Hitchhiking Across the Mucosa

Many pathogens, ranging from HIV to SARS-CoV-2, are transmitted through mucosal surfaces and are thought to require a ‘frontline’ immune response in the mucosa as well as ‘backup’ defense in the blood for effective protection. Yet, traditional parenteral vaccines that are administered by subcutaneous or intramuscular injection typically elicit poor mucosal immunity. While vaccination at mucosal surfaces is known to activate mucosal immunity, development of mucosal vaccines has long been plagued by poor uptake and delivery of vaccine components across mucosal barriers, resulting in weak immune responses. As a result, only a small number of mucosal vaccines have reached the clinic, most of which are based on live attenuated pathogens that naturally infect mucosal surfaces, which cannot be used by transplant patients and others who are immunocompromised. Thus, development of technologies to overcome barriers to mucosal delivery while meeting safety and efficacy requirements of prophylactic vaccines remains an urgent unmet need. 

While mucosal barriers are very good at keeping most vaccine components out, the endogenous protein albumin is known to be very good at getting in. Albumin, a major blood protein also present in mucosal fluids that functions in vivo as a fatty acid transporter, is constitutively transcytosed across mucosal barriers by interactions with the neonatal Fc receptor (FcRn) expressed on mucosal epithelial cells. We are developing intranasal amphiphile vaccines that exploit this biology by 'hitchhiking' on albumin across mucosal barriers for more efficient uptake into the underlying nasal-associated lymphoid tissue (NALT). 'Amph vaccines', consisting of immunogens conjugated to an albumin-binding amphiphilic tail, exhibit enhanced FcRn-dependent uptake into the nasal mucosal tissue leading to enhanced germinal center responses in the NALT. Compared to soluble protein antigens or parenterally administered antigens, intranasal amph vaccines prime significant systemic and mucosal humoral immune responses across multiple mucosal tissues. We are working on continued development of this vaccine platform against a range of mucosally-transmitted pathogens, correlating vaccine design parameters with mucosal uptake and functional immune protection. Employing amph-vaccines to deliver antigen across the mucosal epithelium is a promising strategy to promote mucosally-transmitted pathogens such as HIV, SARS-CoV-2, influenza, and cytomegalovirus (CMV).

[1] Intranasal vaccine uptake is typically impeded by rapid mucociliary clearance, degradation in the mucosal layer, and lack of diffusive uptake across mucosal epithelium. Amph vaccines use endogenous albumin, which is trancytosed across mucosal epithelium by binding the neonatal Fc receptor (FcRn), as a chaperone to hitchhike across mucosal barriers for uptake into the underlying nasal-associated lymphoid tissue (NALT) in order to activate 'frontline' mucosal immune protection.

[2] 'Albumin hitchhiking' enables enhanced retention of amph vaccines in the nasal passage (B), efficient FcRn-mediated uptake into nasal epithelium and underlying submucosa (C), and activation of germinal center (GC) B cells in the NALT (D). (Adapted from Hartwell et al. Science Translational Medicine, 2022.)

Supporting publications:

Hartwell BL, Melo MB, Xiao P, Lemnios AA, Li N, Chang JYH, Yu J, Gebre MS, Chang A, Maiorino L, Carter C, Moyer TJ, Dalvie NC, Rodriguez-Aponte SA, Rodrigues KA, Silva M, Suh H, Adams J, Fontenot J, Love JC, Barouch DH, Villinger F, Ruprecht RM, Irvine DJ. “Intranasal vaccination with lipid-conjugated immunogens promotes antigen transmucosal uptake to drive mucosal and systemic immunity.” Science Translational Medicine. 2022 Jul 20;14(654):eabn1413. DOI: 10.1126/scitranslmed.abn1413.

Szoka FC Jr. “A hitchhiker's guide to mucosal and systemic immunity.” Science Translational Medicine. 2022 Jul 20;14(654):eadc8697. doi: 10.1126/scitranslmed.adc8697.

Tuning Immunity Through the Mucosa

Delivery of vaccines (antigen + adjuvant) to mucosal surfaces is known to be an effective strategy to promote protective immunity at barrier tissues through initiation of immune responses in underlying mucosa-associated lymphoid tissues (MALT). MALT includes the nasal-associated lymphoid tissue (NALT), bronchial-associated lymphoid tissue (BALT), and gut-associated lymphoid tissue (GALT). These tissues play a critical role in the resulting immune response (analogous to lymph nodes in the periphery) as sites where antigen-specific immune responses are orchestrated against mucosally-encountered antigens. Delivery of antigens to the MALT can drive programming of mucosa-specific lymphocyte function and mucosal tissue homing. Yet although well-motivated by the biology of mucosal immunity, delivery of vaccine components across mucosal barriers has presented a major challenge for mucosal vaccine development. 

At the same time, certain mucosal sites such as the gut and lungs are inherently predisposed to induce tolerance upon antigen exposure in the absence of costimulatory signals (i.e. adjuvant), making these tissue sites an attractive target for antigen delivery to treat autoimmunity. Oral antigen delivery to GALT, for instance, preferentially induces systemic tolerance through a specialized mechanism known as oral tolerance.  Despite a long history of using oral tolerance to treat animal models of autoimmunity, this approach has yet to successfully translate into humans. Like mucosal vaccination, a major barrier to clinical translation has been the high dose of antigen required to overcome limited uptake across mucosal barriers in the gut. Mucosal antigen delivery systems that avoid degradation and clearance while achieving efficient uptake at lower doses may address a need for safe and efficacious antigen-specific immunotherapies to treat autoimmunity.

Our lab uses delivery platforms that we have developed for efficient transmucosal uptake, such as albumin-hitchhiking amphiphile vaccines, as tools to explore strategies for 'tuning immunity through the mucosa'. The location of antigen exposure and orchestration of the immune response can be tailored by varying route of administration (i.e., intranasal / intrapulmonary / oral), while the context of antigen presentation can be tailored by varying costimulatory signals (i.e., +/- adjuvant) and microbial background (i.e. dirty vs. specific pathogen free mice). Through these projects we aim to elucidate mechanisms that drive immune activation versus tolerance in the mucosal immune compartment, providing insights that may be used to guide the design of mucosal vaccines and antigen-specific immunotherapies.

Tuning immunity through the mucosa by tailoring location of antigen exposure (in terms of mucosal barrier tissue and the underlying mucosa-associated lymphoid tissue) and context of antigen presentation (in terms of microbial background and costimulatory cues).

Supporting publications:

Hartwell BL*, Martin JT*, Kumarapperuma S*, Melo MB, Carnathan DG, Cossette BJ, Adams J, Gong S, Zhang W, Tokatlian T, Menis S, Schiffner T, Franklin CG, Goins B, Fox PT, Silvestri G, Schief WR, Ruprecht RM, Irvine DJ. “Combined PET and whole-tissue imaging of lymphatic-targeting vaccines in non-human primates.” Biomaterials. 2021; 275:120868. DOI: 10.1016/j.biomaterials.2021.120868

Engineering Immune Tolerance with Targeted Immunotherapies

Many current autoimmune disease therapies act through nonspecific suppression of the immune response, resulting in global immunosuppression and deleterious off-target effects. Antigen-specific immunotherapies that target and suppress only offending autoreactive immune cells to restore autoantigenic tolerance would address a pressing need for safer and more effective therapies, yet remain elusive. Endogenous mechanisms for maintaining immunological tolerance and homeostasis suggest however that restoration of immune tolerance is plausible. For example, mucosal tolerance against ingested antigens is maintained through mechanisms of oral tolerance, where antigens are taken up, processed, and presented in gut-associated lymphoid tissues (GALT) in a highly tolerogenic context. Peripheral tolerance is maintained primarily through a state of antigen unresponsiveness called anergy which occurs when lymphocytes (B or T cells) mount an initial response to primary antigenic signal but do not receive sufficient secondary costimulatory signals to sustain activation. B cells in particular, which contribute to autoimmune pathogenesis through antigen presentation and effector functions, have potential for direct antigen-specific targeting through the B cell receptor (BCR) and provide a promising target for tolerogenic immunotherapies. Multivalent nanomaterials (such as soluble antigen arrays, or cSAgAs) that present multiple copies of covalently-conjugated autoantigen can be tailored to induce anergy in autoimmune B cells through high avidity binding of the BCR in the absence of costimulatory signals to modulate BCR signaling. Design and development of effective cell-targeted antigen-specific immunotherapies should take into account appropriate physicochemical properties of delivered antigen in light of known molecular, cellular, and transport mechanisms for inducing peripheral tolerance. This project seeks to build upon endogenous mechanisms of tolerance to design targeted antigen-specific immunotherapies to treat autoimmune diseases.

Proposed mechanism for inducing B cell anergy with multivalent autoantigen arrays. (Adapted from Hartwell et al. Journal of Autoimmunity, 2018.)

Supporting publications:

Hartwell BL, Pickens CJ, Leon M, Northrup L, Christopher MA, Griffin JD, Martinez-Becerra F, Berkland C. “Soluble antigen arrays disarm antigen-specific B cells to promote lasting immune tolerance in EAE.” J Autoimmunity. 2018; 93(9):76-88. DOI: 10.1016/j.jaut.2018.06.006

Hartwell BL, Pickens CJ, Leon M, Berkland C. “Multivalent antigen arrays exhibit high avidity binding and modulation of B cell receptor-mediated signaling to drive efficacy against EAE.” Biomacromolecules. 2017; 18(6):1893-1907. DOI: 10.1021/acs.biomac.7b00335

Hartwell BL, Martinez-Becerra F, Chen J, Shinogle H, Sarnowski M, Moore D, Berkland C. “Antigen-specific binding of multivalent soluble antigen arrays induces receptor clustering and impedes B cell receptor mediated signaling.” Biomacromolecules. 2016; 17(3): 710-22. DOI: 10.1021/acs.biomac.5b01097

Hartwell BL, Smalter Hall A, Swafford D, Sullivan BP, Garza A, Sestak JO, Northrup L, Berkland C. “Molecular dynamics of multivalent soluble antigen arrays support two-signal codelivery mechanism in treatment of experimental autoimmune encephalomyelitis.” Molecular Pharmaceutics. 2016; 13(2): 330-43. DOI: 10.1021/acs.molpharmaceut.5b00825

Hartwell BL, Antunez L, Sullivan BP, Thati S, Sestak JO, Berkland C. “Multivalent nanomaterials: learning from vaccines and progressing to antigen-specific immunotherapies.” Journal of Pharmaceutical Science. 2015; 104(2): 346-61. Epub 2014. DOI: 10.1002/jps.24273

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