Pfs25 is a malaria transmission-blocking vaccine antigen candidate, but its apparently limited immunogenicity in humans has hindered clinical development. Here, we show that recombinant, his-tagged Pfs25 can be mixed at the time of immunization with pre-formed liposomes containing cobalt-porphyrin-phospholipid (CoPoP), resulting in spontaneous nanoliposome antigen particleization (SNAP). Antigens are stably presented in uniformly-oriented display via his-tag insertion in the CoPoP bilayer, without covalent modification or disruption of antigen conformation. SNAP immunization of mice and rabbits is well tolerated with minimal local reactogenicity and results in orders-of-magnitude higher functional antibody generation compared to other “mix-and-inject” adjuvants. Serum-stable antigen-binding during transit to draining lymph nodes leads to enhanced antigen uptake by phagocytic antigen presenting cells, with subsequent generation of long-lived, antigen-specific plasma cells. Seamless multiplexing with four additional his-tagged Plasmodium falciparum polypeptides induces strong and balanced antibody production, illustrating the simplicity of developing multi-stage particulate vaccines with SNAP immunization.

Prompt membrane permeabilization is a requisite for liposomes designed for local stimuli-induced intravascular release of therapeutic payloads. Incorporation of a small amount (i.e. 5 molar percent) of an unsaturated phospholipid, such as dioleoylphosphatidylcholine (DOPC), accelerated near infrared (NIR) light-triggered release in porphyrin-phospholipid (PoP) liposomes by an order of magnitude. In physiological conditions in vitro, 90% of the loaded drug could be released in a minute with NIR irradiation, while liposomes maintained serum stability in the absence of NIR irradiation. This enabled rapid laser-induced drug release using remarkably low amounts of PoP (i.e. 0.3 molar percent). Light-triggered drug release occurred concomitantly with DOPC and cholesterol oxidation, as detected by mass spectrometry. In the presence of an oxygen scavenger or an antioxidant, light-triggered drug release was inhibited, suggesting the light triggered release mechanism was caused by singlet oxygen mediated oxidization of unsaturated lipids. Despite the irreversible modification of lipid composition, DOPC-containing PoP liposome permeabilization was transient. Human pancreatic xenograft growth in mice was significantly delayed with a single chemophototherapy treatment following intravenous administration of 6 mg/kg doxorubicin, loaded in liposomes containing small amounts of DOPC and PoP.

Surfactant-stripped micelles are formed from a commercially available cyanine fluroalkylphosphate salt dye (CyFaP) and used for high contrast photoacoustic imaging in the second near infrared window (NIR-II). The co-loading of Coenzyme Q10 into surfactantstripped CyFaP (ss-CyFaP) micelles improves yield, storage stability and results in a peak absorption wavelength in the NIR-II window close to the 1064 nm output of Nd-YAG lasers used for photoacoustic imaging. Aqueous ss-CyFaP dispersions exhibit intense NIR-II optical absorption, calculated to be greater than 500 at 1064 nm. ss-CyFaP is detected through 12 cm of chicken breast tissue with PAI. In preclinical animal models, ss-CyFaP is visualized in

Doxorubicin (Dox)-loaded stealth liposomes (similar to those in clinical use) can incorporate small amounts of porphyrin-phospholipid (PoP) to enable chemophototherapy (CPT). PoP is also an intrinsic and intrabilayer Cu chelator, although how radiolabeling impacts drug delivery has not yet been assessed. Here, we show thatCu can radiolabel the stable bilayer of preformed Dox-loaded PoP liposomes with inclusion of 1% ethanol without inducing drug leakage. Dox-PoP liposomes labeled with intrabilayer copper behaved nearly identically to unlabeled ones in vitro and in vivo with respect to physical parameters, pharmacokinetics, and CPT efficacy. Positron emission tomography and near-infrared fluorescence imaging visualized orthotopic mammary tumors in mice with passive liposome accumulation following administration. A single CPT treatment with 665 nm light (200 J/cm) strongly inhibited primary tumor growth. Liposomes accumulated in lung metastases, based on NIR imaging. These results establish the feasibility of CPT interventions guided by intrinsic multimodal imaging of Dox-loaded stealth PoP liposomes.

Short major histocompatibility complex (MHC) class I (MHC-I)-restricted peptides contain the minimal biochemical information to induce antigen (Ag)-specific CD8+ cytotoxic T cell responses but are generally ineffective in doing so. To address this, we developed a cobalt–porphyrin (CoPoP) liposome vaccine adjuvant system that induces rapid particleization of conventional, short synthetic MHC-I epitopes, leading to strong cellular immune responses at nanogram dosing. Along with CoPoP (to induce particle formation of peptides), synthetic monophosphoryl lipid A (PHAD) and QS-21 immunostimulatory molecules were included in the liposome bilayer to generate the “CPQ” adjuvant system. In mice, immunization with a short MHC-I-restricted peptide, derived from glycoprotein 70 (gp70), admixed with CPQ safely generated functional, Ag-specific CD8+ T cells, resulting in the rejection of multiple tumor cell lines, with durable immunity. When cobalt was omitted, the otherwise identical peptide and adjuvant components did not result in peptide binding and were incapable of inducing immune responses, demonstrating the importance of stable particle formation. Immunization with the liposomal vaccine was well-tolerated and could control local and metastatic disease in a therapeutic setting. Mechanistic studies showed that particle-based peptides were better taken up by antigen-presenting cells, where they were putatively released within endosomes and phagosomes for display on MHC-I surfaces. On the basis of the potency of the approach, the platform was demonstrated as a tool for in vivo epitope screening of peptide microlibraries comprising a hundred peptides.