Liposome-based in situ antigen-modification strategy for "universal" T-cell-receptor engineered T cell in cancer immunotherapy.

T-cell receptor (TCR) engineered T-cell therapy, unlike chimeric antigen receptor T-cell therapy, relies on the inherent ability of TCRs to detect a wider variety of antigenic epitopes, such as protein fragments found internally or externally on cells. Hence, TCR-T-cell therapy offers broader possibilities for treating solid tumors. However, because of the complicated process of identifying specific antigenic peptides, their clinical application still encounters significant challenges. Thus, we aimed to establish a novel "universal" TCR-T "artificial antigen expression" technique that involves the delivery of the antigen to tumor cells using DSPE-PEG-NY-ESO-1157-165 liposomes (NY-ESO-1 Lips) to express TCR-T-cell-specific recognition targets. In vitro as well as in vivo studies revealed that they could accumulate efficiently in the tumor area and deliver target antigens to activate the tumor-specific cytotoxic T-cell immune response. NY-ESO-1 TCR-T therapy, when used in combination, dramatically curbed tumor progression and extended the longevity of mice. Additionally, PD-1 blockage enhanced the therapeutic effect of the aforementioned therapy. In conclusion, NY-ESO-1 Lips "cursed" tumor cells by enabling antigenic target expression on their surface. This innovative technique presents a groundbreaking approach for the widespread utilization of TCR-T in solid tumor treatment.

Membrane fusogenic nanoparticle-based HLA-peptide-addressing universal T cell receptor-engineered T (HAUL TCR-T) cell therapy in solid tumor.

T cell receptor-engineered T (TCR-T) cell therapy has demonstrated therapeutic effects in basic research and clinical trials for treating solid tumors. Due to the peptide-dependent recognition and the human leukocyte antigen (HLA)-restriction, TCR-T cell therapy is generally custom designed to target individual antigens. The lack of suitable universal targets for tumor cells significantly limits its clinical applications. Establishing a universal TCR-T treatment strategy is of great significance. This study designed and evaluated the HLA-peptide-addressing universal (HAUL) TCR-T cell therapy based on HLA-peptide (pHLA) loaded membrance fusogenic deliver system. The pHLA-NP-based tumor cell membrane modification technology can transfer the pHLA onto the surface of tumor cells through membrane fusogenic nanoparticles. Then tumor cells are recognized and killed by TCR-T cells specifically. The HAUL TCR-T cell therapy technology is a universal technology that enables tumor cells to be identified and killed by specific TCR-T cells, regardless of the HLA typing of tumor cells.

A novel ferroptosis-related microRNA signature with prognostic value in osteosarcoma.

The induction of ferroptosis is suggested to be a potential therapeutic strategy for cancers. MicroRNAs (miRNAs) are reported to play an important role in cell death processes. This study aims to construct and validate a risk model based on ferroptosis-related miRNAs (FR_miRNAs) to predict prognosis and identify novel therapeutic targets for osteosarcoma. Data from the Therapeutically Applicable Research to Generate Effective Treatments database are used as the training cohort. A prognostic signature based on two FR_miRNAs (miR-635 and miR-593) is developed using univariate Cox regression, least absolute shrinkage and selection operator regression, and multivariate Cox regression analyses. The area under the curve values of the prognostic signature to predict the 1-year, 2-year, 3-year, and 5-year overall survival rates in patients with osteosarcoma are 0.782, 0.781, 0.722, and 0.777, respectively, indicating a good predictive ability. Based on the risk score, patients are divided into low-risk and high-risk groups. Patients with high-risk scores are associated with poor survival. The risk level is determined to be an independent prognostic factor. A nomogram is established for predicting prognosis. The expression levels of PRNP (miR-635-related ferroptosis-related gene (FRG); P=0.024) and HILPDA (miR-593-related FRG; P=0.025) are significantly different between the low-risk and high-risk groups. All results are validated in an external cohort (GSE39040). The results of the functional assay reveal that miR-635 mimics inhibit osteosarcoma (OS) cell proliferation and migration, whereas miR-593 overexpression exerts the opposite effect. In conclusion, miR-635 and miR-593 exert contrasting regulatory effects on OS cell proliferation and migration.

Neoantigen-targeted TCR-T cell therapy for solid tumors: How far from clinical application.

T cell receptor-engineered T (TCR-T) cells targeting neoantigens present potential immunotherapy for solid tumors. With the continuous optimization of the entire production procedures, the manufacturing process of TCR-T cells is becoming more efficient and productive. However, clinical-scale manufacturing of TCR-T cells still encounters tremendous challenges. Here, we summarize the latest progress of neoantigen-targeted TCR-T cell therapy and focus on the technical difficulties in preparing personalized neoantigen-targeted TCR-T cells and the challenges in clinical applications. Possible approaches for improving TCR-T cell therapy are discussed as well in this review.

Lipophilic recombinant-protein insertion endows lymphocytes with enhanced targeting-infiltration ability in EGFR positive cancer.

Adoptive T cell transfer is one of the most promising ways to combat solid tumors. However, the weak infiltration of T cells into tumor sites has restricted their antitumor efficacy. To overcome this obstacle, we used the lipophilic protein painting strategy to improve tumor targeting and penetrating capacity of lymphocytes for the first time. We synthesized the lipid anchor consisting of a bispecific recombinant protein iRGD-antiEGFR and DSPE-PEG derivates, then successfully inserted it into the membranes of T cells. This surface modification was non-invasive and could efficiently improve the infiltration ability of T cells into multicellular spheroids and tumor masses. The surface modified T cells also displayed superior antitumor activities in EGFR-positive tumor xenografts via systematic infusion. Moreover, the permeability and antitumor efficacy of these surface painted T cells could be remarkably enhanced when used in combination with local low-dose irradiation.

Bifunctional iRGD-anti-CD3 enhances antitumor potency of T cells by facilitating tumor infiltration and T-cell activation.

BACKGROUND: Poor infiltration and limited activation of transferred T cells are fundamental factors impeding the development of adoptive cell immunotherapy in solid tumors. A tumor-penetrating peptide iRGD has been widely used to deliver drugs deep into tumor tissues. CD3-targeting bispecific antibodies represent a promising immunotherapy which recruits and activates T cells. METHODS: T-cell penetration was demonstrated in tumor spheroids using confocal microscope, and in xenografted tumors by histology and in vivo real-time fluorescence imaging. Activation and cytotoxicity of T cells were assessed by flow cytometry and confocal microscope. Bioluminescence imaging was used to evaluate in vivo antitumor effects, and transmission electron microscopy was used for mechanistic studies. RESULTS: We generated a novel bifunctional agent iRGD-anti-CD3 which could immobilize iRGD on the surface of T cells through CD3 engaging. We found that iRGD-anti-CD3 modification not only facilitated T-cell infiltration in 3D tumor spheroids and xenografted tumor nodules but also induced T-cell activation and cytotoxicity against target cancer cells. T cells modified with iRGD-anti-CD3 significantly inhibited tumor growth and prolonged survival in several xenograft mouse models, which was further enhanced by the combination of programmed cell death protein 1 (PD-1) blockade. Mechanistic studies revealed that iRGD-anti-CD3 initiated a transport pathway called vesiculovacuolar organelles in the endothelial cytoplasm to promote T-cell extravasation. CONCLUSION: Altogether, we show that iRGD-anti-CD3 modification is an innovative and bifunctional strategy to overcome major bottlenecks in adoptive cell therapy. Moreover, we demonstrate that combination with PD-1 blockade can further improve antitumor efficacy of iRGD-anti-CD3-modified T cells.

Enhanced and Prolonged Antitumor Effect of Salinomycin-Loaded Gelatinase-Responsive Nanoparticles via Targeted Drug Delivery and Inhibition of Cervical Cancer Stem Cells.

BACKGROUND: Cervical cancer stem cells (CCSCs) represent a subpopulation of tumor cells that possess self-renewal capacity and numerous intrinsic mechanisms of resistance to conventional chemotherapy and radiotherapy. These cells play a crucial role in relapse and metastasis of cervical cancer. Therefore, eradication of CCSCs is the primary objective in cervical cancer therapy. Salinomycin (Sal) is an agent used for the elimination of cancer stem cells (CSCs); however, the occurrence of several side effects hinders its application. Nanoscale drug-delivery systems offer great promise for the diagnosis and treatment of tumors. These systems can be used to reduce the side effects of Sal and improve clinical benefit. METHODS: Sal-loaded polyethylene glycol-peptide-polycaprolactone nanoparticles (Sal NPs) were fabricated under mild and non-toxic conditions. The real-time biodistribution of Sal NPs was investigated through non-invasive near-infrared fluorescent imaging. The efficacy of tumor growth inhibition by Sal NPs was evaluated using tumor xenografts in nude mice. Flow cytometry, immunohistochemistry, and Western blotting were used to detect the apoptosis of CSCs after treatment with Sal NPs. Immunohistochemistry and Western blotting were used to examine epithelial-mesenchymal transition (epithelial interstitial transformation) signal-related molecules. RESULTS: Sal NPs exhibited antitumor efficacy against cervical cancers by inducing apoptosis of CCSCs and inhibiting the epithelial-mesenchymal transition pathway. Besides, tumor pieces resected from Sal NP-treated mice showed decreased reseeding ability and growth speed, further demonstrating the significant inhibitory ability of Sal NPs against CSCs. Moreover, owing to targeted delivery based on the gelatinase-responsive strategy, Sal NPs was more effective and tolerable than free Sal. CONCLUSION: To the best of our knowledge, this is the first study to show that CCSC-targeted Sal NPs provide a potential approach to selectively target and efficiently eradicate CCSCs. This renders them a promising strategy to improve the therapeutic effect against cervical cancer.

Immune-Mediated Antitumor Effect By VEGFR2 Selective Inhibitor For Gastric Cancer.

BACKGROUND: It was previously reported that targeting vascular epithelial growth factor (VEGF)/VEGFR could modulate the antitumor immunity. VEGFR2 inhibitor YN968D1 is a highly selective VEGFR2 inhibitor and was approved for the treatment of late-stage gastric cancer in 2014, but its role in antitumor immunity remains unknown. MATERIALS AND METHODS: In this study, we investigated the effects of YN968D1 on the function of T cells in vitro by testing the cytotoxicity and cytokine production. Next, we constructed peritoneal dissemination and subcutaneous gastric cancer mouse model to assess the cytotoxicity of YN968D1-treated T cells in vivo, respectively. RESULTS: We found that the use of YN968D1 in CD8+ T cells could reduce the expression levels of inhibitory checkpoints, such as Lag-3, PD-1, and Tim3, escalate the production of IFN-γ and IL-2 and promote the cytotoxicity of T cells dramatically in vitro. The transfer of YN968D1-treated T cells achieved better tumor control compared to DMSO-treated T cells or control in both peritoneal dissemination and subcutaneous gastric cancer mouse models. CONCLUSION: Our results indicate that YN968D1 can enhance the T cell-mediated antitumor immunity.

Neoantigen identification strategies enable personalized immunotherapy in refractory solid tumors.

BACKGROUND: Recent genomic and bioinformatic technological advances have made it possible to dissect the immune response to personalized neoantigens encoded by tumor-specific mutations. However, timely and efficient identification of neoantigens is still one of the major obstacles to using personalized neoantigen-based cancer immunotherapy. METHODS: Two different pipelines of neoantigens identification were established in this study: (1) Clinical grade targeted sequencing was performed in patients with refractory solid tumor, and mutant peptides with high variant allele frequency and predicted high HLA-binding affinity were de novo synthesized. (2) An inventory-shared neoantigen peptide library of common solid tumors was constructed, and patients' hotspot mutations were matched to the neoantigen peptide library. The candidate neoepitopes were identified by recalling memory T-cell responses in vitro. Subsequently, neoantigen-loaded dendritic cell vaccines and neoantigen-reactive T cells were generated for personalized immunotherapy in six patients. RESULTS: Immunogenic neo-epitopes were recognized by autologous T cells in 3 of 4 patients who utilized the de novo synthesis mode and in 6 of 13 patients who performed shared neoantigen peptide library, respectively. A metastatic thymoma patient achieved a complete and durable response beyond 29 months after treatment. Immune-related partial response was observed in another patient with metastatic pancreatic cancer. The remaining four patients achieved the prolonged stabilization of disease with a median PFS of 8.6 months. CONCLUSIONS: The current study provided feasible pipelines for neoantigen identification. Implementing these strategies to individually tailor neoantigens could facilitate the neoantigen-based translational immunotherapy research.TRIAL REGSITRATION. ChiCTR.org ChiCTR-OIC-16010092, ChiCTR-OIC-17011275, ChiCTR-OIC-17011913; ClinicalTrials.gov NCT03171220. FUNDING: This work was funded by grants from the National Key Research and Development Program of China (Grant No. 2017YFC1308900), the National Major Projects for "Major New Drugs Innovation and Development" (Grant No.2018ZX09301048-003), the National Natural Science Foundation of China (Grant No. 81672367, 81572329, 81572601), and the Key Research and Development Program of Jiangsu Province (No. BE2017607).

iRGD synergizes with PD-1 knockout immunotherapy by enhancing lymphocyte infiltration in gastric cancer.

Poor infiltration of activated lymphocytes into tumors represents a fundamental factor limiting the therapeutic effect of adoptive cell immunotherapy. A tumor-penetrating peptide, iRGD, has been widely used to deliver drugs into tumor tissues. In this study, we demonstrate for the first time that iRGD could also facilitate the infiltration of lymphocytes in both 3D tumor spheroids and several xenograft mouse models. In addition, combining iRGD modification with PD-1 knockout lymphocytes reveals a superior anti-tumor efficiency. Mechanistic studies demonstrate that the binding of iRGD to neuropilin-1 results in tyrosine phosphorylation of the endothelial barrier regulator VE-cadherin, which plays a role in the opening of endothelial cell contacts and the promotion of transendothelial lymphocyte migration. In summary, these results demonstrate that iRGD modification could promote tumor-specific lymphocyte infiltration, and thereby overcome the bottleneck associated with adoptive immune cell therapy in solid tumors.

Application of Unilateral C2 Translaminar Screw in the Treatment for Atlantoaxial Instability as an Alternative or Salvage of Pedicle Screw Fixation.

OBJECTIVE: To analyze the clinical outcomes of our experience with the unilateral C2 translaminar screw technique and evaluate its feasibility as an alternative or salvage of the pedicle screw. METHODS: Eleven consecutive adult patients with atlantoaxial instability who underwent hybrid fixation techniques using unilateral translaminar screw combined with contralateral C2 pedicle screw and bilateral C1 LMS via posterior arch from January 2010 to December 2013 were retrospectively investigated. The surgical techniques and treatment procedures were described, and clinical symptoms, neurologic function, and imaging appearance were evaluated. RESULTS: The average age in our series was 40.9 years. No vascular or neurologic injuries occurred. No hardware failure or breach of C2 lamina or pedicle was observed in postoperative radiographs or computed tomography scans. Good bone union was achieved in all patients. There were no cases of screw pullout or pseudarthrosis. CONCLUSIONS: Patients with variant vertebral artery or osseous anomaly are good candidates for the hybrid translaminar screw technique as an alternative to or salvage of C2 pedicle screw.