is necessary for optimal adjuvant effects. These results indicate that these four (K, D, P, and C) types of CpG ODN have advantages and disadvantages; however, the development of an “all-in-one” CpG ODN activating both B cells and pDCs that forms a stable nanoparticle without aggregation has yet to be accomplished. A better strategy, targeting CpG ODN toward antigen-presenting cells (APCs), is desired to improve immunostimulatory specificity and immunotherapeutic efficacy of CpG ODNs.Schizophyllan (SPG), a soluble β-glucan derived from Schizophyllum commune, is a drug that has been approved in Japan as an enhancer of radiotherapy in cervical carcinoma patients for the last three decades (18). It has been shown to form a complex with polydeoxyadenylic acid (dA) as a triple-helical structure (19). Although we previously demonstrated that mouse and humanized CpG ODN with PO poly(dA) at the 5′ end complexed with SPG enhanced cytokine production and acted as an influenza vaccine adjuvant (20, 21), it has been difficult to achieve high yields of the CpG–SPG complex toward its more efficient and cost-effective preclinical as well as clinical development. Recently, when the PS backbone of the dA sequence was linked to CpG ODN, the efficacy of complex formation was elevated by nearly 100% (22). However, a thorough investigation has yet to be conducted to identify the best humanized CpG sequence and optimization of factors to gain all-in-one activities of the four types of CpG ODN.To do this, we sought to optimize a humanized CpG–SPG complex as a vaccine adjuvant and immunostimulatory agent in humans (in vitro), mice (in vitro and in vivo), and nonhuman primates (in vivo). In this study, we identified a novel K CpG ODN (K3) and SPG complex, namely K3-SPG. It forms a higher-order nanoparticle that can be completely solubilized. We found that this all-in-one K3-SPG displayed a more potent activity than, and different characteristics from, any other type of CpG ODN and previous CpG–SPG complexes.     

Age‐related difference of the association of cardiovascular risk factors with the cardio‐ankle vascular index in the Cardiovascular Prognostic Coupling Study in Japan (the Coupling Registry)

The value of the cardio‐ankle vascular index (CAVI) increases with age. All large‐scale studies of the CAVI have investigated patients <80 years old. Thus, the clinical characteristics of high CAVI in patients aged 80 or more remain unclear. Therefore, we investigated (1) the CAVI in very elderly patients and (2) the determinants of a high CAVI in high‐risk patients, including very elderly patients. The Cardiovascular Prognostic Coupling Study in Japan (Coupling Registry) is a prospective observational study of Japanese outpatients with any cardiovascular risk factors. We enrolled 5109 patients from 30 institutions (average age 68.7 ± 11.4 years, 52.4% males). We investigated the determinants of the CAVI by separating the patients into three groups: 970 middle‐aged (<60 years), 3252 elderly (60‐79 years), and 887 very elderly (≥80 years) patients. The CAVI values of the males were significantly higher those of the females in all age groups (<60 years: 7.81 ± 1.11 vs. 7.38 ± 0.99, P < .001; 60‐79 years: 9.20 ± 1.29 vs. 8.66 ± 1.07, P < .001; ≥80 years: 10.26 ± 1.39 vs. 9.51 ± 1.12, P < .001). In all age groups, the CAVI of the patients with diabetes/glucose tolerance disorder was higher than that of the patients without diabetes/glucose tolerance disorder (<60 years: 7.82 ± 1.22 vs 7.58 ± 1.03, P = .002; 60‐79 years: 9.23 ± 1.20 vs 8.78 ± 1.19, P < .001; ≥80 years: 10.04 ± 1.24 vs 9.75 ± 1.32, P = .002). The determinants of the CAVI in these very elderly patients were age, male sex, low BMI, and mean blood pressure. Diabetes/glucose tolerance disorder and glucose were independently associated with the CAVI in the patients aged <60 years and 60‐79 years, but not in those aged ≥80 years after adjusting for other covariates.     

Efficacy of Using the Japanese Version of the Asthma Control Test for Determing the Level of Asthma Control in Clinical Settings

Background: The Asthma Control Test (ACT) is frequently used for the evaluation of asthma control in clinical care setting because it does not require the use of pulmonary function tests, which can be difficult for general practitioners to use. However, few large-scale studies have investigated the efficacy of the Japanese version ACT (J-ACT) in actual use during clinical care.Methods: The aim of this study was to analyze the efficacy of the J-ACT in a clinical care setting. Using data from a 2008 questionnaire survey including the J-ACT by the Niigata Asthma Treatment Study Group, we compared the ACT scores of 2233 patients with respect to multiple parameters, including the severity by Japanese Society of Allergology and the attack frequency. Using the definition of asthma control partially referred to Global Initiative for Asthma (GINA) guidelines from the survey data, the accuracy screening and determination of optimal ACT cutpoints were performed by retrospective analysis.Results: Cronbach's a for the J-ACT was 0.785. Patients with more severe asthma and more frequent asthma attacks had lower ACT scores than did patients with less severe, less frequent attacks. The optimal ACT cutpoints were 24 for the controlled asthma and 20 for the uncontrolled asthma.Conclusions: Our study, the first large-scale investigation of the efficacy of the J-ACT, determined that this evaluation tool is highly efficacious in establishing the level of asthma control. However, the determination of accurate cutpoints for the J-ACT will require more clear definitions of asthma control in future prospective studies.     

Perceived age of facial features is a significant diagnosis criterion for age‐related carotid atherosclerosis in Japanese subjects: J‐SHIPP study

Aim: Vascular aging is known to be a major determinant of life expectancy. Recently, perceived age was reported to be a better predictor for mortality than chronological age. Based on these findings, we investigated whether or not perceived age was related to atherosclerosis in a general population. Methods: The participants were 273 individuals aged ≥50 years who participated in the Skin‐doc in Anti‐Aging Doc program. Facial photos were taken under a shadowless lamp from three directions (antero‐posterior, and 60° right and left oblique projection) using a high‐resolution digital camera. Perceived age was assessed either by 19 professional nurses in the geriatric ward or using facial identification program software. Carotid intima‐media thickness (IMT), radial augmentation index (AI) and brachial‐ankle pulse wave velocity (baPWV) were measured as indices for atherosclerosis. Results: The perceived age difference (expressed as the difference between perceived age and chronological age), when estimated either by nurses or software, was significantly and negatively associated with chronological age. Subjects who were evaluated by nurses to be younger than their chronological age had significantly lower carotid IMT after adjustment for chronological age. Conversely, carotid IMT was an independent and negative determinant of looking young, as perceived by nurses. Similar observations were also made between perceived age using facial identification software and carotid IMT. Radial AI and baPWV were not associated with perceived age. Conclusion: These findings show that carotid atherosclerosis is related to perceived age. This association might underlie previous findings showing that perceived age predicts life expectancy. Geriatr Gerontol Int 2012; ??: ??–??.     

Usefulness of extent analysis for statistical parametric mapping with asymmetry index using inter-ictal FGD-PET in mesial temporal lobe epilepsy

Objective

Inter-ictal ¹⁸F-2-fluoro-deoxy-d-glucose-positron emission tomography (FDG-PET) is widely used for preoperative evaluation to identify epileptogenic zones in patients with temporal lobe epilepsy. In this study, we combined statistical parametric mapping (SPM) with the asymmetry index and volume-of-interest (VOI) based extent analysis employing preoperative FDG-PET in unilateral mesial temporal lobe epilepsy (MTLE) patients. We also evaluated the detection utility of these techniques for automated identification of abnormalities in the unilateral hippocampal area later confirmed to be epileptogenic zones by surgical treatment and subsequent good seizure control.

Methods

FDG-PET scans of 17 patients (9 males, mean age 35?years, age range 16?C60?years) were retrospectively analyzed. All patients had been preoperatively diagnosed with unilateral MTLE. The surgical outcomes of all patients were Engel class 1A or 1B with postoperative follow-up of 2?years. FDG-PET images were spatially normalized and smoothed. After two voxel-value adjustments, one employing the asymmetry index and the other global normalization, had been applied to the images separately, voxel-based statistical comparisons were performed with 20 controls. Peak analysis and extent analysis in the VOI in the parahippocampal gyrus were conducted for SPM. For the extent analysis, a receiver operating characteristic (ROC) curve was devised to calculate the area under the curve and to determine the optimal threshold of extent.

Results

The accuracy of the method employing the asymmetry index was better than that of the global normalization method for both the peak and the extent analysis. The ROC analysis results, for the extent analysis, yielded an area under the curve of 0.971, such that the accuracy and optimal extent threshold of judgment were 92 and 32.9%, respectively.

Conclusion

Statistical z-score mapping with the asymmetry index was more sensitive for detecting regional glucose hypometabolism and more accurate for identifying the side harboring the epileptogenic zone using inter-ictal FDG-PET in unilateral MTLE than z-score mapping with global normalization. Moreover, the automated determination of the side with the epileptogenic zone in unilateral MTLE showed improved accuracy when the combination of SPM with the asymmetry index and extent analysis was applied based on the VOI in the parahippocampal gyrus.     

Reduced blood flow and preserved vasoreactivity characterize oxygen hypometabolism due to incomplete infarction in occlusive carotid artery diseases.

Recent studies have clarified that hemodynamically compromised patients are at high risk for subsequent stroke. The acetazolamide test is widely used to detect the patients with hemodynamic compromise due to occlusive carotid artery disease. Previous studies have suggested that patients with impaired reactivity to acetazolamide had an increased oxygen extraction fraction (OEF) on PET. However, the underlying pathophysiology has not been defined in patients with reduced blood flow and preserved reactivity to acetazolamide due to carotid occlusive diseases regardless of a normal appearance on MRI. This study aimed to clarify hemodynamic and metabolic parameters in such patients, using (15)O gas and (11)C-flumazenil (FMZ) PET. METHODS: Our study included 15 patients who had reduced cerebral blood flow (CBF) and preserved cerebrovascular reactivity (CVR) to acetazolamide in the ipsilateral middle cerebral artery territory due to occlusive carotid diseases on N-isopropyl-p-(123)I-iodoamphetamine ((123)I-IMP) SPECT. We determined the CBF, cerebral metabolic rate for oxygen (CMRO(2)), cerebral blood volume (CBV), and OEF using (15)O gas PET. The binding potential for (11)C-FMZ was also measured in 5 patients. All patients were medically treated and followed-up during a mean period of 2.7 y. RESULTS: (15)O gas PET scans revealed that the ipsilateral CBF and CMRO(2) were reduced to 80% +/- 11% (P < 0.0001) and 78% +/- 8% (P < 0.0001) of the contralateral side, respectively. However, there was no significant side-to-side difference in the CBV and OEF. The ipsilateral binding potential for (11)C-FMZ was also significantly reduced to 82% +/- 2% of the contralateral side (P < 0.05), being very similar to the asymmetry of the CBF and CMRO(2). No patients suffered further ischemic stroke in the ipsilateral hemisphere during the follow-up period. CONCLUSION: Our results strongly suggest that a reduced CBF and a normal CVR characterize oxygen hypometabolism probably due to ischemia-related neuronal loss-namely, incomplete infarction. Such an ischemic lesion is not hemodynamically compromised and is at very low risk for a subsequent ischemic stroke even if the patient is medically treated.