A draft guidance released by the U.S. Food and Drug Administration (FDA) in June 2011 is intended to provide stakeholders in the nanotechnology field with a framework with which they, and the agency, will construct a greater regulatory understanding. Input from nanotechnology stakeholders will be critical to the development of a comprehensive, coherent guidance for industry, and within this announcement is openly encouraged.

The opinions to be received by the FDA in this context will, of course, be varied, and may well range from, “You don’t even have the expertise to do this,” to “The methods are already in place.”

Expressing the latter sentiment is Scott Minick, president and CEO of BIND Biosciences, Cambridge, Mass. “The FDA has been regulating nanoparticles for a long time,” he says, pointing out that Doxil, a liposomal nanoparticle that he helped to develop, was approved in 1995. “In the case of Doxil, and now with our lead product, BIND-014, the FDA is asking the right kinds of questions.” Yes, there are unique manufacturing considerations; and yes, there are unique pharmacologic properties to ponder. But that is what the preclinical and clinical trial processes are for.

The question is whether BIND-014—a targeted polymeric nanoparticle containing the cytotoxic agent docetaxel—is in fact, strikingly new. The Edison Awards, believing that the BIND platform can be used for many types of drugs, honored the technology with a 2011 Science and Medicine Game Changer Award.

That perception of novelty aside, regulatory consideration should not require reinventing the wheel, says Minick; the particle in question is comprised entirely of elements that have been used in humans before, the only difference here is the scale.

Toxicity of scale
Certainly carbon has been used safely in humans before, yet it is widely recognized that carbon nanostructures carry an element of risk, says nanotoxicologist, Peter Wick, PhD, of the Swiss Federal Laboratories for Materials Science and Technology.¹

“The main issue is the ability to cross biological barriers,” he explains. “Nano-sized materials are easily taken up by cells,” which is both good, and possibly bad. The nanoparticle easily gets to its cellular target, but it might not be able to get back out. “Bio-persistence is the problem. If these things are accumulating you could see stress responses, and a sustained response could cause damage.”

The frustration Wick feels is the inability to cut through the uncertainty, largely due to inadequate modeling. “Animal testing is the gold standard so far, and here you assume that a human being is a 70 kg rat. Obviously this is not the case, and so there is only a 70% chance that any toxicity prediction is correct.”

This disconnect is particularly worrisome when considering bio-persistence over long periods of time. “We have no model for long-term exposure,” he says. “Rats live 2 to 3 years and you need information over 10 to 15 years—look at the incubation time of asbestos.”

His advice to the FDA? Ramp up your staffing, and be wary of your results. “Toxicology has always been an interdisciplinary field and nanotoxicology is even more so. People have to know the material, as well as the toxicologic and biologic systems being considered, ” he says. As for the results, Wick has recently completed a study—as yet unpublished—that suggests that even with known standards—a rarity in nanotechnology—testing results can widely vary from lab to lab.

Nanofabrication
Perhaps a facilitator of laboratory standards will be the nano-scale evolution of the so-called, lab on a chip, as driven by recent developments at Nanoink, Skokie, Ill. Nanoink is an innovator in the process of nanolithography.

Launched in 2009, the NLP-2000 System is a desktop nanolithography platform capable of depositing sub-cellular-scale entities under ambient conditions. “We’ve had nanofabrication for years for metal and silicon,” observes Nanoink CEO, James M. Hussey. “But those chips are made in extreme conditions, so they really were not interesting to biology—there was nothing you could do with it.” Yet the drive in biology was to investigate the ever smaller—the cell, the organelle, the protein, the cytokine.

The advent of bio-nanolithography will enable the most nuanced investigations. “Consider that inkjet-based chips deposit in the picoliter range, and we’re working on a 10^-21 liter deposition,” and when bio-fabricating with 20 nm-tipped dip pens the phenomena of scale becomes apparent; rather than propelling a sample to the chip like an inkjet nozzle would, deposition occurs through the aegis of surface tension and viscosity. The result is an extreme accuracy that eliminates the background assay noise observed with inkjets. “When you make things below a micron you can have a tremendously surprising effect,” says Hussey. Data points can number in the thousands on a given slide; the sample required to run an assay may be no more than 2 mL.

There have been other scale-based phenomenon noticed by Hussey that, though unrelated to his product, are the subject of his concern. At the nano-scale, shape matters. As with a protein, conformation is key to activity, he says, and this goes to the point already raised: reference material will have to be generated in order to predict the toxicity of novel physical dimensions for nano-compounds.

Nanofluidics
Nano-scale lithography is also playing a role in creating nanofluidics devices enabling the separation and characterization of DNA. The nanoAnalyzer, developed by BioNanomatrix, San Diego, is an automated, benchtop instrument for high-resolution, multi-color imaging and single-molecule analysis of DNA.

Erik Holmlin, PhD, president and CEO of BioNanomatrix, explains the advantages. “What we’re trying to do is take a picture, an image directly of the genome without breaking it apart.” Typically today, the first step in DNA sequencing is fragmentation of the strand, but as Holmlin points out, by doing that you destroy structural information. “Researchers are desperately interested in understanding the architecture of a genome—how the components are put together. You can’t do that by using so-called next-generation sequencing. Using nanofluidics, the nanoAnalyzer is able to extract data from DNA that has been linearized, but is otherwise intact.”

A typical use of this technology will be sequence finishing. “The genome is very repetitive; we provide a physical measurement that enables researchers to take those small bits of data and assemble them into complete genome sequences,” says Holmlin.

Nano-sensors
Finally, as detailed in the July issue of Nature Nanotechnology, nanotech-sensors can provide information, in vivo, on the day-to-day activities of a single cell.²

Cells carrying sensors monitor the cellular nano- environment in real-time. (Source: Harvard University)

“We immobilized sensors on the surface of cells that can provide insight into cell signaling in real time,” explains Jeffrey Karp, PhD, director of the Laboratory for Advanced Biomaterials and Stem Cell-Based Therapeutics, Harvard-MIT Division of Health Sciences and Technology, Harvard Medical School. Different from other such attempts, this new method conjugates nano-scale reporter constructs to non-specific amine groups on the cell, thus avoiding the problem of a specific receptor or glycan group turnover.

For this investigation, the conjugate was an aptimer—a nano-scale antibody-like molecule—that binds platelet-derived growth factor (PDGF). The aptimer, which contains a fluorophore, changes conformation in the presence of PDGF and thereby produces a signal. “The idea is that we can deliver these sensors to particular microenvironments and then sense changes with high special/temporal resolution,” says Karp.

One application is stem cell transplantation—detecting the fate of a cell, its location and general health, would be a tremendous boon for stem cell science. “If you can take a cell outside the body and functionalize it with a sensor and then deliver that cell, this gives you a unique window to peer in and observe cellular events which were before not possible,” he explains.

Now, given that the material of Karp’s work could be sensationally portrayed as experiments with nanobots and clones, Karp is more than ready to dispense advice to the FDA on how nanotechnology should be regulated. “It’s important to have a process in place,” he says, “Without it, it ends up being chaos because companies that are trying to commercialize the area may waste a lot of time trying to guess what the FDA is looking for.”

As for the current guidance, “The FDA hasn’t said too much here, but it’s a thought-provoking statement to get the discussion started.” And not a moment too soon, as it seems the game is well underway.

About the Author
Neil Canavan is a freelance journalist of science and medicine based in New York.

References
1. Kaiser JP, et al. Carbon nanotubes - curse or blessing. Curr Med Chem. 2011; 18(14):2115-28.
2. Zhao W, et al. Cell-surface sensors for real-time probing of cellular environments. Nat Nanotechnol. 2011; Jul 17. [Epub ahead of print].