Research

Immune Interactions of Carbohydrate Antigens in Health and Disease

Avci Lab is an interdisciplinary research group addressing problems at the interface of immunology and glycobiology. Our objective is to explore treatment of and protection from infectious diseases and cancer by understanding key molecular and cellular interactions between immune system and carbohydrate antigens associated with microbes or cancerous cells. Avci Lab also investigates immunoregulatory properties of glycans associated with symbiotic bacteria inhabiting host gastrointestinal tract to enable development and healthy-functioning of the immune system. Our research program delineates immune mechanisms involved in carbohydrate-mediated effector and regulatory immune responses, and designs and tests prophylactic and therapeutic agents targeting model pathogens, symbionts and cancers. Major research projects investigated in our lab are briefly described below.

1. Molecular Mechanisms for Carbohydrate Presentation to CD4+ T cells by MHCII Pathway. Most pathogenic bacterial surfaces are decorated with capsular polysaccharides (CPSs). Due to their unique structures and accessibility to immune surveillance, CPSs play critical roles in bacterial interactions with the immune system. Exploiting their high antigenicity, in the past three decades CPSs have been used as main components of glycoconjugate vaccines in clinical practice worldwide. While glycoconjugate vaccines have provided great health benefits in controlling bacterial diseases, their chemical conjugations have often been empirical, with weakly controlled conjugation chemistries resulting in poorly characterized, heterogeneous and variably immunogenic glycoconjugate vaccines. Especially problematic has been the inadequate understanding of the mechanisms by which the immune system interacts with these complex antigens. We previously demonstrated a novel mechanism through which uptake of a glycoconjugate vaccine by antigen presenting cells (APCs) results in the presentation of a carbohydrate epitope by the major histocompatibility class II complex (MHCII), thus stimulating carbohydrate-specific CD4+ T-cell clones (Tcarbs). With this discovery, we demonstrated for the first time that T cells could indeed specifically recognize carbohydrates, which had previously been considered to be “T cell independent” antigens.

Our previous discovery left unanswered critical questions on how this new class of T cell epitopes induces adaptive immune activation at the molecular/structural level. Structural insights would greatly assist in the rational design of conjugate vaccines with optimal immunogenic properties. Our NIH funded project delineates the mechanisms of carbohydrate mediated CD4+ T cell dependent immune responses by employing a model bacterial polysaccharide antigen (capsular polysaccharide of type 3 Streptococcus pneumoniae, Pn3P) and its protein or peptide conjugates.

2. Demystifying Adaptive Immune Mechanisms Induced by HIV Envelope Glycoprotein. Human immunodeficiency virus-1 (HIV-1) has been a major threat to human health and a protective acquired immunodeficiency syndrome (AIDS) vaccine has not as yet been developed. Since the start of the pandemic, AIDS has claimed over 40 million lives. The design of the current generation of vaccines does not make use of immune system activation mechanisms to maximize stimulation of critical immune cells (i.e., helper T cells) involved in producing protective antibodies. Most current HIV vaccine strategies have reached saturation and are largely modification of past unsuccessful efforts. A new perspective to HIV vaccine research is much needed. We proposed an innovative approach with the potential of establishing a new paradigm that the human CD4+ T cell repertoire contains a population of carbohydrate-specific T cells (i.e., Tcarbs) that recognize the N-glycan shield of gp120. The HIV-1 surface is decorated with a heavily glycosylated envelope protein called gp120, whose interaction with the CD4 molecule is the key step for the virus’s entry into CD4+ T cells. We hypothesize that recruitment of Tcarbs will not only induce T cell proliferation and memory, but will also induce production of protective, high-affinity antibodies by B cells through mechanisms such as affinity maturation and antibody class-switch. We believe our proposed studies will yield a new platform to develop a new-generation of truly protective future HIV vaccines. 

3. Aberrant Tumor Glycosylation Modulates the Immune Response. Cancer remains one of the most significant, prevalent, and insidious diseases, largely due to the primary lack of curative responses to most forms and/or stages of the developing disease. Hallmarks of cancer include tumor-promoting inflammation and avoiding destruction by the immune system. Therefore, one key problem observed in many cancers is modulation of the immune system to prevent effective anti-cancer responses. Vaccines have been studied to prime the immune system against cancer-associated antigens, thus promoting destruction of the cancer cells via immune cytotoxicity. However, vaccines have largely been ineffective against cancer cells due to problems with antigen targeting. As cancer cells can closely resemble normal host cells, the immune system may regulate itself to prevent cancer-directed cellular cytotoxicity. During malignant transformation of a mammalian cell, dramatic and aberrant alteration in cellular glycosylation is observed. While, the carbohydrate moieties associated with tumor cells (tumor associated carbohydrate antigens, or TACAs) serve as potential cancer vaccine targets, they have been demonstrated to induce immune suppression for cancer cells to avoid being targeted by cytotoxic immune cells. Such TACA-facilitated mechanisms would allow cancer cells to persist and possibly metastasize without generating an effective immune response.

In our studies, we have identified a novel immune cell population that arises in developing tumor tissue and characterized by their surface expression of a specific TACA-binding lectin. We hypothesize that the cellular signaling resulting from this carbohydrate-protein interaction inhibits immune effector mechanisms (i.e., through immunosuppressive cytokine secretion) and promotes tumor persistence and metastasis.  Our studies elucidate many aspects of differential cell signaling, such as how cancer cells can use signaling processes to “turn on” pro-inflammatory or checkpoint mechanisms, and/or “turn off” antibody-dependent cytotoxic pathways.  Furthermore, we identify how these processes compare with tumor tissues from human cancer patients, and the degree to which tumor glycan-immunity crosstalk affects patient outcomes.  Learning how glycosylation affects these pathways could thus provide mechanisms by which the immune system can be modulated to target cancer cells, even those outside of the primary tumor.

4. Immunomodulatory roles of symbiotic bacteria inhabiting host gastrointestinal tract. The importance of gut microbiota in disease progression and/or treatment has gained recognition in recent years. Previous research has indicated a significant role for microbiota in regulating immune cell activity.  However, there is little understanding of what the mechanism of action of these microbes are, which antigens or signaling molecules may be presented, and/or which immune cells are being recruited or suppressed, particularly in the context of disease.  One such immune cell population that may be affected by changing microbiota populations are MGL2+ immune cells.  These cells have been shown to have regulatory functions in the context of disease and cancer, and may be recruited or differentiated in response to antigen changes associated with different strains of bacteria in the gut.  Our preliminary studies have shown that a population of IL10-secreting, CD301b+ immune cells is associated with the symbiotic bacteria inhabiting gastrointestinal tract. We are currently investigating MGL2-expressing immune cells in the context of inflammation-induced colon cancer and the gut microbiome.  We are particularly interested in how these cell populations are affected by altered glycan expression on microbiota populations in developing colon carcinoma and colitis, and how their regulatory functions can be modulated to affect disease outcome and therapies.

5. Structural and Functional Characterization of the Protein Glycosylation in pneumoniae. Despite a century of in-depth investigation of Streptococcus Pneumoniae (Spn), the bacterium remains to have uncharacterized virulence mechanisms as a major human pathogen. Invasive pneumococcal diseases (IPDs) such as pneumonia, otitis media, bacteremia, and meningitis are particularly worrisome with alarming mortality rates. Since the introduction of the 7-valent (PCV7) and 13-valent (PCV13) glycoconjugate vaccines that are effective against the most prevalent serotypes of S. pneumoniae, the incidence rates of IPD in children have been reduced significantly, with limited reduction in morbidity in elderly and immunocompromised individuals. However, a global serotype distribution shift after conjugate vaccine introduction has highlighted the importance of identifying conserved immunogens to include a wider range of serotypes. Moreover, increasing numbers of clinical isolates from IPD patients contain nonencapsulated Spn (NESpn), indicating the emergence of pathogenic NESpn strains. The serotype distribution shift and the emergence of NESpn in the clinical isolates (up to 15% of isolates in multiple reports) necessitate the urgent investigation of serotype-independent, conserved, protective subunit vaccine targets. Spn surface proteins are major virulence factors and conserved immunogens and, therefore, recombinantly expressed Spn proteins have been examined as vaccine candidates without consideration of their glycosylation. Moreover, Spn contains numerous uncharacterized glycosyltransferases (GTs) that are putatively responsible for protein glycosylation. Importantly, a large scale screening of Spn mutant strains revealed that some of the putative GT genes investigated in these studies contribute to Spn virulence. However, our knowledge of the potential glycosylation of surface proteins by Spn and how this posttranslational modification impacts virulence and immunogenicity is surprisingly very limited.

Our lab elucidates Spn protein glycosylation to determine its role in bacterial virulence and host immune response against this pathogen. Thus, we hypothesize that glycan epitopes from Spn surface glycoproteins and the glycosyltransferases responsible for Spn protein glycosylation contribute to the virulence of the bacterium. We also hypothesize that that these glycan epitopes stimulate immune responses to contribute to the clearance of pathogen. We have generated an array of glycosyltransferase (GT) knock-out strains and are currently investigating potential roles these GTs play in Spn virulence and the host immune interactions of Spn glycoproteins. We found putative links between virulence, GTs and protein glycosylation.