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New method delivers Alzheimer's drug to the brain
March 21, 2011 article
(PhysOrg.com) -- Oxford University scientists have developed a new method for delivering complex drugs directly to the brain, a necessary step for treating diseases like Alzheimer’s, Parkinson’s, Motor Neuron Disease and Muscular Dystrophy.
These diseases have largely resisted attempts to over the last 50 years develop new treatments, partly because of the difficulty of getting effective new drugs to the brain to slow or halt disease progression.
The team has successfully switched off a gene implicated in Alzheimer’s disease in the brains of mice by exploiting exosomes – tiny particles naturally released by cells. The exosomes, injected into the blood, are able to ferry a drug across the normally impermeable blood-brain barrier to the brain where it is needed.
Although this is a significant and promising result, there are a number of steps to be taken before this new form of drug delivery can be tested in humans in the clinic. The study, partly funded by the Muscular Dystrophy Campaign, is published in Nature Biotechnology.
"These are dramatic and exciting results. It’s the first time new 'biological' medicines have been delivered effectively across the blood-brain-barrier to the brain," says Dr. Matthew Wood of the Department of Physiology, Anatomy and Genetics at the University of Oxford, who led the work.
Exosomes are small capsules that are produced by most cells in the body in varying amounts. These natural nanoparticles are thought to be one of the ways cells communicate with each other and the body’s immune system. When exosomes break off from the outer walls of cells, they can take various cellular signals and genetic material with them, transporting this material between different cells.
This led the Oxford University researchers to wonder whether exosomes could be adapted for delivering drugs to different cells and tissues of the body.
"This is the first time this natural system has been exploited for drug delivery," says Dr. Wood.
Novel drugs based on antibodies, peptides or more recently RNA molecules have been developed on many occasions to target specific parts of disease pathways. While these have shown good results in the lab, too often it has proved difficult to get the drugs to the right part of the body to see any effect in humans.
Currently, delivering any such type of therapy to the brain would have to involve neurosurgery. Nothing delivered intravenously would be able to cross from the blood into the brain.
"The major barrier for these drugs is delivery," explains Dr. Wood. "This problem becomes even greater when you want to reach the brain. The blood-brain barrier – which stops most things in the blood stream from crossing to our brains – is much too great an obstacle."
The Oxford University team set out to adapt naturally occurring exosomes to deliver a gene therapy. They used an RNA sequence – RNA is a molecule related to DNA that also caries genetic information – that switches off a gene that’s implicated in Alzheimer’s disease.
To be able to make the approach work, they would need to be able to load the exosomes with the RNA, the drug. But they would also need to be able to target the right tissues in the body.
First of all, they produced and purified exosomes from mouse cells. They then developed and patented new methods to both insert RNA molecules into the exosomes and add protein elements into the exosome coat that would target nerve cells.
The exosomes, injected intravenously into mice, crossed the blood-brain barrier and ended up in the brain. Once there, the RNA was able to switch off a gene implicated in the build up of malformed protein in Alzheimer’s disease. This resulted in a 60% decrease in the brain of the problem enzyme encoded by the gene.
"We’ve shown that a natural system could be exploited to deliver drugs across the blood-brain barrier," says Dr. Wood. "We believe we can use this same technology for Alzheimer’s, motor neuron disease, Parkinson’s and Huntington’s. All we need is a different RNA each time.
"The next steps are to test the exosomes in a mouse model of Alzheimer’s disease to see if it makes a difference to disease progression," Dr. Wood explains.
He also notes that other steps would be needed before exosomes could be tested in humans, including safety tests and scaling up the procedures.
"Many of these diseases have not been possible to treat in the last 50 years using standard drugs. New drugs have been developed based on complex biological molecules – antibodies, peptides, and RNA – but all require new ways of delivering the drugs," he says.
"These natural nanoparticles would be administered intravenously, or perhaps even orally, and would still reach the brain."
Genetic Hijacking May Lead to New Tests for Brain Cancer - old article but worth reading
By Aalok Mehta
February 24, 2009
An unusual form of cell hijacking may offer doctors ways to improve blood tests and treatments for one of the deadliest brain cancers.
Scientists have discovered that glioblastoma tumors release microvesicles—tiny membrane-covered sacs containing proteins and forms of ribonucleic acid (RNA)—that help the cancer spread by co-opting the molecular machinery of nearby cells.
Enough of these microvesicles, also known as exosomes, cross the blood-brain barrier to show up on blood tests, researchers report in the December issue of the journal Nature Cell Biology.
Scientists knew that some cancers send off and fuse with microvesicles that contain growth-assisting proteins, though many of the details remain hazy. The new study is the first to characterize the genetic contents of a cancer exosome.
“These microvesicles are filled with messenger RNAs (mRNAs) and micro RNAs, and they’re an indicator of what’s going on genetically inside a cell,” says study senior author Xandra Breakefield, a professor of neurology at Massachusetts General Hospital and Harvard Medical School. “We think normal cells use this for cell communication. But tumor cells use this with a vengeance—probably for taking over their environment.”
The glioblastoma exosomes contained mutated RNAs associated with the growth of new blood vessels, immune suppression and other processes that contribute to cancer cell survival, Breakefield says. In lab tests with healthy cells, these RNAs were transcribed into proteins, suggesting that the cancer cells are coercing assistance directly from their neighbors.
Genetic profiling
To assess potential diagnostic tools, the scientists tested the blood of 25 people with glioblastoma and found evidence of exosomes in many of the samples. The researchers also were able to do detailed genetic workups; for example, in several instances the team found a mutation in the epidermal growth factor receptor (EGFR) that characterizes one common subtype of glioblastoma.
“This is the first study where we enabled mutational profiles in serum from tumors,” says study lead author Johan Skog, a neurology instructor at Harvard Medical School. “Before we had to do a biopsy; now we can do this in blood.”
The blood tests may even be more precise. In two cases, the researchers found EGFR mutations not detected during a biopsy. The authors suggest that many such abnormalities could be missed during sampling because tumors are heterogeneous mixes of different populations of cells; the tiny sample sites might not contain each type of cell.
Since some mutations are linked with vulnerabilities to certain kinds of drugs, better genetic profiles could allow doctors to choose more effective cancer treatment plans. Regular blood testing could also allow scientists to see how cancer cells are responding to therapy, which might prompt refinements in treatment, Breakefield says.
The tests may also be useful in detecting relapse: Many cancer cells return with similar genetic profiles, often months before they can be seen via brain scanning.
“We desperately need ways to tell if a tumor is coming back or not,” Breakefield says.
The team is now planning to study whether exosomes are involved in other types of solid tumors.
Better than the alternatives?
Other scientists are cautiously hopeful about the new techniques, noting that exosomes remain a poorly understood form of cell communication.
“The paper is potentially important because it illuminates a rather underappreciated—but not previously unknown—mechanism by which tumor cells can communicate with each other and with their microenvironment,” says Sean Lawler, a neurosurgery professor and brain tumor researcher at Ohio State University.
“If it is possible to enrich tumor-specific mRNAs, then this would be an important breakthrough, particularly for brain tumors, in which it is difficult to monitor treatment response or recurrence.”
Calcsmart - thank you for the company!
Exosomes used for genetic therapy!
New gene therapy 'may target' Alzheimer's
Published Date: 22 March 2011
"Scientists have successfully switched off a gene thought to cause Alzheimer's by using a new way to deliver drugs directly to the brain," reported the Daily Mirror. It said that researchers have used "tiny particles called exosomes, which are released by cells, to administer drugs into the brains of mice". The laboratory study behind these headlines was carried out in mice. The findings are significant, demonstrating that exosomes could be used to carry gene therapy to parti
The study paves the way for future research and the finding will be of great interest to the scientific community. Exosomes appear to be able to deliver specific ‘cargoes’ to brain cells so the technology has a number of potential applications. Howev
er, this is early research and the technology has not been tested in human cells. There is also a range of technical and ethical issues associated with gene therapy in humans.
Where did the story come from?This study was carried out by researchers from the University of Oxford. The work was funded by Muscular Dystrophy Ireland and the Muscular Dystrophy Campaign. The paper was published in the peer-reviewed medical journal Nature Biotechnology.
The newspapers have covered the study well, however, some headlines and images may give the false impression that it was in humans or that it will change the way that Alzheimer’s disease is currently treated. This is not the case. This is early research and it has yet to
Exosome-driven drug delivery
Vesicles secreted by the body's own cells could be used to effectively transport drugs to hard-to-reach places, such as the brain. In a study from Nature Biotechnology, researchers were able to harvest exosomes (vesicles secreted by a variety of cells to transport proteins in and out of cells) from mice, target them to brain cells by fusing proteins that bind to brain-cell receptors, and pack them with small interfering RNA molecules to silence BACE1, which plays a role in the formation of the myelin sheaths that are hallmarks of Alzheimer's diease. This proof-of-concept study suggests that exosomes may be useful in treating diseases such as Alzheimer's, Parkinson's and muscular dystrophy, and according to BBC News, the researchers expect to begin trials in human patients within five years.
Human cerebrospinal fluid contains exosomes that represent a novel reservoir for
therapeutic biomarker discovery
J.M. Street1, R.T.A. Chalmers2, T.S. Walsh2, D.J. Webb1, J.W. Dear1. 1University of
Edinburgh, Edinburgh, United Kingdom, 2Royal Infirmary of Edinburgh, Edinburgh, United
Kingdom.
One of the pathological hallmarks of neurodegenerative diseases, such as spongiform
encephalopathies (TSEs) and Alzheimer's dementia (AD), is the accumulation of abnormal
proteins within brain tissue (prions in TSEs and b-amyloid in AD). Studies in cell culture have
demonstrated these proteins are released from cells within lipid vesicles termed exosomes. In
vitro, the exosomal protein content changes with drug therapy, which suggests that exosomes
could represent a reservoir for therapeutic biomarker discovery. Our aim was to determine if
exosomes are present in human cerebrospinal fluid (CSF). CSF was collected from patients
undergoing thoraco-abdominal aortic aneurysm repair (with full ethical approval). This group
was chosen as they have a CSF drain inserted peri-operatively as part of routine clinical
management. The CSF was ultracentrifuged (as per established protocols for urinary
exosome isolation) and the presence of exosomes was tested by western blotting for specific
markers, sucrose gradient centrifugation, immunoelectron microscopy and the proteome was
explored by tandem mass spectrometry. The exosomal markers TSG101 and Flotillin-1 were
found to be enriched in the ultracentrifugation pellet over the unfractionated CSF. Using
sucrose gradient centrifugation, the density of the 'microvesicles' was established as 1.14 -
1.20 g.cm-3, consistent with the density previously reported for exosomes. Staining for
Flotillin-1 during electron microscopy revealed the presence of Flotillin-1 on structures
consistent in size and shape with previous reports for exosomes. Alpha-2-macroglobulin,
which is involved in the clearance of A-beta, and filamin-A, which is linked to periventricular
heterotopia, were identified in the exosomes by tandem mass spectrometry. Based on this
evidence, we conclude that human CSF contains exosomes. The existence of exosomes
provides a source for new diagnostic and therapeutic biomarkers and proteomic discovery
studies are ongoing.
Exosomes linked to Parkinson's disease!
Cell-Produced a-Synuclein Is Secreted in a Calcium-Dependent Manner by Exosomes and Impacts Neuronal Survival
Evangelia Emmanouilidou1,
Katerina Melachroinou1,
Theodoros Roumeliotis2,
Spiros D. Garbis2,
Maria Ntzouni3,
Lukas H. Margaritis3,
Leonidas Stefanis1,4, and
Kostas Vekrellis1
+ Author Affiliations
1Divisions of Basic Neurosciences and
2Biotechnology, Biomedical Research Foundation of the Academy of Athens, Athens 11527, Greece,
3Department of Cell Biology and Biophysics, Faculty of Biology, University of Athens, Athens 15701, Greece, and
4Second Department of Neurology, University of Athens Medical School, Athens 11527, Greece
Abstract
a-Synuclein is central in Parkinson's disease pathogenesis. Although initially a-synuclein was considered a purely intracellular protein, recent data suggest that it can be detected in the plasma and CSF of humans and in the culture media of neuronal cells. To address a role of secreted a-synuclein in neuronal homeostasis, we have generated wild-type a-synuclein and ß-galactosidase inducible SH-SY5Y cells. Soluble oligomeric and monomeric species of a-synuclein are readily detected in the conditioned media (CM) of these cells at concentrations similar to those observed in human CSF. We have found that, in this model, a-synuclein is secreted by externalized vesicles in a calcium-dependent manner. Electron microscopy and liquid chromatography–mass spectrometry proteomic analysis demonstrate that these vesicles have the characteristic hallmarks of exosomes, secreted intraluminar vesicles of multivesicular bodies. Application of CM containing secreted a-synuclein causes cell death of recipient neuronal cells, which can be reversed after a-synuclein immunodepletion from the CM. High- and low-molecular-weight a-synuclein species, isolated from this CM, significantly decrease cell viability. Importantly, treatment of the CM with oligomer-interfering compounds before application rescues the recipient neuronal cells from the observed toxicity. Our results show for the first time that cell-produced a-synuclein is secreted via an exosomal, calcium-dependent mechanism and suggest that a-synuclein secretion serves to amplify and propagate Parkinson's disease-related pathology.
Therapeutic uses of exosomes
Cell's own machinery can deliver therapies to the brain
Jessica Wright
13 April 2011
Express delivery: Researchers can use bubbles of cellular membrane, called exosomes, to carry therapies into the brain.
Exosomes, the brain's system for delivering and recycling molecules, can be manipulated to carry therapeutic fragments of RNA or DNA across the blood-brain barrier and into neurons. The ingenious new technique was published 20 March in Nature Biotechnology.
As studies uncover the genetic and molecular causes of autism, gene therapy — silencing the expression or boosting levels of a particular gene product — could emerge as a therapeutic option.
Targeting these genes to brain cells is complicated, however. Injecting constructs into the blood delivers them indiscriminately to all tissues, and the blood-brain barrier prevents most molecules from crossing into the brain at all.
In the new study, researchers solved both problems by coating mouse exosomes — bubbles of membrane that pinch off and merge their contents when they fuse with another membrane — with a brain-targeting protein.
The exosomes are engineered from mouse bone marrow, and purified and filled with short genetic fragments called siRNA, or short interfering RNA. These fragments bind to and silence specific messages that code for protein. When injected into mice, exosomes containing siRNA for BACE1, a protein associated with Alzheimer's disease, lower BACE1 protein levels by 62 percent in brain tissue.
Injecting siRNA for GAPDH — a protein present in all tissues — into blood silences the protein in the liver, spleen and kidney, but not the brain. By contrast, when GAPDH siRNA is delivered in brain-specific exosomes, it has no effect on these tissues and targets only neurons and other brain cells, including microglia.
Although the genetic causes of autism are not yet well enough understood to suggest a clear target for gene therapy, the ability to target molecules specifically to the brain has potentially wide applications for research into all neurodevelopmental disorders.
Exosomes and melanoma - some strong research linking exosomes to the spread of melanoma!
Exosomes released by melanoma cells prepare sentinel lymph nodes for tumor metastasis
Joshua L Hood1,*,
Susana San Roman1, and
Samuel A Wickline2
+ Author Affiliations
1C-TRAIN, Washington University in Saint Louis
2Washington University, St. Louis
*?Corresponding Author:
Joshua L Hood, C-TRAIN, Washington University in Saint Louis, 4320 Forest Park Avenue, Suite 101, Campus Box 8215, Saint Louis, MO, 63108, United States jhood@dom.wustl.edu
Abstract
Exosomes are naturally occurring biological nanovesicles utilized by tumors to communicate signals to local and remote cells and tissues. Melanoma exosomes can incite a pro-angiogenic signaling program capable of remodeling tissue matrices. In this study, we demonstrate exosome-mediated conditioning of lymph nodes and define microanatomic responses that license metastasis of melanoma cells. Homing of melanoma exosomes to sentinel lymph nodes imposes synchronized molecular signals that effect melanoma cell recruitment, extracellular matrix deposition and vascular proliferation in the lymph nodes. Our findings highlight the pathophysiological role and mechanisms of an exosome mediated process of microanatomical niche preparation that facilitates lymphatic metastasis by cancer cells.
Received December 9, 2010.
Revision received March 7, 2011.
Accepted March 27, 2011.
Copyright © 2011, American Association for Cancer Research.
Exosomes and Ovarian cancer
Interaction and uptake of exosomes by ovarian cancer cells
BMC Cancer, 03/29/2011
Escrevente C et al. – Exosomes are internalized by ovarian tumor cells via various endocytic pathways and proteins from exosomes and cells are required for uptake. Exosomes are enriched in specific glycoproteins that may constitute exosome markers. This work contributes to the knowledge about the properties and dynamics of exosomes in cancer.Results •Ovarian carcinoma SKOV3 cell line has been shown to internalize exosomes from same cells via several endocytic pathways that were strongly inhibited at 4 oC, indicating their energy dependence
So although the two new drugs are up for approval next month, it is not so much of a bad news for AEMD. Althogh the SVR are better compared to the current SOC, they are not where they can be with the HP as an adjunct i.e greater than 90% atleast.
Mutation of the virus may be a big thing. This may confer drug resistance. For the HP there is no issue of resistance!
Now this is just a theory of mine - more the virus more chance of mutation, so if the HP is used to bring the virus count to negligible levels then less chance of mutation and the drug can penetrate more.
Less effectiveness of the drug in African Americans - No issue with the HP in terms of race. Also, is the response in Asians any different? Asia also has its high share in terms of HCV infected people.
The new drugs have some side effects - the HP has been used extensively with minimal side effects as per the company.
HCV apprantely is a huge market and that is why people are excited to see new drugs coming in. But although they offer hope they still do not offer complete recovery round the board. That may a possibility with the HP
So although the two new drugs are up for approval next month, it is not so much of a bad news for AEMD. Althogh the SVR are better compared to the current SOC, they are not where they can be with the HP as an adjunct i.e greater than 90% atleast.
Mutation of the virus may be a big thing. This may confer drug resistance. For the HP there is no issue of resistance!
Now this is just a theory of mine - more the virus more chance of mutation, so if the HP is used to bring the virus count to negligible levels then less chance of mutation and the drug can penetrate more.
Less effectiveness of the drug in African Americans - No issue with the HP in terms of race. Also, is the response in Asians any different? Asia also has its high share in terms of HCV infected people.
The new drugs have some side effects - the HP has been used extensively with minimal side effects as per the company.
HCV apprantely is a huge market and that is why people are excited to see new drugs coming in. But although they offer hope they still do not offer complete recovery round the board. That may a possibility with the HP
http://www.medpagetoday.com/InfectiousDisease/Hepatitis/26136
I have highlighted some interesting points which may work out in AEMD's favor.
However, a briefing document released in advance of the meeting did raise concerns about skin rashes and anemia seen in patients receiving telaprevir in clinical studies and the emergence of mutations in the drug's enzyme target that appear to render the virus drug-resistant.
Sustained response rates were lower in patients with previous inadequate responses to the standard treatment, but still much higher than in control groups receiving another round of conventional therapy:
Prior null responders: 31% versus 3%
Prior partial responders: 57% versus 15%
Prior relapsers: 84% versus 24%
Relapse rates were slashed in patients receiving telaprevir relative to controls: 5% versus 26% in previously untreated patients and 10% versus 57% in treatment-experienced patients.
The core dosing recommendation proposed by Vertex for the drug's label is 750 mg of telaprevir three times daily for 12 weeks, combined with peginterferon and ribavirin at standard doses for 24 or 48 weeks, depending on virologic response.
For treatment-naive patients and prior relapsers whose viral loads are undetectable at weeks four and 12, the shorter regimen would be recommended, according to Vertex's application. Other patients would receive the full 48 weeks of therapy.
This type of "response-guided therapy" raised a concern for FDA staff reviewing the data for boceprevir, as some of the trials showed lower response rates for the alternative dosing scheme.
The FDA's briefing document for telaprevir, on the other hand, did not highlight any problems with the response-guided therapy, although the advisory committee will still be asked to consider its appropriateness.
A somewhat greater worry for FDA staff was a finding of mutations in the HCV protease enzyme that appeared to make the virus resistant to telaprevir. Data from the trials indicated that, in patients not maintaining good responses, certain point mutations in the protease had emerged during treatment.
In one of the studies, 90% of treatment failures had treatment-emergent amino acid substitutions in the protease sequence, according to the briefing document.
The significance of this treatment-emergent resistance also will be a topic of discussion for the panel.
Safety concerns center on two toxicities seen during the telaprevir studies -- skin rashes that, in some patients, were severe enough to warrant stopping the drug, and anemia that was substantially more common with telaprevir.
More than half of patients receiving the drug reported rash or pruritus, with 6% discontinuing treatment as a result. These rates were about double those in the control groups.
Similarly, hemoglobin reductions of grade 3 or more were seen in 55% of telaprevir patients versus 25% of control patients.
Anemia is also an issue for boceprevir, but the skin problems are unique to telaprevir.
Advisory committee members will be asked how these side effects affect their risk-benefit assessments.
Other discussion topics will include recommendations for telaprevir's use in specific populations such as blacks -- as with boceprevir, sustained response rates were about 20 percentage points lower in African Americans -- and recommendations for postmarketing studies on the drug's optimal use.
http://www.medpagetoday.com/InfectiousDisease/Hepatitis/26160
Some points from the article -
Panelist Lawrence Friedman, MD, chair of the Department of Medicine at Newton-Wellesley Hospital in Newton, Mass., said when he first started seeing HCV patients, there was no treatment at all.
"To go to 60% or 70% [sustained response] really seems like a dream come true," he said. "I think this is a major advance, so I'm very enthusiastic about this drug."
On Wednesday, the panel discussed Merck's trial data, which suggest that adding boceprevir to the standard peginterferon-ribavirin combination doubles or triples the sustained viral response rates among certain subsets of patients.
The core treatment regimen with boceprevir tested in the trials included a four-week lead-in period with peginterferon and ribavirin with boceprevir then added for 44 weeks. But some of the trials also included a regimen in which the duration of boceprevir treatment could be extended based on how well a patient was responding to treatment after eight and 24 weeks.
Merck would like the drug's label to allow longer treatment for patients with detectable HCV RNA at eight weeks but who achieve a full virologic response at 24 weeks, as some data suggested that such a regimen improves the sustained response rate.
But the panel expressed concern over that type of "response-guided therapy," in part because it would make HCV treatment even more complicated.
Panelists also discussed the hematologic side effects associated with boceprevir -- including anemia, neutropenia, and thrombocytopenia -- all of which were more common in patients treated with boceprevir -- but they were ultimately convinced that anemia is a manageable side effect during treatment with boceprevir and is reversible after the drug is stopped.
The delays are based on the expectation that "the new treatments will be both more effective and administered over shorter courses," Gardenier said in an email to MedPage Today.
Once the drug is approved, it still has to be made and shipped and insurers need to agree to pay for it, he noted. Currently, experts are expecting to see the drug in the clinic by late summer or early fall.
Exosomes in Gyn cancer
Semin Thromb Hemost 2010; 36(8): 925-929
DOI: 10.1055/s-0030-1267046
© Thieme Medical Publishers
Microparticles and Exosomes in Gynecologic Neoplasias
Rienk Nieuwland1, Joris A.M. van der Post2, Christianne A.R. Lok Gemma2, G. Kenter3, Augueste Sturk1
1 Department of Clinical Chemistry, Academic Medical Center, Amsterdam, The Netherlands
2 Department of Obstetrics and Gynaecology, Academic Medical Center, Amsterdam, The Netherlands
3 Department of Gynaecological Oncology, Academic Medical Center, Amsterdam, The Netherlands
ABSTRACT
This review presents an overview of the functions of microparticles and exosomes in gynecologic neoplasias. Growing evidence suggests that vesicles released from cancer cells in gynecologic malignancies contribute to the hypercoagulable state of these patients and contribute to tumor progression by suppressing the immune system, facilitating extracellular matrix degradation and removal of cytostatics from the tumor cell. Exosomes from ovarian carcinoma cells were shown to be present in peripheral blood and to augment tumor growth, suggesting that these vesicles directly support growth of tumor cells.
Very interesting article about how a common virus like the EBV causes oropharyngeal cnacer through exosomes.
http://currentcancer.com/cellular-communicators-for-cancer-virus-identified-by-unc-scientists.html
Exosomes for salivary gland cancer.
Exosomes from human saliva as a source of microRNA biomarkers
Oral Diseases , 07/23/2009
Michael A et al. – The isolation of miRNAs from easily and non–invasively obtained salivary exosomes with subsequent characterization of the miRNA expression patterns is promising for the development of future biomarkers of the diagnosis and prognosis of various salivary gland pathologies.
Exosomes in Neurological Disease - Back to some DD on exosomes - this field is expanding and AEMD seems to be positioned to capitalize from it.
Andrew F Hill
CML – Neurology 2009;25(2):27–32.
Many cell types, including neurons, are known to release small membranous vesicles known as exosomes. In addition to their protein content, these vesicles have recently been shown to contain messenger RNA (mRNA) and micro RNA (miRNA) species. Roles for these vesicles include cell–cell signaling, removal of unwanted proteins, and transfer of pathogens, such as infectious prions, between cells. Prions are the infectious particles that are responsible for the transmission of neurodegenerative diseases such as Creutzfeldt–Jakob disease (CJD) in humans or bovine spongiform encephalopathy (BSE) in cattle. Exosomes are also involved in the processing of amyloid precursor protein (APP), which is associated with Alzheimer’s disease (AD). As exosomes can be isolated from circulating fluids such as serum, urine, and cerebrospinal fluid (CSF), they provide a potential source of biomarkers for neurological conditions. The current review describes the roles that these vesicles play in neurodegenerative diseases and their potential use in diagnosing brain tumors through analysis of their RNA content
From the CEO, dated 6/2010 - Part 2
I suspect most individuals following the HCV treatment industry are not yet aware of a medical device study that demonstrated the mechanical removal of HCV through blood filtration outperforms Telaprevir as an adjunct to SOC therapy. The insight provided by this clinical validation should significantly benefit our endeavors. In a 63 patient study conducted in Japan, Asahi Kasei Kuraray Medical (Asahi) demonstrated that double filtration plasmapheresis (DFPP) when administered at the outset of SOC therapy provided a 77.8% SVR in HCV-infected patients. In patients who previously failed SOC, DFPP treatment provided an average SVR of 71.4% versus the 51% previously referenced in Telaprevir clinical studies. On average, each patient in the Asahi study received three DFPP treatments each lasting 3.14 hours. In the study, DFPP was administered once daily for three consecutive days at the outset of SOC therapy and provided an average viral load reduction of 26.1% during each treatment period. Amazingly, the 71.4% and 77.8% cures rates were achieved without any additional DFPP during the remaining SOC treatment regimen. As a result of DFPP treatment outcomes, Asahi has advanced DFPP beyond treatment candidate status to actively marketing the treatment in Japan as the V-RAD system, which Asahi derives from the phrase "Virus Removal and Eradication by DFPP". Additional information can be accessed online at: www.v-rad.jp/en/index.html. I shall keep my comments directed towards the science underlying the V-RAD system and not the animation you will encounter at this website. Regardless, the website is quite informative.
However, there are significant limitations for DFPP as compared to our Hemopurifer(R). Like other approaches to therapeutic filtration, DFPP relies on multiple pumps and filters to indiscriminately remove particles by molecule size. For this reason, DFPP also extracts particles beyond HCV that are required for patient health. The safety profile of DFPP can be further diminished by the need for replacement fluids. In combination, these factors limit the time an HCV-infected patient can be exposed to DFPP treatment.
The advantages of our Hemopurifier(R) as compared to DFPP include the following:
The Hemopurifier(R) provides a greater reduction of HCV from circulation during treatment. Our data resulting from over 20 HCV treatments indicates an average viral load reduction of 41% during four-hour treatment applications of the Hemopurifier(R).
The Hemopurifier(R) augments the immune response by removing toxic proteins shed from HCV to kill-off immune cells. These proteins are too small to be captured by DFPP.
The Hemopurifier(R) is designed to selectively capture HCV and immunosuppressive proteins versus the indiscriminate removal of particles by DFPP.
The Hemopurifier(R) is one single-use disposable cartridge versus the requirement for two cartridges and multiple pumps with DFPP.
The selective ability of the Hemopurifier(R) to capture targeted viruses and immunosuppressive proteins (versus the removal of needed blood components) allows for a continuous Hemopurifier(R) treatment strategy to rapidly reduce viral load to low to undetectable levels. Thus, increasing the likelihood that HCV infected individual can be cured by SOC therapy.
While we believe our Hemopurifier(R) has obvious advantages over the DFPP system, I wish to expand on point #5 as it provides a foundation to support our treatment goal of increasing HCV cure rates up to 90%. Based on published treatment literature, it is well established that patients who initiate SOC and achieve a rapid viral response (RVR) have significantly higher cure rates. RVR is defined as undetectable viral load at day 30 of SOC treatment. In fact, published literature indicates the small percentage of patients who do achieve a RVR have cure rates that range from 86-92%. Based on our Hemopurifier(R) data, we believe it is possible to achieve undetectable levels of HCV in week one of SOC therapy, not day 30. Based on data analyzed from four-hour Hemopurifier(R) treatments, we project that a patient with a high viral load of 7 million iu/ml might be reduced to undetectable HCV levels after approximately three days of continuous Hemopurifier(R) treatment. This corresponds to a 4.06 log reduction or a 11,000-fold decrease in viral load. An HCV patient with a moderate viral load of 2 million iu/ml would be projected to reach undetectable levels in approximately 2.5 days of continuous treatment. Such outcomes would position us to achieve our 90% cure rate goal and may allow for decreased dosages and duration of SOC therapy.
To leverage our opportunity in HCV care, we are pursuing strategic relationships that will broaden our ability to commercialize in practitioner driven markets, or accelerate our clinical opportunities in the U.S. and European Union. Additionally, we have responded to a grant opportunity to advance a diagnostic based Hemopurifier(R) and have been working on a candidate clinical protocol for a grant proposal related to the use of our Hemopurifier(R) as an adjunct cancer treatment to remove tumor secreted exosomes known to suppress the immune system of cancer patients. The data from these cumulative activities, including recent HIV and HCV treatment outcomes, will cause us to update the investigational device exemption (IDE) we have previously filed with the FDA related to use of our Hemopurifier(R) as a treatment countermeasure against bioterror and pandemic threats. In this regard, we were recently advised that we were a candidate being considered for a contract award from the Biomedical Advanced Research and Development Authority (BARDA). This was related to a multi-agency contract solicitation known as DMID-NIAID-NIHAI20080022BARDA. We have since been advised by BARDA that they will not be granting awards under this solicitation. BARDA has encouraged to update our data collected since our original submission and resubmit a new proposal to a BARDA specific contract solicitation known as BAA-BARDA-09-34. As we believe our Hemopurifier(R) represents the most advanced broad-spectrum treatment strategy to protect our military and civilian populations from viruses considered bioterror and pandemic threats, we plan to provide BARDA our new submission no later than July 31st. Regardless of these opportunities, our primary focus moving forward will be the treatment of Hepatitis-C.
On behalf of our dedicated team at Aethlon Medical, I thank you for your continued support.
Very truly yours,
James A. Joyce
Chairman, CEO
From the CEO in a PR dated 6/10/10 -Part 1
"Our goal in HCV care is to increase patient cure rates up to 90%. We envision two pathways to reach this goal:
Our Hemopurifier(R) as an adjunct treatment to enhance the benefit of SOC therapy; or
Our Hemopurifier(R) in combination with a candidate therapy to replace SOC therapy.
The achievement of our goal would significantly impact the HCV treatment industry as SOC therapy succeeds in providing sustained viral responses (SVR) in only 30% to 50% of patients who initiate treatment. HCV infection is considered cured when a SVR of undetectable viral load is maintained more than six months after completing treatment. Prior to discussing the clinical rationale supporting our treatment goals, I want to clarify the magnitude of the HCV treatment opportunity.
It is estimated that nearly 180 million people worldwide, or approximately 3% of the world's population, are infected with HCV. To provide perspective, this represents a patient population approximately 5-6 times larger than those infected with HIV/AIDS and over 100 times larger than the population of end stage renal disease (ESRD) patients who require kidney dialysis. However, unlike kidney dialysis and HIV therapeutics, we actually have the opportunity to participate in curing HCV-infected individuals. The global market for therapies to treat HCV is projected to reach $9.1 billion by 2015, and in the United States, the annual cost of advanced liver disease resulting from HCV infection is anticipated to jump to $85 billion in the next two decades. As a result, Medicare costs are anticipated to soar 500%, from $5 billion to $30 billion. As the stakes to treat HCV are high, the competition for new drugs is intense with more than sixty treatment candidates reported to be in development. As we target the use of our Hemopurifier(R) as a drug enhancement device in HCV care, we are positioned to improve treatment outcomes as an adjunct to both SOC therapy and new drug candidates that evolve to challenge SOC therapy in the marketplace.
Significant challenges exist for drugs seeking to supplant SOC therapy, as new candidates must demonstrate substantially greater patient benefit in order for the medical community to consider discontinuing administration of the SOC treatment regimen. As an example, Albuferon, an HCV treatment candidate from Human Genome Sciences (HGSI), recently demonstrated phase III treatment outcomes comparable to SOC therapy with half the number of required injections. As a result, the value of HGSI shares was reduced by 57% the day the study data was released. I can't help but wonder how Albuferon might have performed in combination with our Hemopurifier(R)? Regardless, a drug candidate wishing to supplant SOC therapy in the market will need to Bob Beamon (surpassed world long jump record by almost two feet at the 1968 Olympics) beyond the capabilities of SOC therapy.
For this reason, the primary strategy for most HCV drug candidates is to incrementally improve treatment outcomes as an adjunct to SOC therapy. The challenge facing these candidates is the effect of stacking new drug toxicity on top of established SOC toxicity, which is known to trigger fatigue, bone marrow suppression, anemia and neuropsychiatric effects. Many patients fail SOC therapy because they are unable to endure the toxicity of the 24-48 week regimen on its own. Based on clinical data, Telaprevir, a 3x-day oral drug from Vertex Pharmaceuticals is considered the leading adjunct candidate based on outcomes of a recent phase II study, which documented that 51% of patients that previously failed SOC had a sustained virologic response (SVR) when retreated with SOC and Teleprevir in combination. When considering that only 14% of patients in the study control arm responded to SOC alone, there is certainly valid justification for Telaprevir to be considered the lead adjunct drug candidate by the medical and the financial community. This is reinforced by the reality that Telaprevir represents a significant value component of Vertex (VRTX), which as I write this letter is valued at more $5 billion in the public markets. In regards to deal values in the HCV space, VRTX paid almost $400 million in March to acquire ViroChem, a drug developer with two experimental stage HCV drugs. TheStreet.com, who provides excellent HCV market coverage, reports the following on the Telaprevir clinical outcome; "The data keeps Telarevir ahead of its hepatitis C rivals because no other drug has yet shown the ability to improve the cure rates for both patients new to therapy as well as those who have failed prior therapy." The key phrase in that statement is "no other drug".
AEMD with new HCV drugs -
As JJ has always said that the HP would be used with SOC (RBV+PEG). If the new drugs are expensive and with side effects then MD's would prefer HP + current SOC.
Where does AEMD fit in this? Any thoughts?
Not many people on this board unfortunately. This is such a strong company with a product which can be applicable to many fields of medicine - we need some momentum on this board.
HCV- New drugs - Telaprevir and Boceprevir maybe approved soon.
Standard therapy for HCV which is Pegylated alpha i nterferon (PEG) plus Ribavarin (RBV) will stay and the new drugs will be added to improve the Sustained Viral Response (SVR). The therapy still is atleast 24 weeks and costs approx $35,000.
Exosomes as Biomarker Treasure Chests for Prostate Cancer
Diederick Duijvesz a, Theo Luider b, Chris H. Bangma a, Guido Jenster a .
Accepted 20 December 2010, Published online 29 December 2010, pages 823 - 831
Exosomes as biomarker treasure chests
The molecular content of exosomes is dependent on their cell origin and strongly associates with the original cellular conditions [57]. Therefore, the identification of tissue- or disease-specific exosomal proteins and RNAs will enable the use of these vesicles as a source of new biomarkers. Since the late 1990s, an increasing number of studies have investigated exosomes’ protein content and their potential diagnostic and prognostic values in various types of cancer, resulting in a comprehensive database consisting of 64 papers and a total of 2400 different proteins [58]. All of these protein identifications have been obtained by mass spectrometry. In terms of RNAs, the first study on exosomes was performed in 2007 [28]. Using microarray technology, research has shown that exosomes from mouse-derived bone marrow cells contain messenger RNAs (mRNAs) and miRNAs. An increasing number of papers in which microarrays were used have described the potential role of proteins and miRNAs as diagnostic and prognostic tools [59], [60], and [61].
Until 2002, exosomes had been predominantly isolated and analysed from in vitro cell lines. More recent studies have shown that these vesicles can be isolated from body fluids, such as blood, urine, semen, amniotic fluid, malignant and pleural effusions, bronchoalveolar fluid, synovial fluid, saliva, and breast milk. These findings demonstrate that exosomes are present in all body fluids and can be used for determining health status [62].
Exosomes as a diagnostic and prognostic tool for prostate cancer
Diederick Duijvesz a, Theo Luider b, Chris H. Bangma a, Guido Jenster a .
Accepted 20 December 2010, Published online 29 December 2010, pages 823 - 831 European urology.
In terms of PCa, the reports on exosomes are very few. One of the firstExosomes as Biomarker Treasure Chests for Prostate Cancer
studies reported no apparent differences between exosomes of benign origin and malignant origin regarding their synthesis, storage, and release [63]. Most likely, these vesicles differ in biochemical properties. Unfortunately, to date, no high-throughput techniques such as mass spectrometry and microarray have been used to evaluate the differences between exosomes of benign origin and malignant origin to identify new biomarkers. Four studies used mass spectrometry to profile exosomes derived from PCa cell lines, xenografts, and metastases. To search for PCa-secreted proteins, serum from PCa-xenografted mice was analysed by mass spectrometry [25], and [26]. All of the identified proteins were screened for human-specific sequences by extensive database searching. The proteins containing human-specific sequences were of PCa origin. Interestingly, the subcellular localisation of most of these proteins is cytoplasmic, supporting the idea that these proteins are secreted in mouse blood through exosomes. Indeed, proteomic profiling of exosomes derived from human PCa cell lines confirmed the presence of almost all of the previously identified serum proteins [26]. Two other studies analysed vesicles from prostate cell lines and vertebral PCa metastases by mass spectrometry and identified proteins related to angiogenesis, signal transduction pathways, and cancer progression [51], and [64], including caveolin-1, Akt, pyruvate kinase M2, programmed cell death 6 interacting protein, and poly(A)-binding protein 1. Subsequent in vitro functional assays (eg, migration and proliferation assays) demonstrated that these vesicles can influence cancer microenvironment and promote cancer progression. Although these findings are promising, further investigations are needed to fully elucidate the role of PCa exosomes in cancer development.
Aside from these biological studies, exosomes and exosomal content from patient samples have also been evaluated for their potential as biomarkers. Urinary exosomes from 10 organ-confined PCa patients undergoing hormonal therapy prior to radical radiotherapy were analysed [53]. Other than a considerable variation in the quantity of total exosomal proteins, no difference was observed between healthy men and PCa patients. Although these results do not specify which proteins are present in exosomes, it emphasises the technical feasibility of assessing exosomal proteins to evaluate the clinical status of PCa. However, better sample preparation, such as immunoaffinity isolation, and more robust technical approaches are needed to define significant differences with such huge variation.
RNA expression analysis of urine-derived and PCa cell line–derived exosomes revealed that the known RNA markers for PCa, such as the TMPRSS2:ERG fusion gene and prostate cancer antigen 3 (PCA3), can be detected in exosomes by reverse transcriptase–polymerase chain reaction [10]. The TMPRSS2:ERG fusion transcripts were detected in urinary exosomes from two patients with high Gleason scores but not in those from two patients with low Gleason scores [45]. PCA3 mRNA was detected in exosomes derived from all patients. Interestingly, none of the hormone-treated patients showed detectable levels of TMPRSS2:ERG or PCA3 RNAs, suggesting that the response to treatment might reduce the size of PCa tissue and thereby decrease the expression levels of these androgen-responsive genes.
Exosomes drive anticancer immunosuppression - the last paragraph, in bold, emphasizes the role of exosomes in immunosuppression. It would be difficult to develop different drugs targeting all the possible exosomes - but having the Hemo Purifier to take care of all of them would be much more economical.
25 Jan 2010
Myeloid-derived suppressor cells (MDSCs) are immature immune cells with the ability to suppress T-cell activation. This cell population accumulates in patients with tumours and is a major suppressor of antitumour immunity. Exosomes produced by tumour cells contain T-cell activating antigens as well as having immunosuppressive powers. New research has identified a mechanism by which exosomes promote immunosuppression through the activation of MDSCs with the heat shock protein, Hsp72.
Myeloid-derived suppressor cells are a population of immature myeloid cells, which in humans are characterised by their CD11b+CD33+HLA-DR– nature. In humans and mice with tumours this cell population is enlarged, and activated MDSCs are able to induce antigen-specific MHC class I-restricted tolerance. This is in contrast to naïve cells which lack immunosuppressive properties, suggesting that MDSCs require activation signals from tumour cells to support their suppressive function.
Tumour induced MDSC activation and expansion is known to be regulated by soluble factors, such as Stat3, but also by microvesicles – exosomes. Exosomes are endosome-derived organelles which are secreted through exocytosis and are used for intracellular crosstalk and receptor release in both normal and pathologic conditions. Tumour-derived exosomes were initially described as being immunostimulatory due to their containing T-cell stimulating tumour antigens, but recent reports have suggested that they may have a role in immunosuppression via inhibiting T-cell function.
A study by researchers in Dijon, France published online in the Journal of Clinical Investigation has identified a mechanism by which tumour-derived exosomes can promote MDSC immunosuppression. They found that Stat3, a transcription factor known to promote tumour cell survival, was activated in MDSCs in response to tumour-derived exosomes. The activation of Stat3 promoted the immunosuppressive function of MDSCs without triggering population expansion. The authors found that tumour-derived exosomes express Hsp72, a major heat shock protein known to inhibit apoptosis. Hsp72 is also a ligand for TLR2, and exposure of MDSCs to Hsp72 on exosomes leads to TLR2/MyD88 dependent signalling resulting in production of IL-6, a well known activator of Stat3. Importantly they found that decreasing tumour exosome release pharmacologically in vivo, with inhibitors, in mice enhanced the efficacy of the chemotherapeutic drug cyclophosphamide.
The work indicates that tumour induced immunosuppression depends partly on the tumours ability to induce functional MDSCs by releasing Hsp72-expressing exosomes. The work suggests that drugs that interfere with exosome release may enhance the efficacy of current chemotherapy agents.
Reference
F Chalmin, S Ladoire, G Mignot, J Vincent, M Bruchard, J-P Remy-Martin, W Boireau, A Rouleau, B Simon, D Lanneau, A De Thonel, G Multhoff, A Hamman, F Martin, B Chauffert, E Solary, L Zitvogel, C Garrido, B Ryffel, C Borg, L Apetoh, C Rébé, F Ghiringhelli (2010) Membrane-associated Hsp72 from tumor-derived exosomes mediates STAT3-dependent immunosuppressive function of mouse and human myeloid-derived suppressor cells. J Clinical Investigation Online Early
From RedChip Blog
Showcasing Hemopurifier for U.S. government officials: In January, Aethlon was invited by the Department of Health and Human Services (HHS) to present its Hemopurifier technology platform before multi-agency health officials. This presentation led to follow-on meetings and presentations, including a formal presentation to the Biomedical Advanced Research and Development Authority (BARDA) in February.
If recent data is any indication, Aethlon has ample opportunities to participate in government-funded biodefense programs. Freemind Group, a consultancy firm focused on governmental grant identification and application assistance for the biotechnology industry, recently reported that there has been a dramatic surge in biodefense and infectious disease funding opportunities from the U.S. government. They forecast that funding applications submitted during the months of May, June and July will result in cumulative awards of approximately $500 million. The Hemopurifier has potential applications across a wide range of infectious diseases, giving Aethlon a unique advantage over biotechnology companies whose technology platforms are designed to treat only a single disease
From Cardiff Univ.
Exosomes in Cancer Immunology
Introduction
Exosomes are small vesicles (of around 40-90nm diameter), originating from within multivesicular bodies, which are subsequently secreted into the extracellular space. Exosomes are produced my most/all cell types, and have a molecular phenotype which largely reflects that of the parent cell. The exosome membrane is lipid-raft like, comprising a host of transmembrane and GPI-linked molecules. The lumen is also complex, and may act as a means of delivering molecules from cell to cell.
Exosomes express a complex array of membrane and intralumenal proteins. There may also be mRNA encapsulated within the vesicle, which may be functionally important as can be seen in the above picture.
Research Interests
Exosomes and Cancer Immunology:
Exosomes produced by cancer cells are compositionally different from those produced by non-cancer cells, in that they express tumour-associated antigens. Exosomes may be an efficient mode for delivering these antigens to dendritic cells, and as such may be useful as cancer vaccines. Because cancer-exosomes are very complex, we have also been investigating the possible inhibition of correct immune function(s) driven by exosomes, and our research is helping to define exosomes as a major mechanism of cancer immune evasion. We are currently focussing on exosomes isolated from prostate cancer, and pleural malignant mesothelioma. The following picture shows Prostate Cancer Cells (LNCap) in culture.
Exosomes in cell to cell communication:
The surface of exosomes is covered in molecules that may function in delivering signals to target cells. Integrins, for example, expressed by exosomes from diverse cell types may serve such a function. Under conditions that mimic inflammation, some cells (including fibroblasts), express elevated levels of integrin ligands (e.g. ICAM-1). Exosomes are capable of stimulating Calcium Signalling in such cells, through integrin dependent adhesion. Collaborators in such studies include Dr Maurice Hallett, Neutrophil Signalling Group, Dept of Surgery, and Dr Robert Steadman, Institute of Nephrology. Exosome coated beads (yellow) triger transient Calcium flux (increasing green) in fibroblasts following exosome-fibroblast contact are shown in the following picture.
Exosomes as Cancer Biomarkers:
Recent investigations by us and others have shown exosomes can be isolated from body-fluids including urine. It may be that analysis of the protein repertoire of such readily obtained exosomes will provide information that is clinically useful, about disease status (diagnosis, and monitoring). Current studies to evaluate exosomes in bladder cancer are underway, with collaborators; Dr John Staffurth, Oncology and Dr Ian Brewis, under the auspices of the Cardiff Proteomics Centre . Silver stained gel showing purification of urinary exosomes can be seen below.
From the American Soc. of Cl. Oncology Meeting 2010 - This is what AEMD is also focusing on! Read the conclusion.
Circulating exosomes may provide a more sensitive platform to monitor disease progression compared to circulating tumor cells.
Meeting:
2010 ASCO Annual Meeting
Abstract No:
10580
Citation:
J Clin Oncol 28:15s, 2010 (suppl; abstr 10580)
Author(s):
T. L. Pawlowski, D. Spetzler, T. Tinder, J. Kimbrough, T. Deng, J. Kim, P. Ellis, A. Tyrell, P. Kennedy, C. Kuslich; CarisDx, Phoenix, AZ
Abstract:
Background: Circulating tumor cells (CTCs) have been used to monitor disease progression in patients with different types of metastatic cancer. However only 50% of metastatic breast, 57% of metastatic prostate, and 18% of metastatic colon cancer blood specimens have adequate levels of CTCs for clinical laboratory analysis. Levels of exosomes have recently been shown to correlate with tumor progression. Methods: Exosomes from 1 ml of plasma were isolated by ultracentrifugation. CD-81 antibodies were used to capture and measure the exosome level of breast cancer samples (n=14) and healthy controls (n=4). CTCs were measured for all samples using the Cell Search CTC test protocol. Subsequently, EpCam positive exosomes were captured from metastatic breast (n=10), prostate (n=2), and colon cancer (n=3) samples, and compared to healthy controls (n=7). RNA was extracted from these EpCam positive exosomes and microRNA-21 (miR-21) expression was quantified by a qRT-PCR. Results: The preliminary study of breast cancer samples established 11 of the 14 samples (78.6%) had CD-81 specific exosome levels significantly above the level found in the 4 healthy samples (p=0.002). Only 7 of the 14 (50%) specimens analyzed had more than 5 CTCs, the clinical threshold for metastatic breast cancer. Three cancer samples had CD-81 measured exosome levels below the average of normal samples, one of these had >5 CTCs. miR- 21 analysis of 15 additional metastatic cancer specimens, 5 of which had >5 CTCs, found miR-21 averaged 4.2 x 106, 4.82 x 106 and 5.05 x 106 copies in the breast, prostate, and colon cancer samples respectively. Conversely, the plasma specimens from healthy donors collected in EDTA tubes averaged 1.8 x 104 copies of miR21.
Conclusions: This study suggests that exosome analysis from plasma samples may offer a greater opportunity to monitor and track disease than CTC analysis, while acknowledging that further study will be necessary to establish clinical definitions of exosome load. Furthermore, tumor-derived exosomes provide the ability to characterize tumor of origin miR content, which demonstrates additional opportunities for tumor-specific exosome-based biomarker analysis from a blood sample.
Exosomes and nasopharangeal cancers
Human tumor virus utilizes exosomes for intercellular
David G. Meckes, Jr.a,
Kathy H. Y. Shaira,
Aron R. Marquitza,
Che-Pei Kungb,
Rachel H. Edwardsa, and
Nancy Raab-Trauba,b,1
+ Author Affiliations
aThe Lineberger Comprehensive Cancer Center, and
bDepartment of Microbiology–Immunology, University of North Carolina, Chapel Hill, NC 27599
Edited* by Elliott Kieff, Harvard Medical School and Brigham and Women's Hospital, Boston, MA, and approved October 15, 2010 (received for review September 21, 2010)
Abstract
The Epstein–Barr virus (EBV) latent membrane protein 1 (LMP1) is expressed in multiple human malignancies and has potent effects on cell growth. It has been detected in exosomes and shown to inhibit immune function. Exosomes are small secreted cellular vesicles that contain proteins, mRNAs, and microRNAs (miRNAs). When produced by malignant cells, they can promote angiogenesis, cell proliferation, tumor-cell invasion, and immune evasion. In this study, exosomes released from nasopharyngeal carcinoma (NPC) cells harboring latent EBV were shown to contain LMP1, signal transduction molecules, and virus-encoded miRNAs. Exposure to these NPC exosomes activated the ERK and AKT signaling pathways in the recipient cells. Interestingly, NPC exosomes also contained viral miRNAs, several of which were enriched in comparison with their intracellular levels. LMP1 induces expression of the EGF receptor in an EBV-negative epithelial cell line, and exosomes produced by these cells also contain high levels of EGF receptor in exosomes. These findings suggest that the effects of EBV and LMP1 on cellular expression also modulate exosome content and properties. The exosomes may manipulate the tumor microenvironment to influence the growth of neighboring cells through the intercellular transfer of LMP1, signaling molecules, and viral miRNAs.
From the International worshop in Paris Jan/2011
http://janlotvall.wordpress.com/2011/01/22/exosomes-are-amazing-%e2%80%93-%e2%80%9cone-of-the-most-important-biologic-regulatory-mechanisms%e2%80%9d/
Exosome meeting - Int. Workshop/ Sweeden / 2012
http://janlotvall.wordpress.com/2011/03/19/exosome-meeting-in-2012-gothenburg-sweden/
http://janlotvall.wordpress.com/2011/04/06/finding-consensus-on-exosome-methodology/
Exosomes research is gaining momentum. This is an interesting blogspot.
Some info on exosomes in cancer esp. melanoma - AEMD is doing some research on malignant melanoma also.
The role of tumor exosomes in cancer progression is recently emerging, although initial data pointing at these organelles as carriers of tumor antigenic material for DC-mediated T cell cross-priming have supported clinical attempts to use tumor exosomes as anti-cancer vaccines [11]. However, growing evidence concerning a vast array of suppressive effects exerted by these microvesicles on different components of the immune system is clearly supporting the involvement of tumor exosomes in disease progression [3], [12]. In particular, we and others have recently shown that exosomes secreted by human tumor cells of various origins are able to induce apoptosis in activated T cells, through the expression of death ligands (e.g. FasL, TRAIL) [9], [10], [13], inhibit NK functions [14], [15] and promote the generation of myeloid-derived suppressor cells from normal monocytes [10]. These data, together with the reproducible evidence that exosomes of likely tumor origin can be abundantly found in plasma and neoplastic effusions of cancer patients [16]–[18] support a role of tumor exosomes in molding host microenvironment to allow tumor growth and progression [19], [20].
However, the study of the in vivo role of tumor exosomes has been so far penalized by the lack of suitable methods to quantify exosomes from human body fluids, particularly from plasma of cancer patients. The aim of our study was thus to provide a method to detect and quantify exosomes from small amount of human plasma, with the final goal of identifying a tool for assessing the role of tumor exosomes as potential tumor marker and prognostic factor. This might be particularly relevant in melanoma patients, in which sensitive and reliable serum markers are unfortunately still limited while serum LDH (lactate dehydrogenase) levels remain the only prognostic serum factor for assessing disease course and prognosis [21], [22]. Here, we describe an in-house ELISA that allows quantification and characterization of exosomes from different samples, including plasma from tumor-bearing animals and melanoma patients, as well as from tumor cell culture supernatants. These findings suggest that the detection of tumor exosomes in plasma of cancer patients may represent a potential biomarker in the clinical monitoring of tumor malignancies, in particular melanoma.
High Levels of Exosomes Expressing CD63 and Caveolin-1 in Plasma of Melanoma Patients
Mariantonia Logozzi1, Angelo De Milito1, Luana Lugini2, Martina Borghi1, Luana Calabrò3, Massimo Spada4, Maurizio Perdicchio1, Maria Lucia Marino1, Cristina Federici1, Elisabetta Iessi1, Daria Brambilla1, Giulietta Venturi1, Francesco Lozupone1, Mario Santinami5, Veronica Huber6, Michele Maio3,7, Licia Rivoltini6, Stefano Fais1*
1 Unit of Antitumor Drugs, Department of Therapeutic Research and Medicines Evaluation, Istituto Superiore di Sanità, Rome, Italy, 2 Unit of Molecular and Cellular Imaging, Istituto Superiore di Sanità, Rome, Italy, 3 Division of Medical Oncology and Immunotherapy, Department of Oncology, University Hospital of Siena, Istituto Toscano Tumori, Siena, Italy, 4 Unit of Experimental Immunotherapy, Department of Cell Biology and Neurosciences, Istituto Superiore di Sanità, Rome, Italy, 5 Unit of Melanoma and Sarcoma, Fondazione IRCCS Istituto Nazionale Tumori, Milan, Italy, 6 Unit of Immunotherapy of Human Tumours, Fondazione IRCCS Istituto Nazionale Tumori, Milan, Italy, 7 Unit of Cancer Bioimmunotherapy, Department of Medical Oncology, Centro di Riferimento Oncologico IRCCS, Aviano, Italy
Sepsis and vascular dysfunction
Platelet-derived exosomes induce endothelial cell apoptosis through peroxynitrite generation: experimental evidence for a novel mechanism of septic vascular dysfunction.
Gambim MH, do Carmo Ade O, Marti L, Veríssimo-Filho S, Lopes LR, Janiszewski M.
Source
Division of Rheumatology, University of São Paulo School of Medicine, Avenida Doutor Arnaldo, 455, 01246-903, São Paulo, SP. mgambim@gmail.com
CONCLUSION:
We showed that, in sepsis, NO and bacterial elements are responsible for type-specific platelet-derived exosome generation. Those exosomes have an active role in vascular signaling as redox-active particles that can induce endothelial cell caspase-3 activation and apoptosis by generating superoxide, NO and peroxynitrite. Thus, exosomes must be considered for further developments in understanding and treating vascular dysfunction in sepsis.
PMID: 17894858 [PubMed - indexed for MEDLINE] PMCID: PMC2556756
Sepsis and Exosomes
Platelet-derived exosomes from septic shock patients induce myocardial dysfunction.
Azevedo LC, Janiszewski M, Pontieri V, Pedro Mde A, Bassi E, Tucci PJ, Laurindo FR.
Source
Emergency Medicine Research Laboratory, University of São Paulo School of Medicine, Av. Dr. Enéas de Carvalho Aguiar 255, sala 5023, São Paulo, Brazil. lucianoazevedo@uol.com.br
Abstract
INTRODUCTION:
Mechanisms underlying inotropic failure in septic shock are incompletely understood. We previously identified the presence of exosomes in the plasma of septic shock patients. These exosomes are released mainly by platelets, produce superoxide, and induce apoptosis in vascular cells by a redox-dependent pathway. We hypothesized that circulating platelet-derived exosomes could contribute to inotropic dysfunction of sepsis.
METHODS:
We collected blood samples from 55 patients with septic shock and 12 healthy volunteers for exosome separation. Exosomes from septic patients and healthy individuals were investigated concerning their myocardial depressant effect in isolated heart and papillary muscle preparations.
RESULTS:
Exosomes from the plasma of septic patients significantly decreased positive and negative derivatives of left ventricular pressure in isolated rabbit hearts or developed tension and its first positive derivative in papillary muscles. Exosomes from healthy individuals decreased these variables non-significantly. In hearts from rabbits previously exposed to endotoxin, septic exosomes decreased positive and negative derivatives of ventricular pressure. This negative inotropic effect was fully reversible upon withdrawal of exosomes. Nitric oxide (NO) production from exosomes derived from septic shock patients was demonstrated by fluorescence. Also, there was an increase in myocardial nitrate content after exposure to septic exosomes.
CONCLUSION:
Circulating platelet-derived exosomes from septic patients induced myocardial dysfunction in isolated heart and papillary muscle preparations, a phenomenon enhanced by previous in vivo exposure to lipopolysaccharide. The generation of NO by septic exosomes and the increased myocardial nitrate content after incubation with exosomes from septic patients suggest an NO-dependent mechanism that may contribute to myocardial dysfunction of sepsis.
Comment in
Crit Care. 2008;12(1):110.
Even if they do not get the DARPA funds(seems unlikely as they have all the right ingredients), I consider the float too low - they have room for more dilution. The potential is so huge that 10-20 million dilution in the big picture, provided ofcourse that the HP does what it is supposed to, maybe nothing in the next few years.
Death spiral? If they get the DARPA funds , they will get a tremendous amount of recognition with big pharma and then funds should be easy to comeby.
Lot of applications with one product - big pharma is currrently looking for new products. Aemd may have an answer to a lot of diseases.
The company has tremendous potential.