Methods to diagnose and immunize against the virus causing human Merkel cell carcinoma
Inventors
Moore, Patrick S. • Chang, Yuan • Feng, Huichen • Buck, Christopher Brian • Pastrana, Diana V.
Assignees
University of Pittsburgh • US Department of Health and Human Services
Publication Number
US-8524248-B2
Publication Date
2013-09-03
Expiration Date
2028-12-15
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Abstract
The present invention provides isolated or substantially purified polypeptides, nucleic acids, and virus-like particles (VLPs) derived from a Merkel cell carcinoma virus (MCV), which is a newly-discovered virus. The invention further provides monoclonal antibody molecules that bind to MCV polypeptides. The invention further provides diagnostic, prophylactic, and therapeutic methods relating to the identification, prevention, and treatment of MCV-related diseases.
Core Innovation
The invention provides isolated or substantially purified polypeptides, nucleic acids, and virus-like particles (VLPs) derived from a newly-discovered Merkel cell carcinoma virus (MCV). It further provides monoclonal antibody molecules that specifically bind to MCV polypeptides. These components enable diagnostic, prophylactic, and therapeutic methods related to the identification, prevention, and treatment of diseases associated with MCV.
The problem addressed by the invention is the lack of tools and methods to detect and treat Merkel cell carcinoma virus infections, which are associated with the aggressive Merkel cell carcinoma (MCC) skin cancer. Prior to this discovery, no known virus was linked to MCC with the precision of clonal integration, and there were no diagnostic or therapeutic methods based on viral antigens or nucleic acids for MCC. The challenge was to identify and characterize MCV and develop methods for exposure detection, diagnosis, prevention via vaccination, and treatment including immunotherapy.
Claims Coverage
The claims cover methods for assaying Merkel cell carcinoma virus (MCV) exposure and neutralization, with various specific aspects targeted.
Method of assaying for MCV exposure in a patient
Obtaining a tissue or fluid sample from a patient and assaying for the presence of at least one MCV molecule, which can be an MCV nucleic acid, polypeptide, or an antibody specifically binding an MCV polypeptide; a positive test indicates exposure to MCV.
Assaying for MCV exposure by ELISA
Using an enzyme-linked immunosorbent assay (ELISA) wherein the sample is exposed to an MCV polypeptide, particularly VP1, VP2, or VP3 within a virus-like particle (VLP).
Assaying for MCV neutralization using reporter-containing VLPs
Producing MCV virus-like particles containing a reporter construct, exposing them to a sample, infecting permissive cells with this preparation, and assaying for reporter expression; reduced reporter expression indicates neutralizing antibodies.
Applicability to human and non-human patients and sample types
The methods are applicable to human and non-human animals and can include assays such as in situ hybridization when the sample is tissue.
Detection of MCV molecules including nucleic acids and polypeptides
The assay can detect MCV RNA, DNA, or specific polypeptides including large T, small T, VP1, VP2, VP3, or variants T-1 to T-5.
Use of antibodies specifically binding MCV polypeptides
Use of antibodies specific for MCV polypeptides such as large T, small T, VP1, VP2, VP3 or their variants for detection assays.
The claims collectively provide comprehensive methods centered on detecting MCV exposure by targeting viral nucleic acids, proteins, or antibodies, and assessing neutralization using reporter-bearing virus-like particles across various sample types and patient species.
Stated Advantages
Provides specific diagnostic tools to identify MCV infection and exposure in patients.
Enables high sensitivity assays for MCV detection, including ELISA and neutralization assays that significantly improve detection compared to prior art.
Offers potential therapeutic avenues via vaccination with MCV proteins or VLPs and adoptive T cell immunotherapy.
Allows identification of candidate antiviral agents by in vitro assays targeting viral protein-DNA interactions and promoting triplex DNA formation.
Documented Applications
Diagnostic assays for MCV exposure in tissue or fluid samples by detecting MCV nucleic acid, polypeptides, or antibodies.
Use of MCV-derived virus-like particles (VLPs) in ELISA assays to detect antibodies against MCV in human sera.
Development of monoclonal antibodies specific to MCV large T antigen for immunohistochemical analysis.
Neutralization assays using reporter gene-containing MCV VLPs to assess presence of neutralizing antibodies in patient samples or candidate antibody therapeutics.
Vaccination methods for prevention of MCV-related diseases via administration of MCV DNA, polypeptides, or VLPs.
Therapeutic adoptive immunotherapy by ex vivo activation and expansion of patient T lymphocytes with MCV polypeptides followed by reintroduction to the patient.
Screening methods to identify anti-MCV therapeutic agents that attenuate viral infection by interfering with MCV protein-DNA binding or promoting triplex DNA formation.
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