��˾��ֱ�� researchers enlist tiny biomagnets for faster drug discovery

noreen.rasbach — Mon, 23 Sep 2019 20:16:08 +0000

noreen.rasbach Mon, 09/23/2019 - 16:16

Cutline

Shana Kelley is part of a team of researchers who have developed a new platform that brings together genome editing with magnetic cell sorting to reveal new drug targets for cancer and regenerative medicine (photo by John Loper)

Jovana Drinjakovic

Topic

Breaking Research

Cells

Donnelly Centre for Cellular & Biomolecular Research

CRISPR

Faculty of Applied Science & Engineering

Leslie Dan Faculty of Pharmacy

Medicine by Design

Research & Innovation

What started as a hallway conversation between colleagues is now an “engine for the discovery of new therapeutic targets in cells” thanks to Medicine by Design, says Shana Kelley, a University Professor in the Leslie Dan Faculty of Pharmacy at the University of Toronto.

Kelley’s lab was developing a portable, chip-like device that uses tiny magnets to sort large populations of mixed cell types as part of her Medicine by Design team project. She wondered if the device could be coupled with a CRISPR-based gene-editing technology developed by another Medicine by Design team leader, Jason Moffat, a professor in the Donnelly Centre for Cellular and Biomolecular Research.

They reasoned that the two methods together could speed up combing through the human genome for potential drug targets.

“We casually agreed to combine our technologies – and it worked incredibly well,” says Kelley.

“This is the advantage of being part of the dynamic research ecosystem of Toronto and Medicine by Design,” she says. “I would have never known how to position this technology and link it with CRISPR if I did not have all these great people around to talk to.”

The result of their joint effort, also in collaboration with Stephane Angers, a professor at the Leslie Dan Faculty of Pharmacy, and Ted Sargent, University Professor at the department of electrical and computer engineering, is called MICS, for microfluidic cell sorting, described in a study published Monday in the journal Nature Biomedical Engineering.

MICS will enable researchers to scour the human genome faster when searching for genes and their protein products that can be targeted by drugs.

In one hour, MICS can collect precious rare cells, in which CRISPR has revealed promising drug targets, from a large and mixed cell population. The same experiment would take 20 to 30 hours using the gold standard method of fluorescence-based sorting.

Researchers use CRISPR to switch off each of around 20,000 human genes in cells to see how this affects levels of a disease-related protein which, say, helps cancer spread. This can reveal other gene candidates, and the proteins they encode, that work in the same pathway and that could be targeted with drugs to remove the target protein and halt cancer. The caveat is that genetic screens result in mixed cell populations, with a desired effect present in a vanishingly small proportion of cells, which must be scooped out for further study. Most cell-sorting instruments use laser beams to separate fluorescently labelled cells, but this takes time.

MICS works faster thanks to tiny magnets engineered to bind to the target protein, which leaves the cells sprinkled with magnetic particles. About half the size of a credit card, its surface is streaked with strips of magnetic material that ferry the cells from one end of the device to another. Once at the far end, the cells fall into distinct collection channels based on how many particles they carry, as a proxy for the amount of the target protein.

MICS is about half the size of a credit card, and can sort cells much faster than existing technologies thanks to tiny magnets engineered to bind to the target protein (photo courtesy of Kelley lab)

“As many as one billion cells can travel down this highway of magnetic guides at once and we can process that in one hour,” says Kelley.” It’s a huge game-changer for CRISPR screens.”

To test if MICS can reveal new drug targets, the researchers focused on cancer immunotherapy, in which the immune system is engineered to destroy tumour cells. They looked for a way to reduce the levels of the CD47 protein, which sends a “don’t eat me” signal to the immune system and is often hijacked by cancer cells as a way of escaping immune detection. Others have found that blocking CD47 directly has harmful side effects, prompting the Medicine by Design team to look for the genes that regulate CD47 protein levels.

A genome-wide CRISPR screen revealed a gene called QPCTL, which codes for an enzyme that helps camouflage CD47 from the immune system and could be blocked with an off-the-shelf drug.

“If you can modulate CD47 levels by acting on QPCTL, that could be an interesting way to trick the immune system to clear cancer,” says Moffat.

It’s early days yet, but Kelley and Moffat are hopeful about QPCTL’s therapeutic potential in cancer, perhaps to get macrophages to target tumor cells. They are also launching a multi-lab collaboration PEGASUS project, for Phenotypic Genomic Screening at Scale, which will scale up the technology to interrogate a broad range of therapeutic targets.

On the regenerative medicine front, MICS will help reveal the genes that activate stem cells to turn into specialized cell types, which will make it easier to harvest desired cell types for therapy.

Although Kelley’s team initially developed magnetic cell sorting for isolating tumour cells from the blood, its repurposing for drug target discovery could have a wider impact, with MICS already attracting significant interest from the research community and industry.

��˾��ֱ�� scientists help discover how to turn off CRISPR

ullahnor — Fri, 09 Dec 2016 16:55:21 +0000

��˾��ֱ�� scientists help discover how to turn off CRISPR ullahnor Fri, 12/09/2016 - 11:55

Heidi Singer

Author legacy

Heidi Singer

Subheadline

Researchers are part of an international team that has made gene editing safer and more precise, offering promise for new therapies

CRISPR genome editing is quickly revolutionizing biomedical research, but the new technology is not yet exact. The technique can inadvertently make excessive or unwanted changes in the genome and create off-target mutations, limiting safety and efficacy.

Now, researchers at the University of Toronto and University of Massachusetts Medical School have discovered the first known “off-switches” for CRISPR gene-editing activity, providing greater control and a much-needed “safety valve,” according to a new study featured on the cover of Cell.

The scientists found three proteins that block CRISPR, known as anti-CRISPRs.

��˾��ֱ�� Faculty of Medicine's Alan Davidson, a professor of molecular genetics and biochemistry, and Karen Maxwell, an assistant professor of biochemistry, made the discovery with UMass researcher Erik J. Sontheimer.

“CRISPR is very powerful, but we have to be able to turn it off,” says Davidson. “This is a very fundamental addition to the toolbox which should give researchers more confidence to use gene editing.”

Professor Alan Davidson (left) and Assistant Professor Karen Maxwell are part of a research team that has discovered the first known “off-switches” for CRISPR gene editing

A simple and efficient way of editing the genome, CRISPR is changing biomedical research by making it far easier to inactivate or edit genes in a cell line for study. Work that used to take months or years to perform can now be done in weeks.

Scientists are developing CRISPR to target specific cell types, tissues or organs where a disease occurs. But sometimes, CRISPR hits the wrong target, causing unintended damage.

“CRISPR activity in these other cells, tissues or organs is at best useless and at worst a safety risk,” says Sontheimer. “But if you could build an off-switch that keeps Cas9 (the enzyme that cuts the DNA for editing) inactive everywhere except the intended target tissue, then the tissue specificity will be improved.”

The new paper not only identifies that “off switch” but it shows that CRISPR inhibitors have evolved naturally and can be identified and exploited.

The “off switch” will allow researchers to be more precise in their use of CRISPR. If they only want to use it during one stage of a cell’s life – such as when the DNA is replicating – they can turn it off during all other stages, reducing the chance of unwanted consequences.

A major way of delivering CRISPR into the body is through inactivated viruses that can be programmed to attach themselves to target cells. The challenge is that viruses can’t be engineered to be 100 per cent specific.

Researchers in muscular dystrophy, for example, want to target muscle cells. But a particular virus known for its ability to target muscle cells also attaches itself to liver cells, where it could cause unintended damage. The “off switch” could allow researchers to release “anti-CRISPR” proteins into the body to turn off CRISPR activity in liver cells, offering a new layer of protection against mistakes.

“Knowing we have a safety valve will allow people to develop many more uses for CRISPR,” says Maxwell. “Things that may have been too risky previously might be possible now.”

The “off switch” could be used across the board for any application of CRISPR technology to target specific cells or tissues. For the researchers, the next step is to widen the “off switch” to include other types of CRISPR systems.