Study pinpoints how a normally defensive immune response can help HIV - journal commentary included
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May 19, 2010
Researchers have identified how a normal response to infection, one that usually serves to limit the amount of inflammation, actually contributes to disease progression and viral persistence in HIV-infected patients.
The findings, published in the May 19 issue of the journal Science Translational Medicine, offer important opportunities for further research, both for treatment of long-term persistence of HIV in those who are infected and for prevention of infection in those who are not, according to the study team.
The study, led by UCSF researchers, focused on the body's production of an enzyme called indoleamine 2,3-dioxygenase 1 (IDO1). To prevent the harm associated with chronic inflammation, the body typically turns on IDO1, which then serves to suppress inflammation and immune responses. In the setting of HIV infection, the authors found that IDO1 instead acts to alter the balance between two types of T-cells that have opposing functions.
One type of immune cell, called TH17, releases interleukin-17, a cytokine that has a central role in maintaining the integrity of the mucosal barrier in the gut. The other type, named Treg, prevents inflammation in a non-specific manner and can also turn off immune responses against viruses such as HIV.
The authors found that induction of IDO1 by HIV results in loss of TH17 cells and a relative increase in Tregs. This change in the balance of TH17 cells and Tregs allows bacteria to cross the mucosal barrier of the gut, initiating new inflammatory reactions in the process. At the same time, the increased number of Tregs may prevent the immune system from attacking HIV in areas of the body where strong HIV-specific immune responses are most needed. The altered TH17/Treg balance, in sum, leads to an endless cycle of inflammation induced by the invading microbes, more induction of IDO1, and continued loss of TH17 cells.
"In most instances, reducing inflammation following immune system activation to fight infection is beneficial. But, in HIV disease, this can establish a reinforcing cycle that is strongly linked to disease progression and that may help HIV to persist in patients, said study lead co-author, Jeff Mold, PhD, from the UCSF Division of Experimental Medicine. "Mucosal defenses are breached, microbes cross over, and inflammation results. This leads to increasing IDO1 activity, continued changes in the balance of TH17 and Treg cells, further weakening of the mucosal defenses, and even more inflammation."
The findings represent the next step in a series of research studies reported previously by the same group of investigators, showing that SIV infection of monkeys leading to AIDS is associated with a similar change in TH17 and Treg balances. The change in T cell balance was not observed in another primate, African green monkeys, where infection with SIV is harmless and does not cause disease.
In the current study, the investigators looked at IDO1 activity in HIV-infected human subjects at various stages of disease and in healthy non-infected subjects.
"We confirmed that IDO1 activity is associated with HIV disease progression. But we went further and also looked at the TH17 and Treg balance, and found that the change in the ratio leading to decreasing TH17 cells is also associated with HIV disease progression," said study lead co-author, David Favre, PhD, formerly at UCSF, now with the National Immune Monitoring Laboratory, Montreal.
With pharmacological inhibitors of IDO1 in development and currently in clinical trials for cancer immunotherapy, the finding may lead to new therapeutic approaches for assisting in the control of HIV disease, noted the study team.
"Most of an infected person's own immune responses that are known to affect HIV disease outcomes cannot be manipulated or altered clinically and, hence, have not really had much of an impact for patients. This work, however, is very different, as it has uncovered several possible pathways that might be addressed clinically with developing or available therapeutics," said study co-author, Steven Deeks, MD, professor of medicine at the UCSF Division of HIV/AIDS at San Francisco General Hospital.
IDO1 may play a role in the ability of HIV to persist in HIV-infected patients for their lifetimes, notwithstanding effective treatment with antiretroviral therapies.
"Steve Deeks and I are continuing to examine the role of IDO1 through a study recently announced by amfAR, the Foundation for AIDS Research, into whether the disruption of IDO1 will reduce the level of immune activation, which could then lead to a decrease in viral persistence," said senior study author Joseph M. McCune, MD, PhD, chief of the UCSF Division of Experimental Medicine.
In addition to Favre, Mold, Deeks, and McCune, other study co-authors include Peter Hunt, Bittoo Kanwar, Lillian Seu, Jason Barbour, Margaret Lowe, Anura Jayawardene, Francesca Aweeka, Yong Huang, Jeffrey Martin, and Frederick Hecht from UCSF; Daniel Douek and Jason Brenchley from the National Institute of Allergy and Infectious Diseases, NIH, and P'ng Loke from NYU.
The research was funded by grants from the Elizabeth Glaser Pediatric AIDS Foundation, the National Institute of Allergy and Infectious Diseases, the National Institutes for Health, the Harvey V. Berneking Living Trust, the UCSF-GIVI Center for AIDS Research, and the UCSF Clinical and Translational Institute Clinical Research Center.
The UCSF Division of Experimental Medicine and the UCSF Division of HIV/AIDS at SFGH are affiliated with the AIDS Research Institute (ARI) at UCSF. UCSF ARI houses hundreds of scientists and dozens of programs throughout UCSF and affiliated labs and institutions, making ARI one of the largest AIDS research entities in the world.
UCSF is a leading university dedicated to defining health worldwide through advanced biomedical research, graduate level education in the life sciences and health professions, and excellence in patient care.
Insights into Therapy: Tryptophan Oxidation and HIV Infection - Commentary - pdf attached
Sci Transl Med 19 May 2010:
Vol. 2, Issue 32, p. 32ps23
Michael F. Murray
Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA.
New data from Favre and colleagues strengthen the link between activation of the tryptophan oxidation (TOx) pathway-via the indoleamine 2,3-dioxygenase enzymes IDO1 and IDO2-and chronic inflammation in progressive HIV disease. It can now be appreciated that a pathogenic TOx activation cycle exists in HIV. TOx regulation is a therapeutic target for other diseases, such as cancer and autoimmune disorders. Here TOx control is examined with an eye to eventual therapeutic intervention in HIV disease.
HIV AND BEYOND
Microbial products, including lipopolysaccharide (LPS), the immunostimulatory endotoxin from the outer membrane of Gram-negative bacteria, are known to induce the tryptophan oxidation (TOx) pathway, which is the key degradation route for the essential amino acid l-tryptophan (l-Trp). The first (and rate-limiting) step in the pathway-the conversion of l-Trp to N-formyl-kynurenine-is catalyzed by the indoleamine 2,3-dioxygenases IDO1 and IDO2, two enzymes that are synthesized in both immune cells and other tissues and are the gatekeepers of the immune response via the TOx pathway. In this issue of Science Translational Medicine, Favre and colleagues add to a growing body of knowledge linking LPS-induced endotoxemia (the presence of bacterial endotoxins in the blood), activation of the TOx pathway, and the chronic inflammatory state of progressive HIV disease (1). Researchers currently are developing targeted therapies to alter TOx activation in a number of disease contexts, and a richer understanding of pathway regulatory mechanisms will help to guide these drug discovery and development efforts. Here various aspects of TOx regulation are examined from the perspective of potential therapeutic intervention.
TOX PATHWAY PERMUTATIONS
As an essential amino acid, Trp cannot be synthesized in humans and must be obtained through the diet. Dietary Trp feeds into three major metabolic pathways: (i) protein synthesis; (ii) synthesis of the neurotransmitter serotonin; and (iii) TOx, to form either energy [adenosine triphosphate (ATP)], nicotinamide and related compounds [nicotinamide adenine dinucleotide (NAD) and nicotinamide adenine dinucleotide phosphate (NADP)], or other side products (such as xanthurenic acid). In humans, oxidative catabolism of Trp can be initiated by one of three different enzymes: tryptophan-2,3-dioxygenase (TDO2) in the liver, as well as IDO1 and IDO2, which have incompletely overlapping nonhepatic tissue distributions. These three enzymes open the indole ring, causing the irreversible loss of Trp (2). The TOx pathway is interconnected with other metabolic networks (3) and is variously regulated in a number of tissues.
The liver is thought to be the only organ with substantial TDO2 activity, and this enzyme is not believed to drive the immunoregulatory effects of TOx. Instead, TDO2 is induced by dietary protein and corticosteroids, but not by cytokines such as interferon-γ (IFN-γ) (4). Recent work by Schmidt et al. demonstrated an in vitro immunoregulatory effect associated with TDO2 activity, which may implicate the activity of pathway metabolites more than a clinically significant in vivo effect, although a contribution to immunomodulation within the liver has not been excluded (5). The hepatic end products generated via activation of this pathway are ATP, CO2, and water.
The metabolic pathway initiated by the extrahepatic enzymes IDO1 and IDO2 is shown in Fig. 1. IDO1 and IDO2 are inducible and less substrate-specific for Trp than TDO2. While there are many cytokine inducers of IDO-mediated TOx, the most effective pathway inducer appears to be IFN-γ (4). That said, there are important tissue-specific differences in cytokine induction, leading to different tissue responses to the same systemic signals. For example, in the brain it appears that interleukin-6, and not IFN-γ, drives IDO induction (6). Elucidation of the differences in IDO1- versus IDO2-induced cytokine responses will require further study. Many different microbial infections-bacterial, viral, and parasitic-have been implicated in IDO-induced TOx (7), and a number of microbial products can directly induce the pathway. Bacterial LPS and the HIV-Nef and HIV-Tat proteins are among the best-studied but are not likely to be unique in this capability.
The second enzyme in the TOx pathway is arylformamidase (AFMID), which is responsible for converting the dioxygenase product (N-formyl-kynurenine) to l-kynurenine (l-KYN). l-KYN is subsequently converted to 3-hydroxy-l-kynurenine (3-HKA) by kynurenine 3-monooxygenase (KMO), and 3-HKA is metabolized to 3-hydroxy-anthranilic acid (3-HAA) by kynureninase (KYNU). Favre et al. noted that the 3-HKA and 3-HAA metabolites have effects on the T helper 17 cell (TH17)-to-regulatory T cell (Treg) ratio in vitro (1). It appears that 3-HAA is removed from the tissue microenvironment only by the action of another dioxygenase, 3-hydroxyanthranilate 3,4-dioxygenase (HAAO). This means that, in order to prevent the in vivo accumulation of 3-HAA and its potential effects on T cell ratios, the organism is dependent on the relative activity of this specific enzyme. Outside of the central nervous system (CNS), TOx pathway activation does not lead to the accumulation of quinolinic acid (QA), suggesting that the relative activities of the aminocarboxymuconate semialdehyde decarboxylase (ACMSD) and quinolinate phosphoribosyltransferase (QPRT) enzymes are sufficient to divert QA to other products. In a multistep process initiated by QPRT, the downstream metabolism of QA results in the formation of NAD, NADP, and nicotinamide. In general, it is presumed that the QPRT-driven pathway to nicotinamide and nicotinamide nucleotides occurs preferentially in the extrahepatic TOx pathway, and the ACMSD-driven subpathway to ATP appears to occur preferentially by hepatic TOx, although further detailed studies may alter this understanding (4). In the CNS, QA is produced locally in response to either local or systemic inflammatory signals and stimulates the N-methyl-d-aspartate (NMDA) receptor, a key component in learning and memory (8). Overstimulation of NMDA receptors can cause neurotoxic effects that have been implicated in neurological diseases.
In HIV-infected patients, the amounts of QA measured in the brain and cerebrospinal fluid (CSF) are greatly increased compared to those in controls. In one study (8), a 100-fold variation in the amounts of QA in the CSF and brain tissue was detected among HIV patients, and these elevations did not correlate with serum QA concentrations (8). These data suggest that the degradation of Trp via the TOx pathway is stalled in the CNS. This tissue-specific accumulation of QA may result from a relatively low amount of QPRT activity compared to that of HAAO and other upstream enzymes (Fig. 1). Recent work by Connor and colleagues revealed that systemic stimuli such as LPS induce IDO and KMO activities in the mammalian brain but do not alter kynurenine aminotransferase (KAT) activity (6). A critical question is whether QPRT is also inducible and under what circumstances, because this effect would have the potential to draw down the local QA concentrations and interrupt the neurotoxic effects associated with high concentrations of this metabolite.
The new work by Favre et al. builds on the relationship between LPS and TOx first reported in 1978 by Yoshida and Hayaishi (9) and the findings of Brenchley and others (in 2006) that HIV infection is marked by chronically elevated circulating amounts of LPS (10). Favre et al. place the observations of Terness and others regarding TOx metabolites into a new disease-specific context (11, 12) and provide evidence that the intermediate TOx metabolites 3-HKA and 3-HAA are regulators of a specific T lymphocyte subset ratio; namely, the TH17:Treg ratio (1). The loss of a normal TH17:Treg ratio in the gut is associated with increased LPS endotoxemia and persistent induction of TOx overstimulation in the setting of HIV (13). These data add to an impression that chronic TOx induction during HIV infection is a central pathogenic process that results in the disruption of normal T lymphocyte function and is driven by a combination of endogenous cytokines (particularly IFN-γ), viral products (the HIV-Tat and HIV-Nef proteins), and bacterial products (including LPS) (Fig. 2).
The detrimental effects of TOx induction in progressive HIV disease contrast with the demonstrable benefit of TOx activation in the normal placenta and the presumed value of TOx in limiting microbial access to Trp in some nonviral infections (14). In HIV infection, the mediators of TOx pathogenicity appear to be the altered concentrations of the pathway products, in particular Trp, 3-HKA, 3-HAA, and QA.
The growing interest in targeted therapies to alter TOx activation in cancer management (15) needs to be extended to HIV infection, as well as reproduction, organ transplantation, neurodegenerative diseases, and autoimmune syndromes, in which TOx pathway activation has also been implicated in aspects of disease pathogenesis (16-20).
TOX THERAPEUTIC TACTICS
There is accumulating evidence that small molecules that participate in the TOx pathway are capable of acting as immune regulators outside of their direct role in amino acid catabolism. Further study of the tissue-specific bioavailability of 3-HKA and 3-HAA is needed to confirm their in vivo significance in pathogenic immunoregulatory signaling; however, what emerges through the work to date is a model for chronic activation of IDO-associated TOx, leading to T lymphocyte dysregulation in HIV infection (Fig. 2). Given the number of diseases in which TOx activation appears to be linked to pathogenesis, it is likely that multiple clinical trials will emerge to test agents that might therapeutically manipulate this pathway. Oral Trp loading is one approach to avoid in going forward, except perhaps in a carefully controlled research setting, because it has been associated with both an increase in circulating TOx pathway intermediates (21) and the still poorly understood phenomenon of eosinophilia-myalgia syndrome (22).
As with any metabolic pathway, the pharmacological regulation of extrahepatic TOx can be approached in several ways. The approach that is currently being tried is inhibition of the first and rate-limiting step in the TOx pathway, namely IDO activity (both IDO1 and IDO2). This is being pursued with the competitive inhibitor 1-methyl-D-tryptophan (D-1MT), and there are currently at least four D-1MT trials (phase 1 and 2) in the clinical oncology domain (23). D-1MT was also used in a study of SIV-infected macaques, an animal model of HIV, with some benefit (24). A number of other IDO inhibitory molecules have been studied, but they have not yet progressed to clinical trials (25). There also is interest in the use of Trp catabolite mimetics, such as 3,4-DAA [N-(3,4-dimethyoxycinnamoyl) anthranilic acid], which is orally active and may represent a lead compound for inhibiting autoreactive TH1 cells in autoimmune diseases (20). Furthermore, it may be desirable in some therapeutic settings to differentially regulate IDO1 and IDO2 (26). Because a functional IDO2 gene product is not expressed in as many as 25 to 30% of certain human populations (27), the consequences of this natural experiment need to be better understood, so that they may guide our evolving understanding of how to successfully use pharmacological agents to inhibit this pathway.
Other conceptual therapeutic approaches that target TOx include (i) diverting the pathway from toxic intermediates or end products by inducing benign metabolic side-product formation, (ii) inducing downstream pathway activity to drive end-product formation and avoid the accumulation of toxic intermediates, and (iii) feedback inhibition of the pathway via increased end-product concentrations. On the basis of data that suggested that HIV-infected patients display a metabolic drive toward increased available nicotinamide (28), we initiated a very small clinical trial of oral nicotinamide in part to provide potential feedback inhibition of the TOx pathway (29). We found that oral nico-tinamide administration resulted in increased circulating Trp in HIV-infected patients, but followup is needed to determine the clinical implications of this result (4, 29). Nicotinamide and other endogenous TOx-related compounds may ultimately prove to be useful adjuncts to therapy or lead compounds for the development of new pharmacological agents to treat HIV and other diseases associated with TOx-induced pathogenesis.