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Alfredo Garzino-Demo, PhD

Academic Title:

Associate Professor

Primary Appointment:

Microbiology and Immunology

Additional Title:

Laboratory of Virus-Host Interactions, Division of Basic Science, Institute of Human Virology

Location:

Office: IHV S622; Laboratory: IHV S610

Phone (Primary):

(410) 706-4689

Fax:

(410) 706-4694

Education and Training

University of Torino, Italy, Research Doctorate (equivalent to Ph.D.), Human Oncology, 1994

Visiting Fellow, Laboratory of Tumor Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 1991-1995

Visiting Associate, Laboratory of Tumor Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD, 1995-1996

Research Associate, Institute of Human Virology, University of Maryland Biotechnology Institute, 1996

Biosketch

Throughout my career, I have coupled my strength in molecular biology with my ability to interact with clinician colleagues, so that I can investigate both the basic mechanisms underlying our findings, and how our lab observation apply to a “big picture”, i.e. clinical, setting. Therefore, I am extremely interested in translational studies. My training and track record are in molecular virology. During my postdoctoral years, at the Laboratory of Tumor Cell Biology at NCI, NIH, I studied the effect of viral cofactors such as HHV-6 on the progression of HIV infection, and HIV gene expression regulation. The main focus of my research activity for the last 15 year is on the role of host factors in controlling HIV infection. I was a member of the team that originally identified RANTES, MIP-1α and MIP-1ß as suppressive HIV-suppressive factors in 1995. After joining as faculty the Institute of Human Virology of the University of Maryland in January 1996, I participated to the characterization of another chemokine, MDC, with broad HIV-suppressive activity. In addition, I was the key investigator in a large study of antigen-induced chemokine release in a well characterized adult cohort (MACS cohort). That highly-cited manuscript showed my ability to successfully collaborate with clinicians to evaluate the role of immune system factors in HIV infection. I applied this philosophy to subsequent studies on the role of MCP-1 and the HIV gene tat in neurological AIDS. Recently, we described the HIV-inhibitory activity of beta-defensins, hypothesizing that hBD2 inhibits HIV through a novel intracellular antiviral mechanism of likely importance in mucosal immunity. Our publication in Blood shows that this activity is mediated by induction of the antiviral molecule APOBEC3G via CCR6, a shared chemokine/defensin receptor. We are now investigating the molecular mechanism underlying the induction of APOBEC3G by CCR6 ligands, investigating the intracellular pathway activated by that chemokine receptor. We expanded our studies to Th17 cells and macrophages. Recently, we reported on the antiviral and anti-inflammatory activities of a retrocyclin defensin, RC101, resulting in protection of a mouse model of influenza. In summary, my research  interests revolve around mechanistic studies on defensins, and we are aiming, in the case of HIV, in using defensins in translational research as topical microbicide. In the case of Dengue, and Zika viruses, we intend to fully exploit their translational potential, as they are ideal in their broad microbicidal activity against a broad range of pathogens, including select agents.

Research/Clinical Keywords

HIV; Defensins; chemokines; Chemokine receptors; T cells; Macrophages; NeuroAIDS; antiretrovirals

Highlighted Publications

Prantner D, Shirey KA, Lai W, Lu W, Cole AM, Vogel SN, Garzino-Demo A. The θ-defensin retrocyclin 101 inhibits TLR4- and TLR2-dependent signaling and protects mice against influenza infection.. Leukoc Biol. 2017 Jul 20. pii: jlb.2A1215-567RR. doi: 10.1189/jlb.2A1215-567RR. PubMed PMID: 28729359  PubMed Central PMCID: PMC5454423

Lafferty MK, Sun L, Christensen-Quick A, Lu W, Garzino-Demo A. Human Beta Defensin 2 Selectively Inhibits HIV-1 in Highly Permissive CCR6⁺CD4⁺ T Cells. Viruses. 2017 May 16;9(5). pii: E111. doi: 10.3390/v9050111. PubMed PMID: 28509877

Carroll VA, Lafferty MK, Marchionni L, Bryant JL, Gallo RC, Garzino-Demo A. Expression of HIV-1 matrix protein p17 and association with B-cell lymphoma in HIV-1 transgenic mice. Proc Natl Acad Sci U S A. 2016 Nov 15;113(46):13168-13173. PubMed PMID: 27799525; PubMed Central PMCID: PMC5135339.

Christensen-Quick A, Lafferty M, Sun L, Marchionni L, DeVico A, Garzino-Demo A. Human Th17 Cells Lack HIV-Inhibitory RNases and Are Highly Permissive to Productive HIV Infection. J Virol. 2016 Sep 1;90(17):7833-47. PubMed PMID: 27334595; PubMed Central PMCID: PMC4988157.

Lafferty MK, Sun L, DeMasi L, Lu W, Garzino-Demo A. CCR6 ligands inhibit HIV by inducing APOBEC3G. Blood. 2010 Feb 25;115(8):1564-71. PubMed PMID: 20023216; PubMed Central PMCID: PMC2830761.

Sun L, Finnegan CM, Kish-Catalone T, Blumenthal R, Garzino-Demo P, La Terra Maggiore GM, Berrone S, Kleinman C, Wu Z, Abdelwahab S, Lu W, Garzino-Demo A. Human beta-defensins suppress human immunodeficiency virus infection: potential role in mucosal protection. J Virol. 2005 Nov;79(22):14318-29. PubMed PMID: 16254366; PubMed Central PMCID: PMC1280242.

Garzino-Demo A, Moss RB, Margolick JB, Cleghorn F, Sill A, Blattner WA, Cocchi F, Carlo DJ, DeVico AL, Gallo RC. Spontaneous and antigen-induced production of HIV-inhibitory beta-chemokines are associated with AIDS-free status. Proc Natl Acad Sci U S A. 1999 Oct 12;96(21):11986-91. PubMed PMID: 10518563; PubMed Central PMCID: PMC18399.

Conant K, Garzino-Demo A, Nath A, McArthur JC, Halliday W, Power C, Gallo RC, Major EO. Induction of monocyte chemoattractant protein-1 in HIV-1 Tat-stimulated astrocytes and elevation in AIDS dementia. Proc Natl Acad Sci U S A. 1998 Mar 17;95(6):3117-21. PubMed PMID: 9501225; PubMed Central PMCID: PMC19704.

Additional Publication Citations

Complete List of Published Work in My Bibliography:

http://bit.ly/2mZzR8O

Research Interests

1) Immunopathogenesis of HIV infection, and novel therapeutic approaches. In the last few years, we have identified CD4+CCR6+ T cells as key targets of HIV infection. During last year, we published a study where we characterized the susceptibility and permissivity of CCR6+Th17 cells to HIV infection. CCR6+Th17cells play a crucial role in AIDS pathogenesis. HIV-1 infection (and its corresponding pathogenic SIV infection model in macaques) inflicts early and severe immune damage by depleting the IL-17-producing subset of CD4+ T helper (Th17) cells in the gut mucosa. Th17 cells, which are not replenished over time, are also likely depleted in the oral mucosa upon SIV/HIV-1 infection. IL-17 up-regulates epithelial cell production of anti-microbial peptides (AMP), including defensins and histatins which have potent activity against bacteria, fungi, and viruses and therefore, Th17 cells play a key role in mucosal barrier protection. Thus, lower levels of Th17 cells in the gut and oral mucosa compromises barrier integrity and allows translocation of microbial products from the intestinal lumen to the systemic circulation. This process triggers chronic immune activation, a strong predictor of progression to AIDS. In contrast, a decline of Th17 cells is not observed in HIV-1 long-term non-progressors or in the non-pathogenic SIV infected macaque model. We have shown that expression of the AMP human β-defensin 2 (hBD2) and secretion of histatin-5 (Hst-5) are markedly decreased in the oral mucosa and saliva, respectively, in HIV+ subjects. Based on our findings, we have proposed that the decrease in AMPs expression contributes to increased susceptibility to opportunistic infections. Therefore, based on our findings that β-defensins up-regulate the HIV restriction factor APOBEC3G via CCR6 high levels of hBD2 would make CCR6+ cells, including Th17 cells, less permissive to HIV-1 infection.

Based on these combined findings, we hypothesize that depletion of Th17 cells in the oral mucosa due to HIV infection contributes to hBD2 down-regulation in HIV-1 infected patients. This process further aggravates Th17 depletion in a self-perpetuating cycle, hampering immune protection against HIV-1 itself. Subsequently, the depletion of Th17 cells results in decreased production of antimicrobial pep-tides that may help explain why oral manifestations of HIV infection (opportunistic pathogens, such as C. albicans, HPV) are still observed even in the era of combination antiretroviral therapy (ART). Further, Th17 cells are involved in control of Mycobacterium tuberculosis, so that we are exploring the status of Th17 cells in conifection HIV/Mtb. Finally, we are testing approaches to restore hBD2 levels, or pharmacological interventions to trigger CCR6-mediated up-regulation of APOBEC3G to induce resistance to HIV infection.

2) Retrocyclin defensins. Approximately 50% of the world’s population is susceptible to Dengue (DENV), flaviviruses that cause vector-borne diseases, transmitted by Aedes aegypti or Ae. albopticus. Dengue fever is an incapacitating febrile illness, characterized by retro-orbital pain, headache, skin rash and muscle/bone pain. A more severe, potentially fatal condition, dengue hemorrhagic fever (DHF) is observed in individuals who become re-infected, and it is thought that the presence of cross-reactive antibodies increases infectivity. DHF is characterized by low platelet counts, vascular leakage, and hemorrhage. There is no current therapy or vaccine for DENV so that therapeutic approaches are needed. Another flavivirus, Zika virus (ZIKV), is also transmitted by mosquitos and can cause asymptomatic disease, but recently has been associated with increased incidence of Guillan-Barré syndrome, and several complications of pregnancy, including miscarriages and fetal malformation, including microcephaly. The World Health Organization declared Zika fever a Public Health Emergency of International Concern in 2016. Both Dengue and ZIKV are spread chiefly by an arthropod vector which is present in the US. The health impact of Dengue and Zika viruses makes it imperative to identify new treatments or preventive approaches.

Defensins are broad-spectrum antimicrobial peptides (3-5 kD) expressed in all vertebrates. While defensins disrupt membrane integrity of microbes and viruses, their interactions with a variety of microbial and cellular targets also include multiple other mechanisms of activity against a broad range of unrelated microbialpathogens. There are three subfamilies of defensins, defined as alpha (α), beta (ß), and theta (θ). α- and ß defensins share a similar structure that includes three beta strands stabilized by three disulfide bonds. The topology of disulfide bridges distinguishes α- from ß-defensins. θ-defensins were initially identified in the Rhesus macaque, and they are 18-residue cyclic peptides with three disulfide bridges arranged in a ladder pattern. In humans, a stop codon results in a pseudogene that is expressed as mRNA. However, the sequence information of this pseudogene was used to resurrect the putative coding sequence of human θ-defensins, and the resulting peptide has been named “retrocyclin.” This peptide exhibits high potency and a broad range of antimicrobial activities, in the absence of cellular toxicity, including bacteria (e.g., E. coli, S. aureus, L. monocytogenes, Ps. aeruginosa, and B. anthracis (bacilli and spores), viruses (e.g., HIV, influenza, HSV-1 and -2, dengue virus) and fungi (e.g., C. albicans) (1). In addition, θ-defensins suppress inflammatory responses in vivo in murine models of bacteremic sepsis and influenza infection (2). Thus retrocyclin-based treatments, with their dual antimicrobial and anti-inflammatory activities, could be used to treat infections. Recent technical progress in peptide synthesis makes mass production of retrocyclin a reality, so that its use in preventing and treating microbial diseases is desirable and feasible. Furthermore, the ease with which they can be synthesized according to their canonical sequence or in mutated variants permits their use in investigating the structural determinants of their interference with TLR signaling pathways. We aim at developing new therapeutics (antimicrobial and anti-inflammatory) based on retrocyclin defensins.

3) New antiretrovirals. Highly active antiretroviral therapy (HAART) has revolutionized human immunodeficiency virus (HIV) healthcare turning it into a chronic disease although patients can develop comorbidities which impact them. These include neurological and cognitive complications such as HIV associated neurological disorders (HAND) which can result in minor problems with memory to severe dementia-like symptoms. Even though small molecule treatments are highly effective in the periphery, virus can remain in the central nervous system (CNS) and replicate which then results in the neurological disorders, this may be due to the inability of HIV medications to cross the blood brain barrier (BBB) and inhibit HIV in the brain, which is critical. Several reasons may explain why combination antiretroviral therapy (cART) is inefficient at preventing HAND, such as poor CNS penetration and incomplete inhibition of HIV replication in this anatomical compartment, drug resistance, cART neurotoxicity, or irreversible brain damage prior to initiation of cART. Macrophages, particularly in the CNS, are likely components of the persistent reservoir that resist HIV eradication. However, cART has limited effect in macrophages, due to their scarce phosphorylation activity, which limits the activity of nucleoside analogs, and the expression of P-gp transporters, which pump out protease inhibitors. There are also issues relating to drug-drug interactions between HIV treatments and therefore it is important there is no inhibition of P450’s . Recent studies have shown that the HIV non-nucleoside reverse transcriptase inhibitor (NNRTI) efavirenz has the greatest frequency of neurologic side effects among newer HIV drugs and in vitro it has been shown to induce neural stem cells, while in vivo it has also increased apoptosis. HIV protease inhibitors ritonavir or lopinavir have been shown to display reversible dose-dependent decreases in oligodendrocyte maturation. Clearly it would be important to avoid these issues with new drugs. Accordingly, our hypothesis is that optimized, novel non-nucleoside reverse transcriptase inhibitors that cross the BBB are effective at preventing viral replication in the brain and the periphery and will also have lower incidence of HAND. We established an academic-industrial partnership between the Research Center of Biotechnology RAS, University of Maryland School of Medicine, and Collaborations Pharmaceuticals, Inc. We have assembled an outstanding, highly synergistic team of researchers who collectively possess the expertise required to translate the initial findings into a drug discovery program that is able to progress through the entire spectrum of pre-clinical studies, assessment of efficacy and safety evaluation. Playing to our respective strengths, we will use a combination of medicinal chemistry, computational drug discovery, in vitro assessment against HIV and assessment of BBB entry, absorption, distribution, metabolism and excretion (ADME) properties to develop a new generation of NNRTI.

Grants and Contracts

“New generation of non-nucleoside RT HIV inhibitors avoiding HIV-associated neurocognitive disorders” NIH/NINDS R01 NS102164-01