Stuart Orkin in his lab at Boston Children's Hospital in 1985 (Credit: Boston Children's Hospital)

How one lab's decades­long search for the 'holy grail' of sick­le cell led to the first CRISPR ther­a­py

In a 2008 is­sue of Sci­ence, just a page apart, were the two cru­cial dis­cov­er­ies that would form the cure for sick­le cell dis­ease.

Stu­art Orkin, a hema­tol­o­gist at Boston Chil­dren’s Hos­pi­tal, had pub­lished work iden­ti­fy­ing the ge­net­ic switch that could turn on back­up copies of life-sus­tain­ing he­mo­glo­bin pro­teins in sick­le cell pa­tients. And in the very next ar­ti­cle, a pair of mi­cro­bi­ol­o­gists from Illi­nois de­scribed a sys­tem called CRISPR — the tool that could flip that switch, but a mere cu­rios­i­ty at the time.

“Worlds col­lid­ed, right on pa­per,” Orkin re­called in his sev­enth floor of­fice of the hos­pi­tal last week.

The re­search was hailed as the “holy grail” for sick­le cell, a dis­ease that pre­dom­i­nant­ly af­flicts Black peo­ple and for which new treat­ments have been slow to come de­spite un­der­stand­ing its root cause for near­ly 75 years.

“We all knew that fe­tal he­mo­glo­bin was ben­e­fi­cial if you had more of it in sick­le cell or in tha­lassemia,” Orkin said. But for decades, sci­en­tists couldn’t fig­ure out what turned pro­duc­tion of fe­tal he­mo­glo­bin on and off. “We didn’t know what we were look­ing for.”

It would take years for those two dis­cov­er­ies to con­verge, cul­mi­nat­ing a decade and a half lat­er in the first-ever com­mer­cial gene edit­ing ther­a­py, al­ready ap­proved in the UK and was just ap­proved in the US to­day.

And while the drug it­self be­longs to CRISPR Ther­a­peu­tics and Ver­tex Phar­ma­ceu­ti­cals — which made, test­ed, and com­mer­cial­ized it — Cas­gevy is in many ways Orkin’s brain­child, and the out­come of years of work by teams around the globe who pieced to­geth­er the tar­gets and tools that would make it pos­si­ble.

A cause with­out a cure

Sick­le cell dis­ease is char­ac­ter­ized by red blood cells that are mal­formed in­to a sick­le shape, lim­it­ing their abil­i­ty to trans­port oxy­gen. But they are not that way at birth: In 1948, Brook­lyn pe­di­a­tri­cian Janet Wat­son no­ticed that ba­bies who would lat­er go on to de­vel­op sick­le cell were born with most­ly healthy blood cells.

A year lat­er, in 1949, chemist Li­nus Paul­ing dis­cov­ered that the struc­ture of he­mo­glo­bin pro­teins were dif­fer­ent in peo­ple with sick­le cell. The study was cel­e­brat­ed as the first mol­e­c­u­lar de­scrip­tion of a dis­ease, and led bi­ol­o­gist Ver­non In­gram to pin­point the pre­cise mu­ta­tions that cause sick­ling in 1956.

Wat­son hy­poth­e­sized that fe­tal he­mo­glo­bin was re­sis­tant to sick­ling. And there were some peo­ple who car­ried genes for sick­le cell but whose blood cells, for rea­sons un­known, nev­er stopped mak­ing fe­tal he­mo­glo­bin.

The dis­cov­er­ies cap­ti­vat­ed gen­er­a­tions of hema­tol­o­gists, but would frus­trate them as well. Sick­le cell be­came a text­book ex­am­ple of a ge­net­ic dis­ease, where a sin­gle ty­po cre­ates enor­mous trou­ble. Even when CRISPR gene edit­ing de­buted a decade ago, the tool was bet­ter at si­lenc­ing un­want­ed genes than fix­ing bro­ken ones. And cut­ting out the ty­po would on­ly man­gle he­mo­glo­bin fur­ther.

For decades, the so­lu­tion felt like it was in plain sight. Turn back on fe­tal he­mo­glo­bin, and you could give pa­tients a nor­mal life. And there was phar­ma­ceu­ti­cal progress on that front: Hy­drox­yurea, a drug used to slow the growth of can­cer, in the ear­ly 1980s was shown to trig­ger mod­est fe­tal he­mo­glo­bin pro­duc­tion in mon­keys, and the drug was even­tu­al­ly ap­proved for treat­ing sick­le cell pa­tients in 1998. But it doesn’t com­plete­ly ad­dress the dis­ease, doesn’t work for every­one, and can cause side ef­fects like nau­sea and vom­it­ing.

A rush to clone and se­quence genes in the 1980s fu­eled a search for the enig­mat­ic switch. Sci­en­tists stared at the ge­net­ic code of dif­fer­ent forms of he­mo­glo­bin look­ing for clues. But it was un­known if a sin­gle pro­tein con­trolled the process, or if there was a more com­plex cir­cuit in­volved.

“Dur­ing the dol­drums, we thought there were prob­a­bly five to 10 fac­tors, and what was the hope we’re ever go­ing to fig­ure this out?” Orkin said. “It was a frus­trat­ing era.”

Stuck, he moved on­to oth­er prob­lems and the search for the switch slow­ly fiz­zled out.

The search for the switch 

Al­most 20 years lat­er in 2004, an MD-PhD stu­dent named Vi­jay Sankaran was in­ter­est­ed in sick­le cell dis­ease and had seen hy­drox­yurea help, but al­so knew its lim­i­ta­tions. Orkin’s lab had long since giv­en up on find­ing the fe­tal he­mo­glo­bin switch, and Sankaran want­ed to re­sume the search.

Orkin of­fered him a dose of re­al­i­ty. “That’s great,” Orkin told him. “It’s re­al­ly the most im­por­tant prob­lem we can work on. But most of the peo­ple who worked on it have ei­ther failed, they’ve gone on to oth­er projects, or they’re dead.”

But one thing had changed. The re­cent com­ple­tion of the Hu­man Genome Project was pro­pelling ge­net­ic sleuthing in­to a new era. One of Sankaran’s pro­fes­sors, the fu­ture Ver­tex ex­ec­u­tive David Alt­shuler, preached that faster and cheap­er DNA se­quenc­ing would al­low sci­en­tists to com­pare hun­dreds, maybe thou­sands, of genomes to fish out ge­net­ic vari­ants linked to dis­ease.

Sankaran be­gan to look for com­mon­al­i­ties across the DNA of peo­ple with sick­le cell dis­ease who had fe­tal he­mo­glo­bin, com­pared to those who didn’t. At the same time, a group led by An­to­nio Cao in Italy was al­ready a step ahead, and had iden­ti­fied a few spots linked to the fe­tal pro­tein us­ing the tac­tic, known as a genome-wide as­so­ci­a­tion study, or GWAS.

In 2007, Cao sub­mit­ted the study to the New Eng­land Jour­nal of Med­i­cine, but the ed­i­tors re­ject­ed it, ask­ing for a sec­ond co­hort for con­fir­ma­tion. Cao’s lab teamed up with Orkin’s group, and with their sick­le cell study, linked a gene called BCL11A to per­sis­tent pro­duc­tion of fe­tal he­mo­glo­bin in adults.

“The jour­nal still re­ject­ed it,” Orkin said. “I have ab­solute­ly no idea why.”

But Sankaran had al­ready moved on to his next, and the most cru­cial dis­cov­ery: that BCL11A was cranked up in adult blood cells, but not fe­tal ones. When he used a syn­thet­ic RNA mol­e­cule to block its pro­duc­tion in blood stem cells, fe­tal he­mo­glo­bin shot up.

The key to treat­ing sick­le cell with fe­tal he­mo­glo­bin wasn’t a com­plex con­stel­la­tion of genes. It was the sin­gle switch they’d been seek­ing. Sankaran was work­ing in the lab late that night and ex­cit­ed­ly called Orkin to tell him the re­sults. The im­pli­ca­tions were im­me­di­ate. Af­ter decades of dead ends, the holy grail of hema­tol­ogy was fi­nal­ly found.

A se­lec­tive strat­e­gy

The land­mark re­sults were pub­lished in Sci­ence in 2008 — next to the short men­tion of CRISPR. Sankaran left to fin­ish med­ical school, and an in­com­ing re­search fel­low, Dan Bauer, would car­ry the torch and help turn the dis­cov­ery in­to a ther­a­peu­tic strat­e­gy.

Vi­jay Sankaran (L) and Dan Bauer (Boston Chil­dren’s Hos­pi­tal)

Click on the im­age to see the full-sized ver­sion

Do­ing so would prove tricky. Break­ing BCL11A in all cells would cause prob­lems — bone mar­row cells, for ex­am­ple, re­quire the pro­tein to give rise to red blood cells and oth­er things cir­cu­lat­ing through­out the body.

They need­ed a way to shut the switch off on­ly in red blood cells. But the vari­ants they need­ed to tar­get were buried in part of the gene called an en­hancer, long lines of code that reg­u­late the pro­duc­tion of the switch it­self.

Us­ing an old­er and more la­bo­ri­ous gene-edit­ing tech­nol­o­gy known as TAL­ENs, Bauer re­moved the en­hancer, which trig­gered an al­most com­plete loss of BCL11A in red blood cells, while spar­ing oth­er cells, and boost­ing fe­tal he­mo­glo­bin in the process.

“It was a much big­ger ef­fect than I ex­pect­ed,” Bauer said.

Fy­o­dor Urnov, a gene edit­ing sci­en­tist who was work­ing on sick­le cell dis­ease at Sang­amo Ther­a­peu­tics at the time, called Bauer and Orkin’s work a “slight­ly mys­ti­cal mo­ment” for the field.

“That gave us a path to­wards in­ter­fer­ing with BCL11A func­tion with­out any side ef­fects,” said Urnov, who is now a di­rec­tor at the In­no­v­a­tive Ge­nomics In­sti­tute at UC Berke­ley and a con­sul­tant to Ver­tex. “If you had told me 15 years ago that we would be gene edit­ing an en­hancer to treat sick­le cell dis­ease, I would have po­lite­ly ques­tioned your san­i­ty.”

Ear­ly adopters

As Bauer’s en­hancer pa­per was go­ing to press in 2013, CRISPR was mov­ing from what had been a cu­rios­i­ty in the 2008 pages of Sci­ence, in­to a rev­o­lu­tion­ary new tech­nol­o­gy. Bauer fig­ured if they could break the en­hancer with a key snip from the mol­e­c­u­lar scis­sors at just the right spot, it would be a sim­pler and safer ap­proach than yank­ing the whole thing out.

Orkin, now 77, was one of CRISPR’s ear­ly adopters and part­nered with one of the tech­nol­o­gy’s in­ven­tors, Feng Zhang at MIT, to fig­ure out where pre­cise­ly with­in the en­hancer’s 13,000 let­ters of code to tar­get.

Stu­art Orkin in 2017 (Boston Chil­dren’s Hos­pi­tal)

Click on the im­age to see the full-sized ver­sion

The team pub­lished in Na­ture in 2015 and filed patents on sev­er­al guide RNA mol­e­cules that are used to di­rect CRISPR to the BCL11A en­hancer sites.

But Orkin said he wasn’t in­ter­est­ed in push­ing the ex­per­i­men­tal ther­a­py in­to clin­i­cal tri­als on his own.

“If you re­al­ly want to bring it to pa­tients, it’s got to be done by phar­ma,” he said. “If we had start­ed a tri­al here, we’d still be on our third pa­tient or some­thing.”

He in­struct­ed the hos­pi­tal to make li­cens­es to the tech­nol­o­gy non-ex­clu­sive, a de­ci­sion that would re­sult in small­er roy­al­ties for Boston Chil­dren’s, but would max­i­mize po­ten­tial ac­cess to the many com­pa­nies eye­balling gene edit­ing ther­a­pies for sick­le cell at the time, in­clud­ing CRISPR, Ed­i­tas, In­tel­lia, Sanag­mo and Ver­tex.

“I didn’t want to bet on one,” Orkin said.

In the mean­time, Alt­shuler — the ge­net­ic pro­fes­sor whose lec­tures in­spired Sankaran’s work — had end­ed up at Ver­tex. He’d shared an of­fice with Zhang in his pre­vi­ous role at the Broad In­sti­tute of MIT and Har­vard, was at­tuned to the po­ten­tial of gene edit­ing, and helped strike a part­ner­ship with CRISPR Ther­a­peu­tics that fall.

CRISPR and Ver­tex looked for their own guide mol­e­cules to go af­ter BCL11A, but ul­ti­mate­ly end­ed up us­ing one of Orkin’s.

“They couldn’t find a bet­ter one,” Orkin said. “What they’ve done is di­rect­ly trans­late what we de­scribed.”

Less than four years lat­er, 34-year-old pa­tient Vic­to­ria Gray got an in­fu­sion of her own blood stem cells, care­ful­ly snipped in the BCL11A en­hancer to boost fe­tal he­mo­glo­bin, in hopes that it would fi­nal­ly put an end to the ex­cru­ci­at­ing pain crises from her sick­led cells.

A qual­i­fied land­mark

The rest is his­to­ry. Near­ly all sick­le cell pa­tients who have got­ten Cas­gevy in clin­i­cal tri­als re­main large­ly free of pain crises, and most be­ta tha­lassemia pa­tients no longer re­quire reg­u­lar blood trans­fu­sions.

“There’s no ques­tion it’s a land­mark. The ques­tion is if it’s more im­por­tant as an ex­am­ple of edit­ing, or as a good ther­a­py for this dis­ease?” Orkin said. “The re­sults are great. But it’s go­ing to be hard to im­pact the dis­ease, be­cause there are lots of peo­ple that will nev­er get treat­ed.”

These ther­a­pies re­quire re­mov­ing stem cells from a pa­tient’s bone mar­row, edit­ing the cells in a lab, and us­ing chemother­a­py to wipe out a pa­tient’s dis­eased cells to make room for the al­tered ones. It’s an ar­du­ous, time-con­sum­ing, and cost­ly ap­proach that means these ther­a­pies are un­like­ly to make a dent in the low­er-in­come coun­tries where sick­le cell dis­ease is more preva­lent.

In the mean­time, Orkin is work­ing on a dif­fer­ent ap­proach: a pill that blocks BCL11A. “It does seem kind of back­wards,” he ad­mits.

It’s a chal­lenge. BCL11A be­longs to a class of pro­teins called tran­scrip­tion fac­tors that are no­to­ri­ous­ly dif­fi­cult to drug, nev­er mind the headache of get­ting the com­pound to tar­get red blood cells and not their stem cell pre­cur­sors.

“If you tell me some­thing’s un­drug­gable I want to see if I can drug it. The chal­lenge is part of it,” Orkin said. “It’s prob­a­bly too hard. But I’m hav­ing a rea­son­ably good time try­ing to fig­ure it out.”


Ryan Cross

Senior Science Correspondent