Gregory, J. V. | bioRxiv Science

bioRxiv2021

Systemic delivery of a CXCR4-CXCL12 signaling inhibitor encapsulated in synthetic protein nanoparticles for glioma immunotherapy

Glioblastoma multiforme (GBM) is an aggressive primary brain tumor, with poor prognosis. Major obstacles hampering effective therapeutic response in GBM are tumor heterogeneity, high infiltration of immunosuppressive myeloid cells, and the presence of the blood-brain barrier. The C-X-C Motif Chemokine Ligand 12/ C-X-C Motif Chemokine Receptor 4 (CXCL12/ CXCR4) signaling pathway is implicated in GBM invasion and cell cycle progression. While the CXCR4 antagonists (AMD3100) has a potential anti-GBM effects, its poor pharmacokinetic and systemic toxicity had precluded its clinical application. Moreover, the role of CXCL12/ CXCR4 signaling pathway in anti-GBM immunity, particularly in GBM-mediated immunosuppression has not been elucidated. Here, we developed a synthetic protein nanoparticle (SPNPs) coated with the cell-penetrating peptide iRGD (AMD3100 SPNPs) to target the CXCR4/CXCL12 signaling axis in GBM. We showed that AMD3100 SPNPs effectively blocked CXCR4 signaling in mouse and human GBM cells in vitro as well as in GBM model in vivo. This results in inhibition of GBM proliferation and induction of immunogenic tumor cell death (ICD) leading to inhibition of GBM progression. Our data also demonstrate that blocking CXCR4 sensitizes GBM cells to radiation, eliciting enhanced release of ICD ligands. Combining AMD3100 SPNPs with radiotherapy inhibited GBM progression and led to long-term survival; with 60% of mice remaining tumor-free. This was accompanied by an anti-GBM immune response and sustained immunological memory that prevented tumor recurrence without further treatment. Finally, we showed that systemic delivery of AMD3100 SPNPs decreased the infiltration of CXCR4+ monocytic myeloid-derived suppressor cells to the tumor microenvironment. With the potent ICD induction and reprogrammed immune microenvironment, this strategy has significant potential for future clinical translation. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/457953v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@17c9377org.highwire.dtl.DTLVardef@1b5a4dcorg.highwire.dtl.DTLVardef@5aa4eaorg.highwire.dtl.DTLVardef@1011a86_HPS_FORMAT_FIGEXP M_FIG Immunological mechanism targeting Glioblastoma (GBM) upon blocking CXCR4 signaling pathway with AMD3100-conjugated nanoparticles (SPNPs). (1) Radiotherapy induces glioma cell death, followed by Damage-associated molecular patterns (DAMPs) release. Dendritic cells (DC) are activated by DAMPs and migrate to the regional lymph node where they prime cytotoxic T lymphocyte immune response. Tumor-specific cytotoxic T cells infiltrate the tumor and attack glioma cells. (2) Glioma cells express CXCR4, as well its ligand CXCL12. CXCL12 induces glioma cell proliferation and, (3) as well as mobilization in the bone marrow of CXCR4 expressing myeloid MDSC, which will infiltrate the tumor, and inhibit tumor-specific cytotoxic T cells activity. GEMM of glioma when treated systemically with SPNPs AMD3100 SPNPs plus radiation, nanoparticles block the interaction between CXCR4 and CXCL12, thus (4) inhibiting glioma cell proliferation and (5) reducing mobilization in the bone marrow of CXCR4 expressing myeloid MDSC, (6) generating a reduced MDSC tumor infiltration, as well as releasing MDSC inhibition over tumor specific cytotoxic T cell response. C_FIG

cell biology↗

bioRxiv2020

Supramolecular Organization Predicts Protein Nanoparticle Delivery to Neutrophils for Acute Lung Inflammation Diagnosis and Treatment

Acute lung inflammation has severe morbidity, as seen in COVID-19 patients. Lung inflammation is accompanied or led by massive accumulation of neutrophils in pulmonary capillaries ("margination"). We sought to identify nanostructural properties that predispose nanoparticles to accumulate in pulmonary marginated neutrophils, and therefore to target severely inflamed lungs. We designed a library of nanoparticles and conducted an in vivo screen of biodistributions in naive mice and mice treated with lipopolysaccharides. We found that supramolecular organization of protein in nanoparticles predicts uptake in inflamed lungs. Specifically, nanoparticles with agglutinated protein (NAPs) efficiently home to pulmonary neutrophils, while protein nanoparticles with symmetric structure (e.g. viral capsids) are ignored by pulmonary neutrophils. We validated this finding by engineering protein-conjugated liposomes that recapitulate NAP targeting to neutrophils in inflamed lungs. We show that NAPs can diagnose acute lung injury in SPECT imaging and that NAP-like liposomes can mitigate neutrophil extravasation and pulmonary edema arising in lung inflammation. Finally, we demonstrate that ischemic ex vivo human lungs selectively take up NAPs, illustrating translational potential. This work demonstrates that structure-dependent interactions with neutrophils can dramatically alter the biodistribution of nanoparticles, and NAPs have significant potential in detecting and treating respiratory conditions arising from injury or infections.

bioengineering↗

bioRxiv2019

Systemic Brain Tumor Delivery of Synthetic Protein Nanoparticles for Glioblastoma Therapy

Glioblastoma multiforme (GBM), the most aggressive form of brain cancer, has witnessed very little clinical progress over the last decades, in parts, due to the absence of effective drug delivery strategies. Intravenous injection is the least invasive delivery route to the brain, but has been severely limited by the blood-brain barrier (BBB). Inspired by the capacity of natural proteins and viral particulates to cross the BBB, we engineered a synthetic protein nanoparticle (SPNP) based on polymerized human serum albumin (HSA) equipped with the cell-penetrating peptide iRGD. SPNPs containing siRNA against Signal Transducer and Activation of Transcription 3 factor (STAT3i) result in in vitro and in vivo downregulation of STAT3, a central hub associated with GBM progression. When combined with the standard of care, ionized radiation, STAT3i SPNPs result in tumor regression and long-term survival in 87.5% of GBM bearing mice and primes the immune system to develop anti-GBM immunological memory.

bioengineering↗