Inhibited Lung Metastasis Research Articles

PLK1 phosphorylates RhoGDI1 and promotes cancer cell migration and invasion

BackgroundRho guanine nucleotide dissociation inhibitor 1 (RhoGDI1) plays an important role in diverse cellular processes by regulating Rho guanosine triphosphate (GTP)ases activity. RhoGDI1 phosphorylation regulates the spatiotemporal activation of Rho GTPases during cell migration. In this study, we identified polo-like kinase 1 (PLK1) as a novel kinase of RhoGDI1 and investigated the molecular mechanism by which the interaction between RhoGDI1 and PLK1 regulates cancer cell migration.MethodsImmunoprecipitation, GST pull-down assay, and proximity ligation assay (PLA) were performed to analyze the interaction between RhoGDI1 and PLK1. In vitro kinase assay and immunoprecipitation were performed with Phospho-(Ser/Thr) antibody. We evaluated RhoA activation using RhoGTPases activity assay. Cell migration and invasion were analyzed by transwell assays.ResultsGST pull-down assays and PLA showed that PLK1 directly interacted with RhoGDI1 in vitro and in vivo. Truncation mutagenesis revealed that aa 90-111 of RhoGDI1 are critical for interacting with PLK1. We also showed that PLK1 phosphorylated RhoGDI1 at Thr7 and Thr91, which induces cell motility. Overexpression of the GFP-tagged RhoGDI1 truncated mutant (aa 90-111) inhibited the interaction of PLK1 with RhoGDI1 and attenuated RhoA activation by PLK1. Furthermore, the overexpression of the RhoGDI1 truncated mutant reduced cancer cell migration and invasion in vitro and suppressed lung metastasis in vivo.ConclusionsCollectively, we demonstrate that the phosphorylation of RhoGDI1 by PLK1 promotes cancer cell migration and invasion through RhoA activation. This study connects the interaction between PLK1 and RhoGDI1 to the promotion of cancer cell behavior associated with malignant progression, thereby providing opportunities for cancer therapeutic interventions.

Biomimetic Responsive Nanoconverters with Immune Checkpoint Blockade Plus Antiangiogenesis for Advanced Hepatocellular Carcinoma Treatment.

The first-line treatment for advanced hepatocellular carcinoma (HCC) combines immune checkpoint inhibitors and antiangiogenesis agents to prolong patient survival. Nonetheless, this approach has several limitations, including stringent inclusion criteria and suboptimal response rates that stem from the severe off-tumor side effects and the unfavorable pharmacodynamics and pharmacokinetics of different drugs delivered systemically. Herein, we propose a single-agent smart nanomedicine-based approach that mimics the therapeutic schedule in a targeted and biocompatible manner to elicit robust antitumor immunity in advanced HCC. Our strategy employed pH-responsive carriers, poly(ethylene glycol)-poly(β-amino esters) amphiphilic block copolymer (PEG-PAEs), for delivering apatinib (an angiogenesis inhibitor), that were surface-coated with plasma membrane derived from engineered cells overexpressing PD-1 proteins (an immune checkpoint inhibitor to block PD-L1). In an advanced HCC mouse model with metastasis, these biomimetic responsive nanoconverters induced significant tumor regression (5/9), liver function recovery, and complete suppression of lung metastasis. Examination of the tumor microenvironment revealed an increased infiltration of immune effector cells (CD8+ and CD4+ T cells) and reduced immunosuppressive cells (myeloid-derived suppressor cells and T regulatory cells) in treated tumors. Importantly, our nanomedicine selectively accumulated in both small and large HCC occupying >50% of the liver volume to exert therapeutic effects with minimal systemic side effects. Overall, these findings highlight the potential of such multifunctional nanoconverters to effectively reshape the tumor microenvironment for advanced HCC treatment.

Cancer Cell Membrane-Biomimetic Nanoparticles Based on Gelatin and Mitoxantrone for Synergetic Chemo-Photothermal Therapy of Metastatic Breast Cancer.

The purpose of this paper is to develop a cancer cell membrane biomimetic nanodrug delivery system (NDDS) to achieve an enhanced chemo-photothermal synergistic antitumor effect. The biomimetic NDDSs are composed of mitoxantrone (MIT)-loaded gelatin nanoparticles and IR820-encapsulated 4T1 cancer cell membrane-derived vesicles. The biomimetic NDDS displayed excellent stability and photothermal conversion efficiency. Compared to naked nanoparticles, the cell membrane-coated nanoparticles improved 4T1 cell uptake through homologous targeting and effectively reduced internalization of macrophages. In vivo photothermal imaging results further showed that the NDDS could be enriched at the tumor site for 48 h and could raise the temperature of the tumor area to 60 °C within 5 min under 808 nm laser irradiation. Finally, NDDS successfully inhibited primary tumor growth (over 89% inhibition) and significantly inhibited lung metastasis. This study may provide a new strategy for personalized chemotherapy-photothermal combination therapy of metastatic breast cancer using tumor cell membranes from cancer patients as drug carriers.

MOF@COF Nanocapsules Enhance Soft Tissue Sarcoma Treatment: Synergistic Effects of Photodynamic Therapy and PARP Inhibition on Tumor Growth Suppression and Immune Response Activation.

Soft tissue sarcomas (STS) are highly malignant tumors with limited treatment options owing to their heterogeneity and resistance to conventional therapies. Photodynamic therapy (PDT) and poly-ADP-ribose polymerase (PARP) inhibitors (PARPi) have shown potential for STS treatment, with PDT being effective for sarcomas located on the extremities and body surface and PARPi targeting defects in homologous recombination repair. To address the limitations of PDT and harness the potential of PARPi, herein, a novel therapeutic approach for STS treatment combining nanocapsules bearing integrated metal-organic frameworks (MOFs) and covalent organic frameworks (COFs), i.e., MOF@COF, with PDT and PARPi is proposed. Nanocapsules are designed, referred to as ZTN@COF@poloxamer, which contain a Zr-based MOF and tetrakis (4-carbethoxyphenyl) porphyrin as a photosensitizer, are coated with a COF to improve the sensitizing properties, and are loaded with niraparib to inhibit DNA repair. Experiments demonstrate that this new nanocapsules treatment significantly inhibits STS growth, promotes tumor cell apoptosis, exhibits high antitumor activity with minimal side effects, activates the immune response of the tumor, and inhibits lung metastasis in vivo. Therefore, MOF@COF nanocapsules combined with PARPi offer a promising approach for STS treatment, with the potential to enhance the efficacy of PDT and prevent tumor recurrence.

The safety and efficacy of systemic delivery of a new liver-de-targeted TGFβ signaling inhibiting adenovirus in an immunocompetent triple negative mouse mammary tumor model

Aberrant TGFβ signaling is linked to metastasis and tumor immune escape of many cancers including metastatic triple negative breast cancer (mTNBC). Previously, we have found that oncolytic adenoviruses expressing a TGFβ signaling inhibitory protein (sTGFβRIIFc) induced immune activation in a mouse TNBC (4T1) immunocompetent subcutaneous model with intratumoral injection. Systemic administration of adenoviruses can be a superior route to treat mTNBC but faces the challenges of increased toxicity and viral clearance. Thus, we created a liver-de-targeted sTGFβRIIFc- and LyP-1 peptide-expressing adenovirus (mHAdLyp.sT) with enhanced breast cancer cell tropism. Its safety and immune response features were profiled in the 4T1 model. Our data showed that the systemic administration of mHAdLyp.sT resulted in reduced hepatic and systemic toxicity. mHAdLyp.sT was also effective in increasing Th1 cytokines and anti-tumor cell populations by cytokine analysis, spleen/tumor qRT-PCR, and flow cytometry. We further tested the therapeutic effects of mHAdLyp.sT alone and in combination with immune checkpoint inhibitors (ICIs). mHAdLyp.sT alone and with all ICI combinations elicited significant inhibition of lung metastasis by histological analysis. When mHAdLyp.sT was combined with both anti-PD-1 and anti-CTLA-4 antibodies, primary 4T1 tumor growth was also significantly inhibited. We are confident in advancing this new treatment option for mTNBC.

Clinically approved carbon nanoparticles for enhanced photothermal-immunotherapy toward cancer metastasis

The development of new biomedical applications of approved drugs has several advantages in drug translation. In this study, carbon nanoparticle suspension injection (CNSI), clinically approved for lymph node tracer in China, is explored as a photothermal conversion agents for the treatment of metastatic cancer. Metastasis commonly through lymphatic circulation accounts for the principal cause of death in cancer patients. Up to now, immunotherapy has been already used clinically for metastatic cancer. However, the suppressive tumor immune microenvironment limits the effectiveness of immunotherapy. Herein, we use the photothermal effect of CNSI under near-infrared (NIR) irradiation to eliminate primary tumor and induce the immunogenic cell death (ICD). Further, CNSI under irradiation combined with aPD-1 spur immune response and inhibit the growth of distal tumors. Except for the primary tumor, phototherapy was used to treat metastatic lymph nodes synchronously, which can not only eradicate cancer cells, but also directly activate the immune response inside the lymph nodes. Combining with aPD-1 further enhance the anti-tumor effect to block lymphatic metastases and inhibit lung metastases. This study shows that the carbon nanoparticles have potential clinical application prospects for the treatment of tumor metastasis.

Extracellular Matrix-Trapped Bioinspired Lipoprotein Prolongs Tumor Retention to Potentiate Antitumor Immunity.

The immunomodulatory effects of many therapeutic agents are significantly challenged by their insufficient delivery efficiency and short retention time in tumors. Regarding the distinctively upregulated fibronectin (FN1) and tenascin C (TNC) in tumor stroma, herein a protease-activated FN1 and/or TNC binding peptide (FTF) is designed and an extracellular matrix (ECM)-trapped bioinspired lipoprotein (BL) (FTF-BL-CP) is proposed that can be preferentially captured by the TNC and/or FN1 for tumor retention, and then be responsively dissociated from the matrix to potentiate the antitumor immunity. The FTF-BL-CP treatment produces a 6.96-, 9.24-, 6.72-, 7.32-, and 6.73-fold increase of CD3+CD8+ T cells and their interferon-γ-, granzyme B-, perforin-, and Ki67-expressing subtypes versus the negative control, thereby profoundly eliciting the antitumor immunity. In orthotopic and lung metastatic breast cancer models, FTF-BL-CP produces notable therapeutic benefits of retarding tumor growth, extending survivals, and inhibiting lung metastasis. Therefore, this ECM-trapping strategy provides an encouraging possibility of prolonging tumor retention to potentiate the antitumor immunity for anticancer immunotherapy.

Targeting Erbin-mitochondria axis in platelets/megakaryocytes promotes B cell-mediated antitumor immunity

The roles of platelets/megakaryocytes (MKs), the key components in the blood system, in the tumor microenvironment and antitumor immunity are unclear. In patients with colorectal cancer, the number of platelets was significantly increased in patients with metastasis, and Erbin expression was highly expressed in platelets from patients with metastases. Moreover, Erbin knockout in platelets/MKs suppressed lung metastasis in mice and promoted aggregations of platelets. Mechanistically, Erbin-deficient platelets have increasing mitochondrial oxidative phosphorylation and secrete lipid metabolites like acyl-carnitine (Acar) by abolishing interaction with prothrombotic protein ESAM. Notably, Acar enhanced the activity of mitochondrial electron transport chain complex and mitochondrial oxidative phosphorylation in B cells by acetylation of H3K27 epigenetically. Targeting Erbin in platelets/MKs by a nanovesicle system dramatically attenuated lung metastasis in mice invivo. Our study identifies an Erbin-mitochondria axis in platelets/MKs, which suppresses B cell-mediated antitumor immunity, suggesting a new way for the treatment of metastasis.

Abstract A017: Targeting Tlk2 impairs breast cancer growth and engages immune responses

Abstract The Tousled like kinases 1 and 2 (TLK1 and TLK2) are evolutionarily conserved Ser/Thr kinases that are required for genome stability and normal development in numerous organisms. Both kinases contribute to histone deposition by targeting the histone chaperones ASF1A and ASF1B and are regulated by the DNA damage response (DDR). TLK1/2 expression is ubiquitous and often elevated in human cancers with suppressed immune gene expression. In some cancer types, elevated TLK levels correlate with decreased disease-free survival. In cultured cells, TLK depletion causes replication stress, DNA damage and altered chromatin maintenance. TLK1/2 depletion increases the efficacy of DNA damage checkpoint inhibitors and PARP inhibitors and induces the Alternative-lengthening of telomeres pathway and innate immune signaling. Collectively, this data suggests that TLK activity may be a druggable anti-cancer target. To test this, we generated a conditional Tlk2 allele in mice and introduced the Mouse Mammary Tumor Virus-Polyoma Middle Tumor-antigen (MMTV-PyMT) transgene to generate breast cancers. Deletion of Tlk2 in developing breast cancer resulted in impaired primary tumor growth and a complete inhibition of lung metastasis. Characterization of the tumor microenvironment revealed elevated immune infiltration that includes activated T cells with higher levels of PD-1, which could potentially sensitize these tumors to immune checkpoint blockade. To test this possibility, we established a syngeneic breast cancer model to allow us to combine Tlk2 depletion or deletion with DDR inhibitor and anti-PD-1 therapy in vivo. Preliminary data supports the strategy of targeting Tlk2 in cancer and we will share current data from tumor models and preliminary characterization of small molecule inhibitors. Citation Format: Marina Villamor-Payá, Adrià Caballé, Jordann Smak, Mohamed-Reda Benmebarek, Camille Stephan-Otto Attolini, Travis H. Stracker. Targeting Tlk2 impairs breast cancer growth and engages immune responses [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr A017.

A chemo/chemodynamic nanoparticle based on hyaluronic acid induces ferroptosis and apoptosis for triple-negative breast cancer therapy

Triple-negative breast cancer (TNBC) poses a serious threat to women's life and health due to its high malignancy, strong invasiveness, and propensity for early recurrence and metastasis. Therefore, there is an urgent need to develop a highly effective and low-toxic TNBC treatment scheme to enhance the anti-cancer efficacy and prolong the survival of patients. In this work, we designed and synthesized a chemodynamic therapy (CDT) agent (HA-Fc-Mal). The chemo/chemodynamic (CT/CDT) nanoparticle (HCM@DOX) based on hyaluronic acid induces ferroptosis and apoptotic for TNBC therapy was constructed via self-assembled of HA-Fc-Mal and doxorubicin (DOX). HCM@DOX orderly realized the TNBC targeting, controlled DOX release, GSH depletion and induce ROS erupt. In vivo and in vitro experiments confirmed that HCM@DOX inhibited the growth of 4 T1 tumors through ferroptosis and apoptosis, and the tumor inhibition rate was as high as 81.87 %. In addition, HCM@DOX significantly inhibited lung metastasis and exhibited excellent biosafety. Overall, our findings offer a new strategy for TNBC therapy using a CT/CDT nanoparticle that induces ferroptosis and apoptosis.

Injectable Responsive Hydrogel Delivery Platform: Enabling High Tissue Penetration and Sonogenetic‐Like Potentiating Anti‐Tumor Immunotherapy

AbstractThe characteristics of malignant solid tumors with dense extracellular matrix (ECM) and immunosuppressive microenvironment (IME) seriously restrict the high efficacy and precise treatment of tumors, leading to increased susceptibility to malignant recurrence and metastasis. Herein, an ultrasound‐mediated hydrogel delivery platform HA‐F127@Ti‐MOF‐Au/PEG‐TK‐DOX/PFD (abbr. HFTiDP) based on a “sonogenetic‐like technology” is developed, which encapsulates sonosensitizer (Ti‐MOF‐Au), chemotherapeutic prodrug (PEG‐TK‐DOX), and ECM‐solubilizing drug pirfenidone (PFD), to achieve high tissue penetration and endogenous intracellular regulation of IME. More importantly, exogenous ultrasound stimulation combined with a hydrogel delivery platform can affect drug resistance‐related gene/protein expressions and influence signal transduction pathways in the immune microenvironment, leading to promoting TAAs release to activate immunity, achieving high efficacy in either pancreatic or triple‐negative breast cancer (TNBC), and remarkably inhibiting lung metastasis and splenomegaly. Overall, this work provides a novel strategy for an ultrasound‐mediated niche‐like delivery platform that offers high efficacy and biosafety in localized cancer therapy.

PH-Responsive Charge-Reversal Smart Nanoparticles for Co-Delivery of Mitoxantrone and Copper Ions to Enhance Breast Cancer Chemo-Chemodynamic Combination Therapy.

The poor delivery and limited penetration of nanoparticles into breast cancer tumors remain essential challenges for effective anticancer therapy. This study aimed to design a promising nanoplatform with efficient tumor targeting and penetration capability for effective breast cancer therapy. A pH-sensitive mitoxantrone (MTO) and copper ion-loaded nanosystem functionalized with cyclic CRGDfK and r9 peptide (TPRN-CM) was rationally designed for chemo-chemodynamic combination therapy. TPRN-CM would be quiescent in blood circulation with the CRGDfK peptide on the surface of the nanoparticle to improve its targeting to the tumor. Then, the structure of TPRN-CM changes in the acidic tumor microenvironment, and the r9 peptide can be exposed to make a surface charge reversal to promote deep penetration in the tumor and facilitate their internalization by cancer cells, which was characterized using transmission electron microscopy, dynamic light scattering, flame atomic absorption, etc. The drug release behavior, anti-tumor effects in vivo and in vitro, and the biosafety of the nanoplatform were evaluated. TPRN-CM exhibited remarkable capability to load MTO and Cu2+ with good stability in serum. It can achieve pH-responsive charge reversal, MTO, and Cu2+ release, and can further generate toxic hydroxyl radicals in the presence of glutathione (GSH) and H2O2. In vitro experiments demonstrated that this nanoplatform significantly inhibited proliferation, migration, invasion activities and 3D-tumorsphere growth. In vivo experiments suggested that rationally designed TPRN-CM can be effectively delivered to breast cancer tumors with deep tumor penetration, thereby resulting in a notable reduction in tumor growth and suppression of lung metastasis without causing any apparent side effects. The constructed TPRN-CM nanoplatform integrated tumor targeting, tumor penetration, drug-responsive release, and chemo-chemodynamic combination therapy, thereby providing an intelligent drug delivery strategy to improve the efficacy of breast cancer treatment.

IRGD-mediated liposomal nanoplatforms for improving hepatocellular carcinoma targeted combination immunotherapy and monitoring tumor response via IVIM-MRI.

The combination therapy of targeted treatments and immune checkpoint blockade (ICB) holds great promise for hepatocellular carcinoma (HCC) treatment. However, challenges such as immunogenicity, off-target toxicity of ICB antibodies, low drug co-delivery efficiency, and lack of effective biomarkers to monitor treatment response limit the efficacy of existing targeted immunotherapies. Herein, we synthesized iRGD-modified pH-sensitive liposomal nanoparticles co-encapsulating lenvatinib (Len) and the small molecule PD-1/PD-L1 inhibitor BMS-202 (iRGD-lip@Len/BMS-202) to address issues related to inadequate tumor enrichment and distinct pharmacokinetics of these drugs. Furthermore, intravoxel incoherent motion-magnetic resonance imaging (IVIM-MRI), which is calculated using a biexponential model, can simultaneously reflect both the diffusion of water molecules within the tissue and the microcirculatory perfusion of capillaries. Consequently, we further assessed the feasibility of using IVIM-MRI to monitor the cancer treatment response in nanodrug therapy. These results demonstrated that the iRGD-targeted liposomal nanodrug effectively accumulated in tumors and released in acidic microenvironments. The sustained release of Len facilitated tumor vascular normalization, decreased the presence of Tregs and MDSCs and activated the IFN-γ signaling pathway. This led to increased PD-L1 expression in tumor cells, enhancing the sensitivity of BMS-202. Consequently, there was a synergistic amplification of antitumor immune therapy, resulting in the shrinkage of subcutaneous and orthotopic HCC and inhibition of lung metastasis. Furthermore, IVIM-MRI technology facilitated the non-invasive monitoring of the tumor microenvironment (TME), revealing critical therapeutic response indicators such as the normalization of tumor blood vessels and the degree of hypoxia. Collectively, the combination of Food and Drug Administration (FDA)-approved drugs with iRGD-modified liposomes presents a promising strategy for HCC treatment. Simultaneously, IVIM-MRI provides a non-invasive method to accurately predict the response to this nanodrug.

A novel antagonist of the CCL5/CCR5 axis suppresses the tumor growth and metastasis of triple-negative breast cancer by CCR5-YAP1 regulation

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer (BC) with a high mortality rate, and few effective therapeutic strategies are available. CCL5/CCR5 is an appealing immunotherapeutic target for TNBC. However, its signaling mechanism is poorly understood and its direct antagonists have not been reported. Here, we developed a high-throughput screening (HTS) assay for discovering its antagonists. Verteporfin was identified as a more selective and potent antagonist than the known CCR5 antagonist maraviroc. Without photodynamic therapy, verteporfin demonstrated significant inhibition on TNBC tumor growth through immune regulation, remarkable suppression of lung metastasis by cell-intrinsic mechanism, and a significant extension of overall survival in vivo. Mechanistically, CCR5 was found to be essential for expression of the key hippo effector YAP1. It promoted YAP1 transcription via HIF-1α and exerted further control over the migration of CD8+ T, NK, and MDSC immune cells through chemokines CXCL16 and CXCL8 which were identified from RNA-seq. Moreover, the CCR5-YAP1 axis played a vital role in promoting metastasis by modulating β-catenin and core epithelial-mesenchymal transition transcription factors ZEB1 and ZEB2. It is noteworthy that the regulatory relationship between CCR5 and YAP1 was observed across various BC subtypes, TNBC patients, and showed potential relevance in fifteen additional cancer types. Overall, this study introduced an easy-to-use HTS assay that streamlines the discovery of CCL5/CCR5 axis antagonists. Verteporfin was identified as a specific molecular probe of this axis with great potentials as a therapeutic agent for treating sixteen malignant diseases characterized by heightened CCR5 and YAP1 levels.

Tumor oxygenation nanoliposomes promote deep photodynamic therapy for triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is an aggressive type of breast cancer and has many characteristics including high metastatic rates, poor overall survival, and low response to traditional chemotherapy. Photodynamic therapy (PDT), emerging as a precise treatment modality, has shown promise in improving the antitumor response. However, it still faces challenges such as limited light penetration depth, rapid oxygen consumption, and inadequate targeting ability. In this study, we developed Rose Bengal (RB, photosensitizer) and oxygen co-loaded CREKA-modified UCNP-based nanoliposomes (CLIP-RB-PFOB@UCNP) for tumor targeting and near-infrared (NIR)-triggered deep and long-lasting PDT. Our results demonstrated that CLIP-RB-PFOB@UCNP effectively targeted and accumulated in tumor tissue through the interaction between CREKA and fibronectin, which is overexpressed in tumor cells. Under NIR irradiation, CLIP-RB-PFOB@UCNP exhibited significant destruction of orthotopic tumors, reduced the level of HIF-1α, and efficiently suppressed lung metastasis in a metastatic TNBC model. In conclusion, this study offers new avenues for improving the therapeutic outcomes of PDT for clinical TNBC treatment.

Erbin: an important therapeutic target for blocking tumor metastasis.

Erbin is an adapter protein that interacts with the v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 2 (ERBB2) in epithelial cells. Erbin plays an important role in various signaling pathways, including cell proliferation, apoptosis, and autophagy. Additionally, Erbin is implicated in the pathogenesis and progression of sepsis and various cancers, including breast cancer, acute myeloid leukemia (AML), hepatocellular carcinoma (HCC), and colorectal cancer (CRC). A recent study shows that loss of Erbin increases the release of acyl-carnitine (Acar) through abolishing interaction with prothrombotic protein endothelial cell-specific adhesion molecule (ESAM), promotes mitochondrial oxidative phosphorylation in B cells, and ultimately suppresses lung metastasis of CRC. Accordingly, Erbin provides us with a new potential treatment for tumor metastasis.

Peptide L13S Derived from the BRN2 POU Domain Reduces Metastasis In Vivo and Inhibits Cell Migration and Invasion In Vitro.

The Brain-Specific Homeobox/POU Domain Protein 2 (BRN2) transcription factor supports melanoma progression by regulating the expression of several genes involved in cell migration and invasion. We hypothesized that a peptide designed based on the POU domain of BRN2 could block the BRN2 transcription activity and, consequently, reduce metastasis. Cell viability was accessed by Trypan Blue exclusion dye assay and xCelligence platform. Wound-healing scratch assay and transwell invasion with matrigel membrane assay were performed to analyze cell migration and invasion. The internalization mechanism of the L13S peptide was investigated using confocal microscopy and wound-healing scratch assay. The impact of L13S on cell protein expression was analyzed through western blotting. In vivo assays were conducted to evaluate the protective effect and toxicity of L13S in a metastatic model using murine melanoma cells. Here, we show that the peptide named L13S can inhibit the migration and invasion of murine melanoma cells (B16F10-Nex2) as well as the migration of human melanoma cells (SK-MEL-25 and A375) by regulating the expression of proteins involved in motility. Mechanistically, we found that L13S is internalized by murine melanoma cells via macropinocytosis and binds actin filaments and nuclei. More importantly, in vivo studies indicated that the peptide was able to significantly inhibit lung metastasis in syngeneic models without off-target effects and with virtually no cytotoxicity toward normal organs. L13S peptide is a strong candidate for further development as an anticancer agent for the treatment of melanoma metastasis.

Ultrasound-enhanced nano catalyst with ferroptosis-apoptosis combined anticancer strategy for metastatic uveal melanoma

Uveal melanoma is the most common primary ocular tumor owing to its highly invasive and metastatic characteristics. Currently, standard clinical treatment has an unsatisfied curative effect due to the lack of an effective approach to inhibit the tumor metastasis. Therefore, it is necessary to develop a new strategy that can both restraint local tumors and suppress the ocular tumor metastasis. Herein, we developed ultrasound-responsive nanoparticles (FeP NPs) that can both hinder the growth of in situ ocular tumor and prevent the tumor metastasis through the ferroptosis-apoptosis combined-anticancer strategy. The FeP NPs were assembling by stimulating gallic acid-Fe (III) and paclitaxel, then could be internalized into tumor cells under the cooperative effect of ultrasound, which further activates the intracellular Fenton reaction and generates high reactive oxygen species levels, ultimately leading to mitochondrial damage, lipid per-oxidation, and apoptosis. The FeP NPs can efficiently inhibit the tumor growth in an orthotopic uveal melanoma model. More importantly, the level of the promoting-metastatic factor nerve growth factor receptor (NGFR) secreted by cancer cells is significantly reduced, further limits cancer metastasis to the cervical lymph node and finally inhibits lung metastasis of uveal melanoma. We believe that these designed ultrasound-enhanced nanoparticles possess potential clinical application for preventing the regeneration and metastasis of uveal melanoma.

NHE7 upregulation potentiates the uptake of small extracellular vesicles by enhancing maturation of macropinosomes in hepatocellular carcinoma.

Small extracellular vesicles (sEVs) mediate intercellular communication that contributes to hepatocellular carcinoma (HCC) progression via multifaceted pathways. The success of cell entry determines the effect of sEV on recipient cells. Here, we aimed to delineate the mechanisms underlying the uptake of sEV in HCC. Macropinocytosis was examined by the ability of cells to internalize dextran and sEV. Macropinocytosis was analyzed in Na(+)/H(+) exchanger 7 (NHE7)-knockdown and -overexpressing cells. The properties of cells were studied using functional assays. pH biosensor was used to evaluate the intracellular and endosomal pH. The expression of NHE7 in patients' liver tissues was examined by immunofluorescent staining. Inducible silencing of NHE7 in established tumors was performed to reveal the therapeutic potential of targeting NHE7. The data revealed that macropinocytosis controlled the internalization of sEVs and their oncogenic effect on recipient cells. It was found that metastatic HCC cells exhibited the highest efficiency of sEV uptake relative to normal liver cells and non-metastatic HCC cells. Attenuation of macropinocytic activity by 5-(N-ethyl-N-isopropyl)-amiloride (EIPA) limited the entry of sEVs and compromised cell aggressiveness. Mechanistically, we delineated that high level of NHE7, a sodium-hydrogen exchanger, alkalized intracellular pH and acidized endosomal pH, leading to the maturation of macropinosomes. Inducible inhibition of NHE7 in established tumors developed in mice delayed tumor development and suppressed lung metastasis. Clinically, NHE7 expression was upregulated and linked to dismal prognosis of HCC. This study advances the understanding that NHE7 enhances sEV uptake by macropinocytosis to promote the malignant properties of HCC cells. Inhibition of sEV uptake via macropinocytosis can be exploited as a treatment alone or in combination with conventional therapeutic approaches for HCC.

"Spark" PtMnIr Nanozymes for Electrodynamic-Boosted Multienzymatic Tumor Immunotherapy.

Multienzyme-mimicking redox nanozymes capable of efficient reactive oxygen species (ROS) generation and cellular homeostasis disruption are highly pursued for cancer therapy. However, it still faces challenges from the complicate tumor microenvironment (TME) and high chance for tumor metastasis. Herein, well-dispersed PtMnIr nanozymes are designed with multiple enzymatic activities, including catalase (CAT), oxidase (OXD), superoxide dismutase (SOD), peroxidase (POD), and glutathione peroxidase (GPx), which continuously produce ROS and deplete glutathione (GSH) concurrently in an "inner catalytic loop" way. With the help of electrodynamic stimulus, highly active "spark" species (Ir3+ and Mn3+) are significantly increased, resulting in an effective cascade enzymatic and electrodynamic therapy. Moreover, the cyclic generation of ROS can also facilitate ferroptosis and apoptosis in tumor cells, boosting synergistic therapy. Importantly, lung metastasis inhibition is found, which confirms efficient immunotherapy by the combined effect of immunogenic cell death (ICD) and Mn2+-induced cyclic guanosine monophosphate (GMP)-adenosine monophosphate (AMP) synthase (cGAS)-stimulator of interferon genes (cGAS-STING) pathway, contributing great potential in the treatment of malignant tumors.