Patricia F. Fitzpatrick Dimond Ph.D. Technical Editor of Clinical OMICs President of BioInsight Communications

Scientists believe that if you build it, novel treatments will eventually follow.

What does it take to transform a microbe normally found in the intestinal tract into a cancer-killing machine? These and other questions about the applicability of synthetic biology abound. For cancer research in particular, the field of syn bio could both reveal new insights into the disease and potentially lead to new treatments.

Despite exciting developments in synthetic biology, however, whether the NIH will found and dedicate a specific new program to this field remains to be seen. NCI and the National Institute of General Medical Sciences held a workshop in April 2010 with an eye toward understanding the opportunities for biomedical research in synthetic biology, J. Jerry Li, M.D., Ph.D., Division of Cancer Biology program director, told GEN.

“We invited program managers from other agencies including the National Science Foundation and the Department of Energy. As of 2010, those agencies had dedicated programs to support synthetic biology but NIH as a whole did not,” he remarked. “We asked ourselves whether it was time to put together a synthetic biology centric program.”

NIH held the workshop because recent advances in synthetic biology had created a receptive, anticipatory climate for this new field, Dr. Li added. J. Craig Venter, Ph.D., published work on the first synthetic bacterial genome, and the FDA had just approved Artemisinin, an antimalaria drug produced using engineered bacteria and yeast.

Dr. Li explained that NIH had used the investigator-initiated, nonsolicited R01 programs among its various institutes as the main funding mechanism to support synthetic biology research efforts and continue to “believe it’s the right avenue to get these people supported.”

Signs of Applicability

NCI grants have supported research by J. Christopher Anderson, Ph.D., assistant professor of bioengineering at University of California, Berkeley to develop bacteria as cancer-killing therapeutics. Using bacteria to fight solid tumors is not a new idea. In 2001, writing in Proceedings of the National Academy of Sciences, investigators outlined the requirements for useful bacterial anticancer bombs.

Rakesh K. Jain, Ph.D., and Neil S. Forbes, Ph.D., of Massachusetts General Hospital and Harvard Medical School, said that the anticancer agents should be nontoxic to the host; replicate only within the tumor; be motile and able to disperse evenly throughout a tumor (including hypoxic and necrotic regions); be slowly and completely eliminated from the host; be nonimmunogenic; and be able to cause lysis of tumor cells by direct competition for nutrients, localized production of cytotoxins, or production of therapeutic amplifiers.

As the tools for synthetic biology advanced, in 2010 Dr. Forbes proposed that synthetic biology techniques could be used to “solve many of the key challenges that are associated with bacterial therapies, such as toxicity, stability, and efficiency, and can be used to tune their beneficial features, allowing the engineering of perfect cancer therapies.”

At UC Berkeley, Dr. Anderson along with colleagues at the University of California, San Francisco have been developing such an organism. Dr. Anderson’s laboratory’s emphasis is on advanced DNA assembly, computer-aided manufacture, and therapeutic organisms.

Genetic Devices

While Dr. Anderson’s work is not yet ready for testing in humans, it demonstrates what might be possible with synthetic biology approaches and embodies some very fancy genetic engineering. “Fundamentally the idea was to start with nonpathogenic lab strains of E. coli, then add all of the functionality needed to make a bacterium that can enter the blood stream, invade cancer cells, then kill them,” Dr. Anderson explained to GEN.

“We had this system in mind that has many independent processes inside it that would have to be built by us and then integrated. Some we have successfully built and others not.” The researchers produced their magic microbe by engineering genetic devices, each comprised of multiple genes encoding the instructions for the bacteria to execute specific functions like tumor invasion specifically in response to environmental signals such as oxygen and lactic acid concentrations.

The genes in the devices were borrowed from other microbes and endowed the E. coli with functions it wouldn’t normally possess. For example, the scientists said they used invasin from Y. pseudotuberculosis as an “output module” that could enable the E. coli to invade cancer-derived cells including HeLa, HepG2, and U20S cell lines.

“Basically we showed that you could restrict invasion by monitoring microenvironment cues,” Dr. Anderson said. “We have one gene device that works by environmentally responsive promoters including an oxygen responsive promoter. We have a whole set of these devices that transcriptionally control what the bacteria do with environmental sensors coupled to actuation.” Actuation in this case is what Dr. Anderson calls the “ultimate toxic event,” the expression of RNAase by bacteria in the cancer cells to destroy them.

Key to control of actuation or an event such as the production of an enzyme destructive to a cancer cell is the use of genetic parts to construct a biological version of a near-digital “AND” gate, or two input promoters integrated to activate a single output promoter. An AND gate integrates two input signals into a single output. If both inputs are ON, then the output is ON. If one or both inputs are OFF, then the output is OFF. AND gates are particularly important to program a bacterium to respond to a microenvironment that is not well defined by a single signal (e.g., high lactose and low oxygen).

Expanding the Tool Kit

These microbes are currently being tested in mice, Dr. Anderson said, and he doesn’t disagree that the practical realization of these cancer bomb bacteria may be a ways off. “We have, thus far, made four fully functional devices. This is definitely a project at the leading edge of what you can do with genetic engineering tools today.

“No one has made any system like this. It’s integrating genetic circuits with sensors and actuators, not just the control elements. As soon as you want to actuate something, it’s a whole different ballgame—you are engineering for an environment you have no control over.”

Dr. Anderson noted that his lab engages in three sorts of activities: “out there” applications; “vanilla” foundational projects like new selections for protein engineering; and “weird things.” The projects chosen by his lab, though, all share one thing: they all have existing limitations to the tool kit.

As was also noted at NCI’s syn bio workshop, while a lot of definitions of synthetic biology exist, to truly understand how something works you have to build it. In this sense, organizers said, synthetic biology refers to the design and creation of components of biological systems not found in the natural world as well as the redesign and fabrication of existing biological systems.

As researchers continue to plug away at synthetic biology, the field will no doubt provide some very useful tools for building novel biological entities. Its ultimate clinical practicality and translation into novel therapies remains to be established.

Patricia F. Dimond, Ph.D. ([email protected]), is a principal at BioInsight Consulting.

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