演講MP3+雙語文稿:我們可以通過重寫DNA治療遺傳疾病嗎？

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【演講者及介紹】David R Liu

劉大衛(wèi)——化學(xué)生物學(xué)家。David R. Liu領(lǐng)導(dǎo)了一個研究小組，將化學(xué)和進(jìn)化技術(shù)結(jié)合起來，創(chuàng)造出革命性的新藥。

【演講主題】我們可以通過重寫DNA治療遺傳疾病嗎？

【中英文字幕】

翻譯者psjmz mz 校對者Jin Ge

00:01

The most important gift your mother andfather ever gave you was the two sets of three billion letters of DNA that makeup your genome. But like anything with three billion components, that gift isfragile. Sunlight, smoking, unhealthy eating, even spontaneous mistakes made byyour cells, all cause changes to your genome. The most common kind of change inDNA is the simple swap of one letter, or base, such as C, with a differentletter, such as T, G or A. In any day, the cells in your body will collectivelyaccumulate billions of these single-letter swaps, which are also called"point mutations."

你父母給你的最重要的禮物就是2組包含30億個堿基的DNA，它們構(gòu)成了你的基因組。但就像任何包含太多零件的東西一樣，這個禮物非常脆弱。太陽光、吸煙、不健康的飲食，甚至是細(xì)胞自身出現(xiàn)的錯誤，都能改變你的基因組。最常見的DNA改變就是一個字母，也叫一個堿基，比如C（胞嘧啶），換成了別的堿基，如T(胸腺嘧啶)、 G(鳥嘌呤)或者A(腺嘌呤)。每一天，你身體里的細(xì)胞會累計發(fā)生數(shù)億次單堿基的改變，這也被稱作“點突變”。

00:46

Now, most of these point mutations areharmless. But every now and then, a point mutation disrupts an importantcapability in a cell or causes a cell to misbehave in harmful ways. If that mutationwere inherited from your parents or occurred early enough in your development,then the result would be that many or all of your cells contain this harmfulmutation. And then you would be one of hundreds of millions of people with agenetic disease, such as sickle cell anemia or progeria or muscular dystrophyor Tay-Sachs disease.

大部分點突變是無害的。但時不時，點突變會干擾細(xì)胞的某項重要功能，或者引起細(xì)胞出現(xiàn)異常行為。如果這種變異是從父母遺傳而來的，或者發(fā)生于你生命早期，那么結(jié)果很可能是你的大部分甚至全部細(xì)胞都帶有這種有害變異。你可能就會像其他成千上萬人一樣患上基因疾病，像鐮刀型紅血球病，或者早衰癥，或者肌肉萎縮癥，或者家族黑蒙性癡呆癥。

01:22

Grievous genetic diseases caused by pointmutations are especially frustrating, because we often know the exactsingle-letter change that causes the disease and, in theory, could cure thedisease. Millions suffer from sickle cell anemia because they have a single Ato T point mutations in both copies of their hemoglobin gene. And children withprogeria are born with a T at a single position in their genome where you havea C, with the devastating consequence that these wonderful, bright kids age veryrapidly and pass away by about age 14. Throughout the history of medicine, wehave not had a way to efficiently correct point mutations in living systems, tochange that disease-causing T back into a C. Perhaps until now. Because mylaboratory recently succeeded in developing such a capability, which we call"base editing."

由點基因突變引起的這些不幸的遺傳疾病讓我們尤其沮喪，因為我們往往已經(jīng)知道哪個具體字母（堿基）發(fā)生了突變，從而導(dǎo)致了疾病。因此理論上，我們可以治愈它。數(shù)百萬人被鐮刀型紅血球病折磨，因為他們的血紅蛋白基因中都含有從A到T的點突變。而患有早衰癥的孩子只不過生來就在基因組中的某個位置有一個T，而正常的基因應(yīng)該是C，令人悲傷的是，這些聰明美好的孩子衰老得非?？欤ǔ；畈贿^14歲?？v觀整個醫(yī)藥史，我們還沒有找到有效的方法可以在生命系統(tǒng)中糾正點突變，將引起疾病的T改回正常的C。但現(xiàn)在我們有辦法了。因為我的實驗室最近成功發(fā)明了一種技術(shù)，叫做“堿基編輯”。

02:23

The story of how we developed base editingactually begins three billion years ago. We think of bacteria as sources ofinfection, but bacteria themselves are also prone to being infected, inparticular, by viruses. So about three billion years ago, bacteria evolved adefense mechanism to fight viral infection. That defense mechanism is nowbetter known as CRISPR. And the warhead in CRISPR is this purple protein thatacts like molecular scissors to cut DNA, breaking the double helix into twopieces. If CRISPR couldn't distinguish between bacterial and viral DNA, itwouldn't be a very useful defense system.

關(guān)于我們?nèi)绾伟l(fā)明“堿基編輯”的故事可以追溯到30億年前。我們通常認(rèn)為細(xì)菌是感染源，但其實細(xì)菌本身也容易被感染，特別是被病毒。因此大約30億年前，細(xì)菌進(jìn)化出一種防御機制，來抵抗病毒感染。這種防御機制如今被稱為CRISPR。CRISPR里最強的武器是這種紫色的蛋白質(zhì)，它就像分子剪刀一樣，可以剪斷DNA鏈，將雙螺旋結(jié)構(gòu)剪成2條單螺旋鏈。如果CRISPR分不清細(xì)菌和病毒的DNA，這就不能算是一個好的防御系統(tǒng)。

03:06

But the most amazing feature of CRISPR isthat the scissors can be programmed to search for, bind to and cut only aspecific DNA sequence. So when a bacterium encounters a virus for the firsttime, it can store a small snippet of that virus's DNA for use as a program todirect the CRISPR scissors to cut that viral DNA sequence during a futureinfection. Cutting a virus's DNA messes up the function of the cut viral gene,and therefore disrupts the virus's life cycle.

但CRISPR最神奇之處在于剪刀可以被編輯，專門尋找、鎖定和剪斷特定的DNA片段。所以當(dāng)細(xì)菌首次遇到某個病毒時，它會存儲一小段病毒的DNA以此來引導(dǎo)CRISPR的剪刀，如果將來發(fā)生感染，就剪斷病毒的DNA鏈。剪斷病毒的DNA 會擾亂該病毒基因的表達(dá)功能，從而中斷病毒的生命。

03:46

Remarkable researchers including EmmanuelleCharpentier, George Church, Jennifer Doudna and Feng Zhang showed six years agohow CRISPR scissors could be programmed to cut DNA sequences of our choosing,including sequences in your genome, instead of the viral DNA sequences chosenby bacteria. But the outcomes are actually similar. Cutting a DNA sequence inyour genome also disrupts the function of the cut gene, typically, by causingthe insertion and deletion of random mixtures of DNA letters at the cut site.

許多優(yōu)秀的研究者，比如埃馬紐埃爾·卡彭蒂耶、喬治·丘奇，詹妮佛·杜德納和張鋒，在6年前展示了CRISPR的剪刀可以被編輯，用來剪斷我們選擇的DNA片段，人類的基因片段，而不是細(xì)菌選的病毒的DNA片段。效果是相似的。通過剪斷基因中的DNA片段同樣會影響被剪基因的功能，方法就是在被剪的位置上增加或刪除隨機的DNA堿基組合。

04:24

Now, disrupting genes can be very usefulfor some applications. But for most point mutations that cause geneticdiseases, simply cutting the already-mutated gene won't benefit patients,because the function of the mutated gene needs to be restored, not furtherdisrupted. So cutting this already-mutated hemoglobin gene that causes sicklecell anemia won't restore the ability of patients to make healthy red bloodcells. And while we can sometimes introduce new DNA sequences into cells toreplace the DNA sequences surrounding a cut site, that process, unfortunately,doesn't work in most types of cells, and the disrupted gene outcomes stillpredominate.

在某些情況下，擾亂基因非常有用。但對于大部分引起遺傳疾病的點突變而言，僅僅剪斷已經(jīng)發(fā)生變異的基因，對病人而言并沒有意義，因為這些變異基因的功能需要重置，而不是進(jìn)一步打亂。因此，把那些引起鐮刀型貧血的，已經(jīng)變異的血紅蛋白基因剪斷，并不能恢復(fù)病人的造血功能。有時候我們可以加入一些新的DNA片段到細(xì)胞中，替代被剪斷區(qū)域周圍的DNA鏈，但可惜的是這一過程對大部分細(xì)胞不起作用，被影響的基因仍占主導(dǎo)地位。

05:12

Like many scientists, I've dreamed of afuture in which we might be able to treat or maybe even cure human geneticdiseases. But I saw the lack of a way to fix point mutations, which cause mosthuman genetic diseases, as a major problem standing in the way.

像許多科學(xué)家一樣，我夢想著未來有一天，我們可以治療甚至治愈人類遺傳疾病。但我們?nèi)狈π迯?fù)點突變的方法，而點突變是大部分人類基因疾病的主因，是我們需要解決的主要問題。

05:29

Being a chemist, I began working with mystudents to develop ways on performing chemistry directly on an individual DNAbase, to truly fix, rather than disrupt, the mutations that cause geneticdiseases. The results of our efforts are molecular machines called "baseeditors." Base editors use the programmable searching mechanism of CRISPRscissors, but instead of cutting the DNA, they directly convert one base toanother base without disrupting the rest of the gene. So if you think ofnaturally occurring CRISPR proteins as molecular scissors, you can think ofbase editors as pencils, capable of directly rewriting one DNA letter intoanother by actually rearranging the atoms of one DNA base to instead become adifferent base.

我是一名化學(xué)家，我跟我的學(xué)生們一起研究將化學(xué)反應(yīng)應(yīng)用于單個DNA堿基上的方法，從而真正修復(fù)，而不僅僅是終止引起基因疾病的變異。我們的成果就是分子機器，叫做“堿基編輯器”。堿基編輯器使用的是類似CRISPR剪刀的可編程搜索機制，但與剪斷DNA不同的是，它們直接將一個堿基變成另一個，而不會破壞基因的其他部分。如果將CRISPR蛋白質(zhì)比作分子剪刀的話，堿基編輯器就像鉛筆，它能直接改寫DNA堿基，通過重新排列DNA堿基上的原子，而不是將它變成一個不同的堿基。

06:23

Now, base editors don't exist in nature. Infact, we engineered the first base editor, shown here, from three separateproteins that don't even come from the same organism. We started by takingCRISPR scissors and disabling the ability to cut DNA while retaining itsability to search for and bind a target DNA sequence in a programmed manner. Tothose disabled CRISPR scissors, shown in blue, we attached a second protein inred, which performs a chemical reaction on the DNA base C, converting it into abase that behaves like T. Third, we had to attach to the first two proteins theprotein shown in purple, which protects the edited base from being removed bythe cell. The net result is an engineered three-part protein that for the firsttime allows us to convert Cs into Ts at specified locations in the genome.

堿基編輯器在大自然中并不存在。實際上，我們制造的第一個堿基編輯器，如圖所示，是由3種獨立的蛋白質(zhì)組成，它們甚至都不是來自同一個生物體。我們首先抑制CRISPR剪刀剪斷DNA的功能，并通過編程的方法，保持其搜索和鎖定目標(biāo)DNA片段的能力。在功能被抑制的CRISPR剪刀上，圖中藍(lán)色的部分，我們加上了第2種蛋白質(zhì)，在這里用紅色標(biāo)出，它會與DNA堿基C發(fā)生化學(xué)反應(yīng)，將其轉(zhuǎn)換成與T行為相似的堿基。第3步，我們將圖片中用紫色標(biāo)出的蛋白質(zhì)加在前2種蛋白質(zhì)上，來保護(hù)被編輯過的堿基不被細(xì)胞移除。最終結(jié)果就是制造出一個由3部分組成的蛋白質(zhì)，這也是我們在史上首次將基因組特定位置的堿基C轉(zhuǎn)換為T。

07:21

But even at this point, our work was onlyhalf done. Because in order to be stable in cells, the two strands of a DNAdouble helix have to form base pairs. And because C only pairs with G, and Tonly pairs with A, simply changing a C to a T on one DNA strand creates amismatch, a disagreement between the two DNA strands that the cell has toresolve by deciding which strand to replace. We realized that we could furtherengineer this three-part protein to flag the nonedited strand as the one to bereplaced by nicking that strand. This little nick tricks the cell intoreplacing the nonedited G with an A as it remakes the nicked strand, therebycompleting the conversion of what used to be a C-G base pair into a stable T-Abase pair.

但做到這一步，我們的工作也僅僅完成了一半。因為為了保持細(xì)胞的穩(wěn)定，DNA雙螺旋結(jié)構(gòu)中的兩條鏈必須形成堿基對。因為C只能跟G配對，T只能跟A配對，如果只是將一鏈上的堿基C變成T，會造成DNA雙螺旋的不匹配，要解決這個問題，細(xì)胞需要決定替換哪一條鏈。我們認(rèn)識到可以改進(jìn)這個由3部分組成的蛋白質(zhì)，將未編輯的那條鏈標(biāo)記為要被切割掉。這個小缺口誘騙細(xì)胞用A取代未編輯的G，因為它重新生成了完整的單鏈，這樣就完成了C-G堿基對到穩(wěn)定的T-A堿基對的轉(zhuǎn)變。

08:24

After several years of hard work led by aformer post doc in the lab, Alexis Komor, we succeeded in developing this firstclass of base editor, which converts Cs into Ts and Gs into As at targetedpositions of our choosing. Among the more than 35,000 known disease-associatedpoint mutations, the two kinds of mutations that this first base editor canreverse collectively account for about 14 percent or 5,000 or so pathogenicpoint mutations. But correcting the largest fraction of disease-causing pointmutations would require developing a second class of base editor, one thatcould convert As into Gs or Ts into Cs. Led by Nicole Gaudelli, a former postdoc in the lab, we set out to develop this second class of base editor, which,in theory, could correct up to almost half of pathogenic point mutations,including that mutation that causes the rapid-aging disease progeria.

在實驗室前博士后Alexis Komor 領(lǐng)導(dǎo)的幾年努力工作之后，我們成功地開發(fā)了第一代堿基編輯器，將指定位置的C都轉(zhuǎn)變?yōu)門，G都轉(zhuǎn)變?yōu)锳。在3.5萬多個已知的與點突變有關(guān)的疾病中，第一代堿基編輯器可以逆轉(zhuǎn)的兩種突變總共占致病點突變的 14%或5000種左右。但是，糾正大部分致病點突變需要開發(fā)第二代堿基編輯器，一個可以將A都轉(zhuǎn)變?yōu)镚 或T都轉(zhuǎn)變?yōu)镃的工具。在實驗室前博士后 Nicole Gaudelli的領(lǐng)導(dǎo)下，我們著手開發(fā)了這個第二代堿基編輯器，從理論上講，這樣可以糾正近一半的致病點基因突變，包括導(dǎo)致早衰癥的突變。

09:30

We realized that we could borrow, onceagain, the targeting mechanism of CRISPR scissors to bring the new base editorto the right site in a genome. But we quickly encountered an incredibleproblem; namely, there is no protein that's known to convert A into G or T intoC in DNA. Faced with such a serious stumbling block, most students wouldprobably look for another project, if not another research advisor. (Laughter)But Nicole agreed to proceed with a plan that seemed wildly ambitious at thetime. Given the absence of a naturally occurring protein that performs thenecessary chemistry, we decided we would evolve our own protein in thelaboratory to convert A into a base that behaves like G, starting from aprotein that performs related chemistry on RNA. We set up a Darwiniansurvival-of-the-fittest selection system that explored tens of millions ofprotein variants and only allowed those rare variants that could perform thenecessary chemistry to survive. We ended up with a protein shown here, thefirst that can convert A in DNA into a base that resembles G. And when we attachedthat protein to the disabled CRISPR scissors, shown in blue, we produced thesecond base editor, which converts As into Gs, and then uses the samestrand-nicking strategy that we used in the first base editor to trick the cellinto replacing the nonedited T with a C as it remakes that nicked strand,thereby completing the conversion of an A-T base pair to a G-C base pair.

我們意識到我們可以再次借助，CRISPR剪刀的靶向機制，將新的堿基編輯器帶到基因組的正確位置。但我們很快遇到了一個棘手的難題；具體來說，在DNA中沒有已知的蛋白質(zhì)可以將A轉(zhuǎn)化成G 或者T轉(zhuǎn)化成C。面對如此嚴(yán)重的困難險阻，很多學(xué)生可能會尋找其他方案，而不是咨詢其他研究顧問。（笑聲）但Nicole同意繼續(xù)實施一項當(dāng)時看來雄心勃勃的計劃。鑒于缺乏一種自然產(chǎn)生的蛋白質(zhì)來進(jìn)行必要的化學(xué)反應(yīng)，我們決定在實驗室里進(jìn)化我們自己的蛋白質(zhì)來把A轉(zhuǎn)化成一個像G一樣的堿基，從一種對RNA進(jìn)行相關(guān)化學(xué)反應(yīng)的蛋白質(zhì)開始。我們建立了達(dá)爾文適者生存選擇體系，探索了數(shù)千萬種蛋白質(zhì)變異，只允許那些能夠進(jìn)行必要化學(xué)反應(yīng)的罕見變異存活下來。我們最終得到了這里顯示的蛋白質(zhì)，第一個能把DNA中的A 轉(zhuǎn)化成類似G的堿基。當(dāng)我們把這個蛋白質(zhì)連接到受到抑制的CRISPR剪刀上，這里用藍(lán)色標(biāo)示，第二代堿基編輯器就誕生了，可以把A轉(zhuǎn)變?yōu)镚，然后使用第一代堿基編輯器中同樣的鏈切割策略誘騙細(xì)胞用C取代未編輯的T，當(dāng)它重新生成單鏈后，就完成了A-T堿基對到G-C堿基對的轉(zhuǎn)變。

11:16

(Applause)

（掌聲）

11:18

Thank you.

謝謝。

11:20

(Applause)

（掌聲）

11:23

As an academic scientist in the US, I'm notused to being interrupted by applause.

作為一個美國學(xué)術(shù)科學(xué)家，我還不是很習(xí)慣被掌聲打斷。

11:28

(Laughter)

（笑聲）

11:31

We developed these first two classes ofbase editors only three years ago and one and a half years ago. But even inthat short time, base editing has become widely used by the biomedical researchcommunity. Base editors have been sent more than 6,000 times at the request ofmore than 1,000 researchers around the globe. A hundred scientific researchpapers have been published already, using base editors in organisms rangingfrom bacteria to plants to mice to primates.

我們開發(fā)的這兩代堿基編輯器分別誕生于3年前和1年半前而已。但在這短短的時間里，堿基編輯器已經(jīng)被生物醫(yī)學(xué)團(tuán)隊廣泛使用。堿基編輯器應(yīng)全球超過 1000位研究者的請求已經(jīng)被發(fā)送到全球各地多達(dá)6千次。目前發(fā)表的相關(guān)科研論文多達(dá)百篇，包括了從細(xì)菌到植物，從老鼠到靈長類動物的生物體中使用的堿基編輯器。

12:07

While base editors are too new to havealready entered human clinical trials, scientists have succeeded in achieving acritical milestone towards that goal by using base editors in animals tocorrect point mutations that cause human genetic diseases. For example, acollaborative team of scientists led by Luke Koblan and Jon Levy, twoadditional students in my lab, recently used a virus to deliver that secondbase editor into a mouse with progeria, changing that disease-causing T backinto a C and reversing its consequences at the DNA, RNA and protein levels.

堿基編輯器還太新，尚未進(jìn)入人體臨床試驗，科學(xué)家們已經(jīng)在為之努力了，他們成功使用動物的堿基編輯器來糾正導(dǎo)致人類遺傳疾病的點突變。比如，由Luke Koblan和Jon Levy領(lǐng)導(dǎo)的一個科學(xué)家合作小組，外加我們實驗室的兩個學(xué)生，最近使用了一種病毒將第二代堿基編輯器植入患有早衰癥的老鼠體內(nèi)，把致病的T變回C，并在DNA、RNA和蛋白質(zhì)層面上逆轉(zhuǎn)了其導(dǎo)致的后果。

12:48

Base editors have also been used in animalsto reverse the consequence of tyrosinemia, beta thalassemia, musculardystrophy, phenylketonuria, a congenital deafness and a type of cardiovasculardisease -- in each case, by directly correcting a point mutation that causes orcontributes to the disease. In plants, base editors have been used to introduceindividual single DNA letter changes that could lead to better crops.

堿基編輯器也被用于動物身上來逆轉(zhuǎn)酪氨酸血癥，地中海貧血，肌營養(yǎng)不良，苯丙酮尿癥，某種先天性耳聾和某種類型的心血管疾病——在這些案例中，通過直接糾正導(dǎo)致或者參與致病的點突變就可以逆轉(zhuǎn)病癥。在植物中，堿基編輯器已被用于引入單個DNA字符的改變以帶來更好的收成。

13:22

And biologists have used base editors toprobe the role of individual letters in genes associated with diseases such ascancer. Two companies I cofounded, Beam Therapeutics and Pairwise Plants, areusing base editing to treat human genetic diseases and to improve agriculture.All of these applications of base editing have taken place in less than thepast three years: on the historical timescale of science, the blink of an eye.

生物學(xué)家也使用了堿基編輯器來探索單個堿基在與癌癥等疾病相關(guān)的基因中的作用。我聯(lián)合創(chuàng)辦的兩家公司，Beam Therapeutics和Pairwise Plants，正使用堿基編輯器治療人類基因疾病和改善農(nóng)業(yè)。所有這些對堿基編輯的應(yīng)用都發(fā)生在不到三年的時間里：在科學(xué)的歷史尺度上，這只是一眨眼的功夫。

13:52

Additional work lies ahead before baseediting can realize its full potential to improve the lives of patients withgenetic diseases. While many of these diseases are thought to be treatable bycorrecting the underlying mutation in even a modest fraction of cells in anorgan, delivering molecular machines like base editors into cells in a humanbeing can be challenging. Co-opting nature's viruses to deliver base editorsinstead of the molecules that give you a cold is one of several promising deliverystrategies that's been successfully used. Continuing to develop new molecularmachines that can make all of the remaining ways to convert one base pair toanother base pair and that minimize unwanted editing at off-target locations incells is very important. And engaging with other scientists, doctors, ethicistsand governments to maximize the likelihood that base editing is appliedthoughtfully, safely and ethically, remains a critical obligation.

在堿基編輯器提升基因疾病病人的生命質(zhì)量前，我們?nèi)杂泻芏囝~外的工作要做。盡管許多這些疾病被認(rèn)為是只需要糾正器官中很小一部分細(xì)胞的潛在突變就能治療的，將分子機器(如堿基編輯器) 送入人體細(xì)胞仍然富有挑戰(zhàn)。利用自然界的病毒來傳遞堿基編輯器，而不是讓你感冒的分子來做這個，是幾種已經(jīng)成功實踐的有前景的傳遞策略之一。繼續(xù)研究開發(fā)新的分子機器，找到其他的方法將一個堿基對轉(zhuǎn)變成另一個堿基對，并盡量減少細(xì)胞非目標(biāo)位置上不必要的編輯是非常重要的。與其他科學(xué)家、醫(yī)生、倫理學(xué)家和政府合作，最大限度地提高堿基編輯用于深思熟慮、安全和合乎道德的可能性，仍然是一項重要義務(wù)。

14:57

These challenges notwithstanding, if youhad told me even just five years ago that researchers around the globe would beusing laboratory-evolved molecular machines to directly convert an individualbase pair to another base pair at a specified location in the human genomeefficiently and with a minimum of other outcomes, I would have asked you,"What science-fiction novel are you reading?" Thanks to arelentlessly dedicated group of students who were creative enough to engineerwhat we could design ourselves and brave enough to evolve what we couldn't,base editing has begun to transform that science-fiction-like aspiration intoan exciting new reality, one in which the most important gift we give ourchildren may not only be three billion letters of DNA, but also the means toprotect and repair them.

盡管有這些挑戰(zhàn)，如果你在五年前告訴我全球的研究人員將使用實驗室發(fā)明的分子機器來直接有效地把單個堿基對轉(zhuǎn)變成另一個堿基對，放在特定的基因組位置，而且不會產(chǎn)生其他結(jié)果，我會反問你，“你是不是在讀哪本科幻小說？”感謝我們孜孜不倦的學(xué)生，他們有驚人的創(chuàng)造力來設(shè)計工具，使得我們可以改造自身，并勇敢地去進(jìn)化原本無法進(jìn)化出的特征，堿基編輯已經(jīng)開始將科幻小說般的渴望轉(zhuǎn)變成令人興奮的現(xiàn)實，我們給孩子們最重要的禮物可能不再只是30億DNA個堿基，同時還有保護(hù)和修復(fù)它們的方法。

15:52

Thank you.

謝謝。

15:53

(Applause)

（掌聲）