Making therapeutic proteins last longer

Happy TRAILs to you: PEGylation of proteins through complementary interactions between a His-tag and a Ni2+ complex of nitrilotriacetic acid (NTA, see picture), a well-established practice in protein research, was used to improve the half-life of therapeutic proteins in the blood following systemic administration in vivo. Animal models show that this site-specific modification improves the efficacy of modified TRAIL proteins.

Happy TRAILs to you: PEGylation of proteins through complementary interactions between a His-tag and a Ni2+ complex of nitrilotriacetic acid (NTA, see picture), a well-established practice in protein research, was used to improve the half-life of therapeutic proteins in the blood following systemic administration in vivo. Animal models show that this site-specific modification improves the efficacy of modified TRAIL proteins.

Proteins are responsible for pretty much everything in the human body. When there is a problem with the proteins, it usually leads to disease.

Protein therapy shows enormous potential for treating disease. But sometimes the proteins in a therapeutic treatment break down or are metabolized before they ever reach their target destination.

In a recent paper published in Angewandte Chemie, researchers from the laboratories of Martin Pomper (radiology oncology) and Seulki Lee (radiology, Center for Nanomedicine) at the Johns Hopkins School of Medicine and developed a simple method to validate protein drugs in animal models, said Lee. An illustration related to the paper appeared on the cover of the journal.

“We show that we can extend the half-life, that is, the amount of time the drug stays in the blood, while maintaining the activity of the model protein drug, TRAIL,” said one of the lead authors Maggie Swierczewska. “This has great implications for drug screening and validation methods, especially for the growing protein drug market.”

According to the paper, by attaching a molecule of  polyethylene glycol (PEG) to certain sites on the TRAIL protein drugs through an already well known method, the half-life of the drug could be extended without affecting its beneficial activity.

Authors on this paper include Tae Hyung Kim, Magdalena Swierczewska, Yumin Oh, AeRyon Kim, Dong Gyu Jo, Jae Hyung Park,  Youngro Byun, Scheherazade Sadegh-Nasseri, Martin G. Pomper, Kang Choon Lee, Seulki Lee. Author affiliations include the departments of Radiology and Pathology at the Johns Hopkins School of Medicine, the Johns Hopkins Center of Cancer Nanotechnology Excellence, the Johns Hopkins Institute for NanoBioTechnology, Center for Nanomedicine and collaborators at Sungkyunkwan University and Seoul National University, both in Korea.

Reference: Kim, T. H., Swierczewska, M., Oh, Y., Kim, A., Jo, D. G., Park, J. H., Byun, Y., Sadegh-Nasseri, S., Pomper, M. G., Lee, K. C. and Lee, S. (2013), Mix to Validate: A Facile, Reversible PEGylation for Fast Screening of Potential Therapeutic Proteins In Vivo. Angew. Chem. Int. Ed.. Vol. 52, Issue 27, pages 6880-6884, doi: 10.1002/anie.201302181

Lab coats are summer gear for high school researchers

You don’t think of a lab coat as summer wear for teens, but we don’t quite feel like it’s summer around here until our research interns have arrived. Early in June, INBT’s undergraduate nano-bio researchers arrived. This week our high schoolers in the Summer Academic Research Experience (SARE) scholars got started.

SARE pairs specially selected teens with university mentors who guide them through a mini research project. At the end of their time here, they hold a small poster session. The students gain valuable work skills, learn about scientific careers, get tutoring help, practice their writing, gather data for their projects and earn some cash for the future. Students in the program are recruited from the Boys Hope Girls Home of Baltimore program, The SEED School of Maryland and The Crossroads School, all of which assist in differing ways with in the education, housing, tutoring  and counseling of promising young people from disadvantaged circumstances.

The SARE program was launched in 2009 by Doug Robinson, professor in the cell biology department at the School of Medicine, and is funded jointly by the medical school and Johns Hopkins Institute for NanoBioTechnology.

This year’s SARE scholars include: Diana Bobb is being mentored by Makoto Tanigawa in the Takanari Inoue Lab in the Department of Cell Biology; Kaleel Byrd is being mentored by Ryan Vierling in the Caren Meyers Lab in the Department of Pharmacology; Milan Dower is being mentored by Tom Lampert in the Peter Devreotes Lab in the Department of Cell Biology; Jewel Herndon is being mentored by Herschel Wade in his lab in the Department of Biophysics; De’Sean Markley is being mentored by Hoku West-Foyle in the Douglas Robinson Lab in the Department of Cell Biology

Fraley nets $500K Burroughs Wellcome Fund award for microfluidics work

Stephanie Fraley (Photo: Homewood Photography)

Stephanie Fraley (Photo: Homewood Photography)

A Johns Hopkins research fellow who is developing novel approaches to quickly identify bacterial DNA and human microRNA has won the prestigious $500,000 Burroughs Wellcome Fund (BWF) Career Award at the Scientific Interfaces. The prize, distributed over the next five years, helps transition newly minted PhDs from postdoctoral work into their first faculty positions.

Stephanie Fraley is a postdoctoral fellow working with Samuel Yang, MD, in Emergency Medicine/Infectious Disease at the Johns Hopkins School of Medicine and Jeff Wang, PhD, in Biomedical Engineering with appointments in the Whiting School of Engineering and the medical school. The goal of her work is to develop engineering technologies that can diagnose and guide treatment of sepsis, a leading cause of death worldwide, while simultaneously leading to improved understanding of how human cells and bacterial cells interact.

“Sepsis is an out of control immune response to infection,” Fraley said. “We are developing tools that are single molecule sensitive and can rapidly sort and detect bacterial and host response markers associated with sepsis. However, our devices are universal in that they can be applied to many other diseases.”

Fraley is using lab-on-chip technology, also known as microfluidics, to overcome the challenges of identifying the specific genetic material of bacteria and immune cells. Her technology aims to sort the genetic material down to the level of individual sequences so that each can be quantified with single molecule sensitivity.

“Bacterial DNA is on everything and contamination is everywhere, so trying to find the ones associated with sepsis is like the proverbial search for the needle in the haystack,” Fraley said. “With microfluidics, we can separate out all the bacterial DNA, so instead of a needle in a haystack, we have just the needles.”

Another advantage to Fraley’s novel technology is that it will assess all the diverse bacterial DNA present in a sample, without presuming which genetic material is important. “Bacteria are constantly evolving and becoming drug resistant,” she said. “With this technology, we can see all the bacterial DNA that is present individually and not just the strains we THINK we need to look for.”

Fraley’s award will follow her wherever her career takes her. The first two years of the prize fund postdoctoral training and that last three years help launch her professional career in academia. During the application process, she had to make a short presentation on her proposal to BWF’s panel of experts. “It was like the television show ‘Shark Tank’ but for scientists,” she laughs. “ The panelists gave me many helpful suggestions on my idea.”

Fraley earned her bachelor’s degree in chemical engineering from the University of Tennessee at Chattanooga and her doctorate in chemical and biomolecular engineering with Denis Wirtz, professor and director of Johns Hopkins Physical Sciences-Oncology Center. Wirtz is associate director for the Institute for NanoBioTechnology and Yang and Wang also are INBT affiliated faculty members.

BWF’s Career Awards at the Scientific Interface provides funding to bridge advanced postdoctoral training and the first three years of faculty service. These awards are intended to foster the early career development of researchers who have transitioned or are transitioning from undergraduate and/or graduate work in the physical/mathematical/computational sciences or engineering into postdoctoral work in the biological sciences, and who are dedicated to pursuing a career in academic research. These awards are open to U.S. and Canadian citizens or permanent residents as well as to U.S. temporary residents.

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.

 

 

 

 

Microscopic grippers used successfully in animal biopsies

Tiny, untethered microscale grippers have been successfully used to perform tissue biopsies in live animals, a study in the journal Gastroenterology reports. Researchers affiliated with the Johns Hopkins School of Medicine, Whiting School of Engineering and Institute for NanoBiotechnology developed the self-assembling microgrippers, called mu-grippers. The star-shaped devices use the animal’s own body heat to trigger them to clamp down around tissue to grab a sample like a tiny hand. Because the grippers are magnetic, they can later be retrieved for a minimally invasive procedure.

Dozens of dust-sized surgical mu- grippers in a vial. (Photo by  Evin Gultepe, Gracias Lab, Johns Hopkins University)

Dozens of dust-sized surgical mu- grippers in a vial. (Photo by Evin Gultepe, Gracias Lab, Johns Hopkins University)

David Gracias, the principal investigator for the study and associate professor of chemical and biomolecular engineering, was quoted in a Johns Hopkins press release about the work: ”This is the first time that anyone has used a sub-millimeter-sized device — the size of a dust particle — to conduct a biopsy in a live animal … That’s a significant accomplishment. And because we can send the grippers in through natural orifices, it is an important advance in minimally invasive treatment and a step toward the ultimate goal of making surgical procedures noninvasive.”

Read more here.

 

Regenerative medicine theme of science-writer bootcamp

Screen Shot 2013-04-01 at 3.19.05 PMJohns Hopkins invites you to the fifth annual science-writer boot camp. This year’s topic will be Regenerative Medicine. Join Johns Hopkins experts in regenerative medicine to learn the latest in stem cell research, tissue regeneration and organ transplantation.

Three of the 11 presenters are affiliated faculty members of the Johns Hopkins Institute for NanoBioTechnology. This event is sponsored by the Johns Hopkins Institute for Basic Biomedical Sciences.  There is no cost but reservations are required. Working press as well as freelance writers are invited to attend.

WHAT: Body Building: Recent Advances in Regenerative Medicine

WHEN: Monday, April 29, 2011, 9 a.m. to 4 p.m. (lunch will be provided)

WHERE: Bernstein-Offit Building, room LL7, Johns Hopkins SAIS Campus, 1717 Massachusetts Ave., NW, Washington, D.C. 20036

RSVP: Vanessa McMains at vmcmain1@jhmi.edu or 410-502-9410 by April 19

Confirmed speakers:

  • Gerald Brandacher, M.D. Scientific Director, Composite Tissue Allotransplantation (Reconstructive Transplant) Program
  • Robert Brodsky, M.D. Director, Division of Hematology
  • Jeff Bulte, Ph.D. Director, Cellular Imaging Section, Institute for Cell Engineering (INBT affiliated faculty)
  • Mark Donowitz, M.D. Director, Center for Epithelial Disorders; Director, Conte GI Core Research Center
  • Gary Gerstenblith, M.D. Professor, Medicine
  • Warren Grayson, Ph.D. Assistant Professor, Biomedical Engineering (INBT affiliated faculty)
  • Jun Liu, Ph.D. Professor, Pharmacology and Molecular Sciences
  • Erika Matunis, Ph.D. Associate Professor, Cell Biology
  • Guo-li Ming, M.D., Ph.D. Professor, Neurology and member of the Institute for Cell Engineering (INBT affiliated faculty)
  • Ronald Schnaar, Ph.D. Professor, Pharmacology and Molecular Sciences; Director, Lung Inflammatory Disease Program of Excellence in Glycoscience

We look forward to seeing you on April 29!

Download the color flyer here.

 

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/

 

Nanotech checks on transplanted cell survival

Researchers at Johns Hopkins are using nanotechnology to track the survival and location of transplanted cells. The device, based on nanoscale ph sensors and imaging via magnetic resonance, could help improve outcomes from cell replacement therapies used for conditions such as liver disease or type 1 diabetes.

Cartoon showing nanoscale probe used to detect pH change caused by death of transplanted cell. (McMahon/Nature Materials)

Cartoon showing nanoscale probe used to detect pH change caused by death of transplanted cell. (McMahon/Nature Materials)

“This technology has the potential to turn the human body into less of a black box and tell us if transplanted cells are still alive,” says Mike McMahon, Ph.D., an associate professor of radiology at the Johns Hopkins University School of Medicine principal investigator on the study. “That information will be invaluable in fine-tuning therapies.”

Transplanted cells often fall victim to assault from the body’s immune system, which sees the news cells as foreign invaders. Says McMahon,  ”once you put the cells in, you really have no idea how long they survive.”

When cells die there is a change in the acidity nearby. Using this fact, the researchers developed a nanoparticle sensor that could both sense the change in pH and be detected via MRI. The team tested the sensors on mice and found they they were able to track the location of surviving transplanted cells and determine the proportion that had survived.

“It was exciting to see that this works so well in a living body,” says research fellow Kannie Chan, Ph.D., the lead author on the study, which was published in Nature Materials. This should take a lot of the guesswork out of cell transplantation by letting doctors see whether the cells survive, and if not, when they die,” Chan says. “That way they may be able to figure out what’s killing the cells, and how to prevent it.”

Chan works in the laboratory of Jeff Bulte, Ph.D., the director of cellular imaging at Johns Hopkins’ Institute for Cell Engineering. Bulte and McMahon collaborated on the study. Additional authors include Guanshu Liu, Xiaolei Song, Heechul Kim, Tao Yu, Dian R. Arifin, Assaf A. Gilad, Justin Hanes, Piotr Walczak and Peter C. M. van Zijl, all of the Johns Hopkins University School of Medicine. McMahan, Bulte, Gilad, Hanes and van Zijl are all affiliated faculty members of Johns Hopkins Institute for NanoBioTechnology.

Funding for this study was provided by the National Institute of Biomedical Imaging and Bioengineering (grant numbers R01 EB012590, EB015031, EB015032 and EB007825).

Follow this link to read the paper, MRI-detectable pH nanosensors incorporated into hydrogels for in vivo sensing of transplanted-cell viability, in Nature Materials online http://www.nature.com/nmat/journal/vaop/ncurrent/abs/nmat3525.html

FLC event focuses on Maryland technology

Screen Shot 2013-02-04 at 10.59.42 AMMaryland Technology Past, Present and Future is the topic of a day-long symposium, February 28 at the National Electronics Museum hosted by the Federal Laboratory Consortium Mid-Atlantic Region.

The FLC is a national organization chartered by Congress to foster technology transfer from federal research laboratories and field centers, to other federal agencies; state and local government; academia and the private sector. One of the regional consortium’s efforts has been to conduct a series of one-day forums that highlight specific areas of technology and encourage collaboration and partnership development with federal labs.

Registration is $25 and includes refreshments and lunch. Registration deadline is February 15 and can be made online at this link.

The National Electronics Museum is located at 1745 West Nursery Road in Linthicum, Md. The symposium begins with registration at 8:15 a.m. and adjourns at 3:45 p.m.

In addition to the presentations, the day will offer the opportunity to meet scientists from the regions National Labs such as NASA, NIST, NIH and Goddard as well as representatives of local industry. In addition to the FLC Mid-Atlantic Region, participating organizations for this symposium include Johns Hopkins University and TEDCO.

For further information or if you have difficulty accessing the registration site, please contact John Emond at 301-384-2809 or johnlamaremond@aol.com. You may also contact INBT’s director of corporate partnerships, Tom Fekete at 410-516-8891 or tmfeke@jhu.edu.

A flyer and agenda for the event are below:

Maryland Technology Day Agenda

Maryland Technology Day Flyer

RNA nanotechnology and therapeutics conference registration opens

Mark your calendar. Those affiliated with Johns Hopkins Institute for NanoBioTechnology or Center for Cancer Nanotechnology Excellence may be interested to know that online registration is now open for the 2013 International Conference of RNA Nanotechnology and Therapeutics to be held in Lexington, KY on April 3-5, 2013 at the Crowne Plaza Hotel & Resorts.  The meeting is organized by Peixuan Guo (University of Kentucky CNPP), John Rossi (Beckman Research Institute), Bruce Shapiro (NCI), and Neocles Leontis (Bowling Green State University). Along with invited speakers, there will also be a poster session. Invited speakers are yet to be announced.

Program topics include:

  •  Biophysical and Single Molecule Approaches in RNA Nanotechnology
  • RNA Structure and Folding in Nanoparticles
  • RNA Computation and Modeling
  • RNA Nanoparticle Assembly
  • RNA Nanoparticles in Therapeutics
  • RNA Chemistry for Synthesis, Conjugation, & Labeling of Nanoparticles
  • RNA Systems Biology and Engineering
  • Exosomes and Extracellular RNA Communication

Additional details and registration information can be found at http://nanobio.uky.edu/RNA2013