Irreversible Electroporation: An Emerging Immunomodulatory Therapy on Solid Tumors

Irreversible electroporation (IRE), a novel non-thermal ablation technique, is utilized to ablate unresectable solid tumors and demonstrates favorable safety and efficacy in the clinic. IRE applies electric pulses to alter the cell transmembrane voltage and causes nanometer-sized membrane defects or pores in the cells, which leads to loss of cell homeostasis and ultimately results in cell death. The major drawbacks of IRE are incomplete ablation and susceptibility to recurrence, which limit its clinical application. Recent studies have shown that IRE promotes the massive release of intracellular concealed tumor antigens that become an “in-situ tumor vaccine,” inducing a potential antitumor immune response to kill residual tumor cells after ablation and inhibiting local recurrence and distant metastasis. Therefore, IRE can be regarded as a potential immunomodulatory therapy, and combined with immunotherapy, it can exhibit synergistic treatment effects on malignant tumors, which provides broad application prospects for tumor treatment. This work reviewed the current status of the clinical efficacy of IRE in tumor treatment, summarized the characteristics of local and systemic immune responses induced by IRE in tumor-bearing organisms, and analyzed the specific mechanisms of the IRE-induced immune response. Moreover, we reviewed the current research progress of IRE combined with immunotherapy in the treatment of solid tumors. Based on the findings, we present deficiencies of current preclinical studies of animal models and analyze possible reasons and solutions. We also propose possible demands for clinical research. This review aimed to provide theoretical and practical guidance for the combination of IRE with immunotherapy in the treatment of malignant tumors.

Immunotherapy for Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is characterized by extensive tumor heterogeneity at both the pathologic and molecular levels, particularly accelerated aggressiveness, and terrible metastasis. It is responsible for the increased mortality of breast cancer patients. Due to the negative expression of estrogen receptors, progesterone receptors, and human epidermal growth factor receptor 2, the progress of targeted therapy has been hindered. Higher immune response in TNBCs than for other breast cancer types makes immunotherapy suitable for TNBC therapy. At present, promising treatments in immunotherapy of TNBC include immune checkpoints (ICs) blockade therapy, adoptive T-cell immunotherapy, and tumor vaccine immunotherapy. In addition, nanomedicines exhibit great potential in cancer therapy through the enhanced permeability and retention (EPR) effect. Immunotherapy-involved combination therapy may exert synergistic effects by combining with other treatments, such as traditional chemotherapy and new treatments, including photodynamic therapy (PTT), photodynamic therapy (PDT), and sonodynamic therapy (SDT). This review focuses on introducing the principles and latest development as well as progress in using nanocarriers as drug-delivery systems for the immunotherapy of TNBC.

Multifaceted glycoadjuvant@AuNPs inhibits tumor metastasis through promoting T cell activation and remodeling tumor microenvironment

Abstarct Background Cytosine-phosphate-guanine (CpG) dinucleotides has been used as adjuvants for cancer immunotherapy. However, unmodified CpG are not very efficient in clinical trials. Glucose, ligand of C-type lectin receptors (CLRs), can promote DC maturation and antigen presentation, which is the first step of induction of adaptive immune responses. Therefore, conjugation of type B CpG DNA to glucose-containing glycopolymers may enhance the therapeutic effects against tumor by CpG-based vaccine. Methods gCpG was developed by chemical conjugation of type B CpG DNA to glucose-containing glycopolymers. The therapeutic effects of gCpG-based vaccine were tested in both murine primary melanoma model and its metastasis model. Results gCpG based tumor vaccine inhibited both primary and metastasis of melanoma in mice which was dependent on CD8 + T cells and IFNγ. In tumor microenvironment, gCpG treatment increased Th1 and CTL infiltration, increased M1 macrophages, decreased Tregs and MDSCs populations, and promoted inflammatory milieu with enhanced secretion of IFNγ and TNFα. The anti-tumor efficacy of gCpG was dramatically enhanced when combined with anti-PD1 immunotherapy. Conclusions We confirmed that gCpG was a promising adjuvant for vaccine formulation by activating both tumor-specific Th1 and Tc1 responses, and regulating tumor microenvironments. Graphical Abstract

Synergism between CAR-T Cells and a Personalized Tumor Vaccine in Hematological Malignances

Background: CAR-T cells are a tremendous breakthrough in the treatment of certain type of blood cancer, demonstrating impressive and durable responses. However, mechanisms of resistance and relapse have been reported. Mechanisms contributing to relapse after CAR-T therapy include the downregulation of the CAR-T target antigen and the rapid extinguishing of CAR-T cells. We have developed a personalized cancer vaccine whereby patient derived tumor cells are fused to autologous dendritic cells (DC). DC/tumor vaccine induces a broad anti-tumor immunity capable of preventing relapse but may not be effective in patients with advanced disease. Aims: We sought to overcome relapse and resistance to CAR-T therapy to improve the current response rate to CAR-T cells. To this end, we combined CAR-T cells treatment with our personalized DC/tumor vaccine. We postulated that the DC/tumor vaccine would demonstrate synergy with CAR-T cells in a setting where CAR-T cells reduce the bulky disease and the fusion vaccine prevents relapse by expanding CAR-T cells and tumor antigen specific lymphocytes. Methods: To investigate the effects of the CAR-T/fusion vaccine combination, we used the A20 murine B-cell lymphoma model. Syngeneic T cells were obtained from BALB/c mice and retrovirally transduced with a second-generation CAR construct composed of an antigen binding domain that recognizes murine CD19, the CD3ζ domain, 4-1BB as costimulatory molecule and GFP (m19BBz-GFP). GFP expression was used to assess gene-transfer efficiency and monitor the CAR-transduced T cells. The DC/tumor vaccine was obtained by PEG-mediated fusions of A20 cells and BALB/c DC. In the in vitro experiments, T cells transduced with the m19BBz-GFP CAR or non-transduced T cells were co-cultured in the presence or the absence of the DC/A20 vaccine for 3 days. Tumor killing was measured by quantifying Firefly luciferase activity (WT A20) or Renilla luciferase activity (CD19- A20). Vaccine was removed before starting the killing assay. In the in vivo experiment, B-cell lymphoma was induced in BALB/c mice by tail vein injection of A20 cells. The mice were lymphodepleted and treated with m19BBz-GFP CAR-T. The mice were then subcutaneous injected with the DC/A20 fusion vaccine or with PBS (control group). CD8+ T-cells specific for the A20 idiotype epitope were quantified by MHC Class I tetramer analysis. Results: In vitro co-culture of CAR-T cells and DC/A20 vaccine induced a memory-like CAR-T phenotype and strongly improved CAR-T persistence (measured as % of GFP+ T cells in culture). These results were confirmed in vivo where increased CAR-T percentage was observed in the bone marrow and spleen of vaccinated mice with respect to the unvaccinated control group. Moreover, in the vaccinated mice we detected CD8+ T-cells specific for the A20 idiotype epitope demonstrating the expansion of tumor-specific lymphocytes in response to the fusion vaccine. To assess whether the vaccine-induced persistence of CAR-T cells was translated in a higher tumor killing capacity, we performed an in vitro killing assay. To mimic the presence of CD19- clones in the tumor bulk, we used a mixture of WT A20 and CD19-A20 as CAR-T target cells. Enhanced killing capacity against CD19+ tumors was induced by education of the CAR-T with the vaccine. No effects on the CAR-T killing capacity against CD19- A20 were elicited by the fusion vaccine. However, when the same killing assay was performed using as effector cells a mixture of CAR-T and naïve T cells or a mixture of CAR-T and vaccine-educated T cells, we observed that the addition of vaccine-educated T cell to the CAR-T, strongly reduced both A20 WT and A20 CD19- in the cultures. Thus, vaccine-educated T cells are able to kill tumor cells independently from the presence or the absence of the CAR-T target antigen on the tumor. Conclusions: The combination of CAR-T and DC/tumor vaccine, increasing the persistence of CAR-T cells and evoking a polyclonal T cell response against tumor antigens in the non-CAR-transduced T cells, may represent a novel therapeutic strategy to overcame therapeutic resistance and improve current response rate to CAR-T therapy. Disclosures Capelletti: Caris Life Sciences: Current Employment. Stroopinsky: The Blackstone Group: Consultancy. Kufe: REATA: Consultancy, Current equity holder in publicly-traded company; Genus Oncology: Current equity holder in publicly-traded company; Hillstream BioPharma: Current equity holder in publicly-traded company; Canbas: Consultancy. Themeli: Fate Therapeutics: Patents & Royalties. Rosenblatt: Attivare Therapeutics: Consultancy; Imaging Endpoints: Consultancy; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Parexel: Consultancy; Wolters Kluwer Health: Consultancy, Patents & Royalties; Bristol-Myers Squibb: Research Funding. Sadelain: Minerva Biotechnologies: Patents & Royalties; Juno Therapeutics: Patents & Royalties; Fate Therapeutics: Other: Provision of Services (uncompensated), Patents & Royalties; Mnemo Therapeutics: Patents & Royalties; Takeda Pharmaceuticals: Other: Provision of Services, Patents & Royalties; Ceramedix: Patents & Royalties; NHLBI Gene Therapy Resource Program: Other: Provision of Services (uncompensated); St. Jude Children's Research Hospital: Other: Provision of Services; Atara Biotherapeutics: Patents & Royalties. Avigan: Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Research Funding; Kite Pharma: Consultancy, Research Funding; Juno: Membership on an entity's Board of Directors or advisory committees; Partner Tx: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Aviv MedTech Ltd: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Legend Biotech: Membership on an entity's Board of Directors or advisory committees; Chugai: Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; Parexcel: Consultancy; Takeda: Consultancy; Sanofi: Consultancy.

Cancer Stem Cell for Tumor Therapy

Tumors pose a significant threat to human health. Although many methods, such as operations, chemotherapy and radiotherapy, have been proposed to eliminate tumor cells, the results are unsatisfactory. Targeting therapy has shown potential due to its specificity and efficiency. Meanwhile, it has been revealed that cancer stem cells (CSCs) play a crucial role in the genesis, development, metastasis and recurrence of tumors. Thus, it is feasible to inhibit tumors and improve prognosis via targeting CSCs. In this review, we provide a comprehensive understanding of the biological characteristics of CSCs, including mitotic pattern, metabolic phenotype, therapeutic resistance and related mechanisms. Finally, we summarize CSCs targeted strategies, including targeting CSCs surface markers, targeting CSCs related signal pathways, targeting CSC niches, targeting CSC metabolic pathways, inducing differentiation therapy and immunotherapy (tumor vaccine, CAR-T, oncolytic virus, targeting CSCs–immune cell crosstalk and immunity checkpoint inhibitor). We highlight the potential of immunity therapy and its combinational anti-CSC therapies, which are composed of different drugs working in different mechanisms.

IMMU-03. MULTICENTER RANDOMIZED PLACEBO CONTROLLED PHASE III TRIAL OF AN AUTOLOGOUS FORMALINFIXED TUMOR VACCINE (CELLM-001) FOR NEWLY DIAGNOSED GLIOBLASTOMAS

INTRODUCTION The development of novel treatments for glioblastoma is desired and immunotherapy is theoretically expected for highly invasive glioblastoma. An autologous formalin-fixed vaccine (AFTV) derived from resected tumor tissue is stable, contains multiple tumor peptides, and could induce specific immunity. We have conducted three clinical trials in patients with glioblastoma, and the most recent trial was a double-blind, multicenter, phase IIb trial with 63 case enrollments. Although this Phase IIb study revealed no vaccine effects in the whole cohort (mOS: 25.6 months of AFTV group, 31.5 months of the placebo group), the 3-year PFS for patients with total tumor removal was 81% in the AFTV group versus 46% in the placebo group (P=0.067). AFTV vaccine (Cellm-001) may have an effect on certain patient subgroups, and a Phase III study has started in November 2021 (jRCT2031200153). Based on Phase IIb, the enrolled patients were those who could be completely resected on MRI. Cellm-001 administration to a patient in the placebo group at recurrence (crossover) was prohibited. In addition, photodynamic therapy (PDT) was added as a stratification factor because our retrospective study showed a good prognosis of 19 patients who underwent both PDT and AFTV (mOS 47.7 months). PATIENTS AND METHODS Trial design: double-blind (1: 1), phase III multicenter, registration 4 years, observation 2 years. ESTIMATED ENROLLMENT: 112 patients with primary glioblastoma (18-75 years old) whose contrast-enhanced lesion could be completely removed on the image and who received standard local radiotherapy and temozolomide chemotherapy. STRATIFICATION FACTORS: presence or absence of PDT, age, KPS. ADMINISTRATION METHOD: Intradermal administration 3 times before radiochemotherapy and 6 times in parallel with maintenance chemotherapy after completion. PRIMARY ENDPOINT: OS, secondary endpoints: PFS and adverse events. https://jrct.niph.go.jp/en-latest-detail/jRCT2031200153. CONCLUSION An investigator-initiated phase III trial will investigate the efficacy and safety of unique AFTV immunotherapy.

Exploring Chitosan-Shelled Nanobubbles to Improve HER2+ Immunotherapy Through Dendritic Cells Targeting

Immunotherapy is a valuable approach for the treatment of cancer. Nanotechnology-based delivery systems emerged as a powerful tool for improving immunotherapeutics. Therefore, their association have been proposed to overcome some biopharmaceutical limitations of immunotherapy. This work aims at designing a novel immunotherapeutic nanoplatform for the treatment of HER2+ breast cancer. Here, purposely-tailored chitosan-shelled nanobubbles (NBs) were developed for the loading of DNA vaccine. The NBs were then functionalized with anti-CD1a antibody to target dendritic cells (DCs). The NB formulations showed sizes of about 300 nm and a good physical stability up to 6 months stored at 4 °C. The in vitro characterization confirmed that these NBs were able to load DNA with a good encapsulation efficiency (82%). The antiCD1a-functionalized NBs targeted to DCs demonstrated the capability to induce activation of DCs both in human and mouse models, and elicit a specific immune response able to delay tumor growth in vivo in mice. The results are the proof of concept that DC-targeted chitosan nanobubbles loaded with tumor vaccine may provide an attractive nanotechnology approach for the future immunotherapeutic treatment of cancer.

Phase Ib study of patients with metastatic castrate-resistant prostate cancer treated with different sequencing regimens of atezolizumab and sipuleucel-T

BackgroundCombining an immune checkpoint inhibitor with a tumor vaccine may modulate the immune system to leverage complementary mechanisms of action that lead to sustained T-cell activation and a potent prolonged immunotherapeutic response in metastatic castration resistant prostate cancer (mCRPC).MethodsSubjects with asymptomatic or minimally symptomatic mCRPC were randomly assigned in a 1:1 ratio to receive either atezolizumab followed by sipuleucel-T (Arm 1) or sipuleucel-T followed by atezolizumab (Arm 2). The primary endpoint was safety, while secondary endpoints included preliminary clinical activity such as objective tumor response and systemic immune responses that could identify key molecular and immunological changes associated with sequential administration of atezolizumab and sipuleucel-T.ResultsA total of 37 subjects were enrolled. The median age was 75.0 years, median prostate specific antigen (PSA) was 21.9 ng/mL, and subjects had a median number of three prior treatments. Most subjects (83.8%) had at least one treatment-related adverse event. There were no grade 4 or 5 toxicities attributed to either study drug. Immune-related adverse events and infusion reactions occurred in 13.5% of subjects, and all of which were grade 1 or 2. Of 23 subjects with Response Evaluation Criteria in Solid Tumors measurable disease, only one subject in Arm 2 had a partial response (PR) and four subjects overall had stable disease (SD) at 6 months reflecting an objective response rate of 4.3% and a disease control rate of 21.7%. T-cell receptor diversity was higher in subjects with a response, including SD. Immune response to three novel putative antigens (SIK3, KDM1A/LSD1, and PIK3R6) appeared to increase with treatment.ConclusionsOverall, regardless of the order in which they were administered, the combination of atezolizumab with sipuleucel-T appears to be safe and well tolerated with a comparable safety profile to each agent administered as monotherapy. Correlative immune studies may suggest the combination to be beneficial; however, further studies are needed.Trial registration numberNCT03024216.