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Principal Investigator: Nalinikanth Kotagiri
Organization: UNIVERSITY OF CINCINNATI
Fiscal Year: 2024
Award: $497,870
Funding agency: National Cancer Institute
Project Summary
Adoptive cell transfers constitute a new paradigm in cell-based therapeutics with wide-ranging applications
from neurodegenerative, cardiovascular, autoimmune disorders as well as cancer. Autologous lymphocytes
from the patient are genetically engineered to express receptors to specific antigens on the cell surface of
target cells and reinfused back into the patient for therapeutic action. However, once administered the fate of
cells remains uncertain. In order to answer important questions regarding distribution, turnover and eventual
survival of the cells, techniques to non-invasively monitor these genome-edited cells is necessary. Molecular
imaging, particularly Positron emission tomography is advantageous over traditional diagnostic and imaging
tools because it enables diagnosis and tracking of radiotracers in real-time, is non-invasive and has the highest
sensitivity among clinical imaging modalities. We propose to develop analogues of Green fluorescent
protein/luciferase for nuclear imaging, thus combining the highly desirable elements from both these powerful
modalities towards a chemogenetic nuclear imaging modality. The metallophore-transporter complex found in
bacteria is genetically encoded in the prokaryotic DNA and offers the ideal chemical-biological pair that can be
expressed on mammalian cells to enable nuclear imaging and tracking of these cells. By engineering
mammalian cells to ectopically express the bacterial transporters, we will be able to selectively target and
image the genome -edited cells in vivo using metallophore-radionuclide probes. Bacterial metallophores have
evolved to serve as metal chelators for a wide variety of metals with the majority showing a high binding affinity
for iron. However, several pathogenic bacteria secrete metallophores with the highest binding affinity for
copper(II) (Cu). We, therefore, propose to use 64Cu, a popular radionuclide in Positron Emission Tomography
(PET) imaging to generate metallophore/64Cu complexes as contrast agents. We have identified metallophores
that are able to evade the innate immune system and avoid imminent sequestration. Combined with simple
and one-step processing techniques, they are attractive agents for depth-independent real-time imaging,
tracking and identifying genome-edited cells. Because the native or wild type cells do not express these
transporters, we expect minimal uptake in normal mammalian cells including native bacterial “commensal”
flora, as they remain exclusive to pathogenic bacteria. We hypothesize that through a combination of facile
coordinate complexation chemistry, high selectivity, immune evasiveness and non-endogenous nature,
bacterial metallophore/64Cu will serve as ideal nuclear imaging probes to identify and accurately detect GECs
in vivo. In the first phase we will develop and evaluate the bacteria transporter protein mammalian expression
vector and subsequently will be able to incorporate the vectors into CAR-T-Cells for in vivo reporting. In the
next phase we will determine the in vivo pharmacokinetics, biodistribution and PET imaging of the probes and
genome-edited cells in animal models. In summary this project we will use metallophores as dual role
compounds - as a chelator as well as a targeting ligand for imaging, which is novel and innovative. If
successful, this strategy will enable precise labeling of individual genome-edited cells non-invasively in vivo
and potentially avoiding the shortfall of previous PET reporters such as immunogenicity and low selectivity.
Due to the broad impact and transformative potential of this project we envision a quick and clear path to
clinical trials beyond project phase.
Terms: <Adoptive Cell Transfers><Affinity><Animal Model><Animal Models and Related Studies><Antigens><Autoimmune Diseases><Back><Bacteria><Binding><Biodistribution><Biological><CAR T cells><CAR modified T cells><CAR-T><CAR-Ts><CD19><CD19 gene><CRISPR approach><CRISPR based approach><CRISPR method><CRISPR methodology><CRISPR technique><CRISPR technology><CRISPR tools><CRISPR-CAS-9><CRISPR-based method><CRISPR-based technique><CRISPR-based technology><CRISPR-based tool><CRISPR/CAS approach><CRISPR/Cas method><CRISPR/Cas technology><CRISPR/Cas9><CRISPR/Cas9 technology><Cancers><Cardiovascular><Cardiovascular Body System><Cardiovascular Organ System><Cardiovascular system><Carrier Proteins><Cas nuclease technology><Cell Body><Cell Survival><Cell Therapy><Cell Viability><Cell surface><Cells><Chelating Agents><Chelators><Chemicals><Chemistry><Clinical><Clinical Trials><Clustered Regularly Interspaced Short Palindromic Repeats approach><Clustered Regularly Interspaced Short Palindromic Repeats method><Clustered Regularly Interspaced Short Palindromic Repeats methodology><Clustered Regularly Interspaced Short Palindromic Repeats technique><Clustered Regularly Interspaced Short Palindromic Repeats technology><Complex><Complexons><Contrast Agent><Contrast Drugs><Contrast Media><Copper><Cu element><DNA><Dedications><Deoxyribonucleic Acid><Development><Diagnosis><Dorsum><Drug Kinetics><E coli><E. coli><Elements><Engineering><Escherichia coli><Evaluation><Fe element><Genes><Genetic><Genetic Engineering><Genetic Engineering Biotechnology><Genetic Engineering Molecular Biology><Green Fluorescent Proteins><Heart Vascular><Image><Imaging Device><Imaging Instrument><Imaging Tool><Imaging ligands><Immune><Immune Evasion><Immunes><Individual><Innate Immune System><Iron><Label><Ligands><Liquid substance><Luciferase Immunologic><Luciferases><Malignant Neoplasms><Malignant Tumor><Mammalian Cell><Metals><Modality><Molecular Interaction><Nature><Nerve Degeneration><Neuron Degeneration><PD 1><PD-1><PD1><PET><PET Scan><PET imaging><PETSCAN><PETT><Patients><Pharmacokinetics><Phase><Physiologic><Physiological><Positron Emission Tomography Medical Imaging><Positron Emission Tomography Scan><Positron-Emission Tomography><Process><Rad.-PET><Radioactive Isotopes><Radioisotopes><Radionuclides><Radiopaque Media><Receptor Protein><Recombinant DNA Technology><Reporter><Reporting><Role><S aureus><S. aureus><Staph aureus><Staphylococcus aureus><Surface><System><T cells for CAR><T-Cells><T-Lymphocyte><Techniques><Testing><Therapeutic><Time><Transfection><Transport Protein Gene><Transport Proteins><Transporter Protein><Tumor Tissue><adoptive cell therapy><adoptive cellular therapy><analog><autoimmune condition><autoimmune disorder><autoimmunity disease><autologous lymphocytes><bacteria pathogen><bacterial pathogen><biologic><cell mediated therapies><cell type><cell-based therapeutic><cell-based therapy><cellular therapeutic><cellular therapy><chelation><chimeric antigen T cell receptor><chimeric antigen receptor (CAR) T cells><chimeric antigen receptor T cells><chimeric antigen receptor fusion protein T-cells><chimeric antigen receptor modified T cells><circulatory system><clinical imaging><clinical relevance><clinically relevant><commensal flora><commensal microbes><commensal microbiota><commensal microflora><design><designing><detection limit><developmental><diagnostic tool><expression vector><fluid><fluorescence imaging><fluorescent imaging><genetically engineered><genome editing><genomic editing><image-based method><imaging><imaging method><imaging modality><imaging probe><immune evasive><immunogen><immunogenicity><in vivo><innovate><innovation><innovative><liquid><malignancy><metal chelator><model of animal><molecular imaging><molecule imaging><neoplasm/cancer><neural degeneration><neurodegeneration><neurodegenerative><neurological degeneration><neuronal degeneration><non-invasive monitor><noninvasive monitor><novel><nuclear imaging><pathogenic E coli><pathogenic E. coli><pathogenic Escherichia coli><pathogenic bacteria><positron emission tomographic (PET) imaging><positron emission tomographic imaging><positron emitting tomography><programmed cell death 1><programmed cell death protein 1><programmed death 1><radiolabel><radiolabels><radiotracer><real-time images><realtime image><receptor><sle2><social role><systemic lupus erythematosus susceptibility 2><thymus derived lymphocyte><uptake><vector>