Increased mTORC2 pathway activation in lymph nodes of iMCD‐TAFRO

Idiopathic multicentric Castleman disease (iMCD) is a rare and life‐threatening haematologic disorder involving polyclonal lymphoproliferation and organ dysfunction due to excessive cytokine production, including interleukin‐6 (IL‐6). Clinical trial and real‐world data demonstrate that IL‐6 inhibition is effective in 34–50% of patients. mTOR, which functions through mTORC1 and mTORC2, is a recently discovered therapeutic target. The mTOR inhibitor sirolimus, which preferentially inhibits mTORC1, has led to sustained remission in a small cohort of anti‐IL‐6‐refractory iMCD patients with thrombocytopenia, anasarca, fever, renal dysfunction and organomegaly (iMCD‐TAFRO). However, sirolimus has not shown uniform effect, potentially due to its limited mTORC2 inhibition. To investigate mTORC2 activation in iMCD, we quantified the mTORC2 effector protein pNDRG1 by immunohistochemistry of lymph node tissue from six iMCD‐TAFRO and eight iMCD patients who do not meet TAFRO criteria (iMCD‐not‐otherwise‐specified; iMCD‐NOS). mTORC2 activation was increased in all regions of iMCD‐TAFRO lymph nodes and the interfollicular space of iMCD‐NOS compared with control tissue. Immunohistochemistry also revealed increased pNDRG1 expression in iMCD‐TAFRO germinal centres compared with autoimmune lymphoproliferative syndrome (ALPS), an mTOR‐driven, sirolimus‐responsive lymphoproliferative disorder, and comparable staining between iMCD‐NOS and ALPS. These results suggest increased mTORC2 activity in iMCD and that dual mTORC1/mTORC2 inhibitors may be a rational therapeutic approach.     

Identification and characterization of the retinoic acid response elements in the human RIG1 gene promoter

The expression of retinoic acid-induced gene 1 (RIG1), a class II tumor suppressor gene, is induced in cells treated with retinoids. RIG1 has been shown to express ubiquitously and the increased expression of this gene appears to suppress cell proliferation. Recent studies also demonstrated that this gene may play an important role in cell differentiation and the progression of cancer. In spite of the remarkable regulatory role of this protein, the molecular mechanism of RIG1 expression induced by retinoids remains to be clarified. The present study was designed to study the molecular mechanism underlying the all-trans retinoic acid (atRA)-mediated induction of RIG1 gene expression. Polymerase chain reaction was used to generate a total of 10 luciferase constructs that contain various fragments of the RIG1 5'-genomic region. These constructs were then transfected into human gastric cancer SC-M1 and breast cancer T47D cells for transactivation analysis. atRA exhibited a significant induction in luciferase activity only through the -4910/-5509 fragment of the 5'-genomic region of RIG1 gene relative to the translation initiation site. Further analysis of this promoter fragment indicated that the primary atRA response region is located in between -5048 and -5403 of the RIG1 gene. Within this region, a direct repeat sequence with five nucleotide spacing, 5'-TGACCTctattTGCCCT-3' (DR5, -5243/-5259), and an inverted repeat sequence with six nucleotide spacing, 5'-AGGCCAtggtaaTGGCCT-3' (IR6, -5323/-5340), were identified. Deletion and mutation of the DR5, but not the IR6 element, abolished the atRA-mediated activity. Electrophoretic mobility shift assays with nuclear extract from atRA-treated cells indicated the binding of retinoic acid receptor (RAR) and retinoid X receptor (RXR) heterodimers specifically to this response element. In addition to the functional DR5, the region contains many other potential sequence elements that are required to maximize the atRA-mediated induction. Taken together, we have identified and characterized the functional atRA response element that is responsible for the atRA-mediated induction of RIG1 gene.     

A novel homozygous 10 nucleotide deletion in BBS10 causes Bardet–Biedl syndrome in a Pakistani family

Bardet–Biedl Syndrome is a multisystem autosomal recessive disorder characterized by central obesity, polydactyly, hypogonadism, learning difficulties, rod-cone dystrophy and renal dysplasia. Bardet–Biedl Syndrome has a prevalence rate ranging from 1 in 100,000 to 1 in 160,000 births although there are communities where Bardet–Biedl Syndrome is found at a higher frequency due to consanguinity. We report here a Pakistani consanguineous family with two affected sons with typical clinical features of Bardet–Biedl Syndrome, in addition to abnormal liver functioning and bilateral basal ganglia calcification, the latter feature being typical of Fahr's disease. Homozygous regions obtained from SNP array depicted three known genes BBS10, BBS14 and BBS2. Bidirectional sequencing of all coding exons by traditional sequencing of all these three genes showed a homozygous deletion of 10 nucleotides (c.1958_1967del), in BBS10 in both affected brothers. The segregation analysis revealed that the parents, paternal grandfather, maternal grandmother and an unaffected sister were heterozygous for the deletion. Such a large deletion in BBS10 has not been reported previously in any population and is likely to be contributing to the phenotype of Bardet–Biedl Syndrome in this family.     

鸡减蛋综合征病毒（EDSV—76）末端前体蛋白的基因结构分析

从中国发病鸡群中分离的鸡减蛋综合征病毒弱毒株ＡＡ－２,经常规方法提取其病毒核酸后,组建了完整的限制性内切酶ＰｓｔＩ及ＨｉｎｇⅢ水解片段的基因文库,并对其中ＨｉｎｄⅢ,－ＳａｃⅠ进行了序列测定。同源比较分析证明：其Ｌ链含编码病毒末端前体蛋白,容量为５８０个氨基酸残基的开放读码框架。     

Breast cancer diagnostics based on extracellular DNA and RNA circulating in blood

Extracellular DNA and RNA were extracted from blood plasma and cell surface-bound fractions of healthy women and patients with fibroadenoma and breast cancer. Frequency of methylation of RASSF1A, Cyclin D2, and RARβ2 genes was detected in the extracellular DNA using methylation-specific PCR. Methylation of at least one of these genes was found in plasma of 13% patients with nonmalignant breast fibroadenoma and in 60% of breast cancer patients. Employment cell-surface bound DNA as the substrate for PCR increased the detection frequency of gene methylation up to 87% in patients with fibroadenoma and 95% in breast cancer patients. In clinically healthy women the methylation markers have not been found in extracellular DNA. GAPDH, RASSF8, Ki-67 mRNAs, and 18S rRNA copies were quantified using RT-qPCR of extracellular RNA circulating in blood of patients with breast tumors and healthy controls. The major part of blood extracellular RNA is associated with cell surface. ROC analysis has shown that differences in concentrations 18S RNA, RASSF8, and Ki-67 mRNAs in blood plasma are highly sensitive and specific in discrimination of benign and malignant breast tumors. Thus, analysis of methylated forms of tumor suppressor genes in blood extracellular and quantification of specific extracellular RNA circulating in blood plasma may detect mammary gland tumors and discriminate malignant and benign neoplasms.     

MHC Class I Immunopeptidome: Past,Present, and Future

In the 35 years since the revelation that short peptides bound to major histocompatibility complex class I and II molecules are the secret of the major histocompatibility complex–restricted nature of T-cell recognition, there has been enormous progress in characterizing the immunopeptidome, the repertoire of peptide presented for immunosurveillance. Here, the major milestones in the journey are marked, the contribution of proteasome-mediated splicing to the immunopeptidome is discussed, and exciting recent findings relating the immunopeptidome to the translatome revealed by ribosome profiling (RiboSeq) is detailed. Finally, what is needed for continued progress is opined about, which includes the infusion of talented young scientists into the antigen-processing field, currently undergoing a renaissance; thanks in part to the astounding success of T-cell–based cancer immunotherapy.     

Frequent overexpression of CADM1/IGSF4 in lung adenocarcinoma

We reported comprehensive screening for antigens (Ags) overexpressed on various carcinomas via isolation of human monoclonal antibodies (mAbs) that may be therapeutic in a previous paper (Proc. Natl. Acad. Sci. USA 105, 7287-7292, 2008). Twenty-one distinct Ags highly expressed on several carcinomas were identified and 356 mAbs with unique sequences turned out to bind to one of the 21 Ags. Among them CADM1/IGSF4 which had been originally referred to as tumor suppressor lung cancer 1 (TSLC1) was included. Therefore we examined the expression of CADM1 in lung cancers in this study. Eight different anti CADM1 mAbs were used for immunohistochemical analysis of 29 fresh lung cancer specimens. Staining patterns were categorized to six groups based on the extent of positive staining and the localization of stained portions. While overexpression of CADM1 was observed on the cell surface of adenocarcinomas at a high frequency, around 60%, positive stainings were rarely observed on that of other lung carcinomas including squamous cell carcinomas. Moreover, some clones among the eight mAbs gave different staining patterns from those by the other clones against the same fresh specimen, suggesting presence of variant forms of CADM1 differentiated by mAbs.     

血管生成素样蛋白4和Ⅱ型肺泡细胞表面抗原-6与急性呼吸窘迫综合征严重程度的关系及对预后的评估效能研究

摘要 目的：分析血管生成素样蛋白4(ANGPTL4)和Ⅱ型肺泡细胞表面抗原-6(KL-6)与急性呼吸窘迫综合征严重程度的关系及对预后的评估效能。方法：选择我院自2020年1月至2022年12月收治的120例急性呼吸窘迫综合征患者作为研究对象（观察组）,根据氧合指数（PaO₂/FiO₂）分为轻度组、中度组和重度组;另选120例非急性呼吸窘迫综合征患者作为对照组。检测所有患者血清ANGPTL4和KL-6的表达水平,分析血清ANGPTL4和KL-6与APACHE Ⅱ评分、PaO₂/FiO₂的关系,使用受试者工作特征曲线（ROC）下面积（AUC）评价血清ANGPTL4联合KL-6对急性呼吸窘迫综合征预后的评估效能。结果：对比对照组,观察组血清ANGPTL4、KL-6的表达水平均明显升高（P＜0.05）;血清ANGPTL4、KL-6的表达水平在轻度组、中度组和重度组中差异有统计学意义,且急性呼吸窘迫综合征越严重,升高越明显（P＜0.05）;经Pearson相关性分析,急性呼吸窘迫综合征患者血清ANGPTL4、KL-6的表达水平与PaO₂/FiO₂呈负相关,与APACHE Ⅱ评分呈正相关（P＜0.05）;经ROC曲线分析,血清ANGPTL4联合KL-6预测急性呼吸窘迫综合征患者入院28d内死亡的敏感度为90.14%、特异度为65.74%,AUC为0.900。结论：血清ANGPTL4、KL-6表达水平升高与急性呼吸窘迫综合征严重程度增大密切相关,两者联合在患者预后评估中具有一定价值,可作为判断病情及预后的辅助指标。     

学龄前阻塞性睡眠呼吸暂停低通气综合征儿童肠道菌群特征分析

摘要 目的：探讨学龄前阻塞性睡眠呼吸暂停低通气综合征（OSAHS）儿童与正常儿童肠道菌群的差异。方法：选取2023年7月至2023年11月期间新疆医科大学第一附属医院儿科门诊收治的学龄前OSAHS儿童30例作为OSAHS组,选取同期于新疆医科大学第一附属医院健康管理中心体检健康的儿童30例作为对照组。利用16SrDNA扩增子测序技术对肠道菌群进行分析。采用Spearman法分析睡眠质量与肠道菌群门、属水平丰度的相关性。结果：OSAHS组和对照组共发现2588个扩增子序列变异（ASVs）,OSAHS组检出特有ASVs 1034个,对照组检出特有ASVs 1554个。OSAHS组Chao1指数和Observed otus指数显著低于对照组（P<0.05）,两组间Shannnon指数、Simpson指数、Goods coverage指数、Peilou-e指数均差异无统计学意义（P>0.05）。OSAHS组与对照组间肠道菌群群落结构存在显著差异（P<0.05）。在门水平上,OSAHS组拟杆菌门的相对丰度低于对照组,厚壁菌门的相对丰度、厚壁菌门/拟杆菌门的比例高于对照组（P<0.05）。在属水平上,OSAHS组与对照组组优势菌群相对分度比较差异无统计学意义（P>0.05）。在门水平上,睡眠质量与拟杆菌门呈正相关（P<0.05）。在属水平上,睡眠质量与双歧杆菌属、乳杆菌属呈负相关,与拟杆菌属呈正相关（P<0.05）。阻塞性呼吸暂停低通气指数（OAHI）与肠杆菌属呈负相关（P<0.05）。最低血氧饱和度（LSaO₂）与肠杆菌属呈正相关（P<0.05）。平均血氧饱和度（MSaO₂）与X.Eubacterium._eligens_group呈正相关（P＜0.05）。结论：与正常儿童的肠道菌群的种类和相对丰度相比,学龄前OSAHS儿童的厚壁菌门/拟杆菌门比例升高,可能存在肠道菌群失调。睡眠质量在门、属水平上与拟杆菌门、双歧杆菌属、乳杆菌属明显相关。     

Overcoming resistance to rapalogs in gliomas by combinatory therapies

Glioblastoma is the most common and aggressive brain tumor type, with a mean patient survival of approximately 1 year. Many previous analyses of the glioma kinome have identified key deregulated pathways that converge and activate mammalian target of rapamycin (mTOR). Following the identification and characterization of mTOR-promoting activity in gliomagenesis, data from preclinical studies suggested the targeting of mTOR by rapamycin or its analogs (rapalogs) as a promising therapeutic approach. However, clinical trials with rapalogs have shown very limited efficacy on glioma due to the development of resistance mechanisms. Analysis of rapalog-insensitive glioma cells has revealed increased activity of growth and survival pathways compensating for mTOR inhibition by rapalogs that are suitable for therapeutic intervention. In addition, recently developed mTOR inhibitors show high anti-glioma activity. In this review, we recapitulate the regulation of mTOR signaling and its involvement in gliomagenesis, discuss mechanisms resulting in resistance to rapalogs, and speculate on strategies to overcome resistance. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases (2012).