scholarly journals Engineered probiotics for local tumor delivery of checkpoint blockade nanobodies

2019 ◽  
Author(s):  
Candice Gurbatri ◽  
Courtney Coker ◽  
Taylor E. Hinchliffe ◽  
Ioana Lia ◽  
Samuel Castro ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Targeted Delivery ◽  
Cytotoxic T Lymphocyte ◽  
Checkpoint Inhibitors ◽  
Cancer Therapeutics ◽  
Similar Efficacy ◽  
Checkpoint Blockade ◽  
Syngeneic Mouse ◽  
Syngeneic Mouse Model

ABSTRACTImmunotherapies such as checkpoint inhibitors have revolutionized cancer therapy yet lead to a multitude of immune-related adverse events, suggesting the need for more targeted delivery systems. Due to their preferential colonization of tumors and advances in engineering capabilities from synthetic biology, microbes are a natural platform for the local delivery of cancer therapeutics. Here, we present an engineered probiotic bacteria system for the controlled production and release of novel immune checkpoint targeting nanobodies from within tumors. Specifically, we engineered genetic lysis circuit variants to effectively release nanobodies and safely control bacteria populations. To maximize therapeutic efficacy of the system, we used computational modeling coupled with experimental validation of circuit dynamics and found that lower copy number variants provide optimal nanobody release. Thus, we subsequently integrated the lysis circuit operon into the genome of a probioticE. coliNissle 1917, and confirmed lysis dynamics in a syngeneic mouse model usingin vivobioluminescent imaging. Expressing a nanobody against PD-L1 in this strain demonstrated enhanced efficacy compared to a plasmid-based lysing variant, and similar efficacy to a clinically relevant monoclonal antibody against PD-L1. Expanding upon this therapeutic platform, we produced a nanobody against cytotoxic T-lymphocyte associated protein -4 (CTLA-4), which reduced growth rate or completely cleared tumors when combined with a probiotically-expressed PD-L1 nanobody in multiple syngeneic mouse models. Together, these results demonstrate that our engineered probiotic system combines innovations in synthetic biology and immunotherapy to improve upon the delivery of checkpoint inhibitors.SENTENCE SUMMARYWe designed a probiotic platform to locally deliver checkpoint blockade nanobodies to tumors using a controlled lysing mechanism for therapeutic release.

scholarly journals Engineered probiotics for local tumor delivery of checkpoint blockade nanobodies

2020 ◽  
Vol 12 (530) ◽  
pp. eaax0876 ◽  
Author(s):  
Candice R. Gurbatri ◽  
Ioana Lia ◽  
Rosa Vincent ◽  
Courtney Coker ◽  
Samuel Castro ◽  
...  
Keyword(s):  
Targeted Delivery ◽  
Therapeutic Efficacy ◽  
Tumor Regression ◽  
Checkpoint Inhibitors ◽  
Cancer Therapeutics ◽  
Checkpoint Blockade ◽  
Genetic Circuit ◽  
Release Mechanism ◽  
Syngeneic Mouse ◽  
Syngeneic Mouse Model

Checkpoint inhibitors have revolutionized cancer therapy but only work in a subset of patients and can lead to a multitude of toxicities, suggesting the need for more targeted delivery systems. Because of their preferential colonization of tumors, microbes are a natural platform for the local delivery of cancer therapeutics. Here, we engineer a probiotic bacteria system for the controlled production and intratumoral release of nanobodies targeting programmed cell death–ligand 1 (PD-L1) and cytotoxic T lymphocyte–associated protein-4 (CTLA-4) using a stabilized lysing release mechanism. We used computational modeling coupled with experimental validation of lysis circuit dynamics to determine the optimal genetic circuit parameters for maximal therapeutic efficacy. A single injection of this engineered system demonstrated an enhanced therapeutic response compared to analogous clinically relevant antibodies, resulting in tumor regression in syngeneic mouse models. Supporting the potentiation of a systemic immune response, we observed a relative increase in activated T cells, an abscopal effect, and corresponding increases in systemic T cell memory populations in mice treated with probiotically delivered checkpoint inhibitors. Last, we leveraged the modularity of our platform to achieve enhanced therapeutic efficacy in a poorly immunogenic syngeneic mouse model through effective combinations with a probiotically produced cytokine, granulocyte-macrophage colony-stimulating factor (GM-CSF). Together, these results demonstrate that our engineered probiotic system bridges synthetic biology and immunology to improve upon checkpoint blockade delivery.

scholarly journals Modeling Immune Checkpoint Inhibitor Efficacy in Syngeneic Mouse Tumors in an Ex Vivo Immuno-Oncology Dynamic Environment

2020 ◽  
Vol 21 (18) ◽  
pp. 6478
Author(s):  
Daniel T. Doty ◽  
Julia Schueler ◽  
Vienna L. Mott ◽  
Cassie M. Bryan ◽  
Nathan F. Moore ◽  
...  
Keyword(s):  
Real Time ◽  
Mouse Models ◽  
Immune Checkpoint ◽  
Checkpoint Inhibitors ◽  
Ex Vivo ◽  
Confocal Imaging ◽  
Checkpoint Blockade ◽  
Syngeneic Mouse

The immune checkpoint blockade represents a revolution in cancer therapy, with the potential to increase survival for many patients for whom current treatments are not effective. However, response rates to current immune checkpoint inhibitors vary widely between patients and different types of cancer, and the mechanisms underlying these varied responses are poorly understood. Insights into the antitumor activities of checkpoint inhibitors are often obtained using syngeneic mouse models, which provide an in vivo preclinical basis for predicting efficacy in human clinical trials. Efforts to establish in vitro syngeneic mouse equivalents, which could increase throughput and permit real-time evaluation of lymphocyte infiltration and tumor killing, have been hampered by difficulties in recapitulating the tumor microenvironment in laboratory systems. Here, we describe a multiplex in vitro system that overcomes many of the deficiencies seen in current static histocultures, which we applied to the evaluation of checkpoint blockade in tumors derived from syngeneic mouse models. Our system enables both precision-controlled perfusion across biopsied tumor fragments and the introduction of checkpoint-inhibited tumor-infiltrating lymphocytes in a single experiment. Through real-time high-resolution confocal imaging and analytics, we demonstrated excellent correlations between in vivo syngeneic mouse and in vitro tumor biopsy responses to checkpoint inhibitors, suggesting the use of this platform for higher throughput evaluation of checkpoint efficacy as a tool for drug development.

scholarly journals Virotherapy in Germany—Recent Activities in Virus Engineering, Preclinical Development, and Clinical Studies

Viruses ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1420
Author(s):  
Dirk M. Nettelbeck ◽  
Mathias F. Leber ◽  
Jennifer Altomonte ◽  
Assia Angelova ◽  
Julia Beil ◽  
...  
Keyword(s):  
Cancer Cell ◽  
Targeted Delivery ◽  
Viral Vector ◽  
Checkpoint Inhibitors ◽  
Therapeutic Proteins ◽  
Cancer Therapeutics ◽  
Cell Killing ◽  
Oncolytic Viruses ◽  
Preclinical Research ◽  
Research Activities

Virotherapy research involves the development, exploration, and application of oncolytic viruses that combine direct killing of cancer cells by viral infection, replication, and spread (oncolysis) with indirect killing by induction of anti-tumor immune responses. Oncolytic viruses can also be engineered to genetically deliver therapeutic proteins for direct or indirect cancer cell killing. In this review—as part of the special edition on “State-of-the-Art Viral Vector Gene Therapy in Germany”—the German community of virotherapists provides an overview of their recent research activities that cover endeavors from screening and engineering viruses as oncolytic cancer therapeutics to their clinical translation in investigator-initiated and sponsored multi-center trials. Preclinical research explores multiple viral platforms, including new isolates, serotypes, or fitness mutants, and pursues unique approaches to engineer them towards increased safety, shielded or targeted delivery, selective or enhanced replication, improved immune activation, delivery of therapeutic proteins or RNA, and redirecting antiviral immunity for cancer cell killing. Moreover, several oncolytic virus-based combination therapies are under investigation. Clinical trials in Germany explore the safety and potency of virotherapeutics based on parvo-, vaccinia, herpes, measles, reo-, adeno-, vesicular stomatitis, and coxsackie viruses, including viruses encoding therapeutic proteins or combinations with immune checkpoint inhibitors. These research advances represent exciting vantage points for future endeavors of the German virotherapy community collectively aimed at the implementation of effective virotherapeutics in clinical oncology.

scholarly journals Nanotechnology Approaches for Chloroplast Biotechnology Advancements

2021 ◽  
Vol 12 ◽  
Author(s):  
Gregory M. Newkirk ◽  
Pedro de Allende ◽  
Robert E. Jinkerson ◽  
Juan Pablo Giraldo
Keyword(s):  
Synthetic Biology ◽  
Plant Species ◽  
Targeted Delivery ◽  
Current Knowledge ◽  
Ex Vivo ◽  
Future Research ◽  
Agricultural Field ◽  
Diverse Plant Species ◽  
Diverse Plant

Photosynthetic organisms are sources of sustainable foods, renewable biofuels, novel biopharmaceuticals, and next-generation biomaterials essential for modern society. Efforts to improve the yield, variety, and sustainability of products dependent on chloroplasts are limited by the need for biotechnological approaches for high-throughput chloroplast transformation, monitoring chloroplast function, and engineering photosynthesis across diverse plant species. The use of nanotechnology has emerged as a novel approach to overcome some of these limitations. Nanotechnology is enabling advances in the targeted delivery of chemicals and genetic elements to chloroplasts, nanosensors for chloroplast biomolecules, and nanotherapeutics for enhancing chloroplast performance. Nanotechnology-mediated delivery of DNA to the chloroplast has the potential to revolutionize chloroplast synthetic biology by allowing transgenes, or even synthesized DNA libraries, to be delivered to a variety of photosynthetic species. Crop yield improvements could be enabled by nanomaterials that enhance photosynthesis, increase tolerance to stresses, and act as nanosensors for biomolecules associated with chloroplast function. Engineering isolated chloroplasts through nanotechnology and synthetic biology approaches are leading to a new generation of plant-based biomaterials able to self-repair using abundant CO2 and water sources and are powered by renewable sunlight energy. Current knowledge gaps of nanotechnology-enabled approaches for chloroplast biotechnology include precise mechanisms for entry into plant cells and organelles, limited understanding about nanoparticle-based chloroplast transformations, and the translation of lab-based nanotechnology tools to the agricultural field with crop plants. Future research in chloroplast biotechnology mediated by the merging of synthetic biology and nanotechnology approaches can yield tools for precise control and monitoring of chloroplast function in vivo and ex vivo across diverse plant species, allowing increased plant productivity and turning plants into widely available sustainable technologies.

scholarly journals Emerging role of checkpoint blockade therapy in lymphoma

2016 ◽  
Vol 8 (2) ◽  
pp. 81-90 ◽  
Author(s):  
Natalie Galanina ◽  
Justin Kline ◽  
Michael R. Bishop
Keyword(s):  
Cytotoxic T Lymphocyte ◽  
Checkpoint Inhibitors ◽  
Predictive Biomarkers ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
Treatment Modalities ◽  
Critical Pathways ◽  
Heavily Pretreated ◽  
Relapsed Lymphoma

Following the successful application of immune checkpoint blockade therapy (CBT) in refractory solid tumors, it has recently gained momentum as a promising modality in the treatment of relapsed lymphoma. This significant therapeutic advance stems from decades of research that elucidated the role of immune regulation pathways and the mechanisms by which tumors can engage these critical pathways to escape immune detection. To date, two main pathways, the cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and programmed death 1 (PD-1), have emerged as key targets of CBT demonstrating unprecedented activity particularly in heavily pretreated relapsed/refractory Hodgkin lymphoma and some forms of non-Hodgkin disease. Herein we provide a brief discussion of checkpoint blockade in the context of lymphoma biology with a specific focus on novel checkpoint inhibitors and their therapeutic activity. We discuss current clinical trials and the landscape of CBT to underscore both the remarkable progress and foreseeable limitations of this novel treatment strategy. In particular, we build upon state-of-the-art knowledge and clinical insights gained from the early trials to review potential approaches to how CBT may be integrated with other treatment modalities, including chemoimmunotherapy to improve patient outcomes in the future. Finally, as the role of CBT evolves to potentially become a cornerstone of therapy in refractory/relapsed lymphoma, we briefly emphasize the importance of predictive biomarkers in an effort to select appropriate patients who are most likely to derive benefit from CBT.

scholarly journals CRISPR-Cas9 genome editing using targeted lipid nanoparticles for cancer therapy

2020 ◽  
Vol 6 (47) ◽  
pp. eabc9450 ◽  
Author(s):  
Daniel Rosenblum ◽  
Anna Gutkin ◽  
Ranit Kedmi ◽  
Srinivas Ramishetti ◽  
Nuphar Veiga ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Targeted Delivery ◽  
Gene Editing ◽  
Lipid A ◽  
Cancer Therapeutics ◽  
Intracerebral Injection ◽  
Tumor Cell Apoptosis ◽  
Potential Applications ◽  
Targeted Gene Editing

Harnessing CRISPR-Cas9 technology for cancer therapeutics has been hampered by low editing efficiency in tumors and potential toxicity of existing delivery systems. Here, we describe a safe and efficient lipid nanoparticle (LNP) for the delivery of Cas9 mRNA and sgRNAs that use a novel amino-ionizable lipid. A single intracerebral injection of CRISPR-LNPs against PLK1 (sgPLK1-cLNPs) into aggressive orthotopic glioblastoma enabled up to ~70% gene editing in vivo, which caused tumor cell apoptosis, inhibited tumor growth by 50%, and improved survival by 30%. To reach disseminated tumors, cLNPs were also engineered for antibody-targeted delivery. Intraperitoneal injections of EGFR-targeted sgPLK1-cLNPs caused their selective uptake into disseminated ovarian tumors, enabled up to ~80% gene editing in vivo, inhibited tumor growth, and increased survival by 80%. The ability to disrupt gene expression in vivo in tumors opens new avenues for cancer treatment and research and potential applications for targeted gene editing of noncancerous tissues.

In vivo biotinylation of the vasculature in B-cell lymphoma identifies BST-2 as a target for antibody-based therapy

Blood ◽  
2010 ◽  
Vol 115 (3) ◽  
pp. 736-744 ◽  
Author(s):  
Christoph Schliemann ◽  
Christoph Roesli ◽  
Haruhiko Kamada ◽  
Beatrice Borgia ◽  
Tim Fugmann ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Tumor Stroma ◽  
Bioinformatic Analysis ◽  
Syngeneic Mouse ◽  
Biodistribution Studies ◽  
Syngeneic Mouse Model ◽  
Proteomic Study

Abstract The discovery of accessible markers of lymphoma may facilitate the development of antibody-based therapeutic strategies. Here, we describe the results of a chemical proteomic study, based on the in vivo biotinylation of vascular proteins in lymphoma-bearing mice followed by mass spectrometric and bioinformatic analysis, to discover proteins expressed at the tissue-blood border of disseminated B-cell lymphoma. From a list of 58 proteins, which were more than 10-fold up-regulated in nodal and extranodal lymphoma lesions compared with their levels in the corresponding normal host organs, we validated BST-2 as a novel vascular marker of B-cell lymphoma, using immunochemical techniques and in vivo biodistribution studies. Furthermore, targeting BST-2 with 2 independent monoclonal antibodies delayed lymphoma growth in a syngeneic mouse model of the disease. The results of this study delineate a strategy for the treatment of systemic B-cell lymphoma in humans and suggest that anti–BST-2 antibodies may facilitate pharmacodelivery approaches that target the tumor-stroma interface.

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