圖1 熒光顯微鏡下的類器官（圖片源自網(wǎng)絡(luò)）

　　類器官（Organoids）是一種在體外環(huán)境下培育而成的具備三維結(jié)構(gòu)的微器官（圖1），擁有類似真實器官的復(fù)雜結(jié)構(gòu)，并能部分模擬來源組織或器官的生理功能。借助類器官，研究人員可深入觀察人體組織的變化，更好地理解發(fā)育過程，并可用于再生醫(yī)學(xué)以及藥物的療效篩選。因此，類器官研究具有廣闊的發(fā)展前景。

 圖2 “Organoid Modeling of the Tumor Immune Microenvironment”文章封面

　　生物中心針對2018年12月美國斯坦福大學(xué)Calvin J. Kuo教授團(tuán)隊發(fā)表在國際頂尖雜志Cell的文章“Organoid Modeling of the Tumor Immune Microenvironment”（圖2），特邀諾貝爾醫(yī)學(xué)和生理學(xué)獎獲得者Thomas C. Südhof教授、著名環(huán)境毒理學(xué)及神經(jīng)類器官專家Ellen Fritsche教授就類器官研究的現(xiàn)狀、瓶頸、應(yīng)用價值以及未來的發(fā)展方向展開評述。下面將為讀者呈現(xiàn)訪談內(nèi)容。
　　1. 問：什么是類器官？類器官的主要類型和他們各自的關(guān)鍵特征是什么？
　　Question：What is organoid? What are the main organoid types and their key characteristics?
　　Thomas C. Südhof答：目前有很多該領(lǐng)域的綜述和專著可供參考，以下是我推薦的一些綜述。(編者按：綜述列表見英文回答)
　　Thomas C. Südhof’s answer：There are innumerable review articles and textbooks on organoids that I would suggest you consult.
Here are some reviews:
　　[1] Little MH, Hale LJ, Howden SE, Kumar SV. Generating Kidney from Stem Cells. Annu Rev Physiol. 2019 Feb 10;81:335-357
　　[2] Rowe RG, Daley GQ. Induced pluripotent stem cells in disease modeling and drug discovery. Nat Rev Genet. 2019 Feb 8.
　　[3] Sontheimer-Phelps A, Hassell BA, Ingber DE. Modeling cancer in microfluidic human organs-on-chips. Nat Rev Cancer. 2019 Feb;19(2):65-81.
　　[4] Mittal R, Woo FW, Castro CS, Cohen MA, Karanxha J, Mittal J, Chhibber T, Jhaveri VM. Organ-on-chip models: Implications in drug discovery and clinical applications. J Cell Physiol. 2019 Jun;234(6):8352-8380.
　　[5] Amin ND, Paşca SP. Building Models of Brain Disorders with Three-Dimensional Organoids. Neuron. 2018 Oct 24;100(2):389-405.
　　Ellen Fritsche答：類器官是一種在體外培育而成的具有來源器官顯微解剖特征的多細(xì)胞三維結(jié)構(gòu)。迄今為止，不同組織、疾病模型及模擬發(fā)育的類器官已問世。類器官的工具細(xì)胞主要為組織特異性多能干細(xì)胞。類器官的主要特征包括基于細(xì)胞類別的自我組織及空間限制的定向分化，與體內(nèi)發(fā)育過程相似。他們（類器官）含有多種器官特異性細(xì)胞，這些細(xì)胞的空間組織、排列與來源器官類似。另外，他們（類器官）具有一些來源器官特有的功能。迄今，來源于多種器官的類器官業(yè)已面世，包括腦、腸道、胃、舌、甲狀腺、胸腺、睪丸、肝臟、胰腺、皮膚、肺、腎、心臟及視網(wǎng)膜。除了來源于健康組織的類器官，大量疾病模型（包括腫瘤模型）的類器官也不斷涌現(xiàn)。最后，類器官為科研人員進(jìn)行發(fā)育生物學(xué)研究提供了絕佳模型。
　　Ellen Fritsche’s answer：An organoid is a three-dimensional (3D), multicellular structure with microanatomical features of the organ of origin produced in vitro. So far, organoids of different tissues, disease models, as well as organoids resembling development have been created. Cellular basis for organoids are mainly pluripotent or tissue-specific stem cells. Key features of organoids include their self-organization through cell sorting and spatially restricted lineage commitment in a manner similar to in vivo. They contain multiple, organ-specific cell types which are spatially organized in a manner similar to the organ. In addition, they recapitulate some specific organ functions. Organoids from multiple organs have so far been created. These include brain, intestine, stomach, tongue, thyroid, thymus, testis, liver, pancreas, skin, lung, kidney, heart and retina. In addition to the healthy organoids, a plethora of disease models including tumor models, have been developed. Last, organoids offer researchers an exceptional model to study developmental biology.
　　2. 問：可否談?wù)勵惼鞴僭谏镝t(yī)學(xué)領(lǐng)域的主要應(yīng)用？
　　Question：What are the main applications of the organoids in the field of biomedicine?
　　Thomas C. Südhof答：類器官的價值在于其具有在體外培養(yǎng)環(huán)境下構(gòu)建人類器官疾病模型的潛力。這非常適用于像心臟這樣的組織，在人類早期腦發(fā)育的研究上也逐漸變得可行。但類器官在再生醫(yī)學(xué)上的應(yīng)用依然前路漫漫。
　　Thomas C. Südhof’s answer：The attraction of organoids is that they potentially allow disease modeling of human organs in a dish. This works best for tissues such as heart, and is becoming feasible for early human brain development. The use of organoids in regenerative medicine is still far in the future.
　　Ellen Fritsche答：作為一項重大的技術(shù)突破，類器官目前已被公認(rèn)為生物研究的重要工具,并具有重要的臨床應(yīng)用價值。類器官允許在一個模擬內(nèi)源性細(xì)胞組織和器官結(jié)構(gòu)的環(huán)境中進(jìn)行組織生物學(xué)、發(fā)育、再生、疾病建模 (包括癌癥研究)、器官移植技術(shù)改良、藥物發(fā)現(xiàn)/療效評估以及毒理學(xué)的研究。
　　Ellen Fritsche’s answer：Starting as a major technological breakthrough, organoids are now well-established as an essential tool in biological research and also have important implications for clinical use. Organoids allow research on tissue biology, development, regeneration, disease modeling (including cancer research), improvements in organ transplantation, drug discovery/response as well as toxicological studies in an environment that mimics endogenous cell organization and organ structures.
　　3. 問：在腫瘤生物學(xué)及新藥開發(fā)領(lǐng)域，類器官相對于細(xì)胞系、動物模型的主要優(yōu)勢在哪里？
　　Question：What are the main advantages of using organoids instead of cell lines, or animal models in the field of tumor biology and new drug development?
　　Thomas C. Südhof答：相對于細(xì)胞系而言，類器官構(gòu)建了一個具備三維結(jié)構(gòu)的器官樣組織，盡管并不完全（模擬人類器官）。相較于動物模型，類器官的優(yōu)勢體現(xiàn)在其實現(xiàn)了應(yīng)用人源性組織進(jìn)行實驗研究。
　　Thomas C. Südhof’s answer：The advantage over cell lines is that organoids model a three dimensional organ, although not completely. The advantage over animal models is that organoids enable studies of human material.
　　Ellen Fritsche答：傳統(tǒng)的二維 (2D) 腫瘤細(xì)胞系培養(yǎng)和動物人源性腫瘤異種移植物 (PDTXs) 長期以來一直被用作腫瘤模型, 并為癌癥研究做出了巨大貢獻(xiàn)。然而, 各種缺點阻礙了這些模型的臨床應(yīng)用，這主要是由于與腫瘤治療相關(guān)的藥物開發(fā)是成功率最低的。二維細(xì)胞培養(yǎng)體系不具備免疫細(xì)胞、微環(huán)境、間質(zhì)成分和器官特異性的功能。其他限制包括腫瘤細(xì)胞系經(jīng)多次傳代后缺乏來源腫瘤的遺傳異質(zhì)性, 原因是細(xì)胞在培養(yǎng)皿二維生長的環(huán)境下會發(fā)生優(yōu)勢克隆選擇，但這并不符合生理。此外, PDTX 模型還經(jīng)歷了小鼠特異性的腫瘤演化。在資源方面, 這些模型也是極度的費錢費時。類器官可以克服其中的一些限制。類器官的基因修飾可實現(xiàn)在接近生理環(huán)境的情況下進(jìn)行疾病建模。比如, 將腫瘤性突變導(dǎo)入健康干細(xì)胞可以產(chǎn)生遺傳控制的腫瘤類器官。此外, 類器官可以從患者來源的健康組織和腫瘤組織中迅速培育，從而使患者特異性藥物檢測和個性化治療方案的開發(fā)成為可能。在這種患者特異性的腫瘤類器官中，可觀察到組織穩(wěn)態(tài)(histostasis)，如3D培養(yǎng)保留了與來源患者腫瘤相一致的組織病理學(xué)特征，為未來個性化腫瘤治療的發(fā)展提供了希望。與 PDTX 不同，類器官維護(hù)便利，具有整合免疫細(xì)胞的可能性，易進(jìn)行基因改造 (遺傳性腫瘤建模)，支持匹配對照的研究，并可用于高通量藥物篩選和生物庫的建設(shè)。
　　除了腫瘤學(xué), 類器官也為新藥開發(fā)提供了絕佳模型。新藥開發(fā)的失敗率很高，這在一定程度上是由于動物藥代動力學(xué)和藥效學(xué)的差異或動物疾病模型并不能完全模擬人體病理過程。具有人體特異生理和病理特征的類器官有助于克服這些問題?；谔囟膊?，甚至特定個體，以高通量方式培育的類器官預(yù)計將發(fā)展成為精確治療的強大工具。未來可借助生物庫進(jìn)行篩選，不僅是為了鑒定新藥，還可揭示哪些患者可以從某些 (現(xiàn)有) 藥物的治療中受益。此外，對潛在藥物的重點檢測可為制藥業(yè)提供新的指引。另外，類器官未來可能用于毒理學(xué)檢測, 以作為動物試驗的有力補充(如果不是部分取代的話)。
　　Ellen Fritsche’s answer：Traditional two-dimensional (2D) tumor cell line cultures and patient-derived tumor xenografts (PDTXs) in animals have long been employed as tumor models and have made tremendous contribution to cancer research. However, a variety of drawbacks hamper these models for clinical use as success rates for tumor therapeutics are lowest in the field of drug development. 2D cell line cultures do not contain immune cells, microenvironment, stromal compartments, and organ-specific functions. Other limitations include the lack of genetic heterogeneity of original tumors after many passages for cancer cell lines because clonal selection in the dish happens for superiority in 2D growth, which is not physiologic. Moreover, PDTX models experience mouse-specific tumor evolution. On the resource side, such models are highly money- and time-consuming. Organoids can overcome some of these constraints. Genetic modification of organoids allows disease modeling in a setting that approaches the physiological environment. Here, insertions of tumor mutations into healthy stem cells allow generation of genetically-controlled tumoroids. Additionally, organoids can be grown with high efficiency from patient-derived healthy and tumour tissues, potentially enabling patient-specific drug testing and the development of individualized treatment regimens. In such patient-specific tumoroids, histostasis is observed, i.e. conservation of histopathological traits between 3D cultures and the matched patient tumor, promising advances in personalized tumor therapies in the future. In contrast to PDTX, organoids are of easier maintenance, bear the possibility to integrate immune cells, are amenable to genetic modification (genetic cancer modeling), allow study of matched controls, can be used for high throughput drug screening and biobanking.
　　Besides oncology, organoids are promising models for drug development. Attrition rates in new drug development are high. This is partly reasoned indifferences between animal pharmacokinetics and –dynamics or in animal disease models that do not correctly resemble human pathology. Organoids with human-relevant physiology and pathology are thought to help overcoming these issues. Organoid cultures based on a specific disease and even on a specific individual used in a high-throughput manner are expected to develop into powerful tools for precision therapy. Future screens may be performed using biobanks with the aim of not only identifying new drugs but also revealing which patients may benefit from treatment with certain (existing) drugs. In addition, focused tests of potential drugs should identify new leads for the pharmaceutical industry. Furthermore, organoids may be used in the future for toxicology testing to complement, if not in part replace, animal testing.
　　4. 問：當(dāng)前類器官的局限是什么？為了滿足腫瘤生物學(xué)、干細(xì)胞生物學(xué)、移植、新藥開發(fā)領(lǐng)域的研究需要，類器官需要在哪些方面進(jìn)一步改進(jìn)？
　　Question：What are the limitations of organoids and what aspects of organoids can be further improved to meet the demand for research in tumor biology, stem cell biology, transplantation and drug development?
　　Thomas C. Südhof答: 類器官領(lǐng)域的研究仍在起步階段。即使對于如心臟和肝臟這樣的組織，類器官也很不成熟，僅能部分模擬人體器官。對于腦組織則更甚。許多基本的（腦組織）生理功能，如細(xì)胞生理、生化功能仍有待突破。這將花費數(shù)年的時間。
　　Thomas C. Südhof’s answer：The field of organoid research is still in the beginning. Even for tissues like heart and liver, organoids are very immature and only partly model the human organ. This is worse for brain. Much fundamental biology, such as cell biology and biochemistry, is needed to advance the field. This will take many years.
　　Ellen Fritsche答：類器官是融合了各種器官特異性細(xì)胞類型、組織形態(tài)和功能的組織模型。但類器官僅為有限度的模擬，困擾這項技術(shù)應(yīng)用的一個重要限制是它的體積。當(dāng)類器官體積增加時，缺氧和缺乏可溶性因子所致的組織壞死是亟需解決的問題。解決這個問題的一個可能方案是激活血管生成途徑, 從而使類器官血管化。這已經(jīng)在hiPSC衍生的肝臟類器官上成功實現(xiàn)。類器官領(lǐng)域的另一個挑戰(zhàn)在于一個完整的有機體中所自然存在的器官“對話”。類器官研究可滿足生物工程的要求, 通過培育包含不同類型hiPSC衍生類器官（呈現(xiàn)多個器官系統(tǒng)的結(jié)構(gòu)和功能）的器官芯片設(shè)備，用以在更類似于體內(nèi)的環(huán)境中篩選藥物。另外，通過在類器官中添加免疫細(xì)胞, 還可模擬具有免疫系統(tǒng)的組織間“對話”。另外，在藥理和毒理學(xué)研究中，物質(zhì)的肝臟代謝至關(guān)重要，這可通過以器官芯片的形式包含肝臟代謝來實現(xiàn)。
　　Ellen Fritsche’s answer：Organoids are organ models recapitulating an assortment of organ-specific cell types, tissue morphogenesis and functions. Yet, there are limitations in their mimicry. One important limitation plaguing the application of this technology is their size. When the organoids’ volume increases, the issue of tissue necrosis caused by the lack of diffusion of oxygen and soluble factors needs to be addressed. One solution for this problem might be the activation of angiogenic pathways that will lead to vascularised organoids. This was already succeeded with hiPSC-derived liver organoids. One more challenge of the organoid field lies in organ crosstalk, which is naturally present in an intact organism. Here, organoid research meets bioengineering by producing organ-on-a-chip devices containing different types of hiPSC-derived organoids representing the structure and function of multiple organ systems for screening the effects of drugs in more in vivo-like settings. The crosstalk of tissues with the immune system can be modelled by adding immune cells to the organoids. For pharmacological and toxicological applications, liver metabolism of compounds is crucial. Including such metabolism via an organ-on-a-chip approach can solve this issue.
　　5. 問：當(dāng)前類器官研究的發(fā)展方向如何？
　　Question：What are the current trends for organoids research?
　　Thomas C. Südhof答:（當(dāng)前的現(xiàn)狀是）所有人都在盲目追求應(yīng)用，卻忽略了一個堅實的科學(xué)基礎(chǔ)。我認(rèn)為未來會有數(shù)以百計的公司在那里販賣希望，但他們大多數(shù)將以失敗告終。因為相關(guān)生理學(xué)研究成果并不足以支撐這些應(yīng)用項目。類器官最有前景的應(yīng)用領(lǐng)域應(yīng)是用于肝臟、心臟和腫瘤的藥物篩選。
　　Thomas C. Südhof’s answer：Everybody rushes towards applications, without a solid scientific basis. I think hundreds of companies will be founded that will sell hope, but will mostly fail because the biology isn't there to support applications. Most promising are drug screens in tissue organoids such as liver or heart and in cancer.
　　Ellen Fritsche答：目前類器官研究的趨勢包括建立用于高通量篩選的類器官庫和平臺, 建立其他疾病模型, 以及建立用于整個生物體建模的器官芯片和微流體芯片。在此特別要強調(diào)的是培養(yǎng)基的限制亟待解決。對微流體芯片來講，需要一種芯片上所有類器官均適用的通用培養(yǎng)基。此外，根據(jù)器官系統(tǒng)的不同, 需要開發(fā)與生理過程相關(guān)的來自類器官的高通量數(shù)據(jù)輸出裝置。在臨床方面，為了開發(fā)最佳個體化治療方案，使用源自患者特異性hiPSC類器官的個體化醫(yī)療研究亟需開展。在毒理學(xué)領(lǐng)域，類器官目前已被用來替代動物進(jìn)行毒性測試。
　　Ellen Fritsche’s answer：Current trends for organoid research include generation of organoid banks and platforms for high-throughput screening approaches, generation of additional disease models, and set up of organ-on-a-chip and microfluidics devices for whole organism modeling. Here, especially medium constrictions have to be solved. For microfluidics a common medium for all organoids on the chip is needed. Moreover, depending on the organ system, physiologically relevant high-throughput readouts from organoids need to be developed. On the clinical side, research on personalized medicine using organoids derived from patient-specific hiPSC is warranted for optimal individual treatment regimes. In the toxicology field, organoids as substrates for toxicity testing replacing animals is currently exploited.
　　6. 問：可否預(yù)測一下接下來5年內(nèi)類器官研究領(lǐng)域的發(fā)展？
　　Question：How the organoids research field will be look like in 5 years?
　　Thomas C. Südhof答: 我的預(yù)測是在接下來的5年內(nèi)好的實驗室將學(xué)會如何促進(jìn)類器官的成熟，并明確該方法的局限。我認(rèn)為，盡管類器官為干細(xì)胞研究提供了巨大的機會，如促進(jìn)新發(fā)現(xiàn)和療法的出現(xiàn)；但這將耗費10年或更長的時間來發(fā)展。到那時候，也只有到那時候，走向應(yīng)用才能真正成為可能。在那之前，大量初創(chuàng)公司將會燒掉數(shù)以億計的資金，他們中的少部分將會走向成功，并找到增加營收的新途徑。
　　Thomas C. Südhof’s answer：My prediction is that in 5 years, good science labs will have learned how to mature organoids and the limitations of the approach will have been defined. I think organoids are a tremendous opportunity in stem cell approaches that will enable novel discoveries and therapies, but that this will take at least 10 years to develop. Then and only then will it be possible to rationally move towards applications. Until then, lots of start-ups will have spent hundreds of millions of dollars, and a few of them will have been successful in generating some future avenue of revenue.
　　Ellen Fritsche答：在5年內(nèi)，類器官的遺傳操作與類器官庫相結(jié)合將給生物醫(yī)學(xué)研究帶來翻天覆地的變化。購買來源于具有不同遺傳背景患者的疾病特異性類器官將成為可能。器官芯片平臺將具有特定標(biāo)準(zhǔn)，由合同研究組織（CRO）以與目前動物試驗類似的方式提供。類器官將極大地促進(jìn)藥物療效試驗和安全性測試的開展，因此也將進(jìn)入藥物開發(fā)和化學(xué)安全性評估研究的監(jiān)管領(lǐng)域。
　　Ellen Fritsche’s answer：In 5 years, genetic manipulation of organoids in combination with organoid banking will have revolutionized biomedical research. It will be possible to purchase disease-specific organoids from broad ranges of patients with distinct genetic backgrounds. Organ-on-a-chip platforms will be standard and offered by CROs in a similar manner than currently animal testing. Organoids will have tremendously facilitated drug efficacy and safety testing and thus will have entered also into the regulatory areas of research for drug development as well as chemical safety assessment.

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1.  Thomas C. Südhof教授簡介

　　斯坦福大學(xué)醫(yī)學(xué)院教授、霍華德-休斯醫(yī)學(xué)研究所 (HHMI) 研究員、美國科學(xué)院院士、美國醫(yī)學(xué)科學(xué)院院士、英國皇家學(xué)會外籍院士、2013年諾貝爾醫(yī)學(xué)和生理學(xué)獎獲得者。1955年生于德國哥廷根，1982年獲得哥廷根大學(xué)醫(yī)學(xué)博士學(xué)位。Südhof教授的研究主要聚焦于突觸前神經(jīng)遞質(zhì)釋放的分子機制，為該領(lǐng)域的頂級科學(xué)家。他發(fā)現(xiàn)了囊泡內(nèi)神經(jīng)遞質(zhì)釋放過程中的多種關(guān)鍵蛋白，并闡明了神經(jīng)遞質(zhì)釋放的具體分子機制。鑒于在囊泡轉(zhuǎn)運領(lǐng)域的開創(chuàng)性工作，他先后榮獲拉斯克基礎(chǔ)醫(yī)學(xué)獎及諾貝爾生理學(xué)和醫(yī)學(xué)獎等重要醫(yī)學(xué)獎項。

2.  Ellen Fritsche教授簡介

　　德國IUF-萊布尼茨環(huán)境醫(yī)學(xué)研究所（IUF-Leibniz Research Institute for Environmental Medicine）環(huán)境毒理學(xué)教授，球模型和風(fēng)險評估專家組組長。1998年獲雷根斯堡大學(xué)和杜塞爾多夫大學(xué)醫(yī)學(xué)博士學(xué)位，曾先后在美國國立環(huán)境衛(wèi)生研究所（NIEHS)和IUF-萊布尼茨環(huán)境醫(yī)學(xué)研究所完成博士后研究工作，2009-2012年任亞琛工業(yè)大學(xué)皮膚毒理學(xué)教授。目前為Neurotoxicology雜志副主編、歐洲化學(xué)理事會（cefic）顧問、歐盟地平線2020計劃專家組成員、歐洲替代動物試驗研究中心（CERST-NRW）項目牽頭人、替代法信托大會（ACT）成員和OECD發(fā)育神經(jīng)毒性專家組委員。歷任歐洲替代動物試驗協(xié)會（EUSAAT）副主席、主席。

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