Carnosic acid nanocluster-based framework combined with PD-1 inhibitors impeded tumorigenesis and enhanced immunotherapy in hepatocellular carcinoma.

Clinically, the immune checkpoint inhibitor anti-PD-1 antibody has shown a certain effect in the treatment of hepatocellular carcinoma (HCC), which is limited to a small number of patients with HCC. This study aims to reveal whether carnosic acid nanocluster-based framework (CA-NBF) has a sensitization effect on anti-PD-1 antibody in the treatment of HCC at the cellular and animal levels. MHCC97H cells were treated with CA-NBF, anti-PD-1 and their combination. The effects of CA-NBF and anti-PD-1 on cell proliferation, cell cycle, apoptosis, invasion, and migration were evaluated by MTT assay, flow cytometry, and scratch test. The effects of CA-NBF and anti-PD-1 on Wnt/ß-catenin signaling pathway in MHCC97H cells were detected. A BALB/C nude mouse model of hepatocellular carcinoma was established, and the tumor growth was observed at different time points. The expression of cytotoxic T lymphocyte and helper T lymphocyte markers CD8 and CD4 in tumor tissues was detected by immunohistochemistry. Western blotting was used to detect the Wnt/ß-catenin signaling pathway proteins (Wnt-3a, ß-catenin, and GSK-3ß) level in tumor tissues after CA-NBF and anti-PD-1 treatment. CA-NBF activity was significantly higher than CA, which could prominently reduce the proliferation, migration and invasion of MHCC97H cells and enhance apoptosis by inactivating Wnt/ß-catenin signaling pathway. CA-NBF combined with anti-PD-1 antibody further enhanced cell proliferation, migration, invasion and pro-apoptosis but had no significant effect on Wnt/ß-catenin signaling pathway. CA-NBF in vivo improved the tumor response to PD1 immune checkpoint blockade in HCC, manifested by reducing tumor size and weight, promoting CD4 and CD8 expression. CA-NBF combined with anti-PD-1 have stronger immunomodulatory and anticancer effects without increasing biological toxicity.

Rewiring chaperone-mediated autophagy in cancer by a prion-like chemical inducer of proximity to counteract adaptive immune resistance.

Chaperone-mediated autophagy (CMA), a proteolytic system contributing to the degradation of intracellular proteins in lysosomes, is upregulated in tumors for pro-tumorigenic and pro-survival purposes. In this study, bioinformatics analysis revealed the co-occurrence of upregulated CMA and PD-L1 accumulation in metastatic melanoma with adaptive immune resistance (AIR) to anti-PD1 treatment, suggesting the potential therapeutic effects of rewiring CMA for PD-L1 degradation. Furthermore, this co-occurrence is attributed to IFN-γ-mediated compensatory up-regulation of PD-L1 and CMA, accompanied by enhanced macropinocytosis. Drawing inspiration from the cellular uptake of prions via macropinocytosis, a prion-like chemical inducer of proximity called SAP was engineered using self-assembly of the designed chiral peptide PHA. By exploiting sensitized macropinocytosis, SAP clandestinely infiltrates tumor cells and subsequently disintegrates into PHA, which reprograms CMA by inducing PD-L1 close to HSPA8. SAP degrades PD-L1 in a CMA-dependent manner and effectively restores the anti-tumor immune response in both allografting and Hu-PDX melanoma mouse models with AIR while upholding a high safety profile. Collectively, the reported SAP not only presents an immune reactivation strategy with clinical translational potential for overcoming AIR in cutaneous melanomas but serves as a reproducible example of precision-medicine-guided drug development that fully leverages specific cellular indications in pathological states.

A d-peptide-based oral nanotherapeutic modulates the PD-1/PD-L1 interaction for tumor immunotherapy.

Background: PD-1/PD-L1 immune checkpoint inhibitors are currently the most commonly utilized agents in clinical practice, which elicit an immunostimulatory response to combat malignancies. However, all these inhibitors are currently administered via injection using antibody-based therapies, while there is a growing need for oral alternatives. Methods: This study has developed and synthesized exosome-wrapped gold-peptide nanocomplexes with low immunogenicity, which can target PD-L1 and activate antitumor immunity in vivo through oral absorption. The SuperPDL1exo was characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS), Fourier transform infrared (FTIR), X-ray photoelectron spectroscopy (XPS), and gel silver staining. The transmembrane ability of SuperPDL1exo was evaluated by flow cytometry and immunofluorescence. Cell viability was determined using the Cell Counting Kit-8 (CCK-8) assay. ELISA experiments were conducted to detect serum and tissue inflammatory factors, as well as serum biochemical indicators. Tissue sections were stained with H&E for the evaluation of the safety of SuperPDL1exo. An MC38 colon cancer model was established in immunocompetent C56BL/6 mice to evaluate the effects of SuperPDL1exo on tumor growth in vivo. Immunohistochemistry (IHC) staining was performed to detect cytotoxicity factors such as perforin and granzymes. Results: First, SuperPDL1 was successfully synthesized, and milk exosome membranes were encapsulated through ultrasound, repeated freeze-thaw cycles, and extrusion, resulting in the synthesis of SuperPDL1exo. Multiple characterization results confirmed the successful synthesis of SuperPDL1exo nanoparticles. Furthermore, our data demonstrated that SuperPDL1exo exhibited excellent colloidal stability and superior cell transmembrane ability. In vitro and in vivo experiments revealed that SuperPDL1exo did not cause damage to multiple systemic organs, demonstrating its good biocompatibility. Finally, in the MC38 colon cancer mouse model, it was discovered that SuperPDL1exo could inhibit the progression of colon cancer, and this tumor-suppressive effect was mediated through the activation of tumor-specific cytotoxic T lymphocyte (CTL)-related immune responses. Conclusion: This study has successfully designed and synthesized an oral nanotherapeutic, SuperPDL1exo, which demonstrates small particle size, excellent colloidal stability, transmembrane ability in tumor cells, and biocompatibility. In vivo experiments have shown that it effectively activates T-cell immunity and exerts antitumor effects.

Vps21 Directs the PI3K-PI(3)P-Atg21-Atg16 Module to Phagophores via Vps8 for Autophagy.

Phosphatidylinositol 3-phosphate (PI(3)P) serves important functions in endocytosis, phagocytosis, and autophagy. PI(3)P is generated by Vps34 of the class III phosphatidylinositol 3-kinase (PI3K) complex. The Vps34-PI3K complex can be divided into Vps34-PI3K class II (containing Vps38, endosomal) and Vps34-PI3K class I (containing Atg14, autophagosomal). Most PI(3)Ps are associated with endosomal membranes. In yeast, the endosomal localization of Vps34 and PI(3)P is tightly regulated by Vps21-module proteins. At yeast phagophore assembly site (PAS) or mammalian omegasomes, PI(3)P binds to WD-repeat protein interacting with phosphoinositide (WIPI) proteins to further recruit two conjugation systems, Atg5-Atg12·Atg16 and Atg8-PE (LC3-II), to initiate autophagy. However, the spatiotemporal regulation of PI(3)P during autophagy remains obscure. Therefore, in this study, we determined the effect of Vps21 on localization and interactions of Vps8, Vps34, Atg21, Atg8, and Atg16 upon autophagy induction. The results showed that Vps21 was required for successive colocalizations and interactions of Vps8-Vps34 and Vps34-Atg21 on endosomes, and Atg21-Atg8/Atg16 on the PAS. In addition to disrupted localization of the PI3K complex II subunits Vps34 and Vps38 on endosomes, the localization of the PI3K complex I subunits Vps34 and Atg14, as well as Atg21, was partly disrupted from the PAS in vps21∆ cells. The impaired PI3K-PI(3)P-Atg21-Atg16 axis in vps21∆ cells might delay autophagy, which is consistent with the delay of early autophagy when Atg21 was absent. This study provides the first insight into the upstream sequential regulation of the PI3K-PI(3)P-Atg21-Atg16 module by Vps21 in autophagy.

Erratum: A general-purpose Nanohybrid fabricated by Polymeric Au(I)-peptide precursor to wake the function of Peptide Therapeutics: Erratum.

[This corrects the article DOI: 10.7150/thno.47243.].

Turning a Targeting ß-Catenin/Bcl9 Peptide Inhibitor into a GdOF@Au Core/Shell Nanoflower for Enhancing Immune Response to Cancer Therapy in Combination with Immune Checkpoint Inhibitors.

Combination administration is becoming a popular strategy in current cancer immunotherapy to enhance tumor response to ICIs. Recently, a peptide drug, a protein-protein interaction inhibitor (PPI), that disrupts the ß-catenin/Bcl9 interaction in the tumoral Wnt/ß-catenin pathway has become a promising candidate drug for immune enhancement and tumor growth inhibition. However, the peptide usually suffers from poor cell membrane permeability and proteolytic degradation, limiting its adequate accumulation in tumors and ultimately leading to side effects. Herein, a gadolinium-gold-based core/shell nanostructure drug delivery system was established, where Bcl9 was incorporated into a gadolinium-gold core-shell nanostructure and formed GdOFBAu via mercaptogenic self-assembly. After construction, GdOFBAu, when combined with anti-PD1 antibodies, could effectively inhibit tumor growth and enhance the response to immune therapy in MC38 tumor-bearing mice; it not only induced the apoptosis of cancer cells, but also promoted the tumor infiltration of Teff cells (CD8+) and decreased Treg cells (CD25+). More importantly, GdOFBAu maintained good biosafety and biocompatibility during treatment. Taken together, this study may offer a promising opportunity for sensitizing cancer immunotherapy via metal-peptide self-assembling nanostructured material with high effectiveness and safety.

Nanoparticle delivery of miR-21-3p sensitizes melanoma to anti-PD-1 immunotherapy by promoting ferroptosis.

BACKGROUND: Although anti-programmed cell death protein 1 (PD-1) immunotherapy is greatly effective in melanoma treatment, low response rate and treatment resistance significantly hinder its efficacy. Tumor cell ferroptosis triggered by interferon (IFN)-γ that is derived from tumor-infiltrating CD8+ T cells greatly contributes to the effect of immunotherapy. However, the molecular mechanism underlying IFN-γ-mediated ferroptosis and related potentially promising therapeutic strategy warrant further clarification. MicroRNAs (miRNAs) participate in ferroptosis execution and can be delivered systemically by multiple carriers, which have manifested obvious therapeutic effects on cancer. METHODS: MiRNAs expression profile in IFN-γ-driven ferroptosis was obtained by RNA sequencing. Biochemical assays were used to clarify the role of miR-21-3p in IFN-γ-driven ferroptosis and the underlying mechanism. MiR-21-3p-loaded gold nanoparticles were constructed and systemically applied to analyze the role of miR-21-3p in anti-PD-1 immunotherapy in preclinical transplanted tumor model. RESULTS: MiRNAs expression profile of melanoma cells in IFN-γ-driven ferroptosis was first obtained. Then, upregulated miR-21-3p was proved to facilitate IFN-γ-mediated ferroptosis by potentiating lipid peroxidation. miR-21-3p increased the ferroptosis sensitivity by directly targeting thioredoxin reductase 1 (TXNRD1) to enhance lipid reactive oxygen species (ROS) generation. Furthermore, miR-21-3p overexpression in tumor synergized with anti-PD-1 antibody by promoting tumor cell ferroptosis. More importantly, miR-21-3p-loaded gold nanoparticles were constructed, and the systemic delivery of them increased the efficacy of anti-PD-1 antibody without prominent side effects in preclinical mice model. Ultimately, ATF3 was found to promote miR-21-3p transcription in IFN-γ-driven ferroptosis. CONCLUSIONS: MiR-21-3 p upregulation contributes to IFN-γ-driven ferroptosis and synergizes with anti-PD-1 antibody. Nanoparticle delivery of miR-21-3 p is a promising therapeutic approach to increase immunotherapy efficacy without obvious systemic side effects.

Turing milk into pro-apoptotic oral nanotherapeutic: De novo bionic chiral-peptide supramolecule for cancer targeted and immunological therapy.

Chirality in biomolecules is ubiquitous in our world, but oral nanomedicines constructed from chiral peptides are extremely rare, principally because of the immature nanofabrication and inadequate bioavailability of chiral nanostructures. Methods: To realize the oral administration of chiral peptides and break through their forbidden zone in intracellular space, a chiral-peptide supramolecular (DPAICP) camouflaging with the membrane from milk-derived extracellular vesicles (ME) was developed herein through an aqueous-based growth method of chiral peptide Au(I) infinite covalent polymer (DPAICP) involving in organothiol D-peptides and Au3+, and a feasible camouflage technology using ME. Results: DPAICP@ME possessed favorable pharmaceutical properties to remain stable during the gastrointestinal absorption and blood circulation, and showed the satisfactory tumor accumulation through oral medication. Expectedly, oral DPAICP@ME played its predetermined role in vivo to restore p53 signaling pathway for cancer therapy in B16F10 homograft malignant melanoma model, LLC Lewis orthotopic transplantation model of lung cancer and patient-derived orthotopic xenograft (PDOX) mice model of colon cancer. Moreover, oral DPAICP@ME augmented the action of immunotherapy by Anti-PD1 through the further T-cell activation. Conclusion: The de novo design of the bionic chiral-peptide supramolecule provides a practicable strategy for the construction of biomimetic chiral peptide-derived nanostructures that can be taken orally, and likely boosts chiral nanomedicine discovery efforts for a wider range of diseases including cancer.

A Bionic-Homodimerization Strategy for Optimizing Modulators of Protein-Protein Interactions: From Statistical Mechanics Theory to Potential Clinical Translation.

Emerging protein-protein interaction (PPI) modulators have brought out exciting ability as therapeutics in human diseases, but its clinical translation has been greatly hampered by the limited affinity. Inspired by the homodimerize structure of antibody, the homodimerization contributes hugely to generating the optimized affinity is conjectured. Herein, a statistical-mechanics-theory-guided method is established to quantize the affinity of ligands with different topologies through analyzing the change of enthalpy and the loss of translational and rotational entropies. A peptide modulator for p53-MDM2 termed CPAP is used to homodimerize connecting, and this simple homodimerization can significantly increase the affinity. To realize the cellular internalization and tumor accumulation, Dimer CPAP and Mono CPAP are nanoengineered into gold(I)-CPAP supermolecule by the aurophilic interaction-driven self-assembly. Nano-Dimer CPAP potently suppressed tumor growth in lung cancer allograft model and a patient-derived xenograft model in more action than Nano-Mono CPAP, while keeping a favorable drug safety profile. This work not only presents a physico-mechanical method for calculating the affinity of PPI modulators, but also provides a simple yet robust homodimerization strategy to optimize the affinity of PPI modulators.

Turing miRNA into infinite coordination supermolecule: a general and enabling nanoengineering strategy for resurrecting nuclear acid therapeutics.

BACKGROUND: Clinical translation of therapeutic nuclear acid, particularly those targeting tumor progression, has been hampered by the intrinsic weaknesses of nuclear acid therapeutic including poor systemic stability, rapid clearance, low membrane permeability and lack of targeting ability. Small nuclear acid engineered into carrier-free nanodrugs with structural stability and disease targeting may be viable to overcome pharmaceutical obstacles of nuclear acid. METHODS: A general method through a mild and simple chemistry was established to convert therapeutic miRNA into an infinite Auric-sulfhydryl coordination supramolecular miRNA termed IacsRNA with near-spherical nanostructure, high colloid as well as anti-hydrolysis stability and low macrophage uptakes. RESULTS: IacsRNA presented the increased half-life period in circulation and accumulation at tumor sites in comparison to normal miRNA. Moreover, Iacs-miR-30c showed no toxicity of viscera and sanguis system in the 5-time injection dosage of the treatment. More importantly, Iacs-miR-30c potently suppressed the Wnt signaling pathway in vitro and in vivo, and effectively sensitized both potency of 5-Fu in PDX model of colon cancer and Anti-PD1 in B16F10 homograft model of melanoma. CONCLUSION: Collectively, this work amply confirmed the design of IacsRNA as a general and viable strategy of nano-pharmaceutic to concert flimsy therapeutic miRNA into potential drugs. Considering from a broader perspective, the miRNA-initiated infinite coordination self-assembly strategy has distinct advantages in resurrecting nuclear acid therapeutics, probably bringing new inspiration to RNA-derived therapeutics of a great variety of human diseases including cancer.

A Tumor-Targeting Metal-Organic Nanoparticle Constructed by Dynamic Combinatorial Chemistry toward Accurately Redressing Carcinogenic Wnt Cascade.

Targeted and immunological therapy have revolutionized the malignancy treatment, but is suffering from the dose-limiting side effects and inadequate responsiveness. The emerging nanoscale infinite coordination polymers provide a feasible strategy for tumor targeting and immune sensitization. Herein, a "one-pot" self-assembled strategy based on dynamic combinatorial chemistry (DCC) principle is designed to construct a tumor-targeting metal-organic nanoparticle (MOICP) through a spontaneous co-assembling among three metal-organic coordination polymers tuned by a Wnt-inhibitor carnosic acid (CA). Responding to the tumor microenvironment, MOICP presents an optimized tumor-preferential accumulation and the satisfactory biosafety. MOICP is more active in vitro and in vivo than CA in suppressing of Wnt signaling pathway, and potently inhibits tumor growth in a patient-derived xenograft model of Wnt-activated pancreatic carcinoma. Moreover, MOICP reverses the lack of intratumoral infiltration of T lymphocytes, and hence augments the action of Anti-PD1 (programmed cell death protein 1) immunotherapy in B16F10 melanoma allograft mice model. This clinically viable MOICP can not only be applied to Wnt inhibition for cancer targeted therapy and immunotherapeutic sensitization, but also provides a de novo pattern for nanomedicine architecture with cargo-initiated co-self-assembly guided by DCC, thereby bringing new inspiration in general for disease intervention.

Assembling BH3-mimic peptide into a nanocluster to target intracellular Bcl2 towards the apoptosis induction of cancer cell.

Bcl-2, an anti-apoptotic protein, is always overexpressed in tumor cells to suppress the pro-apoptotic function of Bax, thereby prolonging the life of the tumor. However, BH3 proteins could directly activate Bax via antagonizing Bcl-2 to induce apoptosis in response to the stimulation. Thus, mimicking BH3 proteins with a peptide is a potential strategy for anti-cancer therapy. Unfortunately, clinical translation of BH3-mimic peptide is hindered by its inefficacious cellular internalization and proteolysis resistance. Herein, we translated a BH3-mimic peptide into a peptide-auric spheroidal nanocluster (BH3-AuNp), in which polymeric BH3-Auric precursors [Au1+-S-BH3]narein situself-assembled on the surface of gold nanoparticles by a one-pot synthesis. Expectedly, this strategy could improve the anti-proteolytic ability and cytomembrane penetrability of the BH3 peptide. As a result, BH3-AuNp successfully induced the apoptosis of two cancer cell lines by an order of magnitude compared to BH3. This therapeutic and feasible peptide nano-engineering strategy will help peptides overcome the pharmaceutical obstacles, awaken its biological functions, and possibly revive the research about peptide-derived nanomedicine.

A nano-predator of pathological MDMX construct by clearable supramolecular gold(I)-thiol-peptide complexes achieves safe and potent anti-tumor activity.

As alternatives to small-molecular proteolysis-targeting chimeras (PROTAC), peptide-based molecular glues (MG) are a broad range of dual-functional ligands that simultaneously bind with targetable proteins and E3 ligases by mimicking proteinprotein interaction (PPI) partners. Methods: Herein, we design a peptide-derived MG to target a tumor-driving protein, MDMX, for degradation, and nanoengineered it into a supramolecular gold(I)-thiol-peptide complex (Nano-MP) to implement the proteolysis recalcitrance, cellular internalization, and glutathione-triggered release. To optimize the tumor targeting, a pH-responsive macromolecule termed polyacryl sulfydryl imidazole (PSI) was synthesized to coat Nano-MP. Results: As expected, Nano-MP@PSI induced the MDMX degradation by ubiquitination and subsequently restored the anti-cancer function of p53 and p73. Nano-MP@PSI revealed potent anti-cancer activities in an orthotopic xenograft mouse model of retinoblastoma by intraocular injection and a patient-derived xenograft model of malignant pancreatic cancer by systemic injection, while maintaining a favorable safety profile and showing a highly favorable clearable profile of excretion from the living body. Conclusion: Collectively, this work not only provided a clinically viable paradigm for the treatment of a wide variety of tumors by multiple administration types, but, more importantly, it bridged the chasm between peptides and PROTACs, and likely reinvigorated the development of peptide-derived proteolysis-targeting chimeras for a great variety of diseases.

De Novo Nano-Erythrocyte Structurally Braced by Biomimetic Au(I)-peptide Skeleton for MDM2/MDMX Predation toward Augmented Pulmonary Adenocarcinoma Immunotherapy.

In nature, cells rely on a structural framework called the "cytoskeleton" to maintain their shape and polarity. Based on this, herein a new class of cell-mimicking nanomedicine using bionic skeletons constituted by the oligomeric Au(I)-peptide complex is developed. The peptide function of degrading pathological MDM2 and MDMX is used to synthesize an oligomeric Au(I)-PMIV precursor capable of self-assembling into a clustered spherical bionic skeleton. Through coating by erythrocyte membrane, an erythrocyte-mimicking nano-cell (Nery-PMIV) is developed with depressed macrophage uptakes, increased colloidal stability, and prolonged blood circulation. Nery-PMIV potently restores p53 and p73 in vitro and in vivo by degrading MDM2/MDMX. More importantly, Nery-PMIV effectively augments antitumor immunity elicited by anti-PD1 therapy in a murine orthotopic allograft model for LUAD and a humanized patient-derived xenograft (PDX) mouse model for LUAD, while maintaining a favorable safety profile. Taken together, this work not only presents evidence showing that MDM2/MDMX degradation is a potentially viable therapeutic paradigm to synergize anti-PD1 immunotherapy toward LUAD carrying wild-type p53; it also suggests that cell-mimicking nanoparticles with applicable bionic skeletons hold tremendous promise in offering new therapies to revolutionize nanomedicine in the treatment of a myriad of human diseases.

A general-purpose Nanohybrid fabricated by Polymeric Au(I)-peptide precursor to wake the function of Peptide Therapeutics.

Peptide-derived nanocomposites have been exhibiting fascinating biological advantages, including but not limited to excellent biocompatibility, biological degradation, high targetability and subsequent potent therapeutic efficacy. While some successes have been achieved in the nanoengineering of peptide-based architectures with defined dimensions and medical functions, enormous challenges remain about clinical nano-pharmaceutics of peptides, especially those modulating intracellular protein-protein interactions (PPIs). Methods: We developed a general method to translate intracellular-PPI-targeted peptides into a bioavailable peptide-auric spheroidal nanohybrid (SNH), for which polymeric peptide-Auric precursors [Au1+-S-peptide]n are in-situ reduced on the surface of gold nanoseeds via a simple and mild reaction. As proofs of concept, three cytomembrane-impenetrable peptides with different physicochemical properties were successfully engineered into stable and tumor-specific SNH respectively. Results: To highlight the advantage of SNH, PMI, a hydrophobic and enzyme-intolerant peptide capable of p53 restoration, was selected to challenge the power of SNH in a colon tumor xenografts model. PMI-Au SNH in vivo suppressed tumor growth potently after three administrations: intravenous injection, intraperitoneal injection and gastric perfusion, and maintained a favorable therapeutic safety. Conclusion: This therapeutically feasible strategy of peptide nanoengineering will allow us to fabricate a series of nanomedicines to modulate carcinogenic PPIs that hide and multiply inside cells, and in all likelihood reinvigorate the development of peptide drug against wide varieties of human diseases.

Targeted disruption of the BCL9/ß-catenin interaction by endosomal-escapable nanoparticles functionalized with an E-cadherin-derived peptide.

Abnormal activation of the Wnt/ß-catenin signaling pathway, which underlies multiple malignancies, promotes tumor progression; drugs that can block this pathway are therefore highly attractive candidates for anticancer therapy. Using a therapeutic peptide derived from E-cadherin region V (cECRV), we sought to develop a potent and selective antagonist of ß-catenin that can disrupt the carcinogenic interaction between ß-catenin and BCL9. More importantly, to overcome the pharmacological obstacles of peptide-derived therapeutics (poor nuclease stability and low membrane permeability), a gold nanoparticle (AuNP)-based nanocarrier was designed to deliver cECRV into the cytoplasm to modulate the intracellular interaction of ß-catenin and BCL9. The resultant nanoparticle, pAuNP-cECRV, showed no cytotoxicity towards normal peripheral blood mononuclear cells and induced cycle arrest and subsequent apoptosis of Wnt-hyperactive cancer cells by antagonizing ß-catenin to inhibit the Wnt pathway. Our results indicate that pAuNP-cECRV is very promising for application as an efficient and safe peptide delivery vector for cancer therapy.

Modulating Protein-Protein Interactions In Vivo via Peptide-Lanthanide-Derived Nanoparticles for Hazard-Free Cancer Therapy.

Protein-protein interactions (PPIs) play an important role in almost all vital processes involved in many diseases, especially cancer. Peptides are perfect candidates that modulate PPIs as they can closely mimic principle features of protein. However, the intrinsic drawbacks of peptides, including poor stability and member impenetrability, severely limit the development of peptide-derived therapeutics. Nanotechnology offers a feasible route for anti-cancer peptide delivery, but much remains to be done, especially with respect to the pressing need for a simple method for efficient delivery of peptides into sites of interest towards potent and safe therapy. Herein, we report a one-step method to conjugate lanthanide-doped nanoparticles with p53-activating peptide (PMI: TSFAEYWALLSP), Bcl2-blocking peptide (BIM: IWIAQELRRIGDEFNAYYARR) and CD13-binding peptide (iNGR: CRNGRGPDC) by mercaptogenic self-assembly. The resultant LDN-iNGRPMI-BIM nanoparticles can tumor-specifically accumulate at interest sites, and potently induce apoptosis of cancer cells in vitro and in vivo, while keeping a favorable biosafety profile. Taken together, the general therapeutically viable method reported here will enable us to develop a novel class of peptide-based nanomedicines, and likely reinvigorate peptide drug discovery efforts in general, which will target intracellular protein-protein interactions responsible for initiation and progression of a great variety of human diseases.

Self-Assembly of Therapeutic Peptide into Stimuli-Responsive Clustered Nanohybrids for Cancer-Targeted Therapy.

Clinical translation of therapeutic peptides, particularly those targeting intracellular protein-protein interactions (PPIs), has been hampered by their inefficacious cellular internalization in diseased tissue. Therapeutic peptides engineered into nanostructures with stable spatial architectures and smart disease targeting ability may provide a viable strategy to overcome the pharmaceutical obstacles of peptides. This study describes a strategy to assemble therapeutic peptides into a stable peptide-Au nanohybrid, followed by further self-assembling into higher-order nanoclusters with responsiveness to tumor microenvironment. As a proof of concept, an anticancer peptide termed ß-catenin/Bcl9 inhibitors is copolymerized with gold ion and assembled into a cluster of nanohybrids (pCluster). Through a battery of in vitro and in vivo tests, it is demonstrated that pClusters potently inhibit tumor growth and metastasis in several animal models through the impairment of the Wnt/ß-catenin pathway, while maintaining a highly favorable biosafety profile. In addition, it is also found that pClusters synergize with the PD1/PD-L1 checkpoint blockade immunotherapy. This new strategy of peptide delivery will likely have a broad impact on the development of peptide-derived therapeutic nanomedicine and reinvigorate efforts to discover peptide drugs that target intracellular PPIs in a great variety of human diseases, including cancer.

A Rab5 GTPase module is important for autophagosome closure.

In the conserved autophagy pathway, the double-membrane autophagosome (AP) engulfs cellular components to be delivered for degradation in the lysosome. While only sealed AP can productively fuse with the lysosome, the molecular mechanism of AP closure is currently unknown. Rab GTPases, which regulate all intracellular trafficking pathways in eukaryotes, also regulate autophagy. Rabs function in GTPase modules together with their activators and downstream effectors. In yeast, an autophagy-specific Ypt1 GTPase module, together with a set of autophagy-related proteins (Atgs) and a phosphatidylinositol-3-phosphate (PI3P) kinase, regulates AP formation. Fusion of APs and endosomes with the vacuole (the yeast lysosome) requires the Ypt7 GTPase module. We have previously shown that the Rab5-related Vps21, within its endocytic GTPase module, regulates autophagy. However, it was not clear which autophagy step it regulates. Here, we show that this module, which includes the Vps9 activator, the Rab5-related Vps21, the CORVET tethering complex, and the Pep12 SNARE, functions after AP expansion and before AP closure. Whereas APs are not formed in mutant cells depleted for Atgs, sealed APs accumulate in cells depleted for the Ypt7 GTPase module members. Importantly, depletion of individual members of the Vps21 module results in a novel phenotype: accumulation of unsealed APs. In addition, we show that Vps21-regulated AP closure precedes another AP maturation step, the previously reported PI3P phosphatase-dependent Atg dissociation. Our results delineate three successive steps in the autophagy pathway regulated by Rabs, Ypt1, Vps21 and Ypt7, and provide the first insight into the upstream regulation of AP closure.

TGFBR1 haplotypes and risk of non-small-cell lung cancer.

Transforming growth factor beta (TGF-beta) receptors are centrally involved in TGF-beta-mediated cell growth and differentiation and are frequently inactivated in non-small-cell lung cancer (NSCLC). Constitutively decreased type I TGF-beta receptor (TGFBR1) expression is emerging as a novel tumor-predisposing phenotype. The association of TGFBR1 haplotypes with risk for NSCLC has not yet been studied. We tested the hypothesis that single-nucleotide polymorphisms (SNP) and/or TGFBR1 haplotypes are associated with risk of NSCLC. We genotyped six TGFBR1 haplotype-tagging SNPs (htSNP) by PCR-RFLP assays and one htSNP by PCR-single-strand conformation polymorphism assay in two case-control studies. Case-control study 1 included 102 NSCLC patients and 104 healthy controls from Suzhou. Case-control study 2 included 131 patients with NSCLC and 133 healthy controls from Wuxi. Individuals included in both case-control studies were Han Chinese. Haplotypes were reconstructed according to the genotyping data and linkage disequilibrium status of these seven htSNPs. None of the htSNP was associated with NSCLC risk in either study. However, a four-marker CTGC haplotype was significantly more common among controls than among cases in both studies (P = 0.014 and P = 0.010, respectively), indicating that this haplotype is associated with decreased NSCLC risk {adjusted odds ratio [OR], 0.09 [95% confidence interval (95% CI), 0.01-0.61] and 0.11 [95% CI, 0.02-0.59], respectively}. Combined analysis of both studies shows a strong association of this four-marker haplotype with decreased NSCLC risk (adjusted OR, 0.11; 95% CI, 0.03-0.39). This is the first evidence of an association between a TGFBR1 haplotype and risk for NSCLC.