A quantum leap in biomedicine

Correction of a pathogenic gene mutation in human embryos

Today you should have a look at Nature. The reason is genome editing of human embryos. Although a chinese team performed something similar in 2015, right now the details are clearly explained. The trial has been developed to fix a mutation that causes hypertrophic cardiomyopathy.If you don't want to read the whole article, check the news. Genome editing is gaining momentum as I said some months ago, it is closer than you may think.


PS. Why FBI is concerned about CRISPR? The answer in FT Big Read
PS. The news in FT.
PS. A useful article from Nature 2015:

Genome editing: 7 facts about a revolutionary technology

What everyone should know about cut-and-paste genetics.

• Lucy Odling-Smee,
• Heidi Ledford
• & Sara Reardon

30 November 2015

Article toolsediting is in the spotlight again as a large international meeting on the topic is poised to kick off in Washington DC. Ahead of the summit, which is being jointly organized by the US National Academy of Sciences, the US National Academy of Medicine, the Chinese Academy of Sciences and Britain’s Royal Society and held on 1–3 December, we bring you seven key genome-editing facts.

Yorgos Nikas/Science Photo Library

Human embryos are at the centre of a debate over the ethics of genome editing.

1. Just one published study describes genome editing of human germ cells.

In April, a group led by Junjiu Huang at Sun Yat-sen University in Guangzhou, China, described their use of the popular CRISPR–Cas9 technology to edit the genomes of human embryos. Only weeks before the researchers’ paper appeared in Protein & Cell¹, rumours about the work had prompted fresh debate over the ethics of tinkering with the genomes of human eggs, sperm or embryos, known collectively as germ cells. Huang and colleagues used non-viable embryos, which could not result in a live birth. But in principle, edits to germ cells could be passed to future generations.

2. The law on editing human germ cells varies wildly across the world.

Germany strictly limits experimentation on human embryos, and violations can be a criminal offence. By contrast, in China, Japan, Ireland and India, only unenforceable guidelines restrict genome editing in human embryos. Many researchers long for international guidelines, and some hope that the upcoming summit in Washington DC could be the start of the process to create them.

Ramon Andrade 3Dciencia

The CRISPR-Cas9 system makes editing genomes cheap and easy.

3. You don’t have to be a pro to hack genomes.

The CRISPR–Cas9 technology has made modifying DNA so cheap and easy that amateur biologists working in converted garages or community laboratories are starting to dabble.

4. Cas9 is not the only enzyme in town.

A key ingredient in the CRISPR–Cas9 system is the DNA-cutting enzyme Cas9. But in September, synthetic biologist Feng Zhang at the Broad Institute of MIT and Harvard in Cambridge, Massachusetts, reported the discovery of a protein called Cpf1, which may make it even easier to edit genomes².  (Zhang is one of those who pioneered the use of CRISPR-Cas9 for genome editing in mammalian cells).

BGI

A gene-edited micropig created by the BGI in Shenzen, China. 

5. Pigs are on the front line of genome-editing experiments.

Dogs, goats and monkeys are all part of the growing CRISPR zoo. But pigs in particular have been at the heart of several eye-catching announcements — from micropigs that weigh about six times less than many farm pigs, to super-muscly pigs, to a pig whose genome has been edited in 62 places (the aim being to produce a suitable non-human organ donor).

6. Gates, Google and DuPont want a piece of the genome-editing action.

In August, several high-profile investors, including the Bill & Melinda Gates Foundation and Google Ventures, pumped US$120 million into the genome-editing firm Editas Medicine of Cambridge, Massachusetts. Big Agriculture is following suit: DuPont forged an alliance with the genome-editing firm Caribou Biosciences of Berkeley, California, in October, and announced its intention to use CRISPR–Cas9 technology to engineer crops.

Eloy Alonso/Reuters/Corbis

Microbiologists Emmanuelle Charpentier and Jennifer Doudna.

7. The CRISPR–Cas9 system is at the centre of a patent row.

Zhang was granted a US patent on CRISPR–Cas9 in April 2014. But several months before he filed his application in 2012, molecular biologists Jennifer Doudna at the University of California (UC), Berkeley, and Emmanuelle Charpentier, now at the Max Planck Institute for Infection Biology in Berlin, had filed their own patent. UC Berkeley has since requested that the United States Patent and Trademark Office determine who should get credit for harnessing the CRISPR–Cas9 system, in particular for its application in human cells. And a similar debate is playing out in Europe, where oppositions to a patent that Zhang and his colleagues won in February have been filed. All three scientists co-founded companies that make use of CRISPR–Cas9.

Changing the production function of diagnostic tests (2)

 The Cas proteins behind CRISPR diagnostics

CRISPR diagnostics explained in few words: 

There are many protein tools in the CRISPR toolbox. Each is suited to a particular suite of uses. For example, the common CRISPR protein Cas9 is well suited for genome editing. It is not suited for CRISPR diagnostics. In this blog post, we’ll introduce you to the proteins behind CRISPR diagnostics: Cas12, Cas13, and Cas14.

 CRISPR diagnostics have two key components:

• Protein-guide molecule complexes. These first cut specific nucleic acid sequences that the user wants to detect. After cutting a user-specified sequence, these complexes non-specifically cut other nucleic acids.
• Modified nucleic acids (reporters). These produce a visual signal when cut. They are only cut if the user-specified nucleic acids are cut first. These modified nucleic acids make it easy to observe when the user-specified nucleic acids have been detected (cut).

CRISPR: from lab to the clinic

 This is the year that CRISPR moves from lab to clinic

Jennifer Doudna says:

In 2021, researchers will use CRISPR to enhance our medical response to the Covid-19 pandemic. Teams will continue to collaborate and bring to market vital CRISPR-based diagnostic tools that are accurate, rapid and painless. One currently being developed and scaled by Mammoth Biosciences, a company I co-founded, along with partners at the University of California, San Francisco and the pharmaceutical company GSK, can detect and indicate the presence of SARS-CoV-2 RNA in a similar fashion to a pregnancy test.

 CRISPR will also have an important effect on the way we treat other diseases. In 2021, we will see increased use of CRISPR-Cas enzymes to underpin a new generation of cost-effective, individualised therapies. With CRISPR enzymes, we can cut DNA at precise locations, using specifically designed proteins, and insert or delete pieces of DNA to correct mutations.

This is precisely what is going on. 

 

The long and bumpy road to CRISPR (2)

 Editing Humanity. The CRISPR Revolution and the New Era of Genome Editing

In 2017 I wrote a post about the book by Jennifer Doudna, A Crack in Creation, now Kevin Davies, the editor of the CRISPR journal has published a new book on CRISPR. It is an effort to put all the information and details about CRISPR in one book. Therefore, if you want to now the whole story (or close to) this is the book to read. If you are interested in a general approach, then the Doudna book is better.

It is quite relevant the chapter that explains the role of Francis Mojica in CRISPR (chapter 3), and the chapter 18, on crossing the germline and what happened about the scandal of genome editing by JK.

“When science moves faster than moral understanding,” Harvard philosopher Michael Sandel wrote in 2004, “men and women struggle to articulate their own unease.” The genomic revolution has induced “a kind of moral vertigo.”49 That unease has been triggered numerous times before and after the genetic engineering revolution—the structure of the double helix, the solution of the genetic code, the recombinant DNA revolution, prenatal genetic diagnosis, embryonic stem cells, and the cloning of Dolly. “Test tube baby” was an epithet in many circles but five million IVF babies are an effective riposte to critics of assisted reproductive technology.

With CRISPR, history is repeating itself,

That's it, great book.

 

CRISPR Diagnostics (for COVID-19)

CRISPR–Cas12-based detection of SARS-CoV-2

Applied technologies for detection of COVID are basically PCR molecular assays and immunoassays. However, CRISPR developments are entering into diagnostics and you may find the first example in Nature.

We report development of a rapid (<40 accurate="" and="" as12-based="" assay="" br="" crispr="" detection="" easy-to-implement="" extracts.="" flow="" for="" from="" lateral="" min="" of="" respiratory="" rna="" sars-cov-2="" swab="">We validated our method using contrived reference samples and clinical samples from patients in the United States, including 36 patients with COVID-19 infection and 42 patients with other viral respiratory infections. Our CRISPR-based DETECTR assay provides a visual and faster alternative to the US Centers for Disease Control and Prevention SARS-CoV-2 real-time RT–PCR assay, with 95% positive predictive agreement and 100% negative predictive agreement.

The role of CRISPR in diagnostics tests is going to increase.

Daido Moriyama 

The long and bumpy road to CRISPR

A Crack in Creation:The New Power to Control Evolution

I've read the same book than Diane Coyle this summer. If you want a clear understanding of what's going on in genomic editing, it should be your first choice. A crack in creation is a description and analysis by Jeniffer Doudna the main researcher on the topic. For those that are excited by genome editingit is good to read this statement:

It’s easy to get caught up in the excitement. The fact that gene editing might be able to reverse the course of a disease—permanently—by targeting its underlying genetic cause is thrilling enough. But even more so is the fact that CRISPR can be retooled to target new sequences of DNA and, hence, new diseases. Given CRISPR’s tremendous potential, I’ve grown accustomed over the past several years to being approached by established pharmaceutical companies asking for my help in learning about the CRISPR technology and about how it might be deployed in the quest for new therapeutics.
But therapeutic gene editing is still in its infancy—indeed, clinical trials have only just begun—and there are still big questions about how things will progress from here. The decades-long struggle to make good on the promise of gene therapy should serve as a reminder that medical advances are almost always more complicated than they might seem. For CRISPR, too, the road leading from the lab to the clinic will be long and bumpy.
Deciding what types of cells to target is one of the many dilemmas confronting researchers—should they edit somatic cells (from the Greek soma, for “body”) or germ cells (from the Latin germen, for “bud” or “sprout”)? The distinction between these two classes of cells cuts to the heart of one of the most heated and vital debates in the world of medicine today.
Germ cells are any cells whose genome can be inherited by subsequent generations, and thus they make up the germline of the organism—the stream of genetic material that is passed from one generation to the next. While eggs and sperm are the most obvious germ cells in humans, the germline also encompasses the progenitors of these mature sex cells as well as stem cells from the very early stages of the developing human embryo.
Somatic cells are virtually all the other cells in an organism: heart, muscle, brain, skin, liver—any cell whose DNA cannot be transmitted to offspring.

Therefore, caution is required and ethical implications are huge as I've said before.
Highly recommended.

Diagnosing diseases and editing genes

CRISPR: The gene-editing tool revolutionizing biomedical research
The WIRED Guide to Crispr
A New Startup Wants to Use Crispr to Diagnose Disease

Take your time and read Wired and watch CBSNews. It will provide an exciting perspective of CRISPR. And just to end, imagine that diagnostic tests could be based on CRISPR while you read the last piece of WIRED.
We are closer than you may think.

CRISPR 2020, a breakthrough year

 2020 Was the Turning Point for CRISPR

At an Oregon hospital in March, a patient with a type of inherited blindness became the first to receive a gene-editing injection directly into their eye. It was the first time CRISPR was used in an attempt to edit a gene inside someone’s body. A second person this year also received the experimental treatment, which is designed to snip out a genetic mutation responsible for their severe visual impairment.

 Despite its versatility, CRISPR is still error-prone. For the past few years, scientists have been working on more precise versions of CRISPR that are potentially safer than the original. This year, they made notable progress in advancing these new versions to human patients.

 

Germline genome editing under scrutiny

Societal and Ethical Impacts of Germline Genome Editing: How Can We Secure Human Rights?

Geneva Statement on Heritable Human Genome Editing: The Need for Course Correction

A CRISPR Moratorium Isn't Enough: We Need a Boycott

The Human Right to Science and the Regulation of Human Germline Engineering

The last frontier in genome editing (if it exists) is germline. The special issue of The Crispr journal on bioethics contains an article of special interest and proposes a third process for evaluating individual and societal harms: a Human Rights Impact Assessment.

Human germline alteration is possible, due in part to democratization of genetic tools required for genome editing, and international scientific and legislative bodies are developing frameworks to manage the ramifications of this technology. Common among these frameworks are two pillars: public engagement and foundational principles. These components are necessary for respecting the autonomy of individuals and for fair processes and respecting diverse values.

However, they are not sufficient for protecting the most vulnerable members of society who may not even be in a position to participate in democratic processes. We propose implementing a HRIA, which captures concerns of public health and offers an opportunity to evaluate and anticipate the societal impact of GGE iteratively as the technology advances, public sentiments evolve, and cultural contexts shift. We recognize that this will raise new challenges of how such assessments are shared and implemented and how they can be enforced. We urge regulatory bodies and policy makers to consider this assessment approach in helping to establish robust regulatory frameworks necessary for the global protection of human rights.

And the Geneva Statement on Heritable Human Genome Editing says:

No decision about whether to pursue heritable human genome modification can be legitimate without broadly inclusive and substantively meaningful public engagement and empowerment. Such deliberations may be challenging and messy. They will take time and organizing them will necessitate creativity, hard work, and significant human and financial resources. The course correction proposed here is essential to these efforts.
We must in the meantime respect the predominant policy position against pursuing heritable human genome modification, if we are to prevent individual scientists or small committees from making this momentous decision for us all. This will preserve time to cultivate an informed and engaged public that can consider and discuss the societal consequences of altering the genes of future generations and make wise, democratic decisions about the shared future we aspire to build. 

I agree.

PS. CRISPR in 2020  Two major reports on germline editing, from the National Academies/Royal Society and the World Health Organization, will be released in 2020. We hope the reports will coordinate, with all the voices of CRISPR being heard, so we can build consensual and broadly acceptable frameworks to ensure we use CRISPR responsibly, especially regarding usage in human embryos for germline editing. The public has asked for it, and the community has been working on it. The science versus society gap will be bridged.

Improving CRISPR, a crowd of proteins

Improving CRISPR from Mammoth Biosciences; 

 Genome editing is the process researchers use to make targeted changes to an organism’s DNA (its genome). Scientists have used a variety of technologies for genome editing (see the history of genome editing here). However, since ~2012, CRISPR has made the genome editing processes much easier. CRISPR associated or “Cas” proteins drive this process. They are relatively easy to target to specific DNA sequences. They also work in many organisms.

Yet, the main Cas protein currently used for CRISPR genome editing, SpCas9, has limitations. In this post, we cover SpCas9’s limitations and how newly discovered Cas protein families, Cas14 and CasΦ, potentially overcome these limitations. We hope Cas14 and CasΦ will enable more efficient genome editing in diverse organisms and tissues.

 

Stop covid with CRISPR Diagnostics

With Crispr, a Possible Quick Test for the Coronavirus

Sherlock's quick, CRISPR-based coronavirus test gets emergency nod

STOP COVID

Point-of-care testing for COVID-19 using SHERLOCK diagnostics

Great!

The FDA granted its first emergency authorization for a CRISPR-based test for COVID-19, developed by Sherlock Biosciences, designed to turn results around in about an hour compared to the four to six hours needed for other molecular diagnostics.
The test is based on the company’s namesake technology, SHERLOCK, short for Specific High-sensitivity Enzymatic Reporter unLOCKing, a Cas13a-based CRISPR system that targets RNA rather than DNA. It looks for an RNA sequence specific to SARS-CoV-2, the virus that causes COVID-19, in patient samples taken from the upper airways with a swab or from airways in the lungs known as bronchoalveolar washing.
“If it’s there, it attaches to the Cas13 enzyme and activates it, which leads to the chewing up and cleaving of RNA probes,” Sherlock CEO Rahul Dhanda told FierceMedTech. When cleaved, those RNA molecules release a fluorescent signal to show the virus is present.

Stop Covid with CRISPR Diagnostics (2)

CRISPR–Cas12-based detection of SARS-CoV-2

Mammoth Biosciences (a firm founded by Jennifer Doudna) has partenered with GSK to commercialise a CRISPR Covid test. Therefore, there are right now two firms in the race: Sherlock and Mammoth.
The paper in Nature explains the details:

Here we report the development and initial validation of a CRISPR–Cas12-based assay9 for detection of SARS-CoV-2 from extracted patient sample RNA, called SARS-CoV-2 DNA Endonuclease-Targeted CRISPR Trans Reporter (DETECTR). This assay performs simultaneous reverse transcription and isothermal amplification using loop-mediated amplification (RT–LAMP)14 for RNA extracted from nasopharyngeal or oropharyngeal swabs in universal transport medium (UTM), followed by Cas12 detection of predefined coronavirus sequences, after which cleavage of a reporter molecule confirms detection of the virus. We first designed primers targeting the E (envelope) and N (nucleoprotein) genes of SARS-CoV-2 (Fig. 1a). The primers amplify regions that overlap the World Health Organization (WHO) assay (E gene region) and US CDC assay (N2 region in the N gene)6,15, but are modified to meet design requirements for LAMP. We did not target the N1 and N3 regions used by the US CDC assay, as these regions lacked suitable protospacer adjacent motif sites for the Cas12 guide RNAs (gRNAs). Next, we designed Cas12 gRNAs to detect three SARS-like coronaviruses (SARS-CoV-2 (accession NC_045512), bat SARS-like coronavirus (bat-SL-CoVZC45, accession MG772933) and SARS-CoV (accession NC_004718)) in the E gene and specifically detect only SARS-CoV-2 in the N gene (Supplementary Fig. 1). This design is similar to those used by the WHO and US CDC assays, which use multiple amplicons with probes that are either specific to SARS-CoV-2 or are capable of identifying related SARS-like coronaviruses.

Edward Hopper 

CRISPR therapeutic success

 Treatment by CRISPR-Cas9 Gene Editing — A Proof of Principle

CRISPR-Cas9 Gene Editing for Sickle Cell Disease and β-Thalassemia

Transfusion-dependent β-thalassemia (TDT) and sickle cell disease (SCD) are severe monogenic diseases with severe and potentially life-threatening manifestations. BCL11A is a transcription factor that represses γ-globin expression and fetal hemoglobin in erythroid cells. We performed electroporation of CD34+ hematopoietic stem and progenitor cells obtained from healthy donors, with CRISPR-Cas9 targeting the BCL11A erythroid-specific enhancer. Approximately 80% of the alleles at this locus were modified, with no evidence of off-target editing. After undergoing myeloablation, two patients — one with TDT and the other with SCD — received autologous CD34+ cells edited with CRISPR-Cas9 targeting the same BCL11A enhancer. More than a year later, both patients had high levels of allelic editing in bone marrow and blood, increases in fetal hemoglobin that were distributed pancellularly, transfusion independence, and (in the patient with SCD) elimination of vaso-occlusive episodes. 

A pivotal moment. Great.

The long and bumpy road to CRISPR (3)

 CRISPR People. The Science and Ethics of Editing Humans

A book on the ethics of CRISPR, without a clear prescription, only a review of He horrendous experiment and a personal view of the current situation.

This has been a cautionary tale about scienceand scientists. People can overreach. He Jiankui, driven as far as I can tell by what Macbeth called “vaulting ambition, which o’erleaps itself,”1 and aided by a serious lack of scruples, behaved terribly. He put three lives at needless and reckless risk (and tried to do the same to more). The story is also a cautionary tale for Science. He Jiankui damaged Science by reinforcing the Victor Frankenstein image—the mad, uncontrolled scientist. Most scientists, and hence most science, are much more rule bound, constrained by the needs of getting and keeping jobs, tenure, and most importantly grants, as well as wrapped, in most countries, in many bureaucratic threads of control. But He happened, and Science needs both to act to minimize the harm he has caused and to be seen to be doing so.

A global consensus is a chimera. Seven and a half billion people are not going to agree on this issue—nor will the fraction of those who understand it. Neither will the roughly 200 countries of the world, at least at anything other than a lowest common denominator. Not all countries have agreed to various nuclear weapons or climate change treaties in spite of the apparently obvious need for them. If somehow something close to unanimity were achieved, it would probably be at the cost of precision. Thus, the Council of Europe enacted a convention that banned human cloning, but effectively left open the then-hot question whether it covered just reproductive cloning or “research cloning” (of human embryos for only ex vivo use) by not defining a “human being.” It also left the implantation and enforcement of such a ban up to the member nations, some of whom were probably happy, for political reasons, to sign it but will have little interest in enforcing it. 

And this is the controversial position of the author:

 One might argue that human germline genome changes are irreversible, or less reversible, than some other interventions. But is that true? A mistaken genome change could presumably be reedited, in a living person or in that person’s germline (or embryos), to reverse the error. Or it could be selected against in the individual’s offspring, through PGD or otherwise. It could be too late to avoid harm to the edited person, but that mistake will not have to pass on from generation to generation. Human germline genome editing does raise important questions about safety, coercion, equity, diversity, and  enhancement. These are not unique to editing the human germline genome. They also apply to somatic cell DNA editing, to new drugs, to smartphones, to climate change, and to many other changes from technologies. Questions about reversibility also apply; the social effects of cell phones are probably less reversible than genome edits. The fact that a technological change is in “the human germline genome” should have no special ethical weight.

Really? Security in CRISPR is not unique???

Genome editing, closer than you think

Human Genome Editing Science, Ethics, and Governance

Last week the US patent office ruled that hotly disputed patents on the CRISPR revolutionary genome-editing technology belong to the Broad Institute of Harvard and MIT. In a former post I explained the dispute. Genome editing in my opinion shouldn't be patented and will see exactly the impact of such ruling in US and elsewhere in the next future.
If you want to know in detail what does genome editing means for the future of life sciences, have a look at NASEM book.

It is now possible to insert or delete single nucleotides,interrupt a gene or genetic element, make a single-stranded break in DNA, modify a nucleotide, or make epigenetic changes to gene expression. In the realm of biomedicine, genome editing could be used for three broad purposes: for basic research, for somatic interventions, and for germline interventions.

CRISPR (which stands for clustered regularly interspaced short palindromic repeats) refers to short, repeated segments of DNA originally discovered in bacteria. These segments provided the foundation for the development of a system that combines short RNA sequences paired with Cas9 (CRISPR associated protein 9, an RNA-directed nuclease), or with similar nucleases, and can readily be programmed to edit specific segments of DNA. The CRISPR/Cas9 genome-editing system offers several advantages over previous strategies for making changes to the genome and has been at the center of much discussion concerning how genome editing could be applied to promote human health.

I would just want to say that these patents destroy the soul of science, since access should be available with no barriers for the development of  innovation. Patents are not the incentive for discovery in this case, as I explained in my post, natural processes should'nt be patented. And this is why today is a really sad day.

PS. My posts against patents

Michael Kiwanuka. Home again

Medicine trends

The future of medicine

A new supplement in Nature explains the main trends in Medicine. It is really helpful to have a quick look focused on those approaches that are the more promising for the next future. From the issue, I would pick one article: A CRISPR edit for heart disease, A one-off injection to reduce the risk of cardiovascular disease is now a prospect thanks to advances in gene editing.This is amazing, it changes current perspectives on the first cause of death worldwide (18 million people per year).

 In 2014, Musunuru and his team showed that more than half of Pcsk9 genes in the mouse liver could be silenced with a single injection of an adenovirus containing a CRISPR–Cas9 system directed against Pcsk9. This led to a roughly 90% decrease in the level of Pcsk9 in the blood and a 35–40% fall in blood LDL cholesterol⁴. Next, they used a mouse engineered to contain human liver cells, and tuned the CRISPR–Cas9 payload to target human PCSK9⁵. The team succeeded in showing that the human gene can also be switched off.

This is changing the focus of drug research, and a recent article explains the new approach.  Let's see if finally delivers what they say.

How to change individual letters of your DNA?

Gene editing has made another step forward. And maybe a complementary to the former one, the CRISPR-Cas9,  that was proved viable by Jennifer Doudna and I explained some months ago in this post. No it is indeed more interesting. Two different approaches, base editing and CRISPR-Cas13, have been described in Science and Nature. Adenine base editing allows to correct mutations, it doesn't cut the gene to insert a new one. It is a sharp pencil rather than scisors. With CRISP-Cas13 it is possible to edit RNA, which converts genetic information into proteins. An exciting approach, you correct a book with temporary ink that disappears, rather than making a permanent mark (like in CRISPR-Cas9).
These are exciting times for genetic research, though we'll have to wait for specific clinical applications

Modigliani, now at Tate Gallery

Outperforming CRISPR?

 High-throughput functional variant screens via in vivo production of single-stranded DNA

A new technoique overcomes existing limitations of CRISPR: 

Unlike existing techniques depending on CRISPR-Cas–directed genomic breaks for genome editing, this strategy instead uses single-stranded DNA produced by a retron element for recombineering. This enables libraries of millions of elements to be constructed and offers relaxed design constraints which permit natural DNA or random variation to be used as inputs.

A group of researchers created what they call the "Retron Library Recombineering" (RLR) technique, which could allow scientists to run millions of genetic experiments at the same time. This tool, described in a recent paper in PNAS, employs retrons, which are bacterial DNA segments that undergo reverse transcription to generate single-stranded DNA fragments (ssDNA). RLR produces up to millions of mutations concurrently in bacterial cells and "barcodes" mutant cells, enabling the whole pool to be screened at once. This way large quantities of data can be quickly produced and analyzed.

Changing the production function of diagnostic tests

Next-generation diagnostics with CRISPR

Last week while reading Science I noticed a short and crucial article. Up to now CRISPR technology was focused on gene editing, now we can say that its usefulness is widening into diagnostics. It may change completely molecular diagnostics of "infectious diseases through detection of Zika virus (ZIKV), Dengue virus (DENV), and human papillomavirus (HPV) in human  samples, and noninfectious diseases, such as detection of gene mutations in circulating cell-free DNA from lung cancer patients." The production founction of lab testing would change completely.
Several articles explain details about it. The fight for patents is going to start again on CRISPR diagnostics. And this are unfortunately bad news.
Anyway, Science article reminds us:

These emerging diagnostic tools will by necessity be compared to standard diagnostics to ensure sensitivity and specificity and will need to be field-tested to guarantee performance in patient care settings, as environmental conditions and end-user application might affect performance. Proven assays, if affordable, promise to improve care in resource-limited settings where undifferentiated febrile illness is the norm and where gaps or delays in diagnosis, targeted care, and infection control contribute to infectious disease mortality and spread.

More details in The Verge.

Beyond CRISPR-Cas9

 Expanding the possibilities of CRISPR genome editing with Cas14 and CasΦ

SpCas9 comes from a bacterium called Streptococcus pyogenes (hence “Sp”). S. pyogenes is a human pathogen. There is some evidence that using SpCas9 for genome editing in humans may lead to dangerous immune reactions (Ferdosi et al 2019) although other reports have questioned the importance of this finding.

In addition SpCas9 is quite large. It is 1368 amino acids (aa) long and can be difficult to fit into standard delivery vehicles (learn about CRISPR delivery here). Thus it can be hard to get SpCas9 into target cells and tissues.

Finally, SpCas9 requires the presence of a specific DNA sequence known as a “PAM” to target an adjacent sequence for genome editing. SpCas9’s PAM is 5’-NGG-3’. The need for this sequence restricts the number of sites SpCas9 can edit. This limits SpCas9’s usefulness.