Landmark physical characterization of cancer cells completed

An enormous collaborative effort between a multitude of academic and research centers has characterized numerous physical and mechanical properties on one identical human cancer cell line. Their two-year cooperative study, published online in the April 26, 2013 journal Science Reports, reveals the persistent and agile nature of human cancer cells as compared to noncancerous cells. It also represents a major shift in the way scientific research can be accomplished.

Human breast cancer cells like these were used in the study. (Image created by Shyam Khatau/ Wirtz Lab)

Human breast cancer cells like these were used in the study. (Image created by Shyam Khatau/ Wirtz Lab)

The research, which was conducted by 12 federally funded Physical Sciences-Oncology Centers (PS-OC) sponsored by the National Cancer Institute, is a systematic comparison of metastatic human breast-cancer cells to non-metastatic breast cells that reveals dramatic differences between the two cell lines in their mechanics, migration, oxygen response, protein production and ability to stick to surfaces. They have also discovered new insights into how human cells make the transition from nonmalignant to metastatic, a process that is not well understood.

Denis Wirtz, a Johns Hopkins professor of chemical and biomolecular engineering with joint appointments in pathology and oncology who is the corresponding author on the study, remarked that the work adds a tremendous amount of information about the physical nature of cancer cells. “For the first time ever, scientists got together and have created THE phenotypic signature of cancer” Wirtz said. “Yes, it was just one metastatic cell line, and it will require validation with many other cell lines. But we now have an extremely rich signature containing many parameters that are distinct when looking at metastatic and nonmetastatic cells.”

Wirtz, who directs the Johns Hopkins Physical Sciences-Oncology Center, also noted the unique way in which this work was conducted: all centers used the same human cell line for their studies, which makes the quality of the results unparalleled. And, since human and not animal cells were used, the findings are immediately relevant to the development of drugs for the treatment of human disease.

“Cancer cells may nominally be derived from the same patient, but in actuality they will be quite different because cells drift genetically over just a few passages,” Wirtz said.  “This makes any measurement on them from different labs like comparing apples and oranges.” In this study, however, the genetic integrity of the cell lines were safeguarded by limiting the number times the original cell cultures could be regrown before they were discarded.

The nationwide PS-OC brings together researchers from physics, engineering, computer science, cancer biology and chemistry to solve problems in cancer, said Nastaran Zahir Kuhn, PS-OC program manager at the National Cancer Institute.

“The PS-OC program aims to bring physical sciences tools and perspectives into cancer research,” Kuhn said. “The results of this study demonstrate the utility of such an approach, particularly when studies are conducted in a standardized manner from the beginning.”

For the nationwide project, nearly 100 investigators from 20 institutions and laboratories conducted their experiments using the same two cell lines, reagents and protocols to assure that results could be compared. The experimental methods ranged from physical measurements of how the cells push on surrounding cells to measurements of gene and protein expression.

“Roughly 20 techniques were used to study the cell lines, enabling identification of a number of unique relationships between observations,” Kuhn said.

Wirtz added that it would have been logistically impossible for a single institution to employ all of these different techniques and to measure all of these different parameters on just one identical cell line. That means that this work accomplished in just two years what might have otherwise taken ten, he said.

The Johns Hopkins PS-OC made specific contributions to this work. Using particle-tracking microrheology, in which nanospheres are embedded in the cell’s cytoplasm and random cell movement is visually monitored, they measured the mechanical properties of cancerous versus noncancerous cells. They found that highly metastatic breast cancer cells were mechanically softer and more compliant than cells of less metastatic potential.

Using 3D cell culturing techniques, they analyzed the spontaneous migratory potential (that is, migration without the stimulus of any chemical signal) of cancerous versus noncancerous cells. They also analyzed the extracellular matrix molecules that were deposited by the two cell lines and found that cancerous cells deposited more hyaluronic acid (HA). The HA, in turn, affects motility, polarization and differentiation of cells.  Finally, the Hopkins team measured the level of expression of CD44, a cell surface receptor that recognizes HA, and found that metastatic cells express more CD44.

The next steps, Wirtz said, would be to validate these results using other metastatic cell lines.  To read the paper, which is published in an open access journal, follow this link: http://www.nature.com/srep/2013/130422/srep01449/full/srep01449.html

Excerpts from original press release by Princeton science writer Morgan Kelly were used.

 

 

 

 

Self-assembling drug molecules could fight cancer

A popular method of targeted drug delivery for anti-cancer drugs involves doping another material with the desired pharmaceutical to obtain better targeting efficiency to tumor sites. The problem with this method, researchers have discovered, is that the quantity of drug payload per delivery unit can vary widely and that the materials used for delivery can have toxic side effects.

But what if you could turn the drug molecule itself into a nanoscale delivery system, cutting out the middleman completely?

TEM image of nanotubes formed by self-assembly of an anticancer drug amphiphile. These nanotubes possess a fixed drug loading of 38% (w/w). Image from Cui Lab.

TEM image of nanotubes formed by self-assembly of an anticancer drug amphiphile. These nanotubes possess a fixed drug loading of 38% (w/w). Image from Cui Lab.

Using the process of molecular self-assembly, that is what Honggang Cui, an assistant professor in the Department of Chemical and Biomolecular Engineering at Johns Hopkins University, is attempting to do. His efforts have netted him the prestigious Faculty Early Career Development (CAREER) Award from the National Science Foundation. Cui, an affiliated faculty member of the Johns Hopkins Institute for NanoBioTechnology, will receive the $500,000 award over five years.

Cui explained that a current method of delivering anti-cancer drugs is to enclose them in a nanoscale carrier made of natural or synthetic materials, but this method presents several challenges. “The amount of drug loaded per carrier is very much limited and varies from batch to batch. Even in the same batch, there is a drug loading variation from carrier to carrier. Additionally, the carrier material itself may have toxic side effects,” he said.

Cui’s research seeks to eliminate the need for the carrier by coaxing the drug molecules themselves to form their own carrier through the process of self-assembly. His team is developing new molecular engineering strategies to assemble anti-cancer drugs into supramolecular nanostructures.

“Such supramolecules could carry as much as 100 percent of the drug, would possess a fixed amount of drug per nanostructure and would minimize the potential toxicity of the carrier,” Cui said.

To learn more about research in the Cui lab go to http://www.jhu.edu/cui/

 

Konstantopoulos named BMES fellow

Konstantinos Konstantopoulos (Photo by Mary Spiro)

Konstantinos Konstantopoulos, professor and chair of the Department of Chemical and Biomolecular Engineering at Johns Hopkins University’s Whiting School of Engineering has been named a Fellow of the Biomedical Engineering Society (BMES). Konstantopoulos was one of only nine fellows elected to the Society’s Class of 2012.

BMES states that Konstantopoulos received this honor in recognition of his “seminal bioengineering research contributions involving the discovery and characterization of novel selectin ligands expressed by metastatic tumor cells.”  Formal installation of fellows will take place at the BMES annual meeting  October 24-27 in Atlanta.

Konstantopoulos is an affiliated faculty member of Johns Hopkins Institute for NanoBioTechnology. He is also a project leader with the Johns Hopkins Physical Sciences-Oncology Center. Together with Martin Pomper, a School of Medicine professor of radiology and co-principal investigator of the Johns Hopkins Center of Cancer Nanotechnology Excellence, Konstantopoulos is researching mechanochemical effects on metastasis.

Specifically, his work investigates the effects of fluid mechanical forces at different oxygen tension microenvironments on tumor cell signaling, adhesion and migration. Fluid flow in and around tumor tissue modulates the mechanical microenvironment, including the forces acting on the cell surface and the tethering force on cell-substrate connections. Cells in the interior of a tumor mass experience a lower oxygen tension microenvironment and lower fluid velocities than those at the edges in proximity with a functional blood vessel, and are prompted to produce different biochemical signals. These differential responses affect tumor cell fate that is, whether a cell will live or die, and whether it will be able to detach and migrate to secondary sites in the body.

According to the BMES website, members who demonstrate exceptional achievements and experience in the field of biomedical engineering, as well as a record of membership and participation in the Society, have the opportunity to become fellows. Fellows are selected and conferred  by the BMES board of directors through a highly selective process. Nominations for each of these categories may be made by Society members and the board of directors.

Learn more about research in the Konstantopoulos Lab here.

 

 

Coated nanoparticles move easily into brain tissue

Real-time imaging of nanoparticles green) coated with polyethylene-glycol (PEG), a hydrophilic, non-toxic polymer, penetrate within normal rodent brain. Without the PEG coating, negatively charged, hydrophobic particles (red) of a similar size do not penetrate. Image by Elizabeth Nance, Kurt Sailor, Graeme Woodworth.

Johns Hopkins researchers report they are one step closer to having a drug-delivery system flexible enough to overcome some key challenges posed by brain cancer and perhaps other maladies affecting that organ. In a report published online Aug. 29 in Science Translational Medicine, the Johns Hopkins team says its bioengineers have designed nanoparticles that can safely and predictably infiltrate deep into the brain when tested in rodent and human tissue.

“We are pleased to have found a way to prevent drug-embedded particles from sticking to their surroundings so that they can spread once they are in the brain,” said Justin Hanes, Lewis J. Ort Professor of Ophthalmology and project leader in the Johns Hopkins Center of Cancer Nanotechnology Excellence.

Standard protocols following the removal of brain tumors include chemotherapy directly applied to the surgical site to kill any cancer cells left behind. This method, however, is only partially effective because it is hard to administer a dose of chemotherapy high enough to sufficiently penetrate the tissue to be effective and low enough to be safe for the patient and healthy tissue. Furthermore, previous versions of drug-loaded nanoparticles typically adhere to the surgical site and do not penetrate into the tissue.

These newly engineered nanoparticles overcome this challenge. Elizabeth Nance, a graduate student in chemical and biomolecular engineering, and Johns Hopkins neurosurgeon Graeme Woodworth, suspected that drug penetration might be improved if drug-delivery nanoparticles interacted minimally with their surroundings. Nance achieved this by coating nano-scale beads with a dense layer of PEG or poly(ethylene glycol). The team then injected the coated beads, which had been marked with a fluorescent tag,  into slices of rodent and human brain tissue. They found that a dense coating of PEG allowed larger beads to penetrate the tissue, even those beads that were nearly twice the size previously thought to be the maximum possible for penetration within the brain. They then tested these beads in live rodent brains and found the same results.

Elizabeth Nance. Photo by Ming Yang.

The results were similar when biodegradable nanoparticles carrying the chemotherapy drug paclitaxel and coated with PEG were used. “It’s really exciting that we now have particles that can carry five times more drug, release it for three times as long and penetrate farther into the brain than before,” said Nance. “The next step is to see if we can slow tumor growth or recurrence in rodents.”

Woodworth added that the team “also wants to optimize the particles and pair them with drugs to treat other brain diseases, like multiple sclerosis, stroke, traumatic brain injury, Alzheimer’s and Parkinson’s.” Another goal for the team is to be able to administer their nanoparticles intravenously, which is research they have already begun.

Additional authors on the paper include Kurt Sailor, Ting-Yu Shih, Qingguo Xu, Ganesh Swaminathan, Dennis Xiang, and Charles Eberhart, all from The Johns Hopkins University.

Story adapted from an original press release by Cathy Kolf.

 

Additional news coverage of this research can be found at the following links:

Nanotechnology/Bio & Medicine

Death and Taxes Mag

New Scientist Health

Nanotech Web

Portugese news release (in Portugese)

German Public Radio (in German)

Light-activated synthetic protein illuminates disease destruction

Illustration of collagen’s rope-like structure. Click to watch video. (INBT Animation Studios)

Johns Hopkins researchers have created a synthetic protein that, when activated by ultraviolet light, can guide doctors to places within the body where cancer, arthritis and other serious medical disorders can be detected. The synthetic protein does not zero in directly on the diseased cells. Instead, it binds to nearby collagen that has been degraded by disease or injury.

“These disease cells are like burglars who break into a house and do lots of damage but who are not there when the police arrive,” said S. Michael Yu, a faculty member in the Whiting School of Engineering’s Department of Materials Science and Engineering. “Instead of looking for the burglars, our synthetic protein is reacting to evidence left at the scene of the crime,” said Yu, who was principal investigator in the study.

The technique could lead to a new type of diagnostic imaging technology and may someday serve as a way to move medications to parts of the body where signs of disease have been found. In a study published in the Aug. 27-31 Online Early Edition of Proceedings of the National Academy of Sciences, the researchers reported success in using the synthetic protein in mouse models to locate prostate and pancreatic cancers, as well as to detect abnormal bone growth activity associated with Marfan syndrome.

Collagen, the body’s most abundant protein, provides structure and creates a sturdy framework upon which cells build nerves, bone and skin. Some buildup and degradation of collagen is normal, but disease cells such as cancer can send out enzymes that break down collagen at an accelerated pace. It is this excessive damage, caused by disease, that the new synthetic protein can detect, the researchers said.

A key collaborator was Martin Pomper, a School of Medicine professor of radiology and co-principal investigator of the Johns Hopkins Center of Cancer Nanotechnology Excellence. Pomper and Yu met as fellow affiliates of the Johns Hopkins Institute for NanoBioTechnology. “A major unmet medical need is for a better non-invasive characterization of disrupted collagen, which occurs in a wide variety of disorders,” Pomper said. “Michael has found what could be a very elegant and practical solution, which we are converting into a suite of imaging and potential agents for diagnosis and treatment.”

The synthetic proteins used in the study are called collagen mimetic peptides or CMPs. These tiny bits of protein are attracted to and physically bind to degraded strands of collagen, particularly those damaged by disease. Fluorescent tags are placed on each CMP so that it will show up when doctors scan tissue with fluorescent imaging equipment. The glowing areas indicate the location of damaged collagen that is likely to be associated with disease.

In developing the technique, the researchers faced a challenge because CMPs tend to bind with one another and form their own structures, similar to DNA, in a way that would cause them to ignore the disease-linked collagen targeted by the researchers.

To remedy this, the study’s lead author, Yang Li, synthesized CMPs that possess a chemical “cage” to keep the proteins from binding with one another. Just prior to entering the bloodstream to search for damaged collagen, a powerful ultraviolet light is used to “unlock” the cage and allow the CMPs to initiate their disease-tracking mission. Li is a doctoral student from the Department of Chemistry in the Krieger School of Arts and Sciences at Johns Hopkins. Yu, who holds a joint appointment in that department, is his doctoral adviser.

Yu’s team tested Li’s fluorescently tagged and caged peptides by injecting them into lab mice that possessed both prostate and pancreatic human cancer cells. Through a series of fluorescent images taken over four days, researchers tracked single strands of the synthetic protein spreading throughout the tumor sites via blood vessels and binding to collagen that had been damaged by cancer.

Similar in vivo tests showed that the CMP can target bones and cartilage that contain large amounts of degraded collagen. Therefore, the new protein could be used for diagnosis and treatment related to bone and cartilage damage.

Although the process is not well understood, the breakdown and rebuilding of collagen is thought to play a role in the excessive bone growth found in patients with Marfan syndrome. Yu’s team tested their CMPs on a mouse model for this disease and saw increased CMP binding in the ribs and spines of the Marfan mice, as compared to the control mice.

Funding for the research was provided by the National Science Foundation, the National Institutes of Health and the Department of Defense. The synthetic protein process used in this research is protected by patents obtained through the Johns Hopkins Technology Transfer Office.

Along with Yu, Li and Pomper, co-authors of this study were instructor Catherine A. Foss and medical resident Collin M. Torok from the Department of Radiology and Radiological Science at the Johns Hopkins School of Medicine; Harry C. Dietz, a professor, and Jefferson J. Doyle, a doctoral student, both of the Howard Hughes Medical Institute and Institute of Genetic Medicine at the School of Medicine; and Daniel D. Summerfield a former master’s student in the Department of Materials Science and Engineering.

Adapted from an original press release by Phil Sneiderman.

 

Light-activated synthetic protein illuminates disease destruction

Illustration of collagen’s rope-like structure. Click to watch video. (INBT Animation Studios)

Johns Hopkins researchers have created a synthetic protein that, when activated by ultraviolet light, can guide doctors to places within the body where cancer, arthritis and other serious medical disorders can be detected. The synthetic protein does not zero in directly on the diseased cells. Instead, it binds to nearby collagen that has been degraded by disease or injury.

“These disease cells are like burglars who break into a house and do lots of damage but who are not there when the police arrive,” said S. Michael Yu, a faculty member in the Whiting School of Engineering’s Department of Materials Science and Engineering. “Instead of looking for the burglars, our synthetic protein is reacting to evidence left at the scene of the crime,” said Yu, who was principal investigator in the study.

The technique could lead to a new type of diagnostic imaging technology and may someday serve as a way to move medications to parts of the body where signs of disease have been found. In a study published in the Aug. 27-31 Online Early Edition of Proceedings of the National Academy of Sciences, the researchers reported success in using the synthetic protein in mouse models to locate prostate and pancreatic cancers, as well as to detect abnormal bone growth activity associated with Marfan syndrome.

Collagen, the body’s most abundant protein, provides structure and creates a sturdy framework upon which cells build nerves, bone and skin. Some buildup and degradation of collagen is normal, but disease cells such as cancer can send out enzymes that break down collagen at an accelerated pace. It is this excessive damage, caused by disease, that the new synthetic protein can detect, the researchers said.

A key collaborator was Martin Pomper, a School of Medicine professor of radiology and co-principal investigator of the Johns Hopkins Center of Cancer Nanotechnology Excellence. Pomper and Yu met as fellow affiliates of the Johns Hopkins Institute for NanoBioTechnology. “A major unmet medical need is for a better non-invasive characterization of disrupted collagen, which occurs in a wide variety of disorders,” Pomper said. “Michael has found what could be a very elegant and practical solution, which we are converting into a suite of imaging and potential agents for diagnosis and treatment.”

The synthetic proteins used in the study are called collagen mimetic peptides or CMPs. These tiny bits of protein are attracted to and physically bind to degraded strands of collagen, particularly those damaged by disease. Fluorescent tags are placed on each CMP so that it will show up when doctors scan tissue with fluorescent imaging equipment. The glowing areas indicate the location of damaged collagen that is likely to be associated with disease.

In developing the technique, the researchers faced a challenge because CMPs tend to bind with one another and form their own structures, similar to DNA, in a way that would cause them to ignore the disease-linked collagen targeted by the researchers.

To remedy this, the study’s lead author, Yang Li, synthesized CMPs that possess a chemical “cage” to keep the proteins from binding with one another. Just prior to entering the bloodstream to search for damaged collagen, a powerful ultraviolet light is used to “unlock” the cage and allow the CMPs to initiate their disease-tracking mission. Li is a doctoral student from the Department of Chemistry in the Krieger School of Arts and Sciences at Johns Hopkins. Yu, who holds a joint appointment in that department, is his doctoral adviser.

Yu’s team tested Li’s fluorescently tagged and caged peptides by injecting them into lab mice that possessed both prostate and pancreatic human cancer cells. Through a series of fluorescent images taken over four days, researchers tracked single strands of the synthetic protein spreading throughout the tumor sites via blood vessels and binding to collagen that had been damaged by cancer.

Similar in vivo tests showed that the CMP can target bones and cartilage that contain large amounts of degraded collagen. Therefore, the new protein could be used for diagnosis and treatment related to bone and cartilage damage.

Although the process is not well understood, the breakdown and rebuilding of collagen is thought to play a role in the excessive bone growth found in patients with Marfan syndrome. Yu’s team tested their CMPs on a mouse model for this disease and saw increased CMP binding in the ribs and spines of the Marfan mice, as compared to the control mice.

Funding for the research was provided by the National Science Foundation, the National Institutes of Health and the Department of Defense. The synthetic protein process used in this research is protected by patents obtained through the Johns Hopkins Technology Transfer Office.

Along with Yu, Li and Pomper, co-authors of this study were instructor Catherine A. Foss and medical resident Collin M. Torok from the Department of Radiology and Radiological Science at the Johns Hopkins School of Medicine; Harry C. Dietz, a professor, and Jefferson J. Doyle, a doctoral student, both of the Howard Hughes Medical Institute and Institute of Genetic Medicine at the School of Medicine; and Daniel D. Summerfield a former master’s student in the Department of Materials Science and Engineering.

Adapted from an original press release by Phil Sneiderman.

 

INBT director wins “Provocative Questions” grant

Peter Searson (Photo by Marty Katz)

Peter Searson,  director of Johns Hopkins Institute for NanoBioTechnology, has received a “Provocative Questions” grant from the National Cancer Institute. Among the researchers nationwide who received this award, Searson is one of two at Johns Hopkins University.  Searson is the Joseph R. and Lynn C. Reynolds Professor in the Whiting School of Engineering. He is a professor of materials science and engineering and also co-director of the Johns Hopkins Center of Cancer Nanotechnology Excellence.

A press release issued by Johns Hopkins states:

Two Johns Hopkins scientists are among the first recipients of grants geared to answer “Provocative Questions” in cancer research, a new project funded by the National Cancer Institute (NCI), part of the National Institutes of Health (NIH). Cynthia Sears, M.D., and Peter Searson, Ph.D., will receive more than $500,000 combined in the first of five years of funding.

The federally funded “Provocative Questions” project will distribute more than $22 million this year among 57 grant recipients nationwide. Each grant aims to provide answers to one of 24 questions, solicited from the research community, that address neglected or unsolved areas of cancer research.

Sears and Searson’s projects were chosen by a specially-designed peer-review process from among more than 700 applications submitted by scientists.

Sears will receive $236,480 in the first year of funding to examine how and why certain cancers may be caused by infections. An infectious disease expert, Sears has previously linked some colon cancers to common diarrheal bacteria, and her research will further explore how these and other microbes may cause colon cancer. Sears, a professor of medicine and oncology at Johns Hopkins, says these discoveries may help physicians detect earlier those at risk for colon cancer and prevent it more effectively.

With $316,513 in first-year funding, Searson will develop a new method to study how cancer spreads, also known as “metastasis.” He proposes a device with an artificial blood vessel and other cellular components to study the spread of cancer. The “lab-on-a-chip” approach could replicate processes that occur in metastasis and provide a testing ground for experiments with a method easily reproduced by scientists. Searson is the Joseph R. and Lynn C. Reynolds Professor of Materials Science and Engineering and directs the Johns Hopkins Institute for NanoBioTechnology.

Sears’ grant is titled “Microbial Induction of Colon Cancer and Mechanisms” (NIH, NCI grant #R21CA170492-01), and Searson’s is “An engineered platform for the study of metastasis” (NIH, NCI grant #1R01CA170629-01)

Original press release appears here.

Killing prostate cancer cells with out harming the healthy cells

Experimenting with human prostate cancer cells and mice, cancer imaging experts at Johns Hopkins say they have developed a method for finding and killing malignant cells while sparing healthy ones.

The method, called “theranostic” imaging, targets and tracks potent drug therapies directly and only to cancer cells. It relies on binding an originally inactive form of drug chemotherapy, with an enzyme, to specific proteins on tumor cell surfaces and detecting the drug’s absorption into the tumor. The binding of the highly specific drug-protein complex, or nanoplex, to the cell surface allows it to get inside the cancerous cell, where the enzyme slowly activates the tumor-killing drug.

Researchers say their findings, published in the journal American Chemical Society Nano online Aug. 6, are believed to be the first to show that chemotherapies can be precisely controlled at the molecular level to maximize their effectiveness against tumors, while also minimizing their side effects.

Senior study investigator Zaver Bhujwalla, Ph.D., a professor at the Johns Hopkins University School of Medicine and its Kimmel Cancer Center, notes that a persistent problem with current chemotherapy is that it attacks all kinds of cells and tissues, not just cancerous ones.

In the theranostic imaging experiments, overseen by Bhujwalla and study co-investigator Martin Pomper, M.D., Ph.D., investigators directed drugs only to cancer cells, specifically those with prostate-specific membrane antigen, or PSMA cell surface proteins. Both Pomper and Bhujwalla are affiliated faculty members of Johns Hopkins Institute for NanoBioTechnology.

“Our results show a non-invasive imaging approach to following and delivering targeted therapy to any cancer that expresses PSMA,” says Bhujwalla, who also serves as director of the Johns Hopkins In Vivo Cellular and Molecular Imaging Center (ICMIC), where the theranostic imaging studies were developed.

Bhujwalla says the new technique potentially will work against any cancer in which tumors elevate production of certain cell surface proteins. Examples would include breast cancers with HER-2/neu and CXCR4 proteins, and some liver, lung and kidney cancers also known to express particular proteins. She notes that PSMA is expressed in the vessels of most solid tumors, suggesting that the nanoplex reported in the latest study could be used in general to image and treat a variety of cancers.

In their latest series of experiments, primarily in mice injected with human prostate tumor cells, Bhujwalla and the Johns Hopkins team tested their ability to track with imaging devices the delivery of anti-cancer drugs directly to tumors. Some of the tumors comprised cells with PSMA, while other so-called control tumors had none. Included in the drug nanoplex were small strands of RNA, cell construction acids that can be used instead to block and turn down production of a well-known enzyme, choline kinase, whose levels usually rise with tumor growth. All nanoplex components were imaged inside the tumor, in addition to dropping choline kinase production, which decreased by 80 percent within 48 hours of nanoplex absorption into cells with ample PSMA. When researchers used antibodies to block the action of PSMA, down went the level of nanoplex uptake and drug activation in cancerous cells as measured by dimming of the image.

Different concentrations of the drug nanoplex, tagged with radioactive and fluorescent molecules, were mixed in the lab with prostate cancer tissue cells, some of which had extra PSMA and others which had none. Only those cells with extra PSMA showed nanoplex uptake, as measured by image intensity, which later decreased when PSMA-blocking chemicals were added (back to levels seen in cells with almost no PSMA).

Additional experiments involving injections of three different concentrations of the drug nanoplex showed no damage to other vital mouse organs, such as the kidney and liver, nor any uptick in the mouse immune system response.

“Our theranostic imaging approach shows how the best methods of detection and treatment can be combined to form highly specialized, more potent and safer forms of chemotherapy,” says Pomper, a professor at Johns Hopkins who also serves as an associate director at ICMIC.

He says that an important goal for theranostic imaging is to move it beyond standard chemotherapy that attacks one target molecule at a time.

“With theranostic imaging, we can attack multiple tumor targets, making it harder for the tumor to evade drug treatment,” says Pomper, who is already working with colleagues at Johns Hopkins to identify other molecular targets.

The most recent studies were performed at Johns Hopkins over two years, starting in 2010, with funding support from the National Cancer Institute, part of the National Institutes of Health. The corresponding grant numbers are P50-CA103175, RO1-CA138515 and RO1-CA134675.

In addition to Bhujwalla and Pomper, other Johns Hopkins researchers from the Russell H. Morgan Department of Radiology involved in this imaging study were lead investigators Zhihang Chen, Ph.D., and Mary-France Penet, Ph.D. Additional study co-investigators were Sridhar Nimmagadda, Ph.D.; Li Cong, Ph.D.; Sangeeta Banerjee, Ph.D.; Paul Winnard Jr., Ph.D.; Dmitri Artemov, Ph.D.; and Kristine Glunde, Ph.D.

Press release by David March

Therapeutic nanolex containing multi-modal imaging reporters targeted to prostate specific membrane antigen (PSMA), which is expressed on the cell surface of castrate resistant PCa. (Image by Chen et al. from ACS Nano 2012).

 

Hanes discusses nanoparticle-based drug delivery

Justin Hanes was among the invited speakers at the 2012 NanoBio Symposium.

“Would you poison the entire garden to kill one weed?” asked Justin Hanes at the opening of his talk at the 2012 Johns Hopkins annual NanoBio Symposium. “Unfortunately, that is how most chemotherapy works today.” Hanes is a professor of ophthalmology at Johns Hopkins School of Medicine and an affiliated faculty member of the Institute for NanoBioTechnology.

On average, less than one percent of any chemotherapy cancer treatment will go to a patient’s tumor. The remaining 99 percent circulates through the rest of the patient’s body, kills healthy cells unnecessarily, and causes often unbearable side effects. This alarming statistic has led Hanes and his team to focus on targeted, chemotherapeutic drug delivery using nanoparticles.

Hanes explained that nanoparticles are ideal in cancer treatment because tumors form new blood vessels within themselves to be able to receive nutrients, and these tumor-associated blood vessels are leaky. Thin, leaky blood vessel walls are ideal for drug-loaded nanoparticles, which are on the order of 1-100 nanometers in diameter, to break through to reach tumor cells. The ultimate goal of nanoparticle drug delivery for cancer is to synthesize bio-targeted particles that provide localized delivery straight to the tumor alone, improving drug effectiveness and reducing undesirable side effects.

Many members of the Hanes lab focus on drug delivery in mucus, which exists in the lining of the lungs, vaginal tract, intestines, and many other organs. Although mucus is essential to prevent viruses and bacteria from entering tissues, its sticky consistency also acts as a barrier to drug-loaded nanoparticles. Hanes’s students are synthesizing nanoparticles that can pass through mucosal barriers.

Scientists previously believed that pores within mucus were around 25 nm in size and that nanoparticles would not be able to pass through. However, the Hanes lab’s work shows that these pores may be closer to 400 nm and that particles with a diameter of 500 nm coated with a simple, biodegradable polymer called PEG can pass through. The PEG gives the particles a neutral charge and makes them hydrophilic, or attracted to water, so that they can pass easily through the mucosal meshwork without getting trapped.

The group is now able to encapsulate chemotherapeutic drugs in mucus-penetrating nanoparticles for a variety of different applications, including lung cancer and ovarian cancer. Additionally, Laura Ensign, an INBT-sponsored graduate student in the Hanes lab, showed in a recently-published journal article that her mucus-penetrating particles effectively delivered drugs in the mouse vaginal tract for longer times than previously reported (>24 h). This work could be applied to cervical cancer, where drug-loaded nanoparticles could be administered to travel through the mucosal lining of the reproductive tract for successful treatment.

Overall, the Hanes lab anticipates that their research will contribute to the effectiveness of many types of cancer treatments. Read more about Ensign’s recently published work here.

Story by Allison Chambliss, a Ph.D. student in the Department of Chemical and Biomolecular Engineering with interests in cellular biophysics and epigenetics.

 

Nanoparticles slip through mucus barrier to protect against herpes virus

“Thick, sticky mucus layers limit effectiveness of drug delivery to mucosal tissues. Mucus-penetrating particles or MPPs (in red) are able to penetrate mucus, covering the entire surface of the mouse vagina (in blue). Improved distribution and retention of MPPs led to significantly increased protection in a mouse model for herpes simplex virus infection. Image by Laura Ensign.

Johns Hopkins researchers say they have demonstrated for the first time, in animals, that nanoparticles can slip through mucus to deliver drugs directly to tissue surfaces in need of protection.

The researchers used these mucus-penetrating particles, or MPPs, to protect against vaginal herpes infections in mice. The goal is to create similar MPPs to deliver drugs that protect humans against sexually transmitted diseases or even treat cancer.

“This is the first in vivo proof that MPPs can improve distribution, retention, and protection by a drug applied to a mucosal surface, said Justin Hanes, Ph.D., a professor of ophthalmology at the Johns Hopkins Wilmer Eye Institute and director of the Center for Nanomedicine at the Johns Hopkins University School of Medicine.

Hanes also is a principal investigator with the Johns Hopkins Center of Cancer Nanotechnology Excellence. Results of his team’s experiments are described in the June 13 issue of the journal Science Translational Medicine.

The moist mucosal surfaces of the body, like the eyes, lungs, intestines and genital tract, are protected from pathogens and toxins by layers of moist sticky mucus that is constantly secreted and shed, forming our outermost protective barrier.

“Although many people associate mucus with disgusting cold and cough symptoms, mucus is in fact a sticky barrier that helps keep you healthy,” says Laura Ensign, a doctoral student affiliated with the Center for Nanomedicine at the School of Medicine and with the Department of Chemical and Biomolecular Engineering at Johns Hopkins’ Whiting School of Engineering. She is the lead author of the journal report.

Unfortunately, Ensign noted, mucus barriers also stop helpful drug delivery, especially conventional nanoparticles intended for sustained drug delivery. In a Johns Hopkins laboratory, researchers developed nanoparticles that do not stick to mucus so they can slip through to reach the cells on the mucosal surface, in this case the surface of the mouse vagina, she added.

Ensign explained that conventional nanoparticles actually stick to mucus before releasing their drug payload and are then removed when the mucus is replenished, often within minutes to hours. Working with researchers in the laboratory of Richard Cone, Ph.D., in the Department of Biophysics in the university’s Krieger School of Arts and Sciences, the Hanes team fabricated particles with surface chemistry that mimics a key feature of viruses that readily infect mucosal surfaces.

“Richard Cone’s lab found that viruses, such as the human papilloma virus, could diffuse through human cervical mucus as fast as they diffuse through water. These ‘slippery viruses’ have surfaces that are ‘water-loving,’ ” Hanes said. “In contrast, many nanoparticles intended to deliver drugs to mucosal surfaces are ‘mucoadhesive’ and ‘oil-loving,’ but these nanoparticles stick to the superficial layers of the mucus barrier, the layers that are most rapidly removed.”

To make their mucus-penetrating particles, the team transformed conventional ‘oil-loving’ nanoparticles by coating them with a substance used in many commercial pharmaceutical products: polyethylene glycol. PEG makes the particles “water-loving,” like the viruses that slip right through mucus.

“The key is that the nanoparticles, like viruses, have to be small enough to go through the openings in the mucus mesh, and also have surfaces that mucus can’t stick to. If you think about it,” said Ensign, “mucus sticks to almost everything.”

“Viruses have evolved over millions of years to become slippery pathogens that readily penetrate our protective mucus barriers,” said Cone, “and engineering nanoparticles that penetrate the mucus barrier just like viruses is proving to be a clever way to deliver drugs.”

Hanes emphasized that the MPPs provided greatly improved protective efficacy while at the same time reducing the effective dose of drug needed 10-fold. Furthermore, Hanes added, the MPPs “continue to supply drug for at least a day and provide nearly 100 percent coverage of the mucosal surface of the vagina and ectocervix” in their laboratory mice.

“We’ve shown that mucus-penetrating particles are safe for vaginal administration in mice. Our next move will be to show that they are safe for vaginal administration in humans,” Ensign said. “Now our laboratory currently is working on an MPP formulation of a drug that protects against HIV infection that we hope will be tested in humans.”

Their technology could lead to a once-daily treatment for preventing sexually transmitted diseases, for contraception and for treatment of cervico-vaginal disorders, Ensign said.

Ensign added that MPP technology has the potential to prevent a wide range of mucosal diseases and infections, including chronic obstructive pulmonary disease, lung cancer, and cystic fibrosis,” Ensign said.

Additional authors on the paper include postdoctoral fellow Ying-Ying Wang and research specialist Timothy Hoen from the Department of Biophysics; former master’s student Terence Tse from the Department of Chemical and Biomolecular Engineering; and Benjamin Tang, formerly of Johns Hopkins School of Medicine and currently at the Massachusetts Institute of Technology.

Under a licensing agreement between Kala Pharmaceuticals and the Johns Hopkins University, Hanes is entitled to a share of royalties received by the university on sales of products used in the study.

Hanes and the university own Kala Pharmaceuticals stock, which is subject to certain restrictions under university policy. Hanes is also a founder, a director and a paid consultant to Kala Pharmaceuticals. The terms of this arrangement are being managed by The Johns Hopkins University in accordance with its conflict of interest policies.”

Story by Mary Spiro

Additional news coverage of this research may be found at the following links:

Phys.org

WYPR: The Mucus Ruse

Scientific American