消化器癌(胃癌・大腸癌)の転移の予知とその対策: 生物学的分子腫瘍マーカーよりの検討

金沢大学がん研究所消化器癌の転移能(metastatic potential)を知るべく、病理組織学的多様性、腫瘍マーカーを含めた生化学、発育速度、癌の浸潤・転移に関与する遺伝子異常、血管新生因子などの諸因子などより転移予知の可能性を探っており、その成果は転移予知の重要な指標となりうる事が判明した。また実験的には、ヌードマウスを用いた同所移植による胃癌肝転移モデルの開発に成功し、PCR法を用いた肝微少転移巣の検出を試みているが、この同所性胃癌肝転移モデルの成功は、転移成立機序の解明、潜在的転移能、そして転移抑制実験を行なう上で重要なモデルとなった。一方、浸潤・転移に深くかかわっている細胞外基質分解酵素、マトリックスメタロプロテナーゼ(MMPs)とそのインヒビター(TIMPs)のRNAレベルでの発現、免疫組織学的検討、zymographyによる解析、血清中のMMPs、TIMPsを測定し、潜在性転移能の有無を胃癌組織で検討し、その成果を発表してきた。なかでも当研究所清木教授らにより同定された膜型MMP(MT-MMP)はMMP-2活性化因子として知られ、胃癌組織内でのMT-MMP、MMP-2の共発現している胃癌では、88%に静脈侵襲陽性であり、血行性転移との関連で注目された。また、MMP-7は胃癌組織において癌細胞特異的に発現し、分化型腺癌に優位であり、なかでも癌細胞の癌細胞の脈管内浸襲に重要であると考えられた。一方、胃癌細胞における転移成立に関与するその他の転移関連遺伝子群(血管新生因子、増殖因子、接着因子等)の検討を行なったところ、血行性転移においては血管新生因子vascular endothelial growth factor (VEGF)とMMPsの発現が著名で転移予知可能であることが判明した。最後に治療実験としてヌードマウス胃同所移植肝転移モデルを用いて高転移能を有するヒト胃癌細胞にTIMP-1遺伝子を外来性に導入し、この細胞を同所移植したところ、肝転移は著名に抑制されており、TIMP-1は転移抑制性に働いていることを確認しており、今後臨床レベルでの転移制御のための分子標的として注目に値する。In considering the main steps in the process of tumor invasion and metastasis, the mechanism for invasion of tumor cells through tissue barriers are not well understood, but they appear to involve both mechanical and enzymatic activity. Matrix metalloproteinases (MMPs) play a key role in degradation of the extracellular matrix (ECM) associated with cancer invasion and metastasis. We have proviously studied the production of MMP-1,2,3, and 9 in human gastric carcinomas compared with normal gastric mucosa. Among these MMPs, activation as well as production of the zymogen of MMP-2 (progelatinase A) was well correlated with local invasion and lymphatic permeation and vessel invasion of the gastric cancer. Since proMMP-2 activation is thought to be caused by MT-MMP in vivo, expression of MT-MMP-1 was studied in the same samples. Our data indicated that MT-MMP-1 is exclusively expressed in the carcinoma cells and its expression is well correlated with pro MMP-2 activation.It also is well known that the activation of MMPs is regulated by tissue inbitors of metalloproteinases (TIMPs). In considering negative regulators of TIMPs, we examined inhibition of metastasis in human gastric cancer cells transfected with TIMP-1 gene in vivo, As a metastatic model we used the cell line established from human gastric carcinoma, KKLS at our department. KKLS cell was transfected with exogenous TIMP-1 gene by the Chen-Okayama method and we obtained two clones KT-CL-1 and and KT-CL-14 expressed different levels of TIMP-1. The KKLS cells and these transfectants were orthotopically transplanted into nude mice (murine stomach) and metastasis in the murine liver was detected. Our experimental data showed that KT-CL-1 and KT-CL-14 transfected the cDNA for TIMP-1 gene resulted in dramatic inhibition of metastatic colonies of 46.7% and 26.7% compared with those of parental KKLS cells and K-Neo cell as control. Consequently, the MMPs are therapeutic targets that may evoke cytostatic, as are adhesion molecles, signal transduction pathway, growth factor and angiogenesis. The hope is that by use of selective inhibitors for these targets, we can achieve a halt in tumor invasion and metasitasis without significant toxicity and our encouraging results lead to conclude that they indicates a promising new direction in cnacer therapy.研究課題/領域番号:07457270, 研究期間(年度):1995 – 1997出典：研究課題「消化器癌(胃癌・大腸癌)の転移の予知とその対策: 生物学的分子腫瘍マーカーよりの検討」課題番号07457270（KAKEN：科学研究費助成事業データベース（国立情報学研究所）） （https://kaken.nii.ac.jp/ja/report/KAKENHI-PROJECT-07457270/074572701997kenkyu_seika_hokoku_gaiyo/）を加工して作

Build-up functionalization of anti-EGFR × anti-CD3 bispecific diabodies by integrating high-affinity mutants and functional molecular formats

Designing non-natural antibody formats is a practical method for developing highly functional next-generation antibody drugs, particularly for improving the therapeutic efficacy of cancer treatments. One approach is constructing bispecific antibodies (bsAbs). We previously reported a functional humanized bispecific diabody (bsDb) that targeted epidermal growth factor receptor and CD3 (hEx3-Db). We enhanced its cytotoxicity by constructing an Fc fusion protein and rearranging order of the V domain. In this study, we created an additional functional bsAb, by integrating the molecular formats of bsAb and high-affinity mutants previously isolated by phage display in the form of Fv. Introducing the high-affinity mutations into bsDbs successfully increased their affinities and enhanced their cytotoxicity in vitro and in vivo. However, there were some limitations to affinity maturation of bsDb by integrating high-affinity Fv mutants, particularly in Fc-fused bsDb with intrinsic high affinity, because of their bivalency. The tetramers fractionated from the bsDb mutant exhibited the highest in vitro growth inhibition among the small bsAbs and was comparable to the in vivo anti-tumor effects of Fc-fused bsDbs. This molecule shows cost-efficient bacterial production and high therapeutic potential

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

Impact of COVID-19 on cardiovascular testing in the United States versus the rest of the world

Objectives: This study sought to quantify and compare the decline in volumes of cardiovascular procedures between the United States and non-US institutions during the early phase of the coronavirus disease-2019 (COVID-19) pandemic. Background: The COVID-19 pandemic has disrupted the care of many non-COVID-19 illnesses. Reductions in diagnostic cardiovascular testing around the world have led to concerns over the implications of reduced testing for cardiovascular disease (CVD) morbidity and mortality. Methods: Data were submitted to the INCAPS-COVID (International Atomic Energy Agency Non-Invasive Cardiology Protocols Study of COVID-19), a multinational registry comprising 909 institutions in 108 countries (including 155 facilities in 40 U.S. states), assessing the impact of the COVID-19 pandemic on volumes of diagnostic cardiovascular procedures. Data were obtained for April 2020 and compared with volumes of baseline procedures from March 2019. We compared laboratory characteristics, practices, and procedure volumes between U.S. and non-U.S. facilities and between U.S. geographic regions and identified factors associated with volume reduction in the United States. Results: Reductions in the volumes of procedures in the United States were similar to those in non-U.S. facilities (68% vs. 63%, respectively; p = 0.237), although U.S. facilities reported greater reductions in invasive coronary angiography (69% vs. 53%, respectively; p < 0.001). Significantly more U.S. facilities reported increased use of telehealth and patient screening measures than non-U.S. facilities, such as temperature checks, symptom screenings, and COVID-19 testing. Reductions in volumes of procedures differed between U.S. regions, with larger declines observed in the Northeast (76%) and Midwest (74%) than in the South (62%) and West (44%). Prevalence of COVID-19, staff redeployments, outpatient centers, and urban centers were associated with greater reductions in volume in U.S. facilities in a multivariable analysis. Conclusions: We observed marked reductions in U.S. cardiovascular testing in the early phase of the pandemic and significant variability between U.S. regions. The association between reductions of volumes and COVID-19 prevalence in the United States highlighted the need for proactive efforts to maintain access to cardiovascular testing in areas most affected by outbreaks of COVID-19 infection

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe