An injectable signal-amplifying device elicits a specific immune response against malignant glioblastoma

Despite exciting achievements with some malignancies, immunotherapy for hypoimmunogenic cancers, especially glioblastoma (GBM), remains a formidable clinical challenge. Poor immunogenicity and deficient immune infiltrates are two major limitations to an effective cancer-specific immune response. Herein, we propose that an injectable signal-amplifying nanocomposite/hydrogel system consisting of granulocyte-macrophage colony-stimulating factor and imiquimod-loaded antigen-capturing nanoparticles can simultaneously amplify the chemotactic signal of antigen-presenting cells and the "danger" signal of GBM. We demonstrated the feasibility of this strategy in two scenarios of GBM. In the first scenario, we showed that this simultaneous amplification system, in conjunction with local chemotherapy, enhanced both the immunogenicity and immune infiltrates in a recurrent GBM model; thus, ultimately making a cold GBM hot and suppressing postoperative relapse. Encouraged by excellent efficacy, we further exploited this signal-amplifying system to improve the efficiency of vaccine lysate in the treatment of refractory multiple GBM, a disease with limited clinical treatment options. In general, this biomaterial-based immune signal amplification system represents a unique approach to restore GBM-specific immunity and may provide a beneficial preliminary treatment for other clinically refractory malignancies.

Antitumor synergism between PAK4 silencing and immunogenic phototherapy of engineered extracellular vesicles

Immunotherapy has revolutionized the landscape of cancer treatment. However, single immunotherapy only works well in a small subset of patients. Combined immunotherapy with antitumor synergism holds considerable potential to boost the therapeutic outcome. Nevertheless, the synergistic, additive or antagonistic antitumor effects of combined immunotherapies have been rarely explored. Herein, we established a novel combined cancer treatment modality by synergizing p21-activated kinase 4 (PAK4) silencing with immunogenic phototherapy in engineered extracellular vesicles (EVs) that were fabricated by coating M1 macrophage-derived EVs on the surface of the nano-complex cores assembled with siRNA against PAK4 and a photoactivatable polyethyleneimine. The engineered EVs induced potent PAK4 silencing and robust immunogenic phototherapy, thus contributing to effective antitumor effects in vitro and in vivo. Moreover, the antitumor synergism of the combined treatment was quantitatively determined by the CompuSyn method. The combination index (CI) and isobologram results confirmed that there was an antitumor synergism for the combined treatment. Furthermore, the dose reduction index (DRI) showed favorable dose reduction, revealing lower toxicity and higher biocompatibility of the engineered EVs. Collectively, the study presents a synergistically potentiated cancer treatment modality by combining PAK4 silencing with immunogenic phototherapy in engineered EVs, which is promising for boosting the therapeutic outcome of cancer immunotherapy.

Tat-functionalized Ag-FeO nano-composites as tissue-penetrating vehicles for tumor magnetic targeting and drug delivery

In this paper, we prepared a dual functional system based on dextrin-coated silver nanoparticles which were further attached with iron oxide nanoparticles and cell penetrating peptide (Tat), producing Tat-modified Ag-FeO nanocomposites (Tat-FeAgNPs). To load drugs, an -SH containing linker, 3-mercaptopropanohydrazide, was designed and synthesized. It enabled the silver carriers to load and release doxorubicin (Dox) in a pH-sensitive pattern. The delivery efficiency of this system was assessed using MCF-7 cells, and using null BalB/c mice bearing MCF-7 xenograft tumors. Our results demonstrated that both Tat and externally applied magnetic field could promote cellular uptake and consequently the cytotoxicity of doxorubicin-loaded nanoparticles, with the IC of Tat-FeAgNP-Dox to be 0.63 µmol/L. The delivery efficiency of Tat-FeAgNP carrying Cy5 to the mouse tumor was analyzed using the optical imaging tests, in which Tat-FeAgNP-Cy5 yielded the most efficient accumulation in the tumor (6.7±2.4% ID of Tat-FeAgNPs). Anti-tumor assessment also demonstrated that Tat-FeAgNP-Dox displayed the most significant tumor-inhibiting effects and reduced the specific growth rate of tumor by 29.6% ( = 0.009), which could be attributed to its superior performance in tumor drug delivery in comparison with the control nanovehicles.

Improved method for synthesis of low molecular weight protamine-siRNA conjugate

RNAi technology has aroused wide public interest due to its high efficiency and specificity to treat multiple types of diseases. However, the effective delivery of siRNA remains a challenge due to its large molecular weight and strong anionic charge. Considering their remarkable functions and features that are often desired in drug delivery carriers, biomimetic systems for siRNA delivery become an effective and promising strategy. Based on this, covalent attachment of synthetic cell penetrating peptides (CPP) to siRNA has become of great interest. We developed a monomeric covalent conjugate of low molecular weight protamine (LMWP, a well-established CPP) and siRNA a cytosol-cleavable disulfide linkage using PEG as a crosslinker. Results showed that the conjugates didn't generate coagulation, and exhibited much better RNAi potency and intracellular delivery compared with the conventional charge-complexed CPP/siRNA aggregates. Three different synthetic and purification methods were compared in order to optimize synthesis efficiency and product yield. The methodology using hetero-bifunctional NHS-PEG-OPSS as a crosslinker to synthesize LMWP-siRNA simplified the synthesis and purification process and produced the highest yield. These results pave the way towards siRNA biomimetic delivery and future clinical translation.

Nose-to-brain delivery of macromolecules mediated by cell-penetrating peptides

Brain delivery of macromolecular therapeutics (e.g., proteins) remains an unsolved problem because of the formidable blood-brain barrier (BBB). Although a direct pathway of nose-to-brain transfer provides an answer to circumventing the BBB and has already been intensively investigated for brain delivery of small drugs, new challenges arise for intranasal delivery of proteins because of their larger size and hydrophilicity. In order to overcome the barriers and take advantage of available pathways (e.g., epithelial tight junctions, uptake by olfactory neurons, transport into brain tissues, and intra-brain diffusion), a low molecular weight protamine (LMWP) cell-penetrating peptide was utilized to facilitate nose-to-brain transport. Cell-penetrating peptides (CPP) have been widely used to mediate macromolecular delivery through many kinds of biobarriers. Our results show that conjugates of LMWP-proteins are able to effectively penetrate into the brain after intranasal administration. The CPP-based intranasal method highlights a promising solution for protein therapy of brain diseases.