Interview with Lorena de Oñate, postodoctoral researcher in the Innovative Genomics Institute, led by Prof. Doudna, winner of the Nobel Prize in Chemistry.

Laura Moro, Ph.D.

As you all know, this year Nobel Prize in Chemistry was recently awarded to Prof. Emmanuelle Charpentier and Prof. Jennifer A. Doudna who discovered one of gene technology’s sharpest tools: the CRISPR/Cas9 genetic scissors. We spoke with Lorena de Oñate, PhD, a postdoctoral researcher from Barcelona currently working at the Innovative Genomics Institute from the University of Berkeley, led by Prof. Doudna.

Can you introduce yourself?

I am a biologist by training at the Autonomous University of Barcelona (UAB). I have always been interested in medicine and research so I did my Master at the Pompeu Fabra University (UPF) focused on biomedical research and that was my first contact with human embryonic stem cells and induced pluripotent stem cells for regenerative medicine purposes with Dr. Vassena and Prof. Veiga. Then, I continued my research at the Regenerative Medicine Center in Barcelona (CMRB) with an FPI Grant as a PhD student, working on how to generate heart cells in a dish from different cell sources for heart cell therapy. During my PhD I had the opportunity to do an internship at the Salk Institute in La Jolla, California, where my thesis director, Prof. Izpisúa, also has a laboratory, and I started using CRISPR as a tool for cell engineering. Back to Barcelona, I finished my PhD working at the Institute for Bioengineering of Catalonia with Dr. Montserrat and, after my dissertation, I moved back to California to join Prof. Doudna’s laboratory at the University of California, Berkeley, in order to help Dr. Ross Wilson (a Doudna’s laboratory postdoc at that time) to set up his new laboratory and work on genome editing technologies for cell therapy.

What is your research about?

My current research is on the development of therapeutic delivery of CRISPR/Cas for genome editing in human cells.

I am engineering the two components of CRISPR/Cas9 system: Cas9 protein and the RNA molecule needed to guide Cas9 and find the target sequence in order to enable genome editing in clinically relevant cells.

The idea is targeting the cells in need of genetic correction exclusively, and to do that I use specific cell features as cell locks to enter the cell and molecular targeting as the key to open these locks or entrance doors to the nucleus of the cell. I currently work with liver cells to do genome editing and correct monogenetic diseases, such as hypercholesterolemia or tyrosinemia, and with hematopoietic stem cells, to bring novel therapies for sickle cell anemia and other blood diseases and avoid the harm and risk of  transplantation in patient in the future. We also work with T-cells and brain genome editing in vivo.

How did you feel when you knew about the Nobel prize awarded to Prof. Doudna and how was the celebration at your institute?

Honestly, we have been waiting for this every year and probably this is the first one in which I kind of forgot about it. The phone was burning and woke me up by 5:00 am in the morning. We all watched the announcement at Berkeley on Zoom and I felt thrilled and very proud of both, Prof. Charpentier and Prof. Doudna! Jennifer is an outstanding scientist and an incredible human being. She is an inspiration for all of us, especially women, because of her passion, commitment, persistence and leadership capacities. Looking back, it is a great moment for women and I am very happy that Prof. Doudna will be part of our visible history.

I also admit that I had a bitter sensation because Dr. Francis Mojica -the Spanish microbiologist who first discovered and named CRISPR- was not awarded. We all kind of knew but still hurts and it was a reminder of how science in Spain is still not considered essential and how we are the first ones not promoting and supporting our scientists to expand frontiers and communicate what we can do.

And about celebrations… I only got to say congratulations at the hallway of our building in the morning with a social distance elbow touch and, because of COVID-related restrictions on Campus, they decided to do gather over zoom (lame). Over 300 people attended that Zoom call!

I heard that she promised once that, if she was awarded with the Nobel Prize, she would take all the lab members from that time (2012-2013) to Hawaii, where she grew up.

Can you provide a short description of CRISPR-Cas9 technology and why it is so revolutionary?

CRISPR is and adaptive immune system originally found in bacteria that allows detecting viral DNA and destroy it to protect the cell.  It is composed by the Cas9 protein which has enzymatic or nuclease activity because it can cut nucleotide sequences (DNA) together with a RNA molecule that guides Cas9 to find a precise target site to cut. CRISPR-Cas9 technology stands for the use of this system as a tool to perform gene editing in any organism you can imagine on earth; from humans to tomatoes, from flies to yeast and beyond.

It is so revolutionary because it is very precise compared to previous genome editing technologies, it is easy programmable due to the interchangeable RNA molecule that directs the protein to the DNA sequence of interest and it is cheaper and easier to manufacture than other proteins involving the use of prokaryotes instead of animals like other biologics such as antibodies.

This technology has a tremendous impact in: 1) medicine, because it can cure genetic diseases in humans; 2) in research, because allows scientist to change DNA sequences in a precise manner and observe the outcomes to rewrite and advance cell and molecular biology, embryo development, proteomics, genetics or any discipline you can imagine; 3) agriculture, because precise gene edits in plants and crops can avoid the tedious traditional breeding and generate products that can resist the climate change that we are facing; 4) in the food industry, because it allows to generate new food products that will allow to feed the overpopulated planet stopping animal cruelty for example; 5) energy, because the possibility to engineer novel ways to make biofuel, 6) diagnostics; 7) plagues and pandemic control by gene drive; 8) even technology and so on!

The first time I used Cas9 I felt completely amazed and grateful for its discovery and it literally saved my entire PhD!
Thanks to Cas9 I was able to engineer a human embryonic stem cell line by adding a fluorescent protein to its genome as a so-called “reporter” to understand how cardiac differentiation was happening in vitro and to help me define the best cell culture media to generate human cardiomyocytes in the lab.

What are the main barriers for its application?

In my field, for example, and technically speaking, there are still some barriers to overcome like how to make Cas9 a safe bio-drug that can be injected directly into the patient, because there is the possibility of immunogenicity and also there are physiological and biological barriers that will prevent Cas9 to do its job. Fortunately, those barriers will be solved in the near future with deepest understanding of immunology, directed evolution and proteomics to tune up Cas9.

However, there are other giants that we have to confront before we see CRISPR as a fully accepted technology in society. The only way to overcome those barriers is with education and efficient communication as well as deeper understanding of the technology, trust generation and clear and consensus in regulation because society is scared out of hell about new technologies. It is our nature. CRISPR is something brand new and that is scary. We need to understand the potential of the technology and stick to bioethics to allow its application broadly, but that should be a combined effort and I am very optimistic seen that here at the Innovative Genomics Institute people are fully committed to make that happen.

How do you envisage the future and applications of genome editing? Is CRISPR-Cas9 the last invention that pushed the limits of genetic engineering or there are other methods with similar potential?

CRISPR-Cas9 is just one very effective and easy to use genome editing tool but there are many others among the CRISPR system. Many other Cas proteins are described every year with slightly different activities such as cutting RNA instead of DNA like Cas13. Also, we are engineering them to do other functions such as just activating or inactivating gene expression (CRISPRa, CRISPRi) or to just change a nucleotide precisely by changing its chemistry (CRISPR base editors) thus avoiding to cut the DNA.  We can also engineer Cas proteins to be more compatible with humans and not immunogenic, thus completely safe for cell therapy.

Moreover, this is a system found in bacteria such as Streptococcus Pyogenes, but there are many other orthologs with other features that can provide novel functions such as extreme temperature resistance, fast enzymatic activity or higher fidelity genome editing.

I just think this is the top of the iceberg, and there is way more to discover and explore. Archaea is a fascinating Kingdom to explore, for example, with lots of protein systems still unknown; and thanks to advancements in metagenomics, transcriptomics and synthetic biology we will discover their full and tremendous potential one day.

What are your thoughts on the ethical controversies that surround genome editing?

Every new technology brings new ethical concerns to the table that we might never had to face before, and CRISPR brings that too. We are able to change the code of life with this technology for real!  

The strongest ethical controversies come with the use of CRISPR to edit the genome of human beings at the germline level for enhancement for example. Contrary to genome editing for cell therapy, where CRISPR makes changes in somatic cells like blood cells, muscle, retinal cells to correct a certain mutation that will die with that individual, genome editing in eggs or sperm are transmitted to next generations and that mean that we would be changing our genetics in a driven and controlled way favoring some features over others. That would cause even more disparity and inequality across the planet not only at the economic, education and status level, but also at the genetic level. Racism, discrimination and hate are still pending topics to solve in our society and it is not a matter of using or not a technology like CRISPR. I just see CRISPR as a tool with tremendous potential to advance our society or, by contrary, to perpetuate the dark sides of it like many other technologies did in our history.

Can you give an advice for young scientists that want to pursue a career in academic research?

Do not give up! Always hard work and keep persistent. Be ready to fail very often and bring up your resilience to play.

Remember it is a lifetime job and a lifestyle and that will keep you balanced… I guess 😉 !

Do you want to see how working in a CRISPR lab is? Then do not miss Lorena and her colleagues in this video!