Compositions and methods for treating cancer with anti-ROR1 immunotherapy
Inventors
Orentas, Rimas J. • Schneider, Dina • Dropulic , Boro • Dimitrov, Dimiter S. • Zhu, Zhongyu
Assignees
Lentigen Technology Inc • US Department of Health and Human Services
Publication Number
US-11453719-B2
Publication Date
2022-09-27
Expiration Date
2038-11-02
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Abstract
Chimeric antigen receptors containing ROR1 antigen binding domains are disclosed. Nucleic acids, recombinant expression vectors, host cells, antigen binding fragments, and pharmaceutical compositions, relating to the chimeric antigen receptors are also disclosed. Methods of treating or preventing cancer in a subject, and methods of making chimeric antigen receptor T cells are also disclosed.
Core Innovation
The invention provides chimeric antigen receptors (CARs) containing fully human ROR1 antigen binding domains (ScFv) that exhibit high surface expression on transduced T cells, induce strong cytolysis of ROR1-expressing cells, and result in in vivo expansion and persistence of the transduced T cells. These CARs incorporate nucleotide sequences encoding human anti-ROR1 antibodies or fragments thereof, and include transmembrane domains, intracellular signaling domains such as 4-1BB and CD3 zeta, and linkers or spacers derived from extracellular domains like CD8 or IgG4.
The problem addressed is the significant unmet therapeutic need for improved cancer treatments, particularly for cancers and tumors expressing ROR1, an embryonic protein highly expressed in a wide range of cancers but mostly absent in normal adult tissues. Previous therapeutic strategies including surgery, chemotherapy, and murine-derived CARs have limitations such as poor in vivo expansion, rapid disappearance of CAR-T cells, immunogenicity due to non-human sequences, and toxicity. The invention aims to overcome these issues by providing fully human anti-ROR1 CARs with superior therapeutic properties.
Claims Coverage
The claims cover isolated chimeric antigen receptors (CARs) with fully human anti-ROR1 antigen binding domains, methods of making cells expressing these CARs, and methods of generating RNA-engineered CAR-expressing cells, with specific structural features and sequences.
Fully human anti-ROR1 CAR composition
An isolated CAR comprising at least one extracellular antigen binding domain including a single-chain variable fragment (ScFv) ROR1 antigen binding domain with the amino acid sequence of SEQ ID NO: 2 or 8, a CD8 transmembrane domain, and an intracellular signaling domain comprising functional signaling domains of 4-1BB and CD3 zeta.
Nucleotide encoding of the CAR domains
The antigen binding domain comprising the ScFv ROR1 domain is encoded by the nucleotide sequence of SEQ ID NO: 1 or 7, and the CD8 transmembrane domain comprises the amino acid sequence of SEQ ID NO: 22 or a sequence with at least 85% identity.
Linker or spacer domain connection
The extracellular antigen binding domain and the intracellular signaling domain, or both, are connected to the transmembrane domain by a linker or spacer domain obtained from extracellular domains of IgG4, CD8, or CD28.
Specific CAR amino acid sequences
The CAR comprises one of the specified amino acid sequences selected from SEQ ID NOs: 4, 6, 10, or 12.
Making CAR-expressing T cells by transduction
A method of making a cell by transducing a T cell with a vector encoding the CAR with the described ROR1 ScFv antigen binding domain, CD8 transmembrane domain, and intracellular signaling domains (4-1BB and CD3 zeta).
Generating RNA-engineered CAR-expressing cells
A method of generating a population of RNA-engineered cells by introducing in vitro transcribed or synthetic RNA encoding the CAR with the specified ScFv ROR1 domain, CD8 transmembrane domain, and intracellular signaling domains.
The claims primarily relate to compositions and methods encompassing fully human anti-ROR1 CARs comprising specific extracellular ScFv domains, CD8 transmembrane domains, and intracellular 4-1BB and CD3 zeta signaling domains, as well as methods for making CAR-expressing cells and RNA-engineered cells using these constructs, with specified sequences and linker connections.
Stated Advantages
The CARs exhibit a high surface expression on transduced T cells and a high degree of cytolysis of ROR1-expressing cancer cells.
The CAR-transduced T cells demonstrate robust in vivo expansion and persistence, enhancing therapeutic efficacy.
Use of fully human antigen binding domains reduces the likelihood of host anti-CAR immune responses and CAR T cell elimination.
Different CAR constructs allow tuning of therapeutic activity and specificity, potentially reducing off-tumor toxicity and improving safety.
The CARs show specificity for ROR1-expressing tumor cells without auto-activation in the absence of antigen, indicating controlled activity.
Documented Applications
Treatment or prevention of cancers including hematological cancers such as chronic lymphocytic leukemia (CLL), acute lymphocytic leukemia (ALL), mantle cell lymphoma, non-Hodgkin's lymphoma, chronic myelogenous leukemia (CML), pediatric B cell malignancies, multiple myeloma, and refractory or chemotherapeutic-resistant cancers.
Treatment of solid tumors including lung adenocarcinoma, breast cancer (including triple negative breast cancer), ovarian cancer, pancreatic cancer, glioblastoma, sarcomas (Ewing sarcoma, osteosarcoma, rhabdomyosarcoma, fibrosarcoma), melanoma, and other listed adult and pediatric tumors.
Making of CAR T cells genetically engineered to express the disclosed fully human anti-ROR1 CARs for therapeutic use.
Generating populations of RNA-engineered cells expressing the disclosed CARs for adoptive cell therapy.
Diagnosis, prognosing, or determining risk of ROR1-related diseases by detecting ROR1 expression using the disclosed anti-ROR1 antibodies or fragments.
Inhibition of ROR1-dependent T cell inhibition, altering the tumor microenvironment to inhibit tumor growth, suppressing immunosuppression in anti-tumor immune responses by administering the disclosed anti-ROR1 antibodies or fragments.
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