Presto! Mutated Gene Libraries

Winter 2014


When medical researchers are designing new biological therapies to treat diseases such as cancer, they often have to test hundreds of thousands of DNA mutations in a gene to pick a single winning combination that produces a desired protein to fight an ailment.

The problem? These gene libraries, which can include a million different mutations for a single DNA strand, are expensive and difficult to produce with a precisely tailored mutational composition. It’s like asking a calligrapher to hand-copy a document 1 million times with a single word change in each version. “Researchers wait months for their libraries,” says Elad Firnberg, a Whiting School postdoc in chemical and biomolecular engineering, who works in the laboratory of Professor Marc Ostermeier.

But that may soon change. A Johns Hopkins team, headed by Firnberg, says it has hit upon a faster, more efficient, and more cost-effective way to create these much-needed gene libraries. Called PFunkel, the method enables researchers to create user-defined gene libraries from a single DNA strand in a single day, in a single test tube. How? Firnberg, working with his mentor Ostermeier, has developed a way to trick the DNA strand into photocopying itself many times, with a single mutated change in each copy. Presto! A mutated gene library in a test tube.

The idea looks so promising that they have filed for U.S. patent protection, and Firnberg and four other graduate students working in Ostermeier’s lab developed a business plan and incorporated as Revolve Biotechnologies Inc. “We didn’t set out to start a company, and it’s all happening pretty fast,” says Firnberg, who defended his PhD thesis on the subject in October.

Revolve is one of two Johns Hopkins startups that have received a $100,000 phase I grant from the Maryland Innovation Initiative (MII) for pre-commercialization work. The group is using the fund to transition into the FastForward Accelerator, a Johns Hopkins-run incubator in the Stieff Silver Building established to help commercialize Johns Hopkins intellectual property.

“We are very excited about the potential,” says Ian Tolfree, associate director of FastForward. One upside: The fledgling company has a market. In preliminary work, seven laboratories nationwide requested mutated gene libraries from the startup. Company officials estimate the global market at roughly $100 mil-lion, and primary customers include research universities and corporations making bioengineered products such as drugs, biofuels, chemicals, and new materials.

“We have the sense that there’s a lot of pent-up demand,” says Barrett Steinberg, who serves as the chief executive officer. “If a drug company is looking for a more heat-resistant protein, or a protein that can chew up a sugar more quickly, we can give them 1 million mutations to test.”

The method generated a flurry of interest after Firnberg and Ostermeier published a 2012 academic paper in PLoS ONE. As a proof of principle of the method, they were able to create a library of nearly all possible 18,081 codon mutations of the TEM-1 ß-lactamase gene in a single day. This library led to the discovery of 18 previously unreported adaptive gene variants that provide resistance against clinically used antibiotics.

Such knowledge allows scientists and clinicians to anticipate and respond to the emergence of antibiotic resistant infections in patients.

In early 2014, the postdocs plan to move to the FastForward Accelerator to focus on commercial applications. Firnberg says this entrepreneurial detour wasn’t expected, but it’s welcome. “It seems like we just fell into it,” he says.