Gene editing (CRISPR): Thing to watch and thing to come !?

The other day I talked to our daughter, Janice, about my reading of the development of monoclonal antibody (from historical perspective) and its impact on current clinical medicine.  She alerted me the CRISPR, which is basically a gene editing.

There are two components of gene editing: One is Cas9 enzyme which cuts DNA and the other is a snippet of RNA that guides the molecular scissor to where the researcher wants to cut, using the complementary DNA-RNA sequences.  As the RNA sequences can be custom made, so the scientists can cut the DNA as he/she pleases.

While reading gene editing, my mind went right to sickle cell anemia.  In 1957 (at Cambridge U Cavendish Lab, supported by Max Perutz and Francis Crick) Vernon Ingram discovered the replacement of Glutamic acid by Valine at 6th position of β-globin chain of sickle cell HbS as predicted by Linus Pauling.

In an article published in Science (11/25/1949), Linus Pauling demonstrated the hemoglobin from ones with sickle cell anemia has a different electrophoretic mobility from that of a healthy individual.  Pauling’s quick mind led him to suggest that the difference was the result of different ionizable amino acids sequence.

Pauling was indeed correct.  A replacement of a single amino acids in hemoglobin can wreak such havoc.  The life as we know it is so precise.  This is why medicine has to be exact and precise that science calls for.

This is how sickle cell anemia was the first disease confirmed to be the molecular disease.  Vernon Ingram (born in Germany, left for England, repulsed by Hitler Nazi, spent some years in Rockefeller Institute and Yale U.) was touted as the father of molecular medicine; I would call Pauling the grandfather of molecular medicine.

I initially got excited that gene editing would be the ideal way to “cure” sickle cell anemia by re-replacing the Glutamine back to the position 6 of β-globin chain.  Then I read the news of a gene-editing (CRISPR) summit in December 2015, convened by U.S. National Academies of Sciences and Medicine, Chinese Academy of Science and Royal Society of London.   The summit statement calls for cautious development of medical applications such as correction of the mutation that cause sickle cell anemia or modification of immune cells to target cancer.

This is almost tantamount to a moratorium to ban the gene editing research.  You may be interested in reading the story of “Sidney Brenner and the lift of the ban on recombinant DNA research” that I posted in 2011.  Brenner is about 90 now, this may stir this “lion in winter” again.

http://doctorjytsai.blogspot.com/2011/04/sydney-brenner-and-lift-of-ban-on.html

It is still difficult to predict the use of gene editing in clinical medicine, not only when, but also “if” (can be used).  Scientific progress is inexorable; the research must go on, although caution indeed has to be exercised as we’re tinkering with the “Language of God”.

CRISP (clustered regularly interspaced short palindromic repeats) is a prokaryotic immune system, a way how bacteria acquire “acquired immunity”.  Cas9 was the first nuclease discovered. The scientists got the idea of gene editing by studying how bacteria work to confer resistance to foreign genetic elements.

Sydney Brenner and the lift of the ban on Recombinant DNA research

What is this scrumptious dish got to do with Recombinant DNA or Sydney Brenner?  The advantage of dining alone is one can indulge on the food for both body and mind.  I went to a restaurant while Katy was visiting Taiwan.  I wrote a post about Brenner after I received the book of his biography (see photo) and before reading it.  I couldn’t wait to read the story about the role he played in lifting the ban on recombinant DNA research; I finished Chapter 18 (Getting back to DNA) while eating my lunch.

Seeing this Cedar Plank Salmon (Oven roasted tomatoes, crimini mushrooms, shaved fennel and red onion, goat cheese, sautéed asparagus, and sun-dried tomato pesto—I didn’t even mention salmon), Katy would know where I was eating.  Another reason I put this dish as the leading photo of this post is to attract the potential readers—fine food always acts like a magnet—besides, who is interested in reading Sydney Brenner or recombinant DNA?

Sydney is not only a brilliant scientist in the field that he created (his Nobel Prize attests to it), he is also a charismatic, provocative, and entertaining man in science that renders many tales to tell.  I do not intend to explain recombinant DNA in the interest of the length of this post, except saying a few words about its players: Paul Berg (Nobel Prize in Chemistry for recombinant DNA in 1980; Miao-Chih mentioned last year that he was still at Stanford U Med School; he was actually born in that hospital), Stanley Cohen (whose 1986 Nobel Prize in Medicine was not directly related to recombinant DNA), Herbert Boyer (no Nobel glory for him, but he was the co-founder of Genentech, now under the arm of Roche pharmaceutical giant).

Simply put; recombinant DNA is to use virus to introduce pieces of DNA—foreign gene—into mammalian cells, thus known as genetic engineering; this is tantamount to playing God!  If it works Berg envisioned the possibility that he could use this system to insert useful genes into human cells; isn’t that the concept of gene therapy that we eventually came to know?  Berg was using SV40 (simian virus 40) which had a potential to cause human cancer.  Biohazard anxieties, followed by activist outcry and Cambridge, MA, banned recombinant DNA research and its mayor (Alfred Pellucid) enjoyed his reputation as society’s scientific watchdog.

A letter was published in Science in 1973 to call upon the scientists throughout the world to suspend voluntarily all recombinant researches.  This letter was letter known as “Moratorium Letter”—the dark days in science—was signed by Berg, Cohen, Boyer, and even James Watson, who regretted later (“I found myself feeling deeply frustrated and regretful of my involvement in the Moratorium Letter.” Watson wrote in his DNA: the Secret of Life)

Nature also published an editorial (July 19, 1974) endorsing the ban saying “. . . calls for a temporary halt on two types of genetic engineering research because of the risk of infecting man with bacterial containing hybrid DNA molecules whose biological properties cannot be predicted in advance.”

Entered Sydney Brenner. Berg sought Brenner’s advice on the ethics of genetic engineering work in early 1970s. Brenner played a leading role in 1975 Asilomar Conference which was a turning point in lifting the ban.  He continued to actively engage in the recombinant DNA debates and ensuing legislative considerations until 1978.  Berg composed a glowing tribute and appreciation to Brenner on his 75th birthday.

It was Brenner’s wit, wisdom, passion, and courage that changed the views of many skeptical scientists or even mocked Erwin Chargaff (who discovered there are same amounts of adenine and thymine, and of guanine and cytosine, in DNA, that helped Watson and Crick to figure out DNA structure),

“ People like Erwin Chargaff said that we were tampering with evolution.  How on earth can one ask scientists to be responsible for something that might happen ten million years in the future?”  In talking about he risk of recombinant, he said, “By multiplying a set of small probabilities we could reduce the hazard to very small numbers, say 10-24, that is 10 to he power of -24 (reciprocal of Avogadro’s number) or even 10-50, that is 10 to the power of -50 (about the reciprocal of the number of elementary particles in the universe.)"

In order to prove the bacteria of interest cannot survive outside the laboratory, he once mixed the microbes into a glass of milk and drank it.  He monitored what came out “the other end” and found no evidence that the bacteria colonize his intestine.

This post is mainly about the incomparable Brenner, the benefit of the recombinant DNA research will be another post.

My niece’s (Miao-Chih) article was published in Science 6 August 2010

After receiving her Ph.D. from the University of Cambridge, Cambridge, England, she has been a postdoctoral fellow researcher at Stanford University School of Medicine.  Her article was published in Science August 6, 2010, “Long Noncoding RNA as Modular Scaffold of Histone Modification Complexes”.

If I can explain this to you, it wouldn’t be published in Science—like Nature, Science is the most prestigious and the most competitive scientific journal.  To publish in Science or in Nature must be the dream of everyone in the field of science.

Once when Snoopy quoted Psalm Fifty Verse Twelve to Charlie Brown who couldn’t come up with anything; he then said, “Give me two weeks and I will find a verse to answer you!” So give me two weeks and hopefully I’ll have a thing or two to say.

Let this post serve as my hearty congratulations! I take pride in posting this.

(A few weeks ago when Miao Chih said she was going to mail me the Science journal, I could hardly wait so I went to the Waynesburg University library browsing the Science journals of the past few months thinking her article must just have been published recently. Of course I didn’t find it as it was published last August.  But, I came across a book review about Sydney Brenner—a scientist I never tired of reading.  The book is “Sydney Brenner: a Biography” by Errol Friedberg (I am writing a post of “The Writing Life of James D. Watson” (a book, also authored by Errol Friedberg).  It is of interest that the reviewer wrote, “Surely You’re Joking Mr. Brenner!” This is of course, from “Surely You’re Joking Mr. Feynman!” A book Joy gave me when she was an undergraduate, only after she finished reading; her post-it notes are still on the book.)

The “lost” letters of Francis Crick re-surfaced adding flavor and spice to the DNA story

Our friends Peggy and Harry brought to my attention about the “lost letters” of Francis Crick written during the critical time when the discovery of DNA structure was in the making. They are supposed to be long lost and about 30 letters (1950-1976) re-surfaced mixing in Sydney’s Brenner’s files; about 9 or 10 boxes of which were donated to the Cold Spring Harbor Lab. And the discovery of these “lost” letters was announced in the September issue of Nature.

I suspect the timing may be more than just coincidence of the publication of Sydney Brenner: a biography by Errol Friedberg, M.D. (by Cold Spring Harbor Lab Press at the end of September); it will generate more interest on the story of the discovery of DNA structure in general and on Brenner in particular.

Sydney Brenner is on the list of my “Cultivated Reconnaissance Men in Science”; he established the elegant C. elegans as the model organism in late 1950s. I was very pleased by his Nobel Prize in 2002; a well deserved and long delayed honor; he shared with Robert Horvitz and John Sulston for the programmed death or apoptosis. Sidney shared the same office at Cambridge U. with Crick for twenty some years and followed Crick to the Salk Institute at La Jolla, California. Brenner also had great contribution to unraveling the genetic codes and establishing the safety of recombinant DNA technology. 

These “lost” letters really didn’t reveal any thing that was contradicting to what we’ve known; but it adds more flavor and spice to the many already controversial issues.

It confirms that Watson and Crick was shown Rosalind Franklin’s B form DNA—a form containing more water and revealing the double-helical structure. Thing might not have occurred as we know it had they been shown the A form DNA—less water content, but clearer image and less obvious double-helical structure.

I was surprised to learn that Watson and Crick constructed a triple-helix model in 1951; I was led to believe that it was Linus Pauling who built a triple-helix model and thus put the negatively charged phosphate group inside the hydrophobic environment—a fatal mistake. 

If I do not recall wrong, Miao-Chih’s professor at the National Taiwan University before she went to Cambridge U. for her Ph.D. was a student of Robert Horvitz (at MIT) whose mentor was Sidney Brenner at Cambridge U.  All these scientists, including Miao-Chih, thrived on this worm (Caenorhabditis elegans).  Brenner once quipped this worm should share the Nobel Prize, too.

“Junk” RNA may not be junk after all, Miao-Chih says so in Science

My niece, Miao-Chih, who has been a post-doc researcher at Stanford Medical School the past few years; she had a big day yesterday for her work was just published at Science online (the print version later may have minor revision if needed). Having been published at Science or Nature is the ultimate dream of researchers regardless of what field they are in science. Kudos to Miao-Chih, my hat’s off! I realized how good she was about three years ago when she had a pick of Massachusetts General Hospital or Stanford Medical School. (The following link is for abstract only.)

http://www.sciencemag.org/cgi/content/abstract/science.1192002

Her article is “Long noncoding RNA as Modular Scaffold of Histone Modification Complexes”. If Miao-Chih can explain it in a paragraph or two; it wouldn’t be published at Science.

The great majority of DNA/RNA are non-coding; i.e. not making protein. Recall Frances Crick’s Central Dogma (by the way, Miao-Chih also got her Ph.D. from Cambridge University, England, where Crick and Watson figured out the structure of DNA) of DNA to RNA to protein (DNA to RNA is transcription, and RNA to protein is translation). Those non-coding DNA/RNA or not doing the transcription or translation regulation are traditionally called “junk” DNA/RNA.

Miao-Chih’s work and article is trying to tell us it ain’t so! You just have to trust her; the Science editors have been asking many more questions and more experiments before all questions were answered to their satisfaction.

Just to tell you how frontier a science and what a break through it is; there was a Nobel laureate call her Lab saying he was very interested in their story, but Miao-Chih was one step ahead of him. Non-coding RNA apparently is “hot”; many top-notched scientists are trying to prove the “junk” RNA is not junk at all; I take pride in knowing Miao-Chih is contributing in this frontier of science.

I called my second brother and sister-in-law congratulating them, trying to let them know what kind of a scientific journal/magazine Science is; I do not know whether I succeeded or not.

陳老薑 Congratulations. it's a great achievement indeed.

Lawrence Cancelmi what an accomplished woman!..i will make an attempt at reading the article in science...it seems everything revolves around crick and watson. thank you miao-chih.

Cheng-Hung Lin Amazing!

In defense of Rosalind Franklin

In my post at Sermo about the discovery of DNA structure I failed to mention Rosalind Franklin, a doctor protested saying “With all due respect, you missed Rosalind Franklin, whom J.D. Watson, in The Double Helix credited with the actual x-ray crystallography that sealed the hypothesis.” To which I responded,

I admit it is an unforgivable oversight of not mentioning The Dark Lady of DNA (Rosalind Franklin's biography by Brenda Maddox) when talking about the discovery of DNA structure. Her untimely death in 1958 must make the Nobel Committee much easier to give the awards in 1962. But I have campaigned for her tirelessly over the years. James Watson once suggested that Nobel Prizes should be given in 1962 to him and Crick in Medicine and Wilkins and Franklin in Chemistry, all for the DNA structure. That might be a way to solve the problem had Franklin not passed away then.

Max Perutz and John Kendrew shared the Prize in Chemistry that year; I believe Watson initially went to Kendrew's lab where he hooked up with Crick; that was the golden days for Cavendish Lab.

John Steinbeck was there for Literature that year and even Linus Pauling was there for the Peace Prize or may be the Nobel Committees felt Pauling could or should be (almost) there for the DNA structure, too.

Dr. James Watson in Taiwan

I learn that Dr. James Watson is visiting Taiwan from Edwin Tsai’s Facebook page. The College of Life Science of the National Tsing Hua University is having a 3-day conference (April 1-3, 2010) boasting Dr. James Watson; his mere presence will make it an extraordinary one, even though he is now a Lion in Winter. I do not know whether he’ll be there 3 days in a row or just Today’s (April 3) lecture.

On Feb. 25 I had the following post at Sermo and one doctor remarked that he met Watson at a conference a couple of years ago and “I was totally shaking and star struck. I have never felt that way meeting somebody before. Forget Brad Pitt. Now there is a STAR.”

Another doctor wrote, “Got to see Francis Crick at Northwestern University in 1974, speaking about his subsequent research. Spoke at the Technological Institute [a huge complex; it was said it is bigger than the Pentagon, I visited Joy there on many occasions during her undergraduate years] Auditorium, which seats about 900. I’d suspect there were easily 1,300 or more in the audience.”

The following is what I posted at Sermo on Feb. 28.

Today (Feb. 28) in 1953 Francis Crick went to Eagle pub (nearby Cavendish Lab) to tell everyone within hearing distance that he and James Watson had found the secret of life. And molecular biology was born.

Sidney Brenner once said Crick was the only one he knew can make a living by being a theoretic biologist. He devoted much of later part of his life in “the scientific search for the soul” (the subtitle of his book, Astonishing Hypothesis). Watson is a more prolific and better writer, though.

Crick and Maurice Wilkins (The third man of the double helix) passed away in 2004 and Watson is now a lion in winter. We haven't heard much from him since he made a so-called “racist” remark while promoting his book (Avoid Boring People) in London in late 2007. His memoir came in segments; "The Genes, Girls, and Gmow" ended at his getting married, and "Avoid Boring People" told his life in science up to the time when he left Harvard in 1976. He still has much to say and he shouldn't be silenced.

There is no trace of any racism in “Avoid Boring People”; the outspoken Honest Jim is absolutely not a racist at heart, it is his brutal insensitivity and loose tongue that got him into trouble.

The untimely death of Rosalind Franklin

Speaking of the untimely death of Rosalind Franklin, the following is my comment on her in a post about the early detection of ovarian cancer.

Doctors had difficulty detecting Rosalind Franklin ovarian cancer back in 1956, and we're still facing almost the same dilemma nowadays.

Before leaving for the United States in the summer of 1956 for a 2-month stay at various Labs, including Woods Hole Lab at Cape Cod, Cal Tech, and a 3-week stay at UCLA virus Lab at Berkeley (where she celebrated her 36th birthday), she had a medical check up by a doctor at the University College London's student health; no problems were found. This was a routine physical required of those exposed to radiation risks in their work.

During the last week of her stay, she had trouble zipping her skirt and her "midriff began to bulge". She went to see a doctor; "You're not pregnant?" asked the doctor, "I wish I were," was her answer. The doctor suspected ovarian trouble and the surgery at London confirmed ovarian cancer (the right ovary was the size of a croquet ball) on September 4. She died of abdominal carcinomatosis on April 16, 1958.

The life time risk for ovarian cancer is 1.4% in the US (1 in 71 women), it is 39-46% among the carriers of BRCA1 mutation and 12-20% for BRCA2 mutation. The BRCA mutations are particularly prevalent among Ashkenazi Jews, and I believe Rosalind Franklin was an Ashkenazi Jew, not 100% sure, though.

Immodest Francis Crick--Theoretic Biologist

(While writing my version of Crick’s obituary, I received an e-mail from Janice informing me that Francis Crick succumbed to colon cancer--the was originally written in 2004)

“I have never seen Francis Crick (see photo) in a modest mood.” So James Watson began his story of Double Helix.  And Crick had every reason not to be in that mood.  Janice, I bet it was quite a surprise when his grand-daughter told you that “He is my grand-father!” at Harvard computer room some years ago.)  Reading about him for so many years, I feel like I knew him by heart.

“It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.”--Letter to the Nature (April 25, 1953). Almost every molecular biology professor put this passage in the lecture notes.  This was a compromised statement between Watson and Crick who was 12 years senior to Watson.  The aggressive, quick-witted Crick wanted to elaborate the detailed copying mechanism on the original article of April 25, 1953 (Molecular structure of nucleic acids), the younger, but more cautious Watson was afraid if their hypothesis of copying mechanism were not quite true, it might ruin the whole thing.  They quickly wrote the follow up article (Genetic implications of the structure of deoxyribonucleic acid) that showed up on May 30, 1953 issue of Nature.  It has turned out Crick was correct in the first place.

Sydney Brenner once said Crick was the only one he knew that can make a living by being a theoretic biologist (Crick was working on the PhD in physics when WWII broke out, he then worked on the magnetic and acoustic mine in the British Admiralty.)  After moving to Salk Institute, Crick devoted his life in ‘the scientific search for the soul’ (this is the subtitle of his book Astonishing Hypothesis)—hypothetic neuroscience.

When Watson and Crick constructed that famous DNA model in April 1953, Sidney Brenner (originally from South Africa, born of Jewish parents) was in Oxford University and went to Cambridge to see it.  Brenner and Crick were also together in the Woods Hole Lab and Cold Spring Harbor Lab (in Long Island) in 1954.  Brenner joined Crick at Cavendish Lab in 1956 and “spent 20 years sharing an office with Francis Crick.”  Crick moved to Salk Institute at La Jolla, in 1976 and Brenner rejoined him there again in 2001. Brenner began to work on the ‘elegant’ C. elegans in late 1950s and for which he just received the Nobel Prize (programmed cell death or apoptosis) in Medicine in 2002—exactly 40 years after Watson and Crick did.  Brenner’s Nobel Lecture, Nature’s Gift to Science, talking about the importance of choosing the model organism for the intended research.

It is now almost a ‘common knowledge’ that how DNA uses its four bases to pass along the ‘secrets of life’ to the making of proteins.  It was Crick that first proposed his ‘adaptor’ tRNA (transfer RNA) hypothesis.  It was actually never published; it was just a letter to the then so-called RNA Tie Club. He also worked with Vernon Ingram (who discovered the replacement of Glu by Val at 6^th position of β-globin chain in sickle cell HbS, as predicted by Linus Pauline) and Sydney Brenner in this field.  Brenner was twice a visiting professor at Harvard, as was Crick.

“Crick is very different: brighter than Watson, but he talks a lot, and so he talks a lot of nonsense.” This was from Erwin Chargaff whose discovery of ‘A = T’ and ‘C = G’, along with Rosalind Franklin and Maurice Wilkin’s X-ray crystography of DNA (see photo) were the most important hints to Watson & Crick, in my view.  I have read many reports that Chargaff had a lot of resentments (hence sour grape) that he was ‘so close’ and yet he let the ‘glory’ slip away. He was one of the 20 members of RNA Tie Club.  By the way, Chargaff was another Jew who fled Hitler’s Nazi. (my favorite topic that I mentioned in summer reading.2_2004)

It took many years before the DNA structure hypothesis was firmly accepted.  Matthew Meselson’s semiconservative replication provided the first confirmation.  Meselson took his Ph.D. from Linus Pauling at Cal Tech and who got an honorary degree at Joy’s graduation at Northwestern in 2003.

A few years ago, Time published 100 most influential people of the last century; both Watson and Crick are on the list.  Watson gave an introductory talk on the 2001 Nobel Symposia (Beyond genes) in Sweden, and Crick was no where in sight.  Throughout 2003 there were numerous activities celebrating the 50^th anniversary of the discovery of DNA structure.  Watson savored the glories on many functions, and Crick none.  He was not in his best health then, but that was not the main reason.  He preferred to read and think and hypothesize his science.