Browsing by Subject "Cancer immunotherapy"

Initiation and maintenance of a productive anti-tumor immune response requires a functional T-cell repertoire. Disruptions to T-cell function contribute to tumor immune escape, and to failure of the anti-tumor immune response in cancer patients. T-cell dysfunction is particularly severe in certain types of cancers such as glioblastoma (GBM), which is the most common primary malignant brain tumor in adults and is extremely lethal. Despite near universal confinement to the intracranial compartment, GBM frequently depletes both the number and function of systemic T-cells. A lack of understanding of the mechanisms underlying T-cell dysfunction poses challenges to the goal of developing appropriate and meaningful therapeutic platforms. Currently available treatments, including immunotherapies, for GBM and other intracranial diseases have proven ineffective in part because of underlying T-cell dysfunction. Thus, there is an unmet need for therapies that effectively address T-cell dysfunction. In this dissertation, we explore bone marrow T-cell sequestration, a novel mode of T-cell dysfunction present in GBM and other intracranial tumors.

Chapter 1 provides a comprehensive review of the epidemiology, clinical manifestation and diagnosis, and current standard of care for GBM. Chapter 2 outlines immunotherapeutic strategies under investigation for GBM. Chapter 3 describes the fading notion of traditional brain immune privilege but provides the current understanding of how the brain remains immunologically distinct. In Chapter 4, we explore bone marrow T-cell sequestration, and how this mechanism is usurped by GBM and other intracranial tumors to prevent anti-tumor efficacy of T-cell based immunotherapeutic modalities. In Chapter 5, we propose β-arrestin 2 (BARR2) depletion as a strategy to overcome bone marrow T-cell sequestration. In summary, this original work provides encouraging insights for the development of strategies to enhance anti-tumor efficacy of T-cell based immunotherapy for GBM, reversal of bone marrow T-cell sequestration.

Viruses naturally engage innate immunity, induce antigen presentation, and mediate CD8 T cell priming against foreign antigens. Polioviruses can provide a context optimal for generating antigen-specific CD8 T cells, as they have natural tropism for dendritic cells, preeminent inducers of CD8 T cell immunity; elicit Th1-promoting inflammation; and lack interference with innate or adaptive immunity. However, notorious genetic instability and underlying neuropathogenicity has hampered poliovirus-based vector applications. We devised a strategy based on the polio:rhinovirus chimera PVSRIPO, devoid of viral neuropathogenicity after intracerebral inoculation in human subjects, for stable expression of exogenous antigens. PVSRIPO vectors infect, activate, and induce epitope presentation in DCs in vitro; recruit and activate DCs with Th1-dominant cytokine profiles at the injection site in vivo. They efficiently prime tumor antigen-specific CD8 T cells in vivo, induce CD8 T cell migration to the tumor site, delay tumor growth and enhance survival in murine tumor models.

Despite aggressive surgical resection, cytotoxic chemotherapy, and external beam radiotherapy, most cases of glioblastoma (GBM) remain recalcitrant. These outcomes necessitate novel developmental therapeutics that spare normal tissue. Immunotherapy is a promising novel adjuvant treatment that can harness the cytotoxic capacity of the immune system against tumor-associated antigens with exquisite specificity. To circumvent the challenges associated with the advancement of adoptive cellular immunotherapy, we developed a novel treatment platform, which leverages the use of commercially available and clinically translatable nanoparticles (NPs) that can be combined with tumor derived RNA to peripherally activate T cells against GBM antigens. Although cancer vaccines have suffered from weak immunogenicity, we have advanced a NP vaccine formulation that can reshape a host’s immune profile through combinatorial delivery of RNAs encoding for tumor antigens and RNAs encoding for immunomodulatory molecules to mediate long-lived T cell persistence.

We sought to assess if vaccination with amplified tumor derived RNA encapsulated in lipophilic NPs could be assembled to transfect antigen presenting cells (APCs) in vivo and induce therapeutic anti-tumor immunity in pre-clinical murine tumor models. We hypothesized that RNA encapsulated nanoliposomes would localize to reticuloendothelial organs such as the spleen and liver, transfect APCs therein and induce peripheral antigen specific T cell immunity against GBM. Since activated T cells can cross the blood brain barrier and exert their effector functions against GBM antigens, peripheral transfection of APCs by RNA-NPs represents an attractive vaccination approach for priming endogenous immunity against refractory brain tumors.

We screened several translatable NP formulations for their ability to transfect dendritic cells (DCs) in vitro with GFP mRNA. We demonstrated that the NP DOTAP was the most promising translatable formulation compared to alternative cationic liposomal preparations and linear polyethylenimine NPs with and without DC targeting mannose receptors. RNA-NP vaccines formulated in DOTAP were shown to induce in vivo gene expression and preserve RNA stability over time. We determined that intravenous (IV) injection of RNA-NPs was requisite for inducing functional antigen specific immunity, which was superior to standard peptide vaccines formulated in complete Freund’s adjuvant (CFA). IV administered RNA-NPs localized to splenic and hepatic white blood cells (WBCs); these cells expanded antigen specific T cells when transferred to naïve immunocompetent mice. RNA-NPs induced increased percentages of B7 co-stimulatory molecules, but also elicited compensatory PD-L1 expression. We enhanced the immunogenicity and anti-tumor efficacy of RNA-NP vaccines by combining RNA-NPs with immune checkpoint blockade against PD-L1. We also enhanced the immunogenicity and efficacy of this platform by simply combining mRNAs encoding for immunomodulatory cytokines (i.e. GM-CSF). Finally, we demonstrated that RNA-NP vaccines mediate anti-tumor efficacy against intracranial and subcutaneous melanomas and engender therapeutic anti-tumor efficacy in a cellular immunotherapy model against a radiation/temozolomide resistant invasive murine high-grade glioma.

GBM remains invariably associated with poor patient outcomes thus necessitating development of more targeted therapeutics. Clinically translatable RNA-NPs form stable complexes making them amenable to overnight shipping. They induce potent immune responses when administered systemically and mediate robust anti-tumor efficacy that can be enhanced through co-delivery of immunomodulatory RNAs.

This technology can simultaneously bypass the complexity of cellular therapeutics while cutting down the time to generation of personalized vaccines. Since RNA-NP vaccines can be made within days from a tumor biopsy, providing near immediate immune induction against GBM, these formulations can provide a more feasible and effective therapy with a wide range of applicability for all malignancies that can be targeted using RNA obtained from surgical resection of solid tumors.

Therapeutic cancer vaccines have the potential to revolutionize cancer treatment by providing systemic control of both local and metastatic malignancies. However, stimulating antitumor immune responses in patients with cancer has proven difficult and the success rate associated with cancer vaccines is low. Therefore, there is an urgent need to develop novel cancer vaccine strategies to overcome immunosuppression and produce robust anticancer immunity. Addressing this problem will require the development of new tools to achieve improved localized control of the microenvironment in which cancer antigens are present. Motivated by this rationale, we developed a toolbox of sustained-release immunostimulatory fusions to create cancer vaccines that provide tunable spatiotemporal control of immunostimulatory signals. The backbone of these immunostimulatory fusions is a set of protein biopolymers, elastin-like polypeptides (ELPs), that can undergo a thermally triggered phase transition to form a depot upon injection in vivo for localized and sustained delivery of their payload. To create the toolbox, a set of ELPs were covalently fused to the cytokines, granulocyte-macrophage colony-stimulatory factor (GM-CSF), and interleukin-12 (IL-12), the antigenic peptides SVYFFDWL and SIINFEKL, and the immune adjuvant fibronectin III extra domain A (Fn3EDA). The CpG oligodeoxynucleotide (ODN) adjuvant was bound to an ELP with an oligolysine tail by electrostatic complexation. This toolbox of immunostimulatory, depot-forming ELP fusions was used to develop two new cancer vaccines to improve anticancer immunity: 1) an intratumoral (i.t.) in situ vaccine, consisting of a previously developed ELP-Iodine-131 (131I-ELP) radioisotope conjugate for localized radiotherapy combined with the ELP fusion to locally deliver CpG ODN and 2) a subcutaneous (s.c.) subunit vaccine, consisting of ELP fusions to provide sustained release of an antigenic peptide, GM-CSF, and CpG. Characterization studies demonstrated the depot-forming phase behavior and immunostimulatory activity for each fusion in the toolbox, the ability of the ELP-GM-CSF fusion to recruit antigen-presenting cells (APCs) in vivo, and the ability of an ELP-oligolysine fusion to prolong retention and enhance the activity of electrostatically complexed CpG. Furthermore, we demonstrated that an in situ i.t. depot vaccine improves the local and systemic control of 4T1 mammary carcinoma, leading to a synergistic improvement in survival. Finally, we demonstrated that an optimized s.c. depot vaccine augments CD8 T cell response to both a model antigen and a cancer neoantigen and provides protection from melanoma in a tumor challenge experiment. Altogether, these studies establish a versatile delivery platform to spatiotemporally control immune signaling that advances the development of cancer vaccines.