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great article cjg,
This was nice to read -
However, because of the expedited process in developing products under the government's animal efficacy rule, the viral hemorrhagic fevers indication potentially could be the first approved use for Bavituximab. The FDA's animal efficacy rule, finalized in May 2002, applies to the development & testing of drugs that reduce or prevent serious and life-threatening conditions caused by exposure to lethal or permanently disabling toxic agents or diseases where human efficacy trials are not feasible or ethical, like in the case of Ebola or other viral hemorrhagic fevers. Under the rule, drug regulators can rely on evidence from animal studies to provide substantial evidence of product effectiveness when there is a reasonably well-understood mechanism for the toxicity of the agent and its amelioration or prevention by the product and the effect is demonstrated in more than one animal species expected to react with a response predictive for humans or a well-characterized animal species model. "It would be virtually impossible to run standard Phase I, II, III studies on hemorrhagic fevers, because the incidence is so low under normal circumstances that to enroll patients would not be practical," King explained.
The DTRA development program, he said, "naturally dovetails" with Peregrine's ongoing development efforts for bavituximab in treating viruses. Part of the biodefense contract funds, King said, will aid Peregrine in scaling up its production and testing for the antibody, which he said will assist in advancing bavituximab's other clinical areas. The other portion of the funding is geared directly toward development and testing specifically for the viral hemorrhagic fevers indication, he added. King said it is expected that the military may stockpile Bavituximab as part of its biodefense program, which potentially could result in a long-term partnership for Peregrine with the government and future deals.
---------
j
Impec,
I've read quite a few good pertinent papers lately, but haven't posted them yet since they'd get buried in the recent post frenzy. The primary important thing that's been coming to light is that immunosuppression is coming to be seen as occurring less on a cellular level, and more on a "vesicular" level. I think we'll read more about this new perspective in future Bart Haynes papers.
and yes - I think Peregrine's anti-PS platform will take center-stage in terms of therapeutics that address this microvesicle-facilitated immunosuppression / (PS-facilitated pathogenesis).
j
Paul,
You should substitute "Haynes" for "jazz" in your statement.
Haynes clearly believes the role of PS in disease is a major discovery.
I'm just following it all. It's incredibly exciting, and it's been coming together very quickly in the past few months with the Haynes patent applications showing up on the USPTO & WIPO databases.
j
NIAID Director Anthony Fauci, QUOTE from his Mexico City speech last week -------
"A recent paper just a few days ago from Bart Haynes’ group in CHAVI and Duke showed that not only is a reservoir formed early, but byproducts of CD4 positive T-cell deaths increase significantly within days and are capable of suppressing the human immune response to the virus. So, we have a double whammy. We have a reservoir that almost immediately is formed and we have products of the death of cells suppressing the immune response that would hopefully prevent the establishment of that reservoir.
---------
j
FAUCI in Mexico City ---------
NIH’s Fauci Finds Hope Amid Challenges in AIDS Research
by Marilyn Chase
The NIH’s Anthony Fauci, who once cautioned that there might never be a traditional vaccine to prevent HIV infection and recently pulled the plug on a troubled vaccine trial, sounded a cautiously optimistic note at the 17th International AIDS Conference yesterday.
“The future for AIDS research looks bright and promising,” said Fauci, singling out recent work by Barton Haynes of Duke and Robert Siliciano of Johns Hopkins in illuminating how the virus hides inside the body and suppresses the immune system within days of infection.
--------
j
jess,
Yes, it's the talk of the town.
Haynes and colleagues' discovery that PS-exposing microparticles are what is responsible for shutting down the B cell (antibody) response to the virus was a very important discovery in the history of HIV research. It changes everything. It explains a lot. It got Tony Fauci optimistic. It may have 'raised' the (HIV vaccine) hurdle, but it finally much more clearly described the hurdle.
From reading my posts over the past couple years, you should already know that PS shifts the response of macrophages, altering the cytokine environment, blunting the maturation and antigen-presenting function of dendritic cells, resulting in a weak T cell response.
But Haynes and team have now shown (as explained in the paper in this month's Virology) that PS also destroys the B cell response. That was a VERY important discovery.
When the Director of the Duke Human Vaccine Institue, who happens to be the overall Director of CHAVI, who happens to be a principle investigator in CAVD, who happens to have given the plenary presentation at last year's Gates HIV vaccine conf. (where he spoke about PS-exposing microparticles as the immunosuppressive culprit in early HIV infection :), who happens to be the most-funded NIH research scientist in the country, - When he says that exposed PS is responsible for stopping the antibody (B cell) response to HIV infection, yes, it's gotten a lot of people talking.
Further, when he offers up SOLUTIONS, in patent applications, like saying that inducing anti-PS abs via a vaccine is needed, to block the overwhelming immune-suppression caused by PS-exposing microparticles, and also saying that anti-PS mabs are safe therapy for people already infected with the virus, it's pretty important, and yes, we can obviously assume it will be detailed in a published paper as to WHY he is saying that anti-PS, specifically what appears to be Peregrine's anti-PS mab, can be safe therapy for those already infected.
More importantly, when every single anti-PS mab he's mentioned in all of the recent papers and patent applications ALL appear to be Peregrine's, (via Thorpe and Chen, Peregrine's two anti-PS Core Technology contributors), it makes me very happy.
j
thanks. I think it points to a new unifying model of disease based on apoptotic mimicry.
j
Barton Haynes Recently Published Papers and Patent Applications
Haynes recent paper in the August issue of the journal Virology has generated much discussion in the HIV research community, as well as
several popular news articles.
Examples of follow-on popular news stories:
HIV Conquers Immune System Faster Than Previously Realised
http://www.docguide.com/news/content.nsf/news/852571020057CCF68525748A00661CD7
HIV Overpowers Immune System Quicker than Previously Thought
http://www.genengnews.com/news/bnitem.aspx?name=38938669
These popular articles do a nice job of summarizing the basic insights found by the Haynes team which shed light on what is responsible
for the rapid immune cell death found in early HIV infection. Haynes paper as well as his several patent
applications that have recently appeared in the various US and international patent databases provide the all-important details. Haynes work
discusses exposed phosphatidylserine (PS) on microparticles overwhelming immune cells, facilitating the rise in viral counts.
The following information details this fascinating area of research, and points to this PS-induced immune suppression pathway as being an
important mechanism involved in the pathogenesis of a broad range of diseases.
First, some background
Recent insights into enveloped virus infection, from the journal Science
In the April 2008 journal Science, authors Mercer and Helenius discuss data showing how vaccinia viruses (the pox family of viruses), utilize
phosphatidylserine (PS) exposed on its surface to facilitate entry into cells. Besides PS aiding viral entry, the authors also discuss recent
research showing that exposed PS also prevents an immune response to the virus. They use the term "apoptotic mimicry", (masquerading
as a dying native cell), a term showing up more often in recent research involving how various unrelated pathogens use similar means to
evade the cells of the immune system, thereby surviving and thriving in the body.
Vaccinia Virus Uses Macropinocytosis and Apoptotic Mimicry to Enter Host Cells
http://www.ncbi.nlm.nih.gov/pubmed/18436786?ordinalpos=1&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVDocSum
short quotes from the paper in Science -
“The induction of blebs, the endocytic event, and infection were all critically dependent on the presence of exposed phosphatidylserine in the
viral membrane, which suggests that vaccinia virus uses apoptotic mimicry to enter cells.”
“The Mature Virion membrane has been shown to be enriched in PS and PS is required for infectivity”
The Mercer and Helenius paper spawned several well-written layman's articles on the new insight into viral infection. For instance:
Trojan Horse Of Viruses Revealed
http://www.sciencedaily.com/releases/2008/04/080425065354.htm
excerpts:
ScienceDaily (Apr. 25, 2008) — Viruses use various tricks and disguises to invade cells. Researchers have now discovered yet another
strategy used by viruses: the vaccinia virus disguises itself as cell waste, triggers the formation of evaginations in cells and is suspected to
enter the cell interior before the immune defense even notices.
The invasion strategy
In order to infiltrate a cell, the vaccinia virus exploits the cellular waste disposal mechanism. When a cell dies, other cells in the vicinity ingest
the remains, without needing waste disposal experts such as macrophages. The cells recognize the waste via a special molecule,
phosphatidylserine, which sits on the inner surface of the double membrane of cells.
This special molecule is pushed out as soon as the cell dies and is broken into parts. The vaccinia virus itself also carries this official waste
tag on its surface.
"The substance accumulates on the shell of vaccinia viruses", Jason Mercer explained. The pathogen disguises itself as waste material and
tricks cells into digesting it, just as they normally would with the remains of dead cells. As the immune response is simultaneously
suppressed, the virus can be ingested as waste without being noticed.
Dirty Rotten Poxviruses
http://sciencenow.sciencemag.org/cgi/content/full/2008/425/1
excerpts:
"poxviruses have engineered a way to sneak into cells through the garbage chute."
"the virus disguises itself as junk"
"the team wondered if the virus was playing dead. They found that the virus's surface was studded with phosphatidylserine, a lipid that also
flags dead cells as garbage.
Removing lipids from the virus's surface stopped infection, and recoating the virus with phosphatidylserine
restarted it.
The results suggest that the virus is "more clever than originally thought" because it exploits a garbage-collection process found
in almost all cells, says Mercer.
before proceeding further, two words to understand -
Phosphatidylserine: A phospholipid found on the inside of healthy cell walls. As our cells die, they lose the ability to maintain
phosphatidylserine (PS) on the inside of the cell wall. PS becomes exposed on our dying cells, and is perceived by cells of the immune system as native / "self" debris, not as a foreign pathogen.
Microparticles: Small vesicles shed from dying cells. Microparticles are made of cell membrane phospholipid molecules. Microparticles
expose PS.
A brief primer on phosphatidylserine's role in cell death and immune clearance
As multicellular organisms, we need an immune system that is flexible enough to react differently to foreign invaders as compared to
dying native cells. Over the past decade, researchers have shed light on the "fork in the road" which dictates which general type of response
our immune system pursues. Phosphatidylserine (PS) is a lipid typically found lining the interior of all healthy cell walls. PS flips to the
exterior of the cell membrane as a result of various stresses, and is an early and universal sign of a dying native cell. It is not surprising
that exposed PS has recently been shown to provide the fundamental signal for an immune response steered toward
homeostasis, a self / maintenance type of immune response. Recent data now illustrates that
exposed PS is the impetus that alters the behavior of all the major cell types of the immune system into NOT mounting a specific attack.
Nature finds a way
Scientists have also recently discovered a commonality between cancer, viral, and protozoan
parasitic infections. These distinctly different pathogens all exploit exposed PS to aid their survival and proliferation in the body. This
common strategy is no surprise when considered with the new understanding of the evolution of metazoan PS-mediated immune
suppression. Taken together, these recent discoveries suggest that successful pathogens independently evolved to exploit a similar
evasion mechanism because it provides the crucial advantage (exposed PS) needed to avoid the immune system's attack.
Where does Barton Haynes fit in?
Over the past couple years, in his work looking into the pathogenesis of HIV, Barton Haynes has been following the hypothesis that the
reason HIV manages to proliferate so quickly after infection, and the reason no vaccines have yet been effective, have both been due to a
massive amount of "apoptotic debris" (microparticles from dying cells) which occurs early in HIV infection. This debris contains exposed
phosphatidylserine (PS), and is responsible for the depletion of CD4+ T cells, as well as an overall blunting of immune response to the virus.
Haynes puts forth the idea (documented below) that the fundamental culprit is phosphatidylserine (PS). He cites recent research (Hoffman et al, Interaction between Phosphatidylserine and the Phosphatidylserine Receptor Inhibits Immune
Responses In Vivo. The Journal of Immunology, 174: 1393-1404), showing how exposed PS alters the function of macrophages, dendritic cells, and T cells. In the August Journal of Virology he
also provides new data showing how microparticles with exposed PS alter the fuctions of antibody producing B cells.
here are earlier excerpts illustrating Haynes/CHAVI overall working hypothesis:
Haynes / CHAVI / Gates hypothesis example 1:
"Their [Haynes group] current hypothesis is that HIV induces a massive apoptosis before and during viral ramp up, and that plasma
microparticles (fragments of apoptotic CD3 and T cells) have a suppressive effect on Ab generation, thereby amplifying the apoptotic
cascade. "
May 22, 2007
http://www3.niaid.nih.gov/research/topics/HIV/vaccines/advisory/avrs/PDF/AVRS_May07_Summary.pdf
Haynes / CHAVI / Gates hypothesis example 2:
Conclusion: These results demonstrate that at the time of viral load ramp-up in acute HIV-1 infection, there are elevations in plasma levels of
TRAIL, FAS Ligand, and TNFR2 that were associated with the presence of microparticles from apoptotic T cells. The presence of these
apoptotic markers suggests apoptosis occurring at the time of initial HIV-1 viral load ramp-up.
That PS+ apoptotic cells and microparticles have been reported to suppress antigen specific immune responses suggests the hypothesis
that immune cell apoptosis in the very earliest stages of acute HIV-1 infection may delay the onset of potentially protective anti-HIV-1
immune responses."
August 20, 2007
http://www.hivvaccineenterprise.org/_dwn/poster_sessions.pdf
Haynes recent publication
The following are excerpts from Bart Haynes' new paper in the August 2008 Journal of Virology, which discuss detailed data showing how
exposed PS on microparticles is responsible for the weak immune response to HIV.
Induction of plasma (TRAIL), TNFR-2, Fas ligand, and plasma microparticles after human immunodeficiency virus type 1 (HIV-1)
transmission: implications for HIV-1 vaccine design.
http://jvi.asm.org/cgi/content/abstract/82/15/7700
Excerpts:
("MP" = MicroParticles)
“In this study, we raise the hypothesis that in addition to gut CD4 T cell loss, delay in HIV-1 protective immune responses early on after
HIV-1 transmission may involve the production of elevated levels of immunosuppressive moieties such as TRAIL, TNFR2 and Fas ligand as
well as plasma microparticles.
Microparticles (MPs) are small membrane-bound vesicles that are released from the surface of apoptotic cells
by exocytic or budding processes; as such, MPs bear cell surface markers and can bind annexin V because of the expression of
phosphatidylserine (32-44, 39).
MPs, which circulate in the blood in many clinical conditions, are part of a spectrum of subcellular structures
that are released from cells and can be distinguished from exosomes which are released from multivesicular bodies during activation.
“MPs have immunomodulatory activities and can promote immune cell death; exosomes are also immunologically active, can suppress
immune responses (20,34,42,55), and have been reported elevated in chronic HIV-1 (4).”
“suppression of immune responses can be mediated by T cell MPs (32,34,35). CXCR4+ and CCR5+ MPs can transfer co-receptors to coreceptor
negative cells, making them susceptible to infection by HIV-1 (48,57).
Phagocytosis of MPs by macrophages releases TGF-beta,
prostaglandin E2 and IL-10 that can inhibit antigen specific T and B cell responses (20,35,42). In this regard, Estes et al. have shown
dramatic increases in lymph node TGF-beta and IL-10 on day 12 following SIV infection (22).
Importantly, we have demonstrated that PBMC
and tonsillar cell MPs can directly inhibit memory B cell activation (Figure 8).”
“it is likely that MPs are responsible for the observed B cell suppressive activity seen in vitro in Figure 8.
In the setting of HIV-1 infection
where both activation and apoptosis occur, however, MPs and exosomes may act concomitantly, with exosomes suppressing immune
responses (2,7,15,61), and MPs contributing to both immune suppression and cell death (20,32,34,35,39,42,55).”
August 2008, Journal of Virology
http://jvi.asm.org/cgi/content/abstract/82/15/7700
Where Does Haynes Go From Here? Patents Point The Way...
We can see where Haynes is going in his work by looking at two recent patent applications.
In one recent patent application, in the World Intellectual Property Organization database, Haynes discusses what a future successful
HIV vaccine must do. He specifically states that any future successful vaccine must stop exposed PS from blunting immune response. He
states that an essential goal of a future vaccine is to induce anti-PS antibodies, which would bind/block the PS from downregulating the
immune response.
In Haynes second recent patent application, in the US database, he discusses how monoclonal anti-PS antibodies can be safe therapy in
people already infected with HIV.
excerpts and links to the two recent patent applications:
The “Vaccine” Patent -
- Thus, HTV virions and HIV envelope can directly induce T cell death in AHI, soluble TRAIL can bind to uninfected cells and induce death in
AHI, and with both HTV infection of cells and with massive apoptosis, high levels of phosphatidylserine containing cells and particles likely
abound in AHI.
Phosphatidylserine
(PS) on the surface of HIV infected cells and virions has been found (Figure 7) and Callahan et al have found PS is a cofactor for HIV
infection of monocytes (Callahan, J. Immunol 170:4840 (2003)).
PS-dependent ingestion of apoptotic cells promotes TGF-βl secretion
(Huynh et al, J. Clin. Invest. 109:41 (2002)) and interaction between PS and PS receptor inhibits antibody responses in vivo (Hoffman et al,
J. Immunol. 174:1393 (2005)).
There
are increases in PS+ shed membrane particles in chronic HIV infection (Aupeix et al, J. Clin. Invest. 99:1546 (1997)), and apoptotic
microparticles modulate macrophage immune responses (Distler et al, Apoptosis 10:731 (2005)).
The present invention relates to a multicomponent vaccine that addresses problems resulting from the diversity of HIV by the use consensus
and/or mosaic HIV genes coupled with strategies designed to break immune tolerance to allow for induction of the desired specificity of
neutralzing antibodies at mucosal sites (e.g., through the use of T regulatory cell inhibition and/or TLR-9 agonist adjuvants), and strategies
designed to overcome HIV-I induced apoptosis (e.g., induction of anti- phosphatidylserine (PS) antibodies, anti-CD36 antibodies, and/or antitat
antibodies).
WHAT IS CLAIMED IS:
1. A method of inducing the production of an immune response against HIV-I in a mammal comprising administering to said mammal: i) a
centralized HIV-I gene sequence, ii) an agent that breaks mammalian immune tolerance, and iii) an agent that inhibits HIV-I -induced
apoptosis or an immunosuppressive effect of HIV-I -induced apoptosis, wherein (i), (ii) and (iii) are administered in amounts sufficient to
effect said production.
22. The method according to claim 1 wherein said agent that inhibits HIV-1-induced apoptosis induces anti-phosphatidylserine (PS)
antibodies, anti- CD36 antibodies, or anti-HIV tat antibodies.
http://www.wipo.int/pctdb/en/wads.jsp?IA=US2007024122&LANGUAGE=EN&ID=id00000006427540&VOL=87&DOC=0084d3&WO=08/063
586&WEEK=22/2008&TYPE=A2&DOC_TYPE=PAMPH&PAGE=1
The “therapeutic” patent
Here Haynes discusses the use of anti-phospholipid monoclonal antibodies as HIV therapy. Haynes has moved from seeing the lipid binding
characteristics of the well-known broadly-neutralizing HIV abs (2F5, 4E10) as unfortunate, and now sees the lipid-binding characteristics as
“the key” to broad and safe HIV neutralization. Haynes goes so far as to say that there is no need for an HIV neutralizing antibody to bind a
viral epitope. The idea is presented that binding a host-cell 'tag-along' phospholipid results in successful HIV neutralization.
WHAT IS CLAIMED IS:
1. A method of treating HIV comprising administering to a patient in need thereof an antibody derivable from a normal subject or from an
autoimmune disease subject that binds to a lipid on the surface of HIV or on the surface of HIV-infected cells and thereby neutralizes HIV-
1, wherein said antibody is administered in an amount sufficient to effect said treatment.
2. The method according to claim 1 wherein said antibody is derivable from an anti-phospholipid syndrome subject.
3. The method according to claim 1 wherein said antibody is non-pathogenic.
4. The method according to claim 1 wherein said antibody is IS1, IS4 or IS6, or binding fragment thereof.
5. The method according to claim 1 wherein said antibody is IS1, or binding fragment thereof.
[0105] ....That IS1 neutralized HIV evidences the facts that: a) humans can make non-pathogenic anti-lipid antibodies that neutralize HIV,
and b) IS1 is an antibody that can be safely used as a therapeutic Mab for treatment of HIV infected subjects or in the setting of postexposure
prophylaxis of subjects following needle, sexual or other exposure to HIV or HIV infected materials.
http://appft1.uspto.gov/netacgi/nph-Parser?Sect1=PTO2&Sect2=HITOFF&u=%2Fnetahtml%2FPTO%2Fsearchadv.
html&r=2&p=1&f=G&l=50&d=PG01&S1=%28%22haynes+barton%22.IN.%29&OS=in/%22haynes+barton%22&RS=IN/%22haynes+bart
on%22
The Big Picture
In the Haynes "therapeutic patent" above, one additional quote stands out:
[0051] It will be appreciated from a reading of the foregoing that if HIV has evolved to escape the host immune response by making the
immune system blind to it, other infectious agents may have evolved similarly. That is, this may represent a general mechanism of escape.
That being the case, approaches comparable to those described herein can be expected to be useful in the treatment of such other agents
well.
In their recent paper in Science, (the top of this post) Mercer and Helenius also speculate on this broad-based pathogenic immuneevasion
mechanism:
Also, in the same issue of Science, Fairn and Grinstein discuss the broader implications of Mercer and Helenius's findings:
A One-Sided Signal
http://www.sciencemag.org/cgi/content/full/320/5875/458
Gregory D. Fairn and Sergio Grinstein
excerpts:
"Because the infected cells undergo apoptosis, and thus experience scrambling of plasma membrane lipids, the budding virus also acquires
an envelope that exposes phosphatidylserine on its external surface"
"the presence of exofacial phosphatidylserine is required for viral entry. "
"The involvement of phosphatidylserine may not be limited to infection by vaccinia viruses."
"It is therefore conceivable that HIV similarly requires phosphatidylserine for infection"
"The role of phosphatidylserine in the entry of HIV and other viruses will surely be explored in greater detail now”
ADDENDUM - JULY 24, 2008:
On July 24, 2008, another Haynes patent application went public on the World Intellectual Property Organization database.
Pertinent excerpts follow:
"The time of appearance of antibodies in the development of acute HIV infection has been recently mapped and it has been shown that most
of the antibodies arise after a delay in the peak response to HIV envelope epitopes of approximately two to three weeks. Indeed, the most
protective antibodies, those that neutralize autologous virus, can be delayed for up to a year."
"To begin to understand the "delay" in induction of antibodies at the time of HIV transmission, the first question to be addressed was whether
there are immunosuppressive events, such as massive apoptosis, with release of phosphatidylserine microparticles at the time of viral load
ramp up during acute HIV infection."
"Apoptotic microparticles are the products of either activated or apoptotic cells, that are increased in the plasma of a number of diseases,
including autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis, Crohn's disease, coronary artery disease
and other forms of heart disease, and chronic HIV-I infection."
"Apoptotic microparticles can bind to non-apoptotic cells and induce apoptosis, are procoagulant, proinflammatory, and can be
immunosuppressive for T and B cell responses to specific antigen."
"Thus, the massive apoptosis that occurs with acute HIV infection with resulting release of TRAIL, mediation of apoptosis via FAS-FASL
interactions, and release of PS containing viral and other particles, all conspire to initially immuno suppress the host, preventing rapid
protective B cell responses."
http://www.wipo.int/pctdb/en/wads.jsp?IA=US2008000412&LANGUAGE=EN&ID=id00000006622149&VOL=89&DOC=00fca1&WO=08/0887
47&WEEK=30/2008&TYPE=A2&DOC_TYPE=PAMPH&PAGE=1
-------------------
j
have a look through here -
http://www.clinicaltrials.gov/ct2/results?term=HIV+AND+Duke
j
"The IS1 mab from Pojen is PPHM's per agreement."
Thanks jake for the extra corroboration.
Now can we add this to Haynes' quotes? -
PEREGRINE PHARMACEUTICALS IS1 mab "can be safely used as a therapeutic Mab for treatment of HIV infected
subjects".
PEREGRINE PHARMACEUTICALS IS1 mab can "broadly neutralize HIV in an unprecedented manner".
I like the sound of that.
Bonus Questions:
Q: How many of the 25 or so approved HIV drugs went through trials at Duke, on NIAID money?...
Q: How many of the current HIV therapeutic trials are sponsored by NIAID?
j
"I would consider that a brilliant move by PPHM management"
I consider it a brilliant move, and a smoking gun.
j
Thorpe's CRISP grant ended on January 31, 2008
The work was to conclude with primate work using the real Lassa Fever virus in the biosafety level 4 lab in Galveston TX.
It is now 6 months past the conclusion of Thorpe's big Lassa fever project.
I think we'll see a paper ;)
Actually, Paul Lytle included it on a slide in his NYC presentation:
Published Anti-viral papers from Thorpe/UTSW as well as Duke.
but- MORE TELLING IS THE FACT THAT
The DTRA contract with Peregrine Pharmaceuticals, which is for Lassa and other HFV's, awarded in June 2008, is obviously a good sign as to how Thorpe's work went!!!!
:)
IT'S HAPPENING,
j
----------------------------
Abstract
Grant Number: 5U01AI056412-05
Project Title: Novel Anti-Viral Agents for Treating Lassa Fever
PI Information: Name Email Title
THORPE, PHILIP E. philip.thorpe@utsouthwestern.edu
Abstract: DESCRIPTION (provided by applicant): Our goal is to develop new, broad-spectrum anti-viral agents for treating viral infections caused by hostile dissemination of viral pathogens. The new agents act by targeting normally-internal phospholipids (e.g. phosphatidylserine, PS) that become exposed on the surface of virally-infected cells. Exposure of these phospholipids on infected host cells appears to be necessary for viral maturation or egress. We have raised monoclonal antibodies to PS and other anionic phospholipids that block the spread of several viruses to uninfected cells in vitro, essentially completely. The viruses we have tested thus far include an arenavirus (Pichinde virus), a paramyxovirus (respiratory syncytial virus) and a herpes virus (cytomegalovirus). The phospholipids that they recognize have the same structure and cellular distribution in different mammalian species, simplifying the transition from experimental animals into humans. The antibodies are not toxic to mice, even when administered in high doses for prolonged periods of time. One of the antibodies has been converted into a human-mouse chimeric antibody (ch3G4) and is being produced in a GMP facility by a commercial partner in anticipation of human use. We propose to develop ch3G4 first for treating Lassa fever. Development studies will be conducted in a guinea pig model, which uses the arenavirus, Pichinde, to mimic Lassa fever in humans. The animal model will enable us to safely investigate the mechanism of the anti-viral effect and to establish basic efficacy, safety and pharmacokinetic parameters. Key findings will then be validated for Lassa fever virus itself and with other arenaviruses that cause hemorrhagic fevers in a BSL-4 containment facility at UTMB Center for Biodefense, Galveston. The information generated by these studies may also extend to other viruses that could be used for bioterrorism purposes.
Public Health Relevance:
This Public Health Relevance is not available.
Thesaurus Terms:
antiviral agent, biomaterial development /preparation, immunologic substance development /preparation, monoclonal antibody, phospholipid, virus disease
Lassa virus, cooperative study, disease /disorder model, drug screening /evaluation, pharmacokinetics
biotechnology, electron microscopy, enzyme linked immunosorbent assay, guinea pig, immunocytochemistry, northern blotting
Institution: UNIVERSITY OF TEXAS SW MED CTR/DALLAS
DALLAS, TX 753909105
Fiscal Year: 2007
Department: PHARMACOLOGY
Project Start: 15-AUG-2003
Project End: 31-JAN-2008
ICD: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
IRG: ZAI1
Duke / Gates CAVD / CHAVI is working towards an HIV vaccine.
Thorpe is officially involved under the Gates CAVD, not CHAVI.
(Although the most promising work of both organizations is now centered around Bart Haynes work which focuses on PS as the big bad immuno-suppressive culprit.)
In their work towards an HIV vaccine, Haynes & colleagues are using Peregrine's anti-PS mabs, therapeutically.
If the mabs work, in passive therapy experiments, (preventing HIV from blooming, or from blooming as much, thereby lowering the viral "set-point", or preventing infection all together), then Haynes will have achieved the step of clarifying one goal of an HIV vaccine.
In recently published patent applications, Haynes is now saying that in order for an HIV vaccine to be successful, it must interrupt PS-mediated immuno-suppressive signaling, which they proved is the culprit that has been preventing any previous experimantal viral immunogens from evoking effective T and B cell responses. To do this, Haynes proposes a vaccine that induces anti-PS abs.
Haynes vaccine work with Peregrine's anti-PS mabs have immediate implications for treating people already infected with HIV.
Haynes is now saying in his patent applications that:
* successful vaccines must induce anti-PS abs to block the overwhelming PS-mediated immunosuppression caused by a huge amount of rapidly accumulating shed debris from the membranes of infected T cells.
* anti-PS "can be safely used as a therapeutic Mab for treatment of HIV infected
subjects".
*anti-PS can "broadly neutralize HIV in an unprecedented manner".
Haynes has yet to publish any papers that would explain the above verbatim Haynes quotes.
Jazzbeerman concludes that Haynes is saying the above quotes in his recently published patent applications because he has performed experiments that cause him to conclude that:
* successful vaccines must induce anti-PS abs.
* anti-PS "can be safely used as a therapeutic Mab for treatment of HIV infected
subjects".
*anti-PS can "broadly neutralize HIV in an unprecedented manner".
This is not peripheral work, to be weighed equally with dozens of other research findings.
These are basic, fundamental statements, regarding what must be achieved in HIV vaccine design, as well as what is the most promising HIV immunotherapy to date.
In the history of the CAVD and CHAVI work, the organizations have offered one potential new HIV therapeutic candidate:
anti-PS.
It is currently buried in a Haynes U.S. patent application.
Jazzbeerman thinks he'll see a published paper explaining why haynes is saying,
* anti-PS "can be safely used as a therapeutic Mab for treatment of HIV infected
subjects".
*anti-PS can "broadly neutralize HIV in an unprecedented manner".
In the history of CAVD and CHAVI publications and patent applications, Haynes (or anyone else) has mentioned by name the names of the following anti-PS mabs in the work:
* Tarvacin
* Bavituximab
* IS1
The anti-PS mab IS1 was isolated at UC by Pojen Chen.
Pojen Chen appeared on PPHM's "Core Technologies" list, last summer.
PPHM licensed anti-PS mabs from UC, last summer.
PPHM CEO SK says that Duke is using more of PPHM's anti-PS mabs than just Bavituximab.
You may not see the above as a "smoking gun", but the convesrse would be as follows:
PPHM licensed mabs from UC, but not Pojen Chen's IS1.
PPHM then invites Pojen Chen onto their "Core Technologies" team, even though they licensed other mabs than his.
PPHM has invited "the competition onto their "Core Technology" team.
PPHM is sending a check to the competition.
Peregrine's and Chen's mab are the only mabs specifically mentioned in Haynes work.
Again,
In the history of the CAVD and CHAVI work, the organizations have offered one potential new HIV therapeutic candidate:
anti-PS.
In the history of CAVD and CHAVI publications and patent applications, Haynes (or anyone else) has mentioned by name the names of the following anti-PS mabs in the work:
* Tarvacin
* Bavituximab
* IS1
The anti-PS mab IS1 was isolated at UC by Pojen Chen.
Pojen Chen appeared on PPHM's "Core Technologies" list, last summer.
PPHM licensed anti-PS mabs from UC, last summer.
PPHM CEO SK says that Duke is using more of PPHM's anti-PS mabs than just Bavituximab.
-------------
j
Peregrine Pharmaceuticals Doses First Patient in Phase II Trial of Bavituximab in Patients With Advanced Breast Cancer
Monday August 11, 8:00 am ET
- Peregrine Achieves Milestone as Third Phase II Trial in Its Bavituximab Cancer Program Begins Patient Enrollment and Dosing -
TUSTIN, Calif., Aug. 11 /PRNewswire-FirstCall/ -- Peregrine Pharmaceuticals, Inc. (Nasdaq: PPHM - News) today announced that patient screening and dosing has begun in a Phase II trial designed to evaluate the safety and efficacy of bavituximab in combination with chemotherapy in patients with advanced breast cancer. The new trial is the second Phase II study evaluating bavituximab in advanced breast cancer patients, and Peregrine also is conducting a third Phase II combination therapy trial of bavituximab in non-small cell lung cancer (NSCLC) patients. The primary objective of the new breast cancer study is to assess the overall tumor response rate to the combination of bavituximab with carboplatin and paclitaxel.
"The combination of bavituximab with the chemotherapy drugs carboplatin and paclitaxel performed well in an earlier Phase I study, and we look forward to learning more about bavituximab's potential in this larger breast cancer study," said Steven W. King, president and CEO of Peregrine. "With three Phase II studies now underway, we look forward to significant clinical data being generated throughout the rest of this year."
In the trial's two-stage design, up to 15 patients with advanced breast cancer will be enrolled initially. The study will then be expanded up to a total of 46 patients if promising results are observed. Secondary objectives of the trial include measuring time to tumor progression, duration of response, overall patient survival and safety parameters. Tumor response will be evaluated using Response Evaluation Criteria in Solid Tumors (RECIST) parameters. Patients may continue to receive bavituximab alone after completion of chemotherapy as long as the cancer does not progress and side effects are acceptable. The trial is being conducted in India according to International Conference on Harmonization (ICH) and Good Clinical Practices (GCP) standards.
Peregrine recently reported that a Phase II trial assessing the combination of bavituximab and docetaxel in patients with metastatic breast cancer had achieved the pre-specified Stage A primary endpoint needed to expand the trial to the second stage. Seven of the 14 evaluable patients achieved partial tumor responses and seven had stable disease at week eight according to RECIST criteria.
"Bavituximab showed promising signs of anti-tumor activity in metastatic breast cancer patients in combination with chemotherapy in a clinical study we helped conduct last year," said Dr. Raghunadharao Digumarti, professor of medical oncology at the Nizams Institute of Medical Sciences in Hyderabad, India and a principal investigator of the bavituximab Phase II breast cancer trial. "We look forward to assessing the results from this trial in a larger population of patients with breast cancer, the most commonly diagnosed cancer in women worldwide."
According to the World Health Organization, breast cancer is the most commonly diagnosed cancer in women, and is second only to lung cancer as a leading cause of female cancer deaths. The National Cancer Institute estimates that approximately 182,460 U.S. women will be diagnosed with breast cancer in 2008 and 40,480 women will die of the disease.
Bavituximab is a monoclonal antibody that binds to the cellular membrane component phosphatidylserine (PS) that is usually located inside cells, but which becomes exposed on the outside of the cells that line the blood vessels of tumors, creating a specific target for anti-cancer treatments. By binding to PS, bavituximab is believed to help mobilize the body's immune system to destroy the tumor and the tumor blood vessels. Bavituximab currently is in two separate Phase II combination therapy trials for the treatment of advanced breast cancer and a Phase II combination therapy trial for the treatment of non-small cell lung cancer. Peregrine recently reported that a Phase II trial assessing the combination of bavituximab and docetaxel in patients with metastatic breast cancer had achieved the pre-specified Stage A primary efficacy endpoint needed to expand the trial to the second stage. A Phase I bavituximab monotherapy trial in advanced solid cancers is also continuing.
---------
j
moby,
It's not charity work.
Actually, the bio-manufacturing portion of the $44 million govt funded bio-defense anti-viral research work is obviously at a "cost-plus" amount to Avid, so yes, to whatever extent, it will provide revenue to Avid.
Additionally, some of the $44 million govt funded bio-defense anti-viral research work will be things that will overlap into work that the company would have needed to do- on their own- to advance the anti-PS platform, so that too, to whatever extent, will likely be lowering Peregrine's burn.
(For instance), the official press release mentions a new, humanized/human/whatever anti-PS which obviously needs to go through the wringer in terms of ADME(T) testing, as well as animal efficacy testing. This carries immediate benefits to Peregrine, who would have been paying for the work to advance the new mab.
ALSO, with respect to Peregrine's lead anti-PS candidate, bavituximab: Peregrine would probably never pursue ebola / marburg / hanta / rift valley / lassa / etc! without the goverment's money. The work will cover further characterization of the mab, as well as a whole lotta animal efficacy work, in hopes of fulfilling the "animal rule", govt stockpiling, etc.
In short, to think that the $44 million govt funded bio-defense anti-viral research funding will not benefit Peregrine, Peregrine's bottom line, Peregrine's burn, is short-sighted.
BTW - Again, to put things into perspective,
here's the DTRA's awards over $100K for the past four months again -
Pretty impressive numbers :)
----------
j
HAYNES - THORPE, SMOKING GUN --------
jess,
THE ONLY SCIENTIST SPECIFICALLY NAMED BY THE BILL AND MELINDA GATES FOUNDATION AS WORKING UNDER BARTON HAYNES ON EXPERIMENTS USING ANTI-PS MABS TO INVESTIGATE HOW WELL THEY PROTECT FROM HIV IS - PHILIP THORPE.
"Philip Thorpe is determining the role of lipid binding of anti-HIV antibodies and anti-phosphatidylserine(PS) autoantibodies to protection from HIV infection."
http://www.cavd.org/progressAbstracts.shtml
--------------
Based on what Haynes is saying in his recent papers and patent applications, I'm thinking Thorpe's work went "well".
recent Bart Haynes patent application:
"A method of treating HIV comprising administering to a patient in need thereof an antibody derivable from a normal subject or from an autoimmune disease subject that binds to a lipid on the surface of HIV or on the surface of HIV-infected cells and thereby neutralizes HIV-1, wherein said antibody is administered in an amount sufficient to effect said treatment."
"these antibodies can broadly neutralize HIV in an unprecedented manner."
--------
Now we see Haynes has the Keynote address at the upcoming Gates-funded HIV vaccine conference, and his presentation is titled,
KEYNOTE ADDRESS: What Does a Successful HIV-1 Vaccine Need To Do? Clues from Acute HIV Infection
j
Haynes is upcoming Keynote speaker at Keystone HIV conf ------------
Barton F. Haynes, Duke University Medical Center, USA
KEYNOTE ADDRESS: What Does a Successful HIV-1 Vaccine Need To Do? Clues from Acute HIV Infection
http://www.keystonesymposia.org/Meetings/ViewMeetings.cfm?MeetingID=991
-------------------------
This title refers to his recent work, and his recent paper in the August Journal of Virology, in which he discusses why any future successful HIV vaccine must interrupt immuno-suppressive PS signaling. To do this, Haynes proposes to create a vaccine that induces anti-PS. He also goes on to say, (in a recent patent application, in conclusions which must be based on experiments which have yet to be published in the journals), that anti-PS mabs can be safe therapy for people already infected with HIV.
----------
It's happening,
j
coagulation cascade caused by PS exposing malaria-infected red blood cells -------
Plasmodium falciparum-infected erythrocytes induce tissue factor expression in endothelial cells and support the assembly of multimolecular coagulation complexes.
J Thromb Haemost. 2007 Jan
Francischetti IM, Seydel KB, Monteiro RQ, Whitten RO, Erexson CR, Noronha AL, Ostera GR, Kamiza SB, Molyneux ME, Ward JM, Taylor TE.
Vector Biology Section, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH), Bethesda, MD
OBJECTIVES: We have asked whether interaction of parasitized red blood cells (pRBC) with EC induces tissue factor (TF) expression in vitro and in vivo. The role of phosphatidylserine-containing pRBC to support the assembly of blood coagulation complexes was also investigated. RESULTS: We demonstrate that mature forms of pRBC induce functional expression of TF by EC in vitro with productive assembly of the extrinsic Xnase complex and initiation of the coagulation cascade. Late-stage pRBC also support the prothrombinase and intrinsic Xnase complex formation in vitro, and may function as activated platelets in the amplification phase of the blood coagulation. Notably, post-mortem brain sections obtained from P. falciparum-infected children who died from cerebral malaria and other causes display a consistent staining for TF in the EC. CONCLUSIONS: These findings place TF expression by endothelium and the amplification of the coagulation cascade by pRBC and/or activated platelets as potentially critical steps in the pathogenesis of malaria. Furthermore, it may allow investigators to test other therapeutic alternatives targeting TF or modulators of EC function in the treatment of malaria and/or its complications.
---------
j
exposed PS triggers TGF-beta & immune suppression ------------------
FEBS Lett. 2007 Jan
Involvement of phosphatidylinositol-3-kinase and ERK pathways in the production of TGF-beta1 by macrophages treated with liposomes composed of phosphatidylserine.
* Otsuka M,
* Negishi Y,
* Aramaki Y.
School of Pharmacy, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan.
We explored the involvement of phosphatidylinositol 3-kinase (PI3K) and ERK pathways in the production of TGF-beta1 by macrophages treated with liposomes composed of phosphatidylserine (PS-liposomes). PS-liposomes activated Akt, downstream of the PI3K signal cascade, and ERK which led to the expression of TGF-beta1. PI3K inhibitors, LY294002 and wortmannin, inhibited the activation of Akt and ERK following the treatment with PS-liposomes. These inhibitors also suppressed the production of TGF-beta1. Furthermore, PS-liposomes activated macrophages to induce TGF-beta1 expression through PS-specific receptors. These findings suggested that a PI3K-ERK signaling pathway via the PS-receptor is intimately involved in the production of TGF-beta1 which regulates macrophage functions.
------------------------------------------
Biol Pharm Bull. 2005 Sep
Phosphatidylserine-specific receptor contributes to TGF-beta production in macrophages through a MAP kinase, ERK.
* Otsuka M,
* Goto K,
* Tsuchiya S,
* Aramaki Y.
School of Pharmacy Tokyo University of Pharmacy and Life Science, Tokyo, Japan.
We explored the involvement of the phosphatidylserine (PS)-receptor in the production of TGF-beta by macrophages treated with PS-liposomes. The binding of anti-PS-receptor antibody to macrophages was specifically inhibited by PS-liposomes. The antibody led to an increase in the production of TGF-beta, and also activated ERK, a member of the MAP kinase. But no activations in p38 and JNK were observed. ERK inhibitor, U0126 completely prevented TGF-beta production. On the addition of a TGF-beta neutralizing antibody or U0126, the inhibitory effect of the anti-PS-receptor antibody on macrophage function, nitric oxide production, was restored. These findings suggested that TGF-beta is one of factors produced by PS-liposomes, and the ERK signaling pathway via the PS-receptor is intimately involved in the production of TGF-beta in macrophages.
------------------------------------------------------------
Cell Death Differ. 2005 Feb
Epithelial cells as phagocytes: apoptotic epithelial cells are engulfed by mammary alveolar epithelial cells and repress inflammatory mediator release.
* Monks J,
* Rosner D,
* Geske FJ,
* Lehman L,
* Hanson L,
* Neville MC,
* Fadok VA.
Program in Cell Biology, D509, Department of Pediatrics, National Jewish Medical and Research Center, Denver, CO 80206, USA.
Clearance of apoptotic cells is critical to tissue homeostasis and resolution of inflammatory lesions. Macrophages are known to remove dying cells and release anti-inflammatory mediators in response; however, many cells traditionally thought of as poor phagocytes can mediate this function as well. In the lactating mammary gland following weaning, alveolar epithelial cell death is massive, yet the gland involutes rapidly, attaining its prepregnancy state in a matter of days. We found histologic evidence of apoptotic cell phagocytosis by viable mammary epithelial cells (MEC) in the involuting mouse mammary gland. Cultured MEC were able to engulf apoptotic cells in vitro, utilizing many of the same receptors used by macrophages, including the phosphatidylserine receptor (PSR), CD36, the vitronectin receptor alpha(v)beta3, and CD91. In addition, MEC, like macrophages, produced TGFbeta in response to stimulation of the PSR by apoptotic cells or the anti-PSR ab 217G8E9, and downregulated endotoxin-stimulated proinflammatory cytokine production. These data support the hypothesis that amateur phagocytes play a significant role in apoptotic cell clearance and its regulation of inflammation.
----------------------------------------------------------
J Clin Invest. 2002 Jan
Phosphatidylserine-dependent ingestion of apoptotic cells promotes TGF-beta1 secretion and the resolution of inflammation.
* Huynh ML,
* Fadok VA,
* Henson PM.
Department of Pulmonary and Critical Care Medicine, University of Colorado Health Sciences Center, Denver, Colorado, USA.
Ingestion of apoptotic cells in vitro by macrophages induces TGF-beta1 secretion, resulting in an anti-inflammatory effect and suppression of proinflammatory mediators. Here, we show in vivo that direct instillation of apoptotic cells enhanced the resolution of acute inflammation. This enhancement appeared to require phosphatidylserine (PS) on the apoptotic cells and local induction of TGF-beta1. Working with thioglycollate-stimulated peritonea or LPS-stimulated lungs, we examined the effect of apoptotic cell uptake on TGF-beta1 induction. Viable or opsonized apoptotic human Jurkat T cells, or apoptotic PLB-985 cells, human monomyelocytes that do not express PS during apoptosis, failed to induce TGF-beta1. PS liposomes, or PS directly transferred onto the PLB-985 surface membranes, restored the TGF-beta1 induction. Apoptotic cell instillation into LPS-stimulated lungs reduced proinflammatory chemokine levels in the bronchoalveolar lavage fluid (BALF). Additionally, total inflammatory cell counts in the BALF were markedly reduced 1-5 days after apoptotic cell instillation, an effect that could be reversed by opsonization or coinstillation of TGF-beta1 neutralizing antibody. This reduction resulted from early decrease in neutrophils and later decreases in lymphocytes and macrophages. In conclusion, apoptotic cell recognition and clearance, via exposure of PS and ligation of its receptor, induce TGF-beta1 secretion, resulting in accelerated resolution of inflammation.
----------------------------------------------------------------
Cancer Biol Ther. 2005 Sep
Cancer cell immune escape and tumor progression by exploitation of anti-inflammatory and pro-inflammatory responses.
* Kim R,
* Emi M,
* Tanabe K.
International Radiation Information Center, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, Japan.
Apoptotic cells can be eliminated by phagocytosis, which is mediated by antigen-presenting cells (APCs), such as macrophages and dendritic cells (DCs), through phosphatidylserine (PS) on apoptotic cells and phosphatidylserine receptor (PSR) on APCs. The phagocytosis of apoptotic cells by macrophages is strictly regulated by not only the inflammatory reaction, but also by an increase in anti-inflammatory factors such as IL-10, TGF-beta, and prostaglandin E2 (PGE2), leading to an anti-inflammatory situation, whereby apoptosis contributes to a noninflammatory response. However, because PS and PSR are expressed in cancer cells, shed soluble phosphatidylserine (sPS) can interact with the PS receptor on macrophages, which promotes an anti-inflammatory response to macrophages that may lead to immune escape. The sPS derived from cancer cells also reacts with the PSR on the cancer cells to produce IL-10, TGF-beta, and PGE2, which can cause suppression of anti-tumor immunity through the anti-inflammatory response to macrophages, which produces tumor-associated macrophages. Furthermore, sPS and TGF-beta inhibit the maturation of immature DCs, resulting in a functional inhibition of DCs. The potential roles of PS and PSR in cancer cells and macrophages in immune escape mediated by sPS and anti-inflammatory factors are discussed, which may explain their dual regulation of anti- and pro-inflammatory responses during tumor progression.
-------------------------------------------------------------
Immunobiology. 2003
Exposure of anionic phospholipids serves as anti-inflammatory and immunosuppressive signal--implications for antiphospholipid syndrome and systemic lupus erythematosus.
* Gaipl US,
* Beyer TD,
* Baumann I,
* Voll RE,
* Stach CM,
* Heyder P,
* Kalden JR,
* Manfredi A,
* Herrmann M.
Institute for Clinical Immunology and Rheumatology, Department of Internal Medicine III, Erlangen, Germany
In contrast to necrotic cells, the clearance of apoptotic ones usually is an anti-inflammatory process which elicits only a marginal immune response. During apoptosis phosphatidylserine (PS) is exposed on the outer leaflet of the cytoplasmic membrane and serves as target for the PS receptor of phagocytes. The latter is responsible for anti-inflammatory signalling and the induction of TGFbeta. We were interested whether the immunogenicity of apoptotic cells can be increased by masking PS. We observed that treatment of xenogeneic apoptotic cells with annexin V (AxV) significantly increased the humoral immune response against surface epitopes of these cells. Furthermore, AxV-coated irradiated tumour cells were able to elicit a long lasting tumour specific cytotoxic T lymphocyte response. AxV efficiently blocked the uptake of irradiated cells by macrophages but not by dendritic cells. Furthermore, AxV skewed the phagocytosis of irradiated cells towards inflammation. Investigation of patients with autoimmune diseases further supported the role of anionic surface phospholipids for anti-inflammatory clearance of apoptotic cells. Impaired clearance and opsonisation with anti-phospholipid-antibodies are discussed to be responsible for the development of systemic lupus erythematosus and anti-phospholipid-syndrome, respectively. Presentation of cryptic epitopes from late apoptotic cells in a proinflammatory context may challenge T cell tolerance. In addition, accumulation of uncleared apoptotic debris in the germinal centres of lymph nodes may result in the survival of autoreactive B cells.
----------------------------------------------------------------
Biochem Biophys Res Commun. 2001 Mar 2
Involvement of TGF-beta in inhibitory effects of negatively charged liposomes on nitric oxide production by macrophages stimulated with lps.
* Matsuno R,
* Aramaki Y,
* Tsuchiya S.
Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Tokyo, Hachioji, 192-0392, Japan.
We examined the role of TGF-beta in the inhibitory effects of negatively charged liposomes composed of phosphatidylserine (PS-liposomes) on nitric oxide (NO) production by macrophages stimulated with LPS. The expression of TGF-beta mRNA increased when mouse peritoneal macrophages were treated with PS-liposomes. The inhibitory effect of PS-liposomes on NO production was restored by treatment with anti-TGF-beta antibody. Furthermore, NO production, iNOS mRNA expression, and iNOS protein induction by LPS were inhibited by treatment of macrophages with TGF-beta as well as PS-liposomes. These results indicated that PS-liposomes down-regulate NO production by macrophages through the induction of TGF-beta and suggested that TGF-beta may suppress NO production upstream of the transcription of iNOS mRNA. Copyright 2001 Academic Press.
-------------------------------------------------------------
J Immunol. 2005 Feb 1
Interaction between phosphatidylserine and the phosphatidylserine receptor inhibits immune responses in vivo.
* Hoffmann PR,
* Kench JA,
* Vondracek A,
* Kruk E,
* Daleke DL,
* Jordan M,
* Marrack P,
* Henson PM,
* Fadok VA.
Department of Pediatrics, Program in Cell Biology, National Jewish Medical and Research Center, Denver, CO 80206, USA.
Phosphatidylserine (PS) on apoptotic cells promotes their uptake and induces anti-inflammatory responses in phagocytes, including TGF-beta release. Little is known regarding the effects of PS on adaptive immune responses. We therefore investigated the effects of PS-containing liposomes on immune responses in mice in vivo. PS liposomes specifically inhibited responses to Ags as determined by decreased draining lymph node tissue mass, with reduced numbers of total leukocytes and Ag-specific CD4(+) T cells. There was also a decrease in formation and size of germinal centers in spleen and lymph nodes, accompanied by decreased levels of Ag-specific IgG in blood. Many of these effects were mimicked by an agonistic Ab-specific for the PS receptor. TGF-beta appears to play a critical role in this inhibition, as the inhibitory effects of PS were reversed by in vivo administration of anti-TGF-beta Ab. PS-containing liposomes did not appear to directly inhibit dendritic cell maturation in vitro in response to a variety of stimuli, nor did it prevent their migration to regional lymph nodes in vivo, suggesting that the inhibitory effects may have resulted from complicated interactions between tissue cells and dendritic cells, subsequently inhibiting their ability to productively activate T lymphocytes.
---------------------------------------------------------------
Cancer Res. 2006 Jun 1
Tumor-driven evolution of immunosuppressive networks during malignant progression.
* Kim R,
* Emi M,
* Tanabe K,
* Arihiro K.
International Radiation Information Center, Research Institute for Radiation Biology and Medicine, Hiroshima University, Japan
Tumors evolve mechanisms to escape immune control by a process called immune editing, which provides a selective pressure in the tumor microenvironment that could lead to malignant progression. A variety of tumor-derived factors contribute to the emergence of complex local and regional immunosuppressive networks, including vascular endothelial growth factor, interleukin-10, transforming growth factor-beta, prostaglandin E(2), and soluble phosphatidylserine, soluble Fas, soluble Fas ligand, and soluble MHC class I-related chain A proteins. Although deposited at the primary tumor site, these secreted factors could extend immunosuppressive effects into the local lymph nodes and the spleen, promoting invasion and metastasis. Vascular endothelial growth factors play a key role in recruiting immature myeloid cells from the bone marrow to enrich the microenvironment as tumor-associated immature dendritic cells and tumor-associated macrophages. The understanding of the immunosuppressive networks that evolve is incomplete, but several features are emerging. Accumulation of tumor-associated immature dendritic cells may cause roving dendritic cells and T cells to become suppressed by the activation of indoleamine 2,3-dioxygenase and arginase I by tumor-derived growth factors. Soluble phosphatidylserines support tumor-associated macrophages by stimulating the release of anti-inflammatory mediators that block antitumor immune responses. Soluble Fas, soluble FasL, and soluble MHC class I-related chain A proteins may help tumor cells escape cytolysis by cytotoxic T cells and natural killer cells, possibly by counterattacking immune cells and causing their death. In summary, tumor-derived factors drive the evolution of an immunosuppressive network which ultimately extends immune evasion from the primary tumor site to peripheral sites in patients with cancer.
----------------------------------------------------------------
Biochem Biophys Res Commun. 2004 Nov 12
Toxoplasma gondii exposes phosphatidylserine inducing a TGF-beta1 autocrine effect orchestrating macrophage evasion.
* Seabra SH,
* de Souza W,
* Damatta RA.
Laboratorio de Biologia Celular e Tecidual, Centro de Biociencias e Biotecnologia, Universidade Estadual do Norte Fluminense, 28013-600 Campos dos Goytacazes, RJ, Brazil.
Toxoplasmosis is a worldwide disease caused by Toxoplasma gondii. Activated macrophages control T. gondii growth by nitric oxide (NO) production. However, T. gondii active invasion inhibits NO production, allowing parasite persistence. Here we show that the mechanism used by T. gondii to inhibit NO production persisting in activated macrophages depends on phosphatidylserine (PS) exposure. Masking PS with annexin-V on parasites or activated macrophages abolished NO production inhibition and parasite persistence. NO production inhibition depended on a transforming growth factor-beta1 (TGF-beta1) autocrine effect confirmed by the expression of Smad 2 and 3 in infected macrophages. TGF-beta1 led to inducible nitric oxide synthase (iNOS) degradation, actin filament (F-actin) depolymerization, and lack of nuclear factor-kappaB (NF-kappaB) in the nucleus. All these features were reverted by TGF-beta1 neutralizing antibody treatment. Thus, T. gondii mimics the evasion mechanism used by Leishmania amazonensis and also the anti-inflammatory response evoked by apoptotic cells.
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J Immunol. 2006 Feb 1
Mimicry of apoptotic cells by exposing phosphatidylserine participates in the establishment of amastigotes of Leishmania (L) amazonensis in mammalian hosts.
* Wanderley JL,
* Moreira ME,
* Benjamin A,
* Bonomo AC,
* Barcinski MA.
Divisao de Medicina Experimental, Coordenacao de Pesquisa, Instituto Nacional de Cancer, Rio de Janeiro, Brazil.
Signaling through exposed phosphatidylserine (PS) is fundamental for the TGFbeta1-dependent, noninflammatory phagocytosis of apoptotic cells. This same mechanism operates in the internalization of amastigotes of Leishmania (L) amazonensis (L(L)a) in a process quoted as apoptotic mimicry. Now we show that the host modulates PS exposure by the amastigotes and, as a consequence, BALB/c mice-derived amastigotes expose significantly more PS than those derived from C57BL/6 mice. Due to this difference in the density of surface PS molecules, the former are significantly more infective than the latter, both in vivo, in F1 (BALB/c x C57BL/6) mice, and in vitro, in thioglycollate-derived macrophages from this same mouse strain. PS exposure increases with progression of the lesion and reaches its maximum value in amastigotes obtained at the time point when the lesion in C57BL/6 mice begins to decrease in size and the lesions in BALB/c mice are still growing in size. Synthesis of active TGFbeta1, induction of IL-10 message, and inhibition of NO synthesis correlate with the amount of surface PS displayed by viable (propidium iodide-negative) infective amastigote. Furthermore, we also show that, similar to what happens with apoptotic cells, amastigotes of L(L)a are internalized by macropinocytosis. This mechanism of internalization is consistent with the large phagolysosomes characteristic of L(L)a infection. The intensity of macrophage macropinocytic activity is dependent on the amount of surface PS displayed by the infecting amastigote.
----------
j
(exposed PS) blood-stage malaria correlates with TGF-beta increase and rapid parasite growth --------
Upregulation of TGF-beta, FOXP3, and CD4+CD25+ regulatory T cells correlates with more rapid parasite growth in human malaria infection.
Center for Clinical Vaccinology and Tropical Medicine, Nuffield Department of Clinical Medicine, Oxford University, Churchill Hospital, Oxford OX3 7LJ, United Kingdom.
Understanding the regulation of immune responses is central for control of autoimmune and infectious disease. In murine models of autoimmunity and chronic inflammatory disease, potent regulatory T lymphocytes have recently been characterized. Despite an explosion of interest in these cells, their relevance to human disease has been uncertain. In a longitudinal study of malaria sporozoite infection via the natural route, we provide evidence that regulatory T cells have modifying effects on blood-stage infection in vivo in humans. Cells with the characteristics of regulatory T cells are rapidly induced following blood-stage infection and are associated with a burst of TGF-beta production, decreased proinflammatory cytokine production, and decreased antigen-specific immune responses. Both the production of TGF-beta and the presence of CD4+CD25+FOXP3+ regulatory T cells are associated with higher rates of parasite growth in vivo. P. falciparum-mediated induction of regulatory T cells may represent a parasite-specific virulence factor.
------------
j
malaria relies on exposed PS ----------
Cytoadherence of Malaria-Infected Red Blood
Cells Involves Exposure of Phosphatidylserine
Introduction
Malaria, one of the most life-threatening infectious
diseases in the world, causes 1.5-2.7 million deaths
annually [1]. Of the four species of human malaria
parasites, Plasmodium falciparum causes the severest of
symptoms and the greatest number of deaths. Adherence
of red cells infected with the mature stages of the parasite
to endothelial cells (EC) lining the postcapillary venules,
especially in the deep tissues, enables the parasites to
avoid splenic clearance and parasite growth is favored
by an hypoxic microenvironment. The adherence of
P. falciparum-infected red cells also results in the
occlusion of microvessels and can lead to
unconsciousness, coma, and in some cases, death. The
virulent nature of P. falciparum has been attributed to
mechanical obstruction by adherent infected red cells [2].
In this context, an understanding of the mechanisms of
the adherence of malaria-infected red cells may lead to
the development of anti-adhesive reagents and new
therapies.
......
The asymmetry of phospholipids in the lipid bilayer
of the plasma membrane is maintained by several membrane
enzymes such as scramblase, translocase and
flippase [28]. Disruption of phospholipid asymmetry
results in the exposure of phosphatidylserine (PS)
molecules, which are normally located in the inner leaflet
of the phospholipid bilayer. We and other investigators
have shown that PS is exposed on the malaria-infected
red cell surface coincident with parasite development [29-
31]. PS liposomes or PS-exposed cells bind to CD36 [32]
and TSP [33] both of which serve as receptors for the
adherence of P. falciparum-infected red cells [34, 35].
Here, we demonstrate that PS exposure on the exofacial
surface of the P. falciparum -infected cell is involved in
the binding to CD36 and TSP.
.......
Discussion
Drastic changes in the red blood cell membrane are
induced during the intracellular maturation of the malarial
parasite, P. falciparum. These include visible changes in
the shape of the red cell, the appearance of electron-dense
protrusions (called knobs) [3, 49], increased membrane
permeability [50, 51], decreased deformability [52],
insertion of parasite-derived proteins [8-10], changes in
the composition and oxidative damage of membrane
lipids [53, 54], a reduction in cholesterol content [30],
as well as degradation of membrane proteins [55].
........
Further,
Facer et al. [63] detected elevated levels of antiphospholipid
antibodies in the serum of P. falciparuminfected
patients using an ELISA method, and found the
highest IgG and IgM binding was to PS and other anionic
phospholipids, indicating that infected red cells expose
PS on their surface in vivo. In that report, the percentage
of anti-PS IgG or IgM positive serum of P. falciparuminfected
patients as 89% and 79%, respectively. Thus,
the accumulating data support PS exposure in P.
falciparum infected red cells.
.......
The precise mechanism of PS exposure on
P.falciparum-infected red cells remains undefined.
Several investigators have demonstrated that treatment
of red cells with various oxidants, such as
phenylhydrazine [64], hydrogen peroxide [65], or
diamide [66], also results in PS exposure. And, PS
exposure has been demonstrated in red cells from patients
with sickle cell anemia [36, 67, 68], thalassemia [69],
diabetes [70], all of which may be under oxidative stress
[71-73]. The erythrocytic stages of Plasmodium as they
mature, digest hemoglobin, and release highly oxidative
iron-containing products. And, oxidized membrane lipids
have been detected in trophozoite- or schizont-infected
red cell membranes [54], coincident with the time we
and other investigators have detected PS exposure [29-
31]. It seems plausible that oxidative stress could act as
a trigger for PS exposure on the surface of the malariainfected
red cell.
.......
Since PS exposure was detected in the various strains of
falciparum malaria we tested, as well as strains used by
other workers, we contend that PS exposure can
contribute to the universal binding of infected cells to
CD36 and TSP. Our findings (i.e., PS exposure and PSmediated
infected-cell binding to TSP) is also compatible
with the suggestion that PS is involved in abnormal sickle
red cell binding to TSP [76, 77].
.......
Further investigation of the structural requirement of
inhibition of PS-mediated infected-cell binding, as well
as a determination of the relative contributions of PfEMP-
1, band 3 and PS to cytoadherence, may assist in the
development of novel and effective anti-adhesive agents.
In conclusion, PS exposure, modification in band 3
protein, and PfEMP-1 all contribute to cytoadherence/
sequestration and disease pathogenesis in P. falciparum
malaria.
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natural anti-PS has anti-malaria action -----------
High levels of anti-phospholipid antibodies in uncomplicated and severe Plasmodium falciparum and in P. vivax malaria.
Facer CA, Agiostratidou G.
London Hospital Medical College, UK.
The majority (75%) of adult patients with uncomplicated Plasmodium falciparum and P. vivax malaria are positive for anti-phospholipid antibodies (aPLA) as demonstrated by ELISA using a panel of anionic and cationic phospholipids. The highest IgG and IgM binding was to the anionic phospholipids, phosphatidylserine (PS), phosphatidic acid (PA) and cardiolipin (CL), but excluding phosphatidylinositol (PI) to which only low antibody levels were found. Comparison of the mean IgG and IgM aPLA showed a trend for anti-PA > CL > PS > PC > PE > PI. Anti-PI levels were compared in two groups of African children, one group with non-severe and the other with severe (cerebral) falciparum malaria. Children with cerebral disease had significantly lower IgM anti-PI. The results are discussed with the view that serum-derived aPLA may have a role in 'anti-disease' immune responses. Their possible role in the opsonization and phagocytosis of parasitized erythrocytes and in thrombocytopenia is also considered.
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another interesting PS microparticles in malaria pathogenesis --------
Cerebral malaria: role of microparticles and platelets in alterations of the blood–brain barrier
Valéry Combes1, Nicolas Coltel2, Dorothée Faille2, Samuel Crocodile Wassmer2 and Georges Emile Grau1
1University of Sydney, Department of Pathology, Medical Foundation Building (K25), 92-94 Parramatta Road, Camperdown, NSW 2042, Australia 2CNRS UMR6020, Immnupathology Group, Faculty of Medicine, IFR48, Université de la Méditerranée, 27, Bd Jean Moulin, F-13385 Marseille Cedex 05, France
Abstract
Brain lesions of cerebral malaria (CM) are characterised by a sequestration of Plasmodium falciparum-parasitised red blood cells (PRBC), leucocytes and platelets within brain microvessels, by an excessive release of pro-inflammatory cytokines as well as by disruption of the blood–brain barrier (BBB). We evaluated the possibility that PRBC and platelets interact and induce functional alterations in brain endothelium. Using an in vitro model of endothelial lesion, we showed that platelets can act as bridges between PRBC and endothelial cells (EC) allowing the binding of PRBC to endothelium devoid of cytoadherence receptors. Furthermore, platelets potentiated the cytotoxicity of PRBC for brain EC by inducing an alteration of the integrity of their monolayer and increasing their apoptosis. These findings provide insights into the mechanisms by which platelets can be deleterious to the brain endothelium during CM. Another aspect of inflammatory and infectious diseases is that they often lead to activation of vascular and blood cells. Such activation results in an enhanced vesiculation, i.e. the release of circulating microparticles (MP). We thus explored plasma levels of endothelial MP in Malawian children with malaria. Plasma MP numbers were markedly increased on admission only in patients with severe malaria complicated with coma. Using the experimental mouse model of CM, we evaluated the pathogenic implications of MP using genetically deficient mice in which the capacity to vesiculate is impaired. Such mice, lacking the ABCA-1 gene, upon infection by Plasmodium berghei ANKA, showed complete resistance to CM. When purified from infected susceptible animals, MP were able to reduce normal plasma clotting time and to significantly enhance tumour necrosis factor release from naïve macrophages. Altogether these data provide a novel insight into the pathogenic mechanisms leading to the neurological syndrome. The finding that ABCA-1 gene deletion confers complete protection against cerebral pathology, linked to an impaired MP production, provides new potential targets for therapeutic amelioration of severe malaria.
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PS microparticles in malaria pathogenesis --------
ABCA1 Gene Deletion Protects against Cerebral Malaria
Potential Pathogenic Role of Microparticles in Neuropathology
Valéry Combes*, Nicolas Coltel*, Mélanie Alibert{dagger}, Miranda van Eck{ddagger}, Cécile Raymond{dagger}, Irène Juhan-Vague§, Georges Emile Grau* and Giovanna Chimini{dagger}
From the Laboratory of Immunopathology,* the Centre d’Immunologie de Marseille Luminy,{dagger} INSERM/Centre National de la Recherche Scientifique/Université de la Méditerranée, and the Haematology Laboratories,§ Faculty of Medicine, IFR48, Marseille, France; and the Division of Biopharmaceutics,{ddagger} Leiden/Amsterdam Center for Drug Research, Gorlaeus Laboratories, Leiden University, The Netherlands
Abstract
The ATP-binding cassette transporter A1 (ABCA1) modulates the transbilayer distribution of phosphatidylserine at the outer leaflet of the plasma membrane. This external exposure of phosphatidylserine is a hallmark of microparticle production and is impaired in ABCA1–/– mice. In this study, we report about the complete resistance to cerebral malaria of these mice. On analysis of histological and systemic parameters we evidenced an impairment of cellular responses to Plasmodium berghei ANKA infection in ABCA1–/– mice, as shown by lower plasma tumor necrosis factor levels, a weaker up-regulation of endothelial adhesion molecules in brain microvessels, a reduced leukocyte sequestration, as well as an ablated platelet accumulation. Besides, the number and the procoagulant activity of microparticles were dramatically reduced in the plasma of ABCA1–/– compared to ABCA1+/+ mice. Moreover, microparticles derived from Plasmodium berghei ANKA-infected ABCA1+/+ mice induced a significant increase of tumor necrosis factor release by noninfected macrophages. In ABCA1–/– mice platelet and macrophage responses to vesiculation agonists were ablated and reduced, respectively. Altogether, by pointing out the ABCA1 transporter as a major element controlling cerebral malaria susceptibility, these data provide a novel insight into its pathophysiological mechanisms and are consistent with a pathogenic role of microparticles in this neurological syndrome.
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Speaking of Georgia -
Here's the chart on how thing's are going based on comments in the July 14 conf call, as compared to standard of care Avastin + Docetaxel.
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Could a vaccine to cholesterol cause a wrinkle in the HIV-1 envelope? --------
PLOS Biology
Shawn J. Green
"In our quest to identify a vaccine strategy directed against HIV-1, the unexpected observation by Haynes et al. revealed that the two broadly neutralizing human antibodies, 2F5 and 4E10, both have binding specificities directed towards phospholipids, including cardiolipin, phosphatidylserine, and other neutral phospholipids [12]. Such observations provoke us to examine the merits of targeting lipid microdomains, in particular, those enriched with cholesterol, cholesterol oxides, and certain phospholipids uniquely displayed in the HIV envelope."
http://biology.plosjournals.org/perlserv/?request=read-response&doi=10.1371/journal.pbio.0040365&ct=1&SESSID=aa14d0dc65460385d0771880479064e0#r1361
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Haynes HIV vaccine WIPO patent application --------
1. A method of inducing the production of an immune response against HIV-I in a mammal comprising administering to said mammal: i) a centralized HIV-I gene sequence, ii) an agent that breaks mammalian immune tolerance, and iii) an agent that inhibits HIV-I -induced apoptosis or an immunosuppressive effect of HIV-I -induced apoptosis, wherein (i), (ii) and (iii) are administered in amounts sufficient to effect said production.
22. The method according to claim 1 wherein said agent that inhibits HIV-1-induced apoptosis induces anti-phosphatidylserine (PS) antibodies, anti- CD36 antibodies, or anti-HIV tat antibodies.
DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a multicomponent, multifunctional HTV vaccine targeted at overcoming: i) HIV diversity, ii) tolerance constraints of neutralizing antibody induction, and iii) apoptotic induced immunosuppression. The invention provides an HIV vaccine comprising centralized HIV gene inserts (consensus, mosaic), a tolerance-breaking component (e.g., TLR-agonists, T regulatory cell innhibition), and a component that can inhibit the immunosuppression of apoptotsis, or inhibit apoptosis itself (e.g., anti-PS, anti- CD36 antibody induction, and/or anti-HTV tat antibody induction)
The present invention includes strategies to prevent apoptosis that include, but are not limited to, the use of PS-containing HIV immunogens, such as PS liposomes, either with or without CON-S or CON-T gpl40 or HTV env epitopes associated with the liposomes, such as 2F5-GTH1 peptide lipid conjugates (Figures 8A and 8B) administered with adjuvants to break tolerance and induce anti-PS antibodies that inhibit PS-CD36 interactions. Alternatively, recombinant CD36 can be targeted in order to raise anti-CD36 antibodies, preferably, both anti-PS or anti-CD36 antibodies are induced at mucosal sites to prevent apoptotic mediated immune suppression.
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Haynes recent U.S. patent application -------
1. A method of treating HIV comprising administering to a patient in need thereof an antibody derivable from a normal subject or from an autoimmune disease subject that binds to a lipid on the surface of HIV or on the surface of HIV-infected cells and thereby neutralizes HIV-1, wherein said antibody is administered in an amount sufficient to effect said treatment.
2. The method according to claim 1 wherein said antibody is derivable from an anti-phospholipid syndrome subject.
3. The method according to claim 1 wherein said antibody is non-pathogenic.
4. The method according to claim 1 wherein said antibody is IS1, IS4 or IS6, or binding fragment thereof.
5. The method according to claim 1 wherein said antibody is IS1, or binding fragment thereof.
[0105] ....That IS1 neutralized HIV evidences the facts that: a) humans can make non-pathogenic anti-lipid antibodies that neutralize HIV, and b) IS1 is an antibody that can be safely used as a therapeutic Mab for treatment of HIV infected subjects or in the setting of post-exposure prophylaxis of subjects following needle, sexual or other exposure to HIV or HIV infected materials.
[0108] Alving and colleagues have made a mouse mab against phosphatidyl inositol phosphate and have shown that it neutralizes HIV in a PBMC assay. What the present studies show is that humans can spontaneously make anti-lipid antibodies and that these antibodies can broadly neutralize HIV in an unprecedented manner.
[0109] Summarizing, autoimmune disease patients can make antibodies that bind to virus-infected cells and, presumably, to budding HIV virions by virtue of their reactivity to HIV membranes and host membranes. Certain anti-lipid antibodies from autoimmune disease patients can also react with the Envelope trimer (such as IS6) but not all of the antibodies react also with the trimer (i.e., IS1 and IS4 do not react). Therefore, reactivity with the HIV envelope is not a prerequisite for neutralization in these antibodies.
[0051] It will be appreciated from a reading of the foregoing that if HIV has evolved to escape the host immune response by making the immune system blind to it, other infectious agents may have evolved similarly. That is, this may represent a general mechanism of escape. That being the case, approaches comparable to those described herein can be expected to be useful in the treatment of such other agents well.
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Haynes quote from Duke PR about his recent microparticle paper -------------
Bart Haynes: (re: HIV vaccine design)
"It is becoming clearer why we have failed in our efforts to date, and what we need to confront to succeed in the future."
http://www.dukemednews.com/news/article.php?id=10364
.......
"Researchers also examined the effects of cell death products upon B cells, another arm of the immune system responsible for the creation of antibodies. Previous studies have shown that the antibody response to HIV-1 is "too little, too late" – appearing after the virus has peaked and after the reservoir of infected T cells has already been established.
Through a series of in vitro laboratory experiments with peripheral blood cells, scientists found that microparticles suppressed levels of IgG and IgA, two classes of antibodies that normally would protect a person against infection. "This is important because many scientists believe that a fast-acting memory B cell response as well as a T cell response will be necessary to fight HIV-1" said Nancy Gasper-Smith, PhD, the lead author of the study."
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Haynes microparticle paper, in the current Journal of Virology ------------
“In this study, we raise the hypothesis that in addition to gut CD4 T cell loss, delay in HIV-1 protective immune responses early on after
HIV-1 transmission may involve the production of elevated levels of immunosuppressive moieties such as TRAIL, TNFR2 and Fas ligand as
well as plasma microparticles.
Microparticles (MPs) are small membrane-bound vesicles that are released from the surface of apoptotic cells
by exocytic or budding processes; as such, MPs bear cell surface markers and can bind annexin V because of the expression of
phosphatidylserine (32-44, 39).
MPs, which circulate in the blood in many clinical conditions, are part of a spectrum of subcellular structures
that are released from cells and can be distinguished from exosomes which are released from multivesicular bodies during activation. Unlike
MPs, exosomes express endosomal markers.
MPs have immunomodulatory activities and can promote immune cell death; exosomes are also immunologically active, can suppress
immune responses (20,34,42,55), and have been reported elevated in chronic HIV-1 (4).
suppression of immune responses can be mediated by T cell MPs (32,34,35). CXCR4+ and CCR5+ MPs can transfer co-receptors to coreceptor
negative cells, making them susceptible to infection by HIV-1 (48,57).
Phagocytosis of MPs by macrophages releases TGF-beta,
prostaglandin E2 and IL-10 that can inhibit antigen specific T and B cell responses (20,35,42).
In this regard, Estes et al. have shown
dramatic increases in lymph node TGF-beta and IL-10 on day 12 following SIV infection (22). Importantly, we have demonstrated that PBMC
and tonsillar cell MPs can directly inhibit memory B cell activation (Figure 8).
both Fas ligand and TRAIL are incorporated into MPs (37,53). Fas ligand expressing MPs can directly induce apoptosis in nearby cells
(20,37,53), and activated T cells can be the target of Fas ligand-mediated proapoptotic microvesicles
it is likely that MPs are responsible for the observed B cell suppressive activity seen in vitro
In the setting of HIV-1 infection
where both activation and apoptosis occur, however, MPs and exosomes may act concomitantly, with exosomes suppressing immune
responses (2,7,15,61), and MPs contributing to both immune suppression and cell death (20,32,34,35,39,42,55).
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Haynes's latest patent application, went public on July 28th on the WIPO database ------------
"The time of appearance of antibodies in the development of acute HIV infection has been recently mapped and it has been shown that most
of the antibodies arise after a delay in the peak response to HIV envelope epitopes of approximately two to three weeks. Indeed, the most
protective antibodies, those that neutralize autologous virus, can be delayed for up to a year."
"To begin to understand the "delay" in induction of antibodies at the time of HIV transmission, the first question to be addressed was whether
there are immunosuppressive events, such as massive apoptosis, with release of phosphatidylserine microparticles at the time of viral load
ramp up during acute HIV infection."
"Apoptotic microparticles are the products of either activated or apoptotic cells, that are increased in the plasma of a number of diseases,
including autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis, Crohn's disease, coronary artery disease
and other forms of heart disease, and chronic HIV-I infection."
"Apoptotic microparticles can bind to non-apoptotic cells and induce apoptosis, are procoagulant, proinflammatory, and can be
immunosuppressive for T and B cell responses to specific antigen."
"Thus, the massive apoptosis that occurs with acute HIV infection with resulting release of TRAIL, mediation of apoptosis via FAS-FASL
interactions, and release of PS containing viral and other particles, all conspire to initially immuno suppress the host, preventing rapid
protective B cell responses."
"Thus, the production of high levels of biologically active plasma mediators and byproducts of cell death during the first two to three weeks of
HIV- 1 transmission raises the notion that the window of opportunity for a preventive vaccine to work may be shorter than previously thought,
ie within the first 14-17 days of transmission, placing considerable constraints on the time available for development of robust anti-HIV-1
immunity following transmission."
"Inhibition of cell death and immunosuppressive MP mediated effects by a vaccine for HIV or other infectious agents may be important as
well. This could be accomplished, for example, by an HIV vaccine component inducing anti-lipid antibodies or antibodies against other
components of microparticles to facilitate clearance of microparticles and/or to block microparticle toxic effects."
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WOW - THANKS! Great post. I second the words of Tony Fauci, and slipperypete! by repeating Fauci's quote -
The NIH’s Anthony Fauci, who once cautioned that there might never be a traditional vaccine to prevent HIV infection and recently pulled the plug on a troubled vaccine trial, sounded a cautiously optimistic note at the 17th International AIDS Conference yesterday.
“The future for AIDS research looks bright and promising,” said Fauci, singling out recent work by Barton Haynes of Duke and Robert Siliciano of Johns Hopkins in illuminating how the virus hides inside the body and suppresses the immune system within days of infection.
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it's happening!
:)
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Phosphatidylserine on HIV Envelope Is a Cofactor for Infection of Monocytic Cells --------
http://www.jimmunol.org/cgi/content/full/170/9/4840
Melissa K. Callahan2,3,*, Paul M. Popernack2,{dagger}, Shigeki Tsutsui4,{ddagger}, Linh Truong{ddagger}, Robert A. Schlegel*,§ and Andrew J. Henderson5,*,{dagger},{ddagger}
Graduate Programs in * Biochemistry, Microbiology, and Molecular Biology and {dagger} Pathobiology, and Departments of {ddagger} Veterinary Science and § Biochemistry and Molecular Biology, Pennsylvania State University, University Park, PA 16802
Abstract
HIV-1 is an enveloped retrovirus that acquires its outer membrane as the virion exits the cell. Because of the association of apoptosis with the progression of AIDS, HIV-1-infected T cells or macrophages might be expected to express elevated levels of surface phosphatidylserine (PS), a hallmark of programmed cell death. Virions produced by these cells would also be predicted to have PS on the surface of their envelopes. In this study, data are presented that support this hypothesis and suggest that PS is required for macrophage infection. The PS-specific protein annexin V was used to enrich for virus particles and to inhibit HIV-1 replication in primary macrophages, but not T cells. HIV-1 replication was also significantly inhibited with vesicles consisting of PS, but not phosphatidylcholine. PS is specifically required for HIV-1 infection because viruses pseudotyped with vesicular stomatitis virus G and amphotropic murine leukemia virus envelopes were not inhibited by PS vesicles or annexin V. These data indicate that PS is an important cofactor for HIV-1 infection of macrophages.
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Tumor-Released Microvesicles as Vehicles of Immunosuppression -------
http://cancerres.aacrjournals.org/cgi/content/abstract/67/7/2912
Roberta Valenti, Veronica Huber, Manuela Iero, Paola Filipazzi, Giorgio Parmiani and Licia Rivoltini
Unit of Immunotherapy of Human Tumors, Fondazione IRCCS Istituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy
Tumor-released microvesicles, or exosomes, which are abundant in the body fluids of patients with cancer, are likely to be involved in tumor progression. We recently showed that microvesicles released by human melanoma and colorectal carcinoma cells can promote the differentiation of monocytes to myeloid-derived suppressor cells which support tumoral growth and immune escape. These findings underscore an important role for these extracellular organelles in remodeling tumor-stromal interactions to promote malignancy.
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Human Colorectal Cancer Cells Induce T-Cell Death Through Release of Proapoptotic Microvesicles: Role in Immune Escape ---------
http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6WFX-4GB4GVS-N&_user=10&_rdoc=1&_fmt=&_orig=search&_sort=d&view=c&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=75380a159a3a7247c6704f122fe3d1bc
Veronica Huberlow asterisk, Stefano Fais‡, Manuela Ierolow asterisk, Luana Lugini‡, Paola Caneselow asterisk, Paola Squarcinalow asterisk, Annamaria Zaccheddulow asterisk, Marisa Colone§, Giuseppe Arancia§, Massimo Gentiledouble vertical bar, Ettore Seregni¶, Roberta Valentilow asterisk, Giuseppina Ballabio¶, Filiberto Belli#, Ermanno Leo#, Giorgio Parmianilow asterisk and Licia Rivoltinilow asterisk, Corresponding Author Contact Information, E-mail The Corresponding Author
low asteriskUnit of Immunotherapy of Human Tumors, Istituto Nazionale Tumori, Milan, Italy ‡Department of Drug Research and Evaluation, Istituto Superiore di Sanità, Rome, Italy §Department of Health and Technology, Istituto Superiore di Sanità, Rome, Italy double vertical barDepartment of Experimental Medicine and Pathology, Virology Section, University “La Sapienza,” Rome, Italy ¶Unit of Nuclear Medicine, Istituto Nazionale Tumori, Milan, Italy #Unit of Colorectal Surgery, Istituto Nazionale Tumori, Milan, Italy
Background & Aims: Normal and neoplastic cells release microvesicles, whose effects on the immune system still need to be elucidated. Because human colorectal cancer cells are hypothesized to escape immune recognition by expressing proapoptotic molecules, we investigated whether microvesicles bearing Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand and inducing apoptosis of activated T cells are secreted by colorectal cancer cells both in vitro and in affected patients. Methods: Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand expression were analyzed in colorectal cancer cells and purified microvesicles by flow cytometry, Western blotting, and immunoelectron microscopy. Microvesicle tumor origin was assessed through simultaneous detection of lysosomal (CD63) and adenocarcinoma (carcinoembryonic antigen) markers. Proapoptotic activity of microvesicles was evaluated by annexin V/propidium iodide staining and caspase activation in T cells, including CD8+ T lymphocytes from colorectal cancer patients. Results: Colorectal cancer cells showed a granular pattern of tumor necrosis factor-related apoptosis-inducing ligand and Fas ligand expression, suggesting a secretory behavior. These proapoptotic molecules were detected on isolated microvesicles, together with class I HLA, CD63, and carcinoembryonic antigen. Microvesicles induced Fas ligand-mediated and tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis of activated CD8+ T cells generated from colorectal cancer patients. Microvesicles with comparable phenotypes and functions were found in plasma from patients with advanced disease, whereas vesicular structures expressing Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand were also detected in colorectal cancer specimens. Conclusions: These data show that colorectal cancer induces T-cell apoptosis through the release of Fas ligand-bearing and tumor necrosis factor-related apoptosis-inducing ligand-bearing microvesicles both in vitro and in vivo. This mechanism of immune escape has potential implications as a prognostic factor and could be targeted for the development of new antitumor therapies in colorectal cancer patients.
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Human Tumor-Released Microvesicles Promote the Differentiation of Myeloid Cells with Transforming Growth Factor-ß–Mediated Suppressive Activity on T Lymphocytes ----------
http://cancerres.aacrjournals.org/cgi/content/full/66/18/9290
Roberta Valenti1, Veronica Huber1, Paola Filipazzi1, Lorenzo Pilla1, Gloria Sovena1, Antonello Villa3,4, Alessandro Corbelli2,3,4, Stefano Fais5, Giorgio Parmiani1 and Licia Rivoltini1
1 Unit of Immunotherapy of Human Tumors, Istituto Nazionale Tumori; 2 Fondazione D'Amico, Milan, Italy; 3 Microscopy and Image Analysis Consortium; 4 Dipartimento di Neuroscienze e Tecnologie Biomediche, Università Milano-Bicocca, Monza, Italy; and 5 Department of Drug Research and Evaluation, Section of Pharmacogenetic, Drug Resistance and Experimental Therapeutics, Istituto Superiore di Sanità, Rome, Italy
Abstract
Human tumors constitutively release endosome-derived microvesicles, transporting a broad array of biologically active molecules with potential modulatory effects on different immune cells. Here, we report the first evidence that tumor-released microvesicles alter myeloid cell function by impairing monocyte differentiation into dendritic cells and promoting the generation of a myeloid immunosuppressive cell subset. CD14+ monocytes isolated from healthy donors and differentiated with interleukin (IL)-4 and granulocyte macrophage colony-stimulating factor in the presence of tumor-derived microvesicles turned into HLA-DR–/low cells, retaining CD14 expression and failing to up-regulate costimulatory molecules, such as CD80 and CD86. These phenotypic changes were paralleled by a significant release of different cytokines, including IL-6, tumor necrosis factor-{alpha}, and transforming growth factor-ß (TGF-ß), and a dose-dependent suppressive activity on activated T-cell–proliferation and cytolytic functions, which could be reversed by anti-TGF-ß–neutralizing antibodies. Microvesicles isolated from plasma of advanced melanoma patients, but not from healthy donors, mediated comparable effects on CD14+ monocytes, skewing their differentiation toward CD14+HLA-DR–/low cells with TGF-ß–mediated suppressive activity on T-cell–functions. Interestingly, a subset of TGF-ß–secreting CD14+HLA-DR– cells mediating suppressive activity on T lymphocytes was found to be significantly expanded in peripheral blood of melanoma patients compared with healthy donors. These data suggest the development in cancer patients of an immunosuppressive circuit by which tumors promote the generation of suppressive myeloid cells through the release of circulating microvesicles and without the need for cell-to-cell contact. Therapeutic interventions on the crucial steps of this pathway may contribute to restore tumor/immune system interactions favoring T-cell–mediated control of tumor growth in cancer patients.
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Tumor Exosomes Inhibit Differentiation of Bone Marrow Dendritic Cells ----------
http://www.jimmunol.org/cgi/content/abstract/178/11/6867
Shaohua Yu*, Cunren Liu*, Kaihong Su*, Jianhua Wang*, Yuelong Liu*, Liming Zhang*, Chuanyu Li*, Yingzi Cong{ddagger}, Robert Kimberly*, William E. Grizzle{dagger}, Carla Falkson§ and Huang-Ge Zhang2,*,¶
* Department of Medicine, Division of Clinical Immunology and Rheumatology, {dagger} Department of Pathology, {ddagger} Department of Medicine, Division of Gastroenterology and Hepatology, and § Department of Medicine, Division of Hematology and Oncology, University of Alabama at Birmingham, Birmingham, Alabama 35294; and ¶ Birmingham Veterans Administration Medical Center, Birmingham, AL 35233
Abstract
The production of exosomes by tumor cells has been implicated in tumor-associated immune suppression. In this study, we show that, in mice, exosomes produced by TS/A murine mammary tumor cells target CD11b+ myeloid precursors in the bone marrow (BM) in vivo, and that this is associated with an accumulation of myeloid precursors in the spleen. Moreover, we demonstrate that TS/A exosomes block the differentiation of murine myeloid precursor cells into dendritic cells (DC) in vitro. Addition of tumor exosomes at day 0 led to a significant block of differentiation into DC, whereas addition at later time points was less effective. Similarly, exosomes produced by human breast tumor cells inhibited the differentiation of human monocytes in vitro. The levels of IL-6 and phosphorylated Stat3 were elevated 12 h after the tumor exosome stimulation of murine myeloid precursors, and tumor exosomes were less effective in inhibiting differentiation of BM cells isolated from IL-6 knockout mice. Addition of a rIL-6 to the IL-6 knockout BM cell culture restored the tumor exosome-mediated inhibition of DC differentiation. These data suggest that tumor exosome-mediated induction of IL-6 plays a role in blocking BM DC differentiation.
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Tumour-released exosomes and their implications in cancer immunity --------
http://www.nature.com/cdd/journal/v15/n1/abs/4402237a.html
M Iero1, R Valenti1, V Huber1, P Filipazzi1, G Parmiani2, S Fais3 and L Rivoltini1
1Unit of Immunotherapy of Human Tumours, Fondazione IRCCS Istituto Nazionale Tumori, Milan, Italy
2Unit of Immunobiotherapy of Solid Tumours, San Raffaele Scientific Institute, Milan, Italy
3Department of Drug Research and Evaluation, Pharmacogenetic, Drug Resistance, and Experimental Therapeutic Section, Istituto Superiore di Sanità, Rome, Italy
Abstract
Tumour cells release vesicular structures, defined as microvesicles or exosomes, carrying a large array of proteins from their originating cell. The expression of antigenic molecules recognized by T cells has originally suggested a role for these organelles as a cell-free antigen source for anticancer vaccines. However, recent evidence shows that tumour exosomes may also exert a broad array of detrimental effects on the immune system, ranging from apoptosis in activated antitumour T cells to impairment of monocyte differentiation into dendritic cells and induction of myeloid suppressive cells. Immunosuppressive exosomes of tumour origin can be found in neoplastic lesions and sera from cancer patients, implying a potential role of this pathway in in vivo tumour progression. Through the expression of molecules involved in angiogenesis promotion, stromal remodelling, delivery of signalling pathways through growth factor/receptor transfer, chemoresistance and genetic intercellular exchange, tumour exosomes could represent a versatile tool for moulding host environment. Hence, their secretion by neoplastic cells may in the future become a novel pathway to target for therapeutic intervention in cancer patients.
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The role of membrane lipids in the induction of macrophage apoptosis by microparticles --------
Lars C. Huber1, 2 Contact Information, Astrid Jüngel1, 2, Jörg H. W. Distler1, 2, Falk Moritz1, 2, Renate E. Gay1, 2, Beat A. Michel1, 2, David S. Pisetsky3, Steffen Gay1, 2 and Oliver Distler1, 2
(1) Center of Experimental Rheumatology, University Hospital Zurich, Gloriastrasse 23, CH-8091 Zurich, Switzerland
(2) Zurich Center for Integrative Human Physiology (ZIHP), Zurich
(3) Division of Rheumatology, Durham VA Hospital and Duke University Medical Center, Durham, NC, USA
Abstract
Microparticles are membrane-derived vesicles that are released from cells during activation or cell death. These particles can serve as mediators of intercellular cross-talk and induce a variety of cellular responses. Previous studies have shown that macrophages undergo apoptosis after phagocytosing microparticles. Here, we have addressed the hypothesis that microparticles trigger this process via lipid pathways. In these experiments, microparticles induced apoptosis in primary macrophage cells or cell lines (RAW 264.7 or U937) with up to a 5-fold increase. Preincubation of macrophages with phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)BP) reduced the microparticle-induced apoptosis in a dose-dependent manner. PtdIns(3,5)BP is a specific inhibitor of the acid sphingomyelinase and thus can block the generation of pro-apoptotic ceramides. Similarly, the pre-incubation of macrophages with PtdIns(3,5)BP prevented microparticle-induced upregulation of caspase 8, which is a major target molecule of ceramide action in the apoptosis pathway. PtdIns(3,5)BP, however, had no effect on the spontaneous rate of apoptosis. To evaluate further signaling pathways induced by microparticles, the extracellular signal regulated kinase (ERK-) 1 was investigated. This kinase plays a role in activating phospholipases A2 which cleaves membrane phospholipids into arachidonic acid; microparticles have been suggested to be a preferred substrate for phospholipases A2. As shown in our experiments, microparticles strongly increased the amount of phosphorylated ERK1/2 in RAW 264.7 macrophages in a time-dependent manner, peaking 15 min after co-incubation. Addition of PD98059, a specific inhibitor of ERK1, prevented the increase in apoptosis of RAW 264.7 macrophages. Together, these data suggest that microparticles perturb lipid homeostasis of macrophages and thereby induce apoptosis. These results emphasize the importance of biolipids in the cellular cross-talk of immune cells. Based on the fact that in clinical situations with excessive cell death such as malignancies, autoimmune diseases and following chemotherapies high levels of circulating microparticles might modulate phagocytosing cells, a suppression of the immune response might occur due to loss of macrophages.
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