The advent of immune checkpoint inhibitor (ICI) therapies has dramatically changed the treatment landscape for non-small cell lung cancer (NSCLC). However, most patients treated with ICIs are either non-responders or develop progressive disease (PD), despite an initial response to therapy. One of the mechanisms behind this, is the action of suppressive immune cell populations within the tumor microenvironment (TME) such as regulatory T cells (Tregs), which act to shield tumors from the patient's immune response. We plan to target and deplete these Tregs specifically within the TME through the C-C Chemokine G Protein-coupled Receptor 8 (CCR8), which is selectively upregulated in activated tumor-resident Tregs and is absent from essential effector T lymphocytes. Selective elimination of CCR8+ Tregs is anticipated to promote a more effective immune response against NSCLC tumors, while avoiding the dangerous autoimmune side effects associated with non-selective depletion. Targeting CCR8 and other G protein-coupled receptors (GPCRs) with antibody therapeutics has historically been a challenge. This is largely due to the intrinsic properties that make them poor antigens, which include their low abundance, poor immunogenicity, and conformational heterogeneity. To address this, Abilita Bio developed the Enabled Membrane Protein (EMP) directed evolution technology, which generates enhanced GPCR variants with transformative improvements in biophysical properties, while preserving their structure and biological relevance. Using the EMP methodology, we evolved a conformationally stabilized version of CCR8, which was used as a protein antigen for llama immunization and the discovery of VHH single-domain antibody families that uniquely target the transmembrane core of the wild-type human receptor. Our goal is to leverage these antibodies' advantageous properties to construct a molecule with best-in-class potential. In the proposed Phase I SBIR research, we will complete a thorough in vitro characterization and optimization of antibody hits, select a lead molecule, and then perform in vivo efficacy studies in a novel CCR8-humanized mouse model of NSCLC in a head-to-head comparison with a competitor's antibody to show superiority. We will accomplish this goal through the execution of the following scientific aims. Aim 1: we will analyze our diverse VHH hit collection for binding characteristics and biophysical stability in vitro, and then select the best molecules for further characterization. Aim 2: we will characterize antibody pharmacology and potency in mechanism of action studies to select a lead antibody for in vivo studies. In Aim 3: we will test in vivo efficacy of our lead antibody in a CCR8- humanized mouse model of NSCLC. This is, to our knowledge, the first in vivo disease model study for a CCR8 therapeutic program, where human CCR8 will be targeted in the context of a functional TME. Successful completion of this Phase I research will provide critical validation of the therapeutic potential for these novel antibodies and evidence to support our best-in-class hypothesis. Results of these studies will support continued program advancement and a future application for an SBIR Phase II grant to support the completion of preclinical development.
Public Health Relevance Statement: Project Narrative Lung cancer is the leading cause of cancer-related mortality in the United States, and more than 80% of cases fall into the non-small cell lung cancer category. The introduction of checkpoint inhibitor immunotherapy has dramatically changed the outlook for some patients, but most fail to benefit. With support from the National Cancer Institute, we are addressing the urgent need to develop new drugs for non-small cell lung cancer. We are utilizing an innovative technology platform to accelerate the discovery of functional therapeutic antibodies for integral membrane proteins. We have applied it to identify a new class of drugs with the potential to augment the effect of checkpoint immunotherapies and extend their benefit to more patients battling this devastating disease.
Project Terms: Acceleration; Antibodies; immunogen; Antigens; Biological Assay; Assay; Bioassay; Biologic Assays; Biophysics; biophysical foundation; biophysical principles; biophysical sciences; Non-Small-Cell Lung Carcinoma; NSCLC; NSCLC - Non-Small Cell Lung Cancer; Non-Small Cell Lung Cancer; Nonsmall Cell Lung Carcinoma; Cells; Cell Body; Clinical Research; Clinical Study; Combined Modality Therapy; Multimodal Therapy; Multimodal Treatment; combination therapy; combined modality treatment; combined treatment; multi-modal therapy; multi-modal treatment; Control Groups; Dangerousness; Disease; Disorder; Engineering; Family; Flow Cytometry; Flow Cytofluorometries; Flow Cytofluorometry; Flow Microfluorimetry; Flow Microfluorometry; flow cytophotometry; Future; Goals; Grant; Head; Heterogeneity; Human; Modern Man; Immunization; Immunologic Surveillance; Immune Surveillance; Immunologic Surveillances; Immunological Surveillance; Immunological Surveillances; Immunosurveillance; Immunotherapy; Immune mediated therapy; Immunologically Directed Therapy; immune therapeutic approach; immune therapeutic interventions; immune therapeutic regimens; immune therapeutic strategy; immune therapy; immune-based therapies; immune-based treatments; immuno therapy; In Vitro; Integral Membrane Protein; Intrinsic Membrane Protein; Transmembrane Protein; Transmembrane Protein Gene; Lead; Pb element; heavy metal Pb; heavy metal lead; Llama; Membrane Proteins; Membrane Protein Gene; Membrane-Associated Proteins; Surface Proteins; Methodology; Study models; Molecular Conformation; Molecular Configuration; Molecular Stereochemistry; conformation; conformational; conformational state; conformationally; conformations; mortality; Mus; Mice; Mice Mammals; Murine; Patients; Pharmacology; Proteins; Research; Signal Transduction; Cell Communication and Signaling; Cell Signaling; Intracellular Communication and Signaling; Signal Transduction Systems; Signaling; biological signal transduction; Specificity; Survival Rate; T-Lymphocyte; T-Cells; thymus derived lymphocyte; Regulatory T-Lymphocyte; Treg; regulatory T-cells; Technology; Testing; United States; Measures; falls; Dissociation; Mediating; arrestin B; ß-arrestin; beta-arrestin; improved; Surface; Clinical; Phase; Variation; Variant; biologic; Biological; Malignant Tumor of the Lung; Pulmonary Cancer; Pulmonary malignant Neoplasm; lung cancer; Malignant neoplasm of lung; Measurement; Disease Progression; C-C Chemokines; CC Chemokines; ß-Chemokines; beta-Chemokines; Chemotactic Cytokines; Homologous Chemotactic Cytokines; Intercrines; SIS cytokines; chemoattractant cytokine; chemokine; directed evolution; Directed Molecular Evolution; Tumor Immune Escape; cancer evasion; cancer immune escape; cancer immune evasion; tumor evasion; tumor immune evasion; Tumor Escape; Immunological response; host response; immune system response; immunoresponse; Immune response; Therapeutic; Reporter; programs; Immune; Immunes; System; Tumor Volume; innovative technologies; Membrane; membrane structure; receptor; Receptor Protein; receptor binding; receptor bound; tumor growth; receptor density; T-Cell Depletion; T-cell depletion therapy; T-lymphocyte depletion therapy; Structure; novel; member; G-Protein-Coupled Receptors; G Protein-Complex Receptor; G Protein-Coupled Receptor Genes; GPCR; Categories; Disease model; disorder model; Property; response; Molecular Interaction; Binding; CCL1 gene; CCL1; I-309; P500; SCYA1; SISe; TCA3; Progressive Disease; Address; Affinity; National Cancer Institute; NCI Organization; in vivo; Cancer Etiology; Cancer Cause; Collection; Knock-in Mouse; KI mice; knockin mice; Small Business Innovation Research Grant; SBIR; Small Business Innovation Research; Transmembrane Domain; TM Domain; Transmembrane Region; Validation; validations; Characteristics; Pathway interactions; pathway; tumor microenvironment; cancer microenvironment; immunogenicity; designing; design; Population; Therapeutic antibodies; new drug treatments; new drugs; new pharmacological therapeutic; new therapeutics; new therapy; next generation therapeutics; novel drug treatments; novel drugs; novel pharmaco-therapeutic; novel pharmacological therapeutic; novel therapy; novel therapeutics; murine model; mouse model; tumor; response to therapy; response to treatment; therapeutic response; therapy response; treatment response; arm; head-to-head analysis; head-to-head comparison; nanobody; sdAb; single domain antibodies; nanobodies; biophysical characteristics; biophysical characterization; biophysical measurement; biophysical parameters; biophysical properties; FOXP3; Forkhead Box P3; JM2; SCURFIN; FOXP3 gene; humanized mice; humanized mouse; patient subgroups; patient subpopulations; patient subtypes; patient subsets; check point immunotherapy; check point inhibitor therapy; check point inhibitory therapy; check point therapy; checkpoint immunotherapy; checkpoint inhibitor therapy; checkpoint inhibitory therapy; immune check point therapy; immune checkpoint therapy; checkpoint therapy; efficacy study; new drug class; novel drug class; pre-clinical development; preclinical development; Checkpoint inhibitor; immune check point inhibitor; Immune checkpoint inhibitor; antitumor immune response; anti-tumor immune response; preservation; in vivo testing; in vivo evaluation; side effect; Autoimmune; technology platform; technology system
