The Future of Nanofabrication and Molecular Scale Devices in Nanomedicine
Research Scientist, Zyvex Corp.
published July 2002
URL: http://www.rfreitas.com/Nano/FutureNanofabNMed.htm
http://www.zyvex.com/Publications/articles/FutureNanofabNMed.html
The original version of this paper was submitted for publication in September 2000 but was not published until July 2002, as Chapter 4 in the book: Renata G. Bushko, ed., Future of Health Technology, IOS Press, Amsterdam, The Netherlands, 2002, pp. 45-59. The abstract is indexed in Medline as: Robert A. Freitas Jr., “The future of nanofabrication and molecular scale devices in nanomedicine,” Studies in Health Technology and Informatics 80(2002):45-59, and this is the correct literature citation for the paper. The document reprinted here differs slightly from the published original.
Nanotechnology is engineering and
manufacturing at the molecular scale, and the application of nanotechnology to
medicine is called nanomedicine.
Nanomedicine subsumes three mutually overlapping and progressively more
powerful molecular technologies. First,
nanoscale-structured materials and devices that can be fabricated today hold
great promise for advanced diagnostics and biosensors, targeted drug delivery
and smart drugs, and immunoisolation therapies. Second, biotechnology offers the benefits of molecular medicine
via genomics, proteomics, and artificial engineered microbes. Third, in the longer term, molecular machine
systems and medical nanorobots will allow instant pathogen diagnosis and
extermination, chromosome replacement and individual cell surgery in vivo, and
the efficient augmentation and improvement of natural physiological
function. Current research is exploring
the fabrication of designed nanostructures, nanoactuators and nanomotors,
microscopic energy sources, and nanocomputers at the molecular scale, along
with the means to assemble them into larger systems, economically and in great
numbers.
1. Nanotechnology and Nanomedicine
“There is a growing sense in the scientific
and technical community that we are about to enter a golden new era,” announced
Richard E. Smalley, winner of the 1996 Nobel Prize in Chemistry, in recent
Congressional testimony [1]. On June
22, 1999, Smalley spoke in support of a new National Nanotechnology Initiative
before the Subcommittee on Basic Research of the U.S. House Science Committee
in Washington, DC. “We are about to be
able to build things that work on the smallest possible length scales, atom by
atom,” Smalley said. “Over the past
century we have learned about the workings of biological nanomachines to an
incredible level of detail, and the benefits of this knowledge are beginning to
be felt in medicine. In coming decades
we will learn to modify and adapt this machinery to extend the quality and
length of life.” Smalley founded the
Center for Nanoscale Science and Technology at Rice University in Texas in
1996. But he became personally interested
in the medical applications of nanotechnology in 1999, after he was diagnosed
with a type of non-Hodgkin’s lymphoma (the same sort that killed King Hussein
of Jordan). Smalley then endured an
apparently successful course of chemotherapy that caused all the hair on his
head to fall out.
“Twenty years ago,” Smalley continued, “without
even this crude chemotherapy I would already be dead. But twenty years from now, I am confident we will no longer have
to use this blunt tool. By then,
nanotechnology will have given us specially engineered drugs which are
nanoscale cancer-seeking missiles, a molecular technology that specifically
targets just the mutant cancer cells in the human body, and leaves everything
else blissfully alone. To do this,
these drug molecules will have to be big enough – thousands of atoms – so that we can code the
information into them of where they should go and what they should kill. They will be examples of an exquisite,
human-made nanotechnology of the future.
I may not live to see it. But,
with your help, I am confident it will happen.
Cancer – at least the type that I have – will be a thing of the past.”
The term “nanotechnology” generally refers to
engineering and manufacturing at the molecular or nanometer length scale. (A nanometer is one-billionth of a meter,
about the width of 6 bonded carbon atoms.)
The field is experiencing an explosion of interest. Nanotechnology is so promising that the U.S.
President, in his January 2000 State-of-the-Union speech, announced that he
would seek $475 million for nanotechnology R&D via the National Nanotechnology
Initiative, effectively doubling federal nanotech funding for FY2001. The President never referred to
“nanotechnology” by name, but he gushed about its capabilities, marveling at a
technology that will someday produce “molecular computers the size of a tear
drop with the power of today’s fastest supercomputers.”
After the President’s speech, Walter
Finkelstein, president and CEO of NanoFab Inc. in Columbia, MD, agreed that it
was conceivable that the technology could be used to develop computers chips so
small that they could be injected into the bloodstream – “Fantastic
Voyage-like,” he said – to locate medical problems. In February 2000, John Hopcroft, dean of the College of
Engineering at Cornell University, announced plans for a new 150,000-square-foot
nanotechnology research center. The
facility already has $12 million per year of earmarked funding and is expected
to support 90 local jobs and approximately 110 graduate students. “The implications of this research are
enormous,” Hopcroft asserted, and include “the development of mechanical
devices that can fight disease within the human body.
In May 2000, the National Cancer Institute
signed an agreement with NASA, the U.S. space agency, to study the medical
potential of nanoparticles. Nanoscience
has also attracted the attention of the U.S. National Institutes of Health
(NIH), which hosted one of the first nanotechnology and biomedicine conferences
in June 2000. In July, the National
Science Foundation (NSF) announced a Nanoscale Science and Engineering
Initiative to provide an estimated $74 million in funding for nanotechnology
research. Northwestern University in
Evanston, Illinois will spend $30 million on a new nanofabrication facility of
its own, joining existing operations such as the Stanford Nanofabrication
Facility (started in 1985 with $15 million of backing from 20 industrial
sponsors) and the Cornell Nanofabrication Facility, expected to attract 450
researchers in 2000, half of them visiting scientists. Cornell is spending $50 million on a new
building for the Facility, and has just won a $20 million, five-year grant from
the NSF to operate a new nanobiotechnology center which will make nanoscale
tools available to biologists.
Burgeoning interest in the medical
applications of nanotechnology has led to the emergence of a new field called
nanomedicine [2, 3]. Most broadly,
nanomedicine is the process of diagnosing, treating, and preventing disease and
traumatic injury, of relieving pain, and of preserving and improving human
health, using molecular tools and molecular knowledge of the human body.
It is most useful to regard the emerging
field of nanomedicine as a set of three mutually overlapping and progressively
more powerful technologies. First, in
the relatively near term, nanomedicine can address many important medical
problems by using nanoscale-structured materials that can be manufactured
today. This includes the interaction of
nanostructured materials with biological systems – in June 2000, the first 12
Ph.D. candidates in “nanobiotechnology” began laboratory work at Cornell
University. Second, over the next 5-10
years, biotechnology will make possible even more remarkable advances in
molecular medicine and biobotics (microbiological robots), some of which are
already on the drawing boards. Third,
in the longer term, perhaps 10-20 years from today, the earliest molecular
machine systems and nanorobots may join the medical armamentarium, finally
giving physicians the most potent tools imaginable to conquer human disease, ill-health,
and suffering.
The initial medical applications of
nanotechnology, using nanostructured materials, are already being tested in a
wide variety of potential diagnostic and therapeutic areas.
For example, fluorescent tags are commonplace
in medicine and biology, found in everything from HIV tests to experiments that
image the inner functions of cells. But
different dye molecules must be used for each color, color-matched lasers are
needed to get each dye to fluoresce, and dye colors tend to bleed together and
fade quickly after one use. “Quantum
dot” nanocrystals have none of these shortcomings. These dots are tiny particles measuring only a few nanometers
across, about the same size as a protein molecule or a short sequence of
DNA. They come in a nearly unlimited
palette of sharply-defined colors, can be excited to fluorescence with white
light, and can be linked to biomolecules to form long-lived sensitive probes to
identify specific compounds. They can
track biological events by simultaneously tagging each biological component
(e.g., different proteins or DNA sequences) with nanodots of a specific color.
Quantum Dot [4], the manufacturer, believes
this kind of flexibility could offer a cheap and easy way to screen a blood
sample for the presence of a number of different viruses at the same time. It could also give physicians a fast
diagnostic tool to detect, say, the presence of a particular set of proteins
that strongly indicates a person is having a heart attack. On the research front, the ability to
simultaneously tag multiple biomolecules both on and inside cells could allow
scientists to watch the complex cellular changes and events associated with
disease, providing valuable clues for the development of future pharmaceuticals
and therapeutics. In mid-2000,
Genentech began evaluating the dots for commercial utility in a variety of
cellular and molecular assays. A
related technology called PEBBLES (Probes Encapsulated by Biologically
Localized Embedding) [5], pioneered by Raoul Kopelman at the University of
Michigan, allows dye-tagged nanoparticles to be inserted into living cells to
monitor metabolism or disease conditions.
2.2 Artificial Molecular Receptors
Another early goal of nanomedicine is to
study how biological molecular receptors work, and then to build artificial
binding sites on a made-to-order basis to achieve specific medical
results. Buddy D. Ratner at the
University of Washington in Seattle has researched the engineering of polymer
surfaces containing arrays of artificial receptors. In a recent series of experiments [6], Ratner and his colleagues
used a new radiofrequency-plasma glow-discharge process to imprint a polysaccharide-like
film with nanometer-sized pits in the shape of such biologically useful protein
molecules as albumin (the most common blood protein), fibrinogen (a clotting
protein), lysozyme and ribonuclease (two important enzymes), and immunoglobulin
(antibodies). Each protein type sticks
only to a pit with the shape of that protein.
Ratner’s engineered surfaces may be used for quick biochemical
separations and assays, and in biosensors and chemosensors, because such
surfaces will selectively adsorb from solution only the specific protein whose
complementary shape has been imprinted, and only at the specific place on the
surface where the shape is imprinted.
The RESIST Group at the Welsh School of Pharmacy at Cardiff University
[7] and others have looked at how molecularly imprinted polymers could be
medically useful in clinical applications such as controlled drug release, drug
monitoring devices, and biological and antibody receptor mimics.
Dendrimers represent yet another
nanostructured material that may soon find its way into medical
therapeutics. Starburst dendrimers are
tree-shaped synthetic molecules with a regular branching structure emanating
outward from a core. Dendrimers form
nanometer by nanometer, so the number of synthetic steps or “generations”
dictates the exact size of the particles in a batch. Each molecule is typically a few nanometers wide but some have
been constructed up to 30 nanometers wide, incorporating more than 100,000
atoms. The peripheral layer of the
dendrimer particle can be made to form a dense field of molecular groups that
serve as hooks for attaching other useful molecules, such as DNA, which hunker
down amongst the outermost branches.
In 1998, James R. Baker Jr. co-founded the
Center for Biologic Nanotechnology at the University of Michigan to bring
together doctors, medical researchers, chemists and engineers to pursue the use
of dendrimers as a safer and more effective genetic therapy agent [8]. For Baker, these nanostructures are
attractive because they can sneak DNA into cells while avoiding triggering an
immune response, unlike viral vectors commonly employed today for
transfection. The dendrimer molecule is
decorated with specific snippets of DNA, then injected into biological tissue. Upon encountering a living cell, dendrimers
of a certain size trigger a process called endocytosis in which the cell’s
outermost membrane deforms into a tiny bubble, or vesicle. The vesicle encloses the dendrimer which is
then admitted into the cell’s interior.
Once inside, the DNA is released and migrates to the nucleus where it
becomes part of the cell’s genome. The
technique has been tested on a variety of mammalian cell types [9], and Baker
hopes to begin clinical human trials of dendrimer gene therapy in 2001. Donald Tomalia, another co-founder of the
Center for Biologic Nanotechnology, recently reported using glycodendrimer
“nanodecoys” to trap and deactivate influenza virus particles [10]. The glycodendrimers present a surface that
mimics the sialic acid groups normally found in the mammalian cell membrane,
causing virus particles to adhere to the outer branches of the decoys instead
of the natural cells.
Medical nanomaterials also may include “smart
drugs” that become medically active only in specific circumstances. A good example is provided by Yoshihisa
Suzuki at Kyoto University, who has designed a novel drug molecule that
releases antibiotic only in the presence of an infection [11]. Suzuki started with the common antibiotic
molecule gentamicin and bound it to a hydrogel using a newly developed peptide
linker. The linker can be cleaved by a
proteinase enzyme manufactured by Pseudomonas aeruginosa, a
Gram-negative bacillus that causes inflammation and urinary tract infection,
folliculitis, and otitis externa in humans. Tests on rats show that when the hydrogel is applied to a wound
site, the antibiotic is not released if no P. aeruginosa bacteria are
present. But if any bacteria of this
type are present, then the proteolytic enzyme that the microbes naturally
produce cleaves the linker and the gentamicin is released, killing the
bacteria. “If the proteinase specific
to each bacterium [species] can be used for the signal,” writes Suzuki,
“different spectra of antibiotics could be released from the same dressing
material, depending on the strain of bacterium.” This specificity of action is highly desirable because the
indiscriminate prophylactic use of antibiotics is associated with the emergence
of strains of drug-resistant bacteria, and most antibiotics apparently have at
least some toxicity for human fibroblasts.
Immunotoxins are another class of smart
drugs, in this case activating only in the presence of cancer cells. An immunotoxin molecule is an engineered
hybrid of functional protein modules fabricated from two different types of
proteins: a toxin and an antibody. Toxin proteins are normally produced and
released by infectious bacteria. The protein
binds to the surface of a host cell, penetrates it, and kills it. Toxin molecules are so potent that just a
few of them can kill a cell. Antibodies
are proteins produced by the immune system to recognize and bind to specific
foreign materials. An immunotoxin
molecule is made by fusing a part of the gene encoding a toxin with a part of
the gene encoding an antibody that recognizes surface features on cancer
cells. This creates a novel gene that
can be used to express a new synthetic protein molecule. This new molecule will bind only to a cancer
cell (via a module from the antibody protein), then penetrate it and kill it
(via modules from the toxin protein).
The first experiments with mice showed that these engineered proteins
successfully eliminated certain tumors.
Then early in 2000, National Cancer Institute researchers confirmed that
an immunotoxin made from a truncated form of Pseudomonas exotoxin was
cytotoxic to malignant B-cells taken from patients with hairy cell leukemia
[12]. A second clinic trial at the
Universitaet zu Koeln in Germany also found that a ricin-based immunotoxin had
moderate efficacy against Hodgkin’s lymphoma in some patients [13].
2.5 Nanopore Immunoisolation Devices
Mauro Ferrari, director of the Biomedical
Engineering Center at Ohio State University and chairman of the BioMEMS
Consortium on Medical Therapeutics, with Tejal Desai has created what could be
considered one of the earliest therapeutically useful nanomedical devices
[14]. Ferrari and his collaborators at
the Biomedical Microdevices Center at the University of California at Berkeley
employed bulk micromachining to fabricate tiny cell-containing chambers within
single crystalline silicon wafers. The
chambers interface with the surrounding biological environment through
polycrystalline silicon filter membranes which are micromachined to present a
high density of uniform nanopores as small as 20 nanometers in diameter. These pores are large enough to allow small
molecules such as oxygen, glucose, and insulin to pass, but are small enough to
impede the passage of much larger immune system molecules such as
immunoglobulins and graft-borne virus particles. Safely ensconced behind this artificial barrier, immunoisolated
encapsulated rat pancreatic cells may receive nutrients and remain healthy for
weeks, happily secreting insulin back out through the pores, while the immune
system remains blissfully unaware of the foreign cells which it would normally
attack and reject.
Ferrari and Desai believe that microcapsules
containing replacement islets of Langerhans cells – most likely
easily-harvested piglet islet cells – could be implanted beneath the skin of
some diabetes patients. This could
temporarily restore the body’s delicate glucose control feedback loop without
the need for powerful immunosuppressants that can leave the patient at serious
risk for infection. Supplying encapsulated
new cells to the body could also be a valuable way to treat other enzyme or
hormone deficiency diseases, including encapsulated neurons which could be
implanted in the brain and then be electrically stimulated to release
neurotransmitters, possibly as part of a future treatment for Alzheimer’s or
Parkinson’s diseases.
2.6 Nanopore Sensors and DNA Sequencing
The flow of materials through nanopores can
also be externally regulated. The first
artificial voltage-gated molecular nanosieve was fabricated by Charles R.
Martin and colleagues [15] at Colorado State University in 1995. Martin’s membrane contains an array of
cylindrical gold nanotubules with inside diameters as small as 1.6 nanometers. When the tubules are positively charged,
positive ions are excluded and only negative ions are transported through the
membrane. When the membrane receives a
negative voltage, only positive ions can pass.
Future similar nanodevices may combine voltage gating with pore size,
shape, and charge constraints to achieve precise control of ion transport with
significant molecular specificity. In
1997, an exquisitely sensitive ion channel switch biosensor was built by an
Australian research group [16]. The
scientists estimated that their sensor could detect a minute change in chemical
concentration equivalent to a single sugar cube tossed into Sidney harbor, or
roughly one part in a billion billion.
Daniel Branton at Harvard University has
conducted an ongoing series of experiments using an electric field to drive a
variety of RNA and DNA polymers through the central nanopore of an
alpha-hemolysin protein channel mounted in a lipid bilayer similar to the outer
membrane of a living cell [17]. As
early as 1996, the researchers had determined that the individual nucleotides
comprising the polynucleotide strands must be passing single-file through the
2.6 nanometer-wide nanopore, and that changes in ionic current could be used to
measure polymer length. By 1998,
Branton had shown that the nanopore could be used to rapidly discriminate
between pyrimidine and purine segments (the two types of nucleotide bases)
along a single RNA molecule. In 2000,
the scientists demonstrated the ability to distinguish between DNA chains of
similar length and composition that differ only in base pair sequence. A similar research effort at the University
of California at Santa Cruz has produced nanopore devices with read rates
potentially up to 1000 bases per second [18].
Because nanopores can rapidly discriminate and characterize DNA polymers
at low copy number, future refinements of this experimental approach may
eventually provide a low-cost high-throughput method for very rapid genome
sequencing.
Biotechnology originally contemplated the
application of biological systems and organisms to technical and industrial
processes, but in recent times the field has expanded to include genetic
engineering and the emerging fields of genomics, proteomics, transcriptomics,
gene chips, artificial chromosomes, and even biobotics. Biotechnology now takes as its ultimate goal
no less than the engineering of all biological systems, even completely
designed organic living systems, using biological instrumentalities or “wet”
nanotechnology. There are many good
summaries of biotechnology elsewhere, so here we focus on efforts to engineer
natural nanomachines to create new cellular devices.
During the 1990s, bioengineered viruses of
various types and certain other vectors routinely were being used in
experimental genetic therapies as “devices” to target and penetrate certain
cell populations, with the objective of inserting therapeutic DNA sequences
into the nuclei of human target cells in vivo.
Retrovirally-altered lymphocytes (T cells) began to be injected into
humans for therapeutic purposes.
Another example was the use, by Neurotech (Paris), of genetically
modified cerebral endothelial cell vectors to attack glioblastoma. This was the first therapeutic use of
genetically engineered endothelial cells in humans; Phase I/II clinical studies were underway in 2000.
Engineered bacteria were also being pursued
by Vion Pharmaceuticals in collaboration with Yale University. In their “Tumor Amplified Protein Expression
Therapy” program [19], antibiotic-sensitive Salmonella typhimurium (food
poisoning) bacteria were attenuated by removing the genes that produce purines
vital to bacterial growth. The tamed
strain could not survive very long in healthy tissue, but quickly multiplied
1000-fold inside tumors which are rich in purines. The engineered bacteria were available in multiple serotypes to
avoid potential immune response in the host, and Phase I human clinical trials
were underway in 2000 using clinical dosages.
The next step would be to add genes to the bacterium to produce anticancer
proteins that can shrink tumors, or to modify the bacteria to deliver various
enzymes, genes, or prodrugs for tumor cell growth regulation.
In 1998, Glen Evans, then at the University
of Texas Southwestern Medical Center, described the possible construction of
synthetic genomes and artificial organisms.
His proposed strategy involved determining or designing the DNA sequence
for the genome, synthesizing and assembling the genome, then introducing the
synthetic DNA into an enucleated pluripotent host cell to create an artificial
organism. Genome engineers could modify
an existing microbe by adding a biochemical pathway borrowed from other
organisms, though this remains a difficult task because tailoring an existing
system to match unique requirements demands detailed knowledge about the
pathway. But ultimately, says Adam P.
Arkin at Lawrence Berkeley National Laboratory, “we want to learn to program
cells the same way we program computers.”
Some genome engineers have started by building the biological equivalent
of the most basic switch in a computer – a digital flip-flop. “Cells switch genes on and off all the
time,” observes MIT’s Thomas F. Knight, Jr., who has pioneered some of this
research. A cellular toggle switch,
made of DNA and some well-characterized regulatory proteins, might be devised
to turn on a specific gene when exposed to a particular chemical. These could be used in gene therapies –
implanted genes might be controlled with single doses of specially selected
drugs, one to switch the gene on, another to switch it off.
Arcady Mushegian of Akkadix Corp. [20] has
looked at the genes present in the genomes of fully sequenced microbes to see
which ones are always conserved in nature.
He concludes that as few as 300 genes are all that may be required for
life, constituting the minimum possible genome for a functional microbe. An organism containing this minimal gene set
would be able to perform the dozen or so functions required for life – manufacturing
cellular biomolecules, generating energy, repairing damage, transporting salts
and other molecules, responding to environmental chemical cues, and
replicating. The minimal microbe – a
basic cellular chassis – could be specified by a genome only 150,000
nucleotides bases in length. Glen
Evans, now at Egea BioSciences, can already produce made-to-order DNA strands
that are 10,000 nucleotide bases in length [21] and is striving to increase
this length by at least a factor of ten.
The engineered full-genome DNA, once synthesized, would then be placed
inside an empty cell membrane – most likely a living cell from which the
nuclear material had been removed.
These artificial biobots could be designed to produce useful vitamins,
hormones, enzymes or cytokines in which the patient’s body was deficient, or to
selectively absorb and metabolize into harmless endproducts harmful substances
such as poisons, toxins, or indigestible intracellular detritus, or even to
perform useful mechanical tasks.
Besides their direct medical applications,
biobots might be employed in molecular construction. Gerald J. Sussman at MIT notes that when computer parts are
reduced to the size of single molecules, engineered microbes could be directed
to lay down complex electronic circuits.
“Bacteria are like little workhorses for nanotechnology; they’re wonderful at manipulating things in
the chemical and ultramicroscopic worlds,” he says. “You could train them to become electricians and plumbers, hire
them with sugar and harness them to build structures for you.”
The
third major branch of nanomedicine – molecular nanotechnology (MNT) or
nanorobotics [2, 22] – takes as its purview the engineering of all complex
mechanical medical systems constructed from the molecular level. Just as biotechnology extends the range and
efficacy of treatment options available from nanomaterials, the advent of
molecular nanotechnology will again expand enormously the effectiveness,
comfort and speed of future medical treatments while at the same time significantly
reducing their risk, cost, and invasiveness.
MNT will allow doctors to perform direct in vivo surgery on individual
human cells.
The
first and most famous scientist to voice these possibilities was the late Nobel
physicist Richard P. Feynman, who worked on the Manhattan Project at Los Alamos
during World War II and later taught at CalTech for most of his professorial
career. In his remarkably prescient
1959 talk “There’s Plenty of Room at the Bottom,” Feynman proposed employing
machine tools to make smaller machine tools, these to be used in turn to make
still smaller machine tools, and so on all the way down to the atomic level
[23]. Feynman prophetically concluded
that this is “a development which I think cannot be avoided.” Such nanomachine tools, nanorobots and
nanodevices could ultimately be used to develop a wide range of atomically
precise microscopic instrumentation and manufacturing tools – that is,
nanotechnology.
Feynman was clearly aware of the potential
medical applications of the new technology he was proposing. After discussing his ideas with a colleague,
Feynman offered [23] the first known proposal for a nanomedical procedure to
cure heart disease: “A friend of mine
(Albert R. Hibbs) suggests a very interesting possibility for relatively small
machines. He says that, although it is
a very wild idea, it would be interesting in surgery if you could swallow the
surgeon. You put the mechanical surgeon
inside the blood vessel and it goes into the heart and looks around. (Of course the information has to be fed
out.) It finds out which valve is the
faulty one and takes a little knife and slices it out. Other small machines might be permanently
incorporated in the body to assist some inadequately functioning organ.” Later in his historic lecture in 1959,
Feynman urged us to consider the possibility, in connection with biological
cells, “that we can manufacture an object that maneuvers at that level!”
Extending nanomedicine to molecular machine
systems will probably require, among many other things, the ability to build
precise structures, actuators and motors that operate at the molecular level,
thus enabling manipulation and locomotion.
For example, in 1992 K. Eric Drexler of the Institute for Molecular Manufacturing
theorized that an efficient nanomechanical bearing could be made by bending two
graphite sheets into cylinders of different diameters, then inserting the
smaller one into the larger one [22].
By 2000, John Cumings and Alex Zettl at U.C. Berkeley had demonstrated
experimentally that nested carbon nanotubes do indeed make exceptionally
low-friction nanobearings [24].
But early mechanical nanorobots might be
made, at least in part, of DNA. The
idea of using DNA to build nanoscale objects has been pioneered by Nadrian
Seeman at New York University [25]. Two
decades ago, Seeman recognized that a strand of DNA has many advantages as a
construction material. First, it is a
relatively stiff polymer. Its
intermolecular interaction with other strands can be readily predicted and
programmed due to the base-pair complementarity of nucleotides, the fundamental
building blocks of genetic material.
DNA also tends to self-assemble.
Arbitrary sequences are readily manufactured using conventional
biotechnological techniques, and DNA is readily manipulated and modified by a
large number of enzymes. During the
1980s, Seeman worked to develop strands of DNA that would zip themselves up into
more and more complex shapes – first tiny squares, then three-dimensional
stick-figure cubes comprised of 480 nucleotides each, then a truncated
octahedron containing 2550 nucleotides.
By the mid-1990s, Seeman could fabricate nanoscale DNA stick figures of
almost any regular geometric shape, by the billions per batch.
In 1999, Seeman reported yet another
breakthrough – the construction of a mechanical DNA-based device that might
serve as the basis for a nanoscale robotic actuator [26]. The mechanism has two rigid double-stranded
DNA arms a few nanometers long that can be made to rotate between fixed
positions by introducing a positively charged cobalt compound into the solution
surrounding the molecules, causing the bridge region to be converted from the
normal B-DNA structure to the unusual Z-DNA structure. The free ends of the arms shift position by
2-6 nanometers during this fully reversible structural conversion, like a hinge
opening and closing. “It’s a very simple
nanomachine,” admits Seeman, “but in the scheme of molecular devices it’s huge
because it generates more than four times the amount of movement produced by
typical molecular devices.” A large
version of the device might function as an elbow, while smaller devices could
serve as finger joints.
Bernard Yurke at Bell Laboratories and Andrew
Turberfield at the University of Oxford synthesized another DNA actuator using
three single strands of artificial DNA which, when placed together, find their
complementary partners and self-assemble to form a V-shaped structure [27]. The open mouth of this nanotweezer can be
made to close by adding a special “fuel” strand which binds to the
single-stranded DNA dangling from the ends of the arms of the tweezers and zips
them closed. A special “removal” strand,
when added, binds to the fuel strand and pulls it away, opening the
nanotweezers again. The cycle may then
be repeated.
In 1999, Philip Kim and Charles Lieber at
Harvard University created the first general-purpose nanotweezer [28]. Its working end is a pair of electrically
controlled carbon nanotubes made from a bundle of multiwalled carbon
nanotubes. To operate the tweezers, a
voltage is applied across the electrodes, causing one nanotube arm to develop a
positive electrostatic charge and the other to develop a negative charge. The
attractive force can be increased or decreased by varying the applied voltage –
8.5 volts completely closes the arms, while lower voltages give different
degrees of grip. Using the tool, Kim
and Lieber have successfully grasped 500-nanometer clusters of polystyrene
spheres, about the same size scale as cellular substructures. They were also able to remove a
semiconductor wire 20 nanometers wide from a mass of entangled wires. At present, each of the tweezer’s arms is
about 50 nanometers wide and 4 microns long.
But by growing single-walled nanotubes directly onto the electrodes, the
researchers hope to produce nanotweezers small enough to grab individual
macromolecules.
Other researchers are developing nanomotors
for future nanorobots. Most notably,
Carlo Montemagno at Cornell University has modified a natural biomotor to
incorporate nonbiological parts, creating the first artificial hybrid nanomotor
[29]. Montemagno started with natural
ATPase, a ubiquitous enzyme found in virtually every living organism and which
helps to convert food into usable energy in living cells. The moving part of an ATPase molecule is a
central protein shaft (or rotor, in electric-motor terms) that rotates in
response to electrochemical reactions with each of the molecule’s three proton
channels (comparable to the electromagnets in the stator coil of an electric
motor). ATP (adenosine triphosphate) is
the fuel that powers the molecular motor’s motion.
Using the tools of genetic engineering,
Montemagno added metal-binding amino acid residues to the ATPase. This allowed each motor molecule to bind
tightly to nanoscale nickel pedestals prepared by electron beam lithography. Properly oriented motor molecules 12
nanometers in diameter were then attached to the pedestals with a precision
approaching 15 nanometers, and a silicon nitride bar a hundred nanometers long
was bound to the rotor subunit of each motor molecule, all by
self-assembly. In a microscopic video
presentation, dozens of bars could be seen spinning like a field of tiny
propellers. The group’s first
integrated molecular motor ran for 40 minutes at 3-4 revolutions per
second. Subsequent motors have been
operated for hours continuously by feeding them plenty of ATP. Montemagno has been measuring things like
horsepower and motor efficiency, simple tests that would be familiar to any
mechanical engineer inspecting a car engine.
Montemagno is also trying to build a solar-powered, biomolecular
motor-driven autonomous nanodevice, wherein light energy is converted into ATP
which then serves as a fuel source for the motor. “We think we’ll be able to make autonomous devices that are
powered by light on a scale of 1 micron or less, smaller than bacteria,” he
says.
Montemagno is developing a chemical means of
switching his hybrid motors on and off reliably. By engineering a secondary binding site tailored to a cell’s
signaling cascade, he plans to use the sensory system of the living cell to
control nanodevices implanted within the cell.
Montemagno envisions tiny chemical factories operating inside living
cells. He speculates that these
nanofactories could be targeted to specific cells, such as those of tumors,
where they would synthesize and deliver chemotherapy agents. Within three years he expects to have a
motor assembled within a living cell, with the cell’s physiology providing the
energy to run it. “My 10-year goal is to make a device that harvests single
molecules within a living cell, maybe a cellular pharmacy that produces a drug,
stores it within the cell, and then based upon some signal, releases it,”
Montemagno said in 2000. “For a
technology that wasn’t expected to produce a useful device before the year
2050, I think we’ve made a pretty good start.
But we have a long way to go before it’s safe to turn these little
machines loose in the human body.”
Nanomotor research is progressing in other
laboratories as well. For instance, a
78-atom chemically-powered rotating motor was synthesized in 1999 as a proof of
principle by chemist T. Ross Kelly at Boston College [30]. Ben Feringa at the University of Groningen
in the Netherlands has built an artificial 58-atom motor molecule that spins
when illuminated by solar energy [31].
Another potential nanorobot power source is a modified microbial fuel
cell – laboratory demonstrations of such cells contain captive bacteria or
immobilized enzymes [32] which, when fed organic material, convert chemical
energy into electricity that could be used to power tiny motors.
Truly effective medical nanorobots may
require onboard computers to allow a physician to properly monitor and control
their work. Molecular electronics or
“moletronics” is a hot research topic in nanotechnology right now. For example, in 2000, a collaborative effort
between UCLA and Hewlett Packard produced the first laboratory demonstration of
completely reversible room-temperature molecular switches that could be
employed in nanoscale memories, using mechanically interlinked ring molecules
called catenanes [33]. Two independent
companies – Molecular Electronics Corp. in Texas and California Molecular
Electronics Corp. in California – have sprung up with the explicit goal of
building the first commercial molecular electronic devices including memories
and other computational components of computers, possibly in the next few
years, using techniques of self-assembly.
As machine structures become more complex,
getting all the parts to spontaneously self-assemble in the right sequence will
be increasingly difficult. To build
such complex structures, it makes more sense to design a mechanism that can
assemble a molecular structure by what is called positional assembly – that is,
picking and placing molecular parts exactly where you want them. A device capable of positional assembly
would work much like the robot arms that manufacture cars on automobile
assembly lines in Detroit, or which insert electronic components onto computer
circuit boards with blinding speed in Silicon Valley. Using the positional assembly approach, the robot manipulator
picks up a part, moves it to the workpiece, installs it, then repeats the
procedure over and over with many different parts until the final product is
fully assembled.
One of the leading proponents of positional
assembly at the molecular scale is Zyvex Corp., a privately-held nanotechnology
research and development corporation headquartered in Richardson, Texas
[34]. Zyvex is the first engineering company
with the explicit goal of creating a molecular assembler that uses positional
assembly to manufacture atomically precise structures. As a first step toward this goal, in 1998
Zyvex demonstrated the ability to use three independently-controlled inch-long
robotic arms to manipulate tiny carbon nanotubes in three dimensions, under the
watchful eye of a scanning electron microscope that can monitor objects and
motions as small as 6 nanometers at near-video scan rates. Zyvex still has a very long way to go before
it can assemble nanoscale parts into useful machines, but its work is a small
step in the right direction and the research continues today. Zyvex engineers are also conceiving and
testing various manufacturing architectures that may someday enable massively
parallel, or exponential, construction of large batches of identical molecular
machines simultaneously. This might
allow vast numbers of nanodevices – ultimately including medical nanorobots –
to be produced relatively inexpensively and to molecular specifications.
4.7 Nanomedical Diagnosis and Treatment
The idea of placing autonomous self-powered
nanorobots inside of us might seem a bit odd, but actually the human body
already teems with such nanodevices.
For instance, more than 40 trillion single-celled microbes swim through
our colon, outnumbering our tissue cells almost ten to one. Many bacteria move by whipping around a tiny
tail, or flagellum, that is driven by a 30-nanometer biological ionic nanomotor
powered by pH differences between the inside and the outside of the bacterial
cell. Our bodies also maintain a
population of more than a trillion motile biological nanodevices called
fibroblasts and white cells such as neutrophils and lymphocytes, each measuring
perhaps 10 microns in size. These
beneficial natural nanorobots are constantly crawling around inside of us,
repairing damaged tissues, attacking invading microbes, and gathering up
foreign particles and transporting them to various organs for disposal from the
body.
The greatest power of nanomedicine will
emerge in a decade or two when we learn to design and construct complete
artificial nanorobots using nanometer-scale parts and subsystems including
sensors, motors, manipulators, power plants, and molecular computers. If we make the reasonable assumption that we
will someday be able to build these complex medical nanorobots, and build them
cheaply enough and in sufficiently large numbers to be useful therapeutically,
then what are the medical implications?
We have space here to describe only a few of the many possibilities [2,
35-39].
One thing that would change dramatically is
clinical diagnostics and treatment.
Consider a patient who goes to his doctor with a mild fever, nasal
congestion, discomfort, and cough. In
the nanomedical era, taking and analyzing microbial samples will be as quick
and convenient as the electronic measurement of body temperature using a
tympanic thermometer in a late 20th-century clinical office or hospital. The physician faces the patient and pulls
from her pocket a lightweight handheld device resembling a pocket
calculator. She unsnaps a
self-sterilizing cordless pencil-sized probe from the side of the device and
inserts the business end of the probe into the patient’s opened mouth in the manner
of a tongue depressor. The ramifying
probe tip contains billions of nanoscale molecular assay receptors mounted on
hundreds of self-guiding retractile stalks.
Each assay receptor is sensitive to the chemical signature of one of
thousands of specific bacterial coats or viral capsids.
The patient says “Ahh,” [35] and a few
seconds later a three-dimensional color-coded map of the throat area appears on
a display panel held in the doctor’s hand.
A bright spot on the screen marks the exact location where the first
samples are being taken. Underneath the
color map scrolls a continuously updated microflora count, listing in the
leftmost column the names of the ten most numerous microbial and viral species
that have been detected, key biochemical marker codes in the middle column, and
measured population counts in the right column. The number counts flip up and down a bit as the physician directs
probe stalks to various locations in the pharynx to obtain a representative
sampling, with special attention to sores or exudate. After a few more seconds, the data for two of the bacterial
species suddenly highlight in red, indicating the distinctive molecular
signatures of specific toxins or pathological variants. One of these two species is a known, and
unwelcome, bacterial pathogen. The
diagnosis is completed and the infectious microbe is promptly exterminated
using a patient-inhaled aerosol of mobile nanorobots which the physician has
programmed to seek out and destroy that one microbial strain. After a few minutes the nanorobots have
finished their work and are retrieved by the doctor. A resurvey with the diagnostic probe reveals no evidence of the
pathogen.
Another major change that nanomedicine will
bring is the ability to dramatically extend natural human capabilities. As a simple example, a few years ago I
designed an artificial mechanical red cell called a “respirocyte” [36]. Still entirely theoretical, the respirocyte
measures 1 micron in diameter and just floats along in the bloodstream. It is a spherical nanorobot made of 18
billion atoms precisely arranged in a diamondoid structure to make a tiny
pressure tank that can be pumped full of up to 3 billion oxygen (O2)
and carbon dioxide (CO2) molecules.
Later on, these gases can be released from the tank in a controlled
manner using tiny molecular pumps.
Gases are stored onboard at pressures up to about 1000 atmospheres.
Respirocytes mimic the action of the natural
hemoglobin-filled red blood cells. Gas
concentration sensors on the outside of each device let the nanorobot know when
it is time to load O2 and unload CO2 (at the lungs), or
vice versa (at the tissues). Each
respirocyte can store and transport 236 times as much gas per unit volume as a
natural red cell. So the injection of a
5 cc therapeutic dose of 50% respirocyte saline suspension, a total of 5
trillion individual nanorobots, into the human bloodstream can exactly replace
the gas carrying capacity of the patient’s entire 5.4 liters of blood. But up to 1 liter of respirocyte suspension
could safely be added to the bloodstream, which could keep a patient’s tissues
safely oxygenated for up to 4 hours in the event a heart attack caused the
heart to stop beating. Or it would
enable a healthy person to sit quietly at the bottom of a swimming pool for
four hours, holding his breath, or to sprint at top speed for at least 15
minutes without breathing.
Similarly, an artificial mechanical platelet
or “clottocyte” [37] could make possible complete hemostasis in just 1 second,
even for moderately large wounds, a response time 100-1000 times faster than
the natural system. The basic
clottocyte is conceived as a serum oxygen/glucose-powered spherical nanorobot,
2 microns in diameter, that contains a compactly-folded fiber mesh. Upon command from its control computer, the
device unfurls its mesh packet in the vicinity of an injured blood vessel –
following, say, a cut through the skin.
Soluble thin films coating certain parts of the mesh dissolve upon
contact with plasma water, revealing sticky sections (e.g., complementary to
blood group antigens unique to red cell surfaces) in desired patterns. Blood cells are immediately trapped in the
overlapping artificial nettings released by multiple neighboring activated clottocytes,
and bleeding halts at once. While up to
300 natural platelets might be broken and still be insufficient to initiate a
self-perpetuating clotting cascade, even a single clottocyte, upon reliably
detecting a blood vessel break, can rapidly communicate this fact to its
neighboring devices [2], immediately triggering a progressive
carefully-controlled mesh-release cascade.
Clottocytes may perform a clotting function that is equivalent in its
essentials to that performed by biological platelets, but at only 0.01% of the
bloodstream concentration of those cells or about 20 nanorobots per cubic
millimeter of serum. Hence clottocytes
appear to be about 10,000 times more effective as clotting agents than an equal
volume of natural platelets.
4.9 Chromosome Replacement Therapy
Medical nanorobots will also be able to
intervene at the cellular level, performing in vivo cytosurgery. The most likely site of pathological
function in the cell is the nucleus – more specifically, the chromosomes. In one simple cytosurgical procedure, a
nanorobot controlled by a physician would extract existing chromosomes from a
diseased cell and insert new ones in their place. This is called chromosome replacement therapy. The replacement chromosomes will be
manufactured to order, outside of the patient’s body in a laboratory benchtop
production device that includes a molecular assembly line, using the patient’s
individual genome as the blueprint. The
replacement chromosomes are appropriately demethylated, thus expressing only
the appropriate exons that are active in the cell type to which the nanorobot
has been targeted. If the patient
chooses, inherited defective genes could be replaced with nondefective
base-pair sequences, permanently curing a genetic disease. Given the speed with which nanorobots can be
administered and their potential rapidity of action, it is possible that an
entire whole-body procedure could be completed in one hour or less. Robert Austin at Princeton University has
also begun early thinking along these lines, hoping someday to design a
nanoprobe capable of identifying biological markers that are specific for
targeted diseases. “Then you just pop
open the cells, remove the bad DNA from that cell, and repair it on a
single-cell level,” he says. “That’s a
long way down the road, but it will happen.”
In the first half of the 21st century,
nanomedicine should eliminate virtually all common diseases of the 20th
century, and virtually all medical pain [38] and suffering as well. Only conditions that involve a permanent
loss of personality and memory information in the brain – such as an advanced
case of Alzheimer’s disease or a massive head trauma – may remain incurable in
the nanomedical era. Because aging is
believed to be the result of a number of interrelated molecular processes and
malfunctions in cells, and because cellular malfunctions will be largely
reversible, middle-aged and older people who gain access to an advanced
nanomedicine can expect to have most of their youthful health and beauty
restored. And they may find few remaining
limits to human longevity in this wonderfully vigorous state. It is a bright future that lies ahead for
medicine, but we shall all have to work very long and very hard to bring it to
fruition.
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––––––––––––––––––––––––––––––––
Robert A. Freitas
Jr. is the author of Nanomedicine,
the first book-length technical discussion of the potential medical applications
of molecular nanotechnology and medical nanorobotics. Volume I was published in October 1999 by Landes
Bioscience while Freitas was a Research Fellow at the Institute for Molecular Manufacturing in Palo Alto,
California. Freitas is completing
Volumes II and III as a Research Scientist at Zyvex
Corp., a nanotechnology R&D startup company headquartered in Richardson,
Texas, and is also consulting on molecular assembler design at Zyvex.
––––––––––––––––––––––––––––––––
Last updated by the author on 19 May 2003