Thymidine kinase gene
Inventors
Levy, John P. • Reed, Rebecca A • MCNULTY, Joseph • Johnson, Jr., Robert G.
Assignees
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Publication Number
US-11364307-B2
Publication Date
2022-06-21
Expiration Date
Abstract
Nucleic acid sequences encoding improved Herpes Simplex Virus Thymidine Kinases are provided, including their use in diagnostic and therapeutic applications. The thymidine kinases may be mutated using conservative mutations, non-conservative mutations, or both. Also provided are gene therapeutic systems, including viral and retroviral particles.
Core Innovation
The disclosed invention relates to a polynucleotide encoding a mutated form of thymidine kinase from human herpes simplex virus type 1 (HSV1-TK), in combination with at least one pro-apoptotic agent, where the encoded HSV1-TK comprises at least two mutated amino acids in a nuclear localization sequence (NLS) relative to a wild-type HSV1-TK amino acid sequence. The NLS comprises amino acid residues 1-33 of SEQ ID NO: 2, and the combination of the mutated HSV1-TK and the at least one pro-apoptotic agent increases cell kill activity relative to a wild-type HSV1-TK.
A retroviral vector format is described for delivery of the polynucleotide encoding the mutated HSV1-TK together with the at least one pro-apoptotic agent. The document also describes therapeutic administration approaches for the retroviral delivery vector, including parenteral administration and intra-arterial infusion approaches such as hepatic arterial infusion, along with systemic administration schedules and therapeutic course structures.
The disclosed embodiments include a course-based regimen with dosing over a therapeutic course and rest periods between courses. The document also includes a Phase 1 clinical trial protocol for Reximmune-C2 (HSV-TK + GM-CSF), describing dose escalation, safety endpoints, imaging-based surrogate expression assessment using [18F]FHBG PET, supportive co-administration (valganciclovir and valacyclovir), and tumor response assessment criteria including irRC and RECIST 1.1.
Claims Coverage
The independent claim coverage centers on a polynucleotide and its encoded mutated HSV1-TK variant with an NLS containing at least two mutated amino acids, together with at least one pro-apoptotic agent, where the combination increases cell kill activity relative to wild-type HSV1-TK. The provided content includes one independent claim and dependent refinements that add sequence constraints, additional sequence signals, specific pro-apoptotic agents, and delivery and administration features.
Mutated HSV1-TK with NLS amino-acid mutations plus pro-apoptotic agent increases cell kill
A polynucleotide encoding a mutated form of HSV1-TK and at least one pro-apoptotic agent, wherein the encoded HSV1-TK comprises at least two mutated amino acids in an NLS relative to a wild-type HSV1-TK amino acid sequence, wherein the NLS comprises amino acid residues 1-33 of SEQ ID NO: 2, and wherein the combination of the mutated HSV1-TK and the at least one pro-apoptotic agent increases cell kill activity relative to a wild-type HSV1-TK.
NLS includes specific mutated residue positions
The NLS includes at least two mutated amino acids corresponding to amino acid residues at positions 25, 26, 32, and 33 of SEQ ID NO: 2.
Nuclear export signal sequence constraint
A nuclear export signal sequence is present with the polynucleotide, defined by LQKKLEELELDG (SEQ ID NO: 24).
Specific pro-apoptotic agents (p53 or Rb)
The cancer suppressor included as the pro-apoptotic agent is p53 or Rb.
Retroviral vector comprises the mutated HSV1-TK polynucleotide and pro-apoptotic agent
A retroviral vector comprises the polynucleotide encoding the mutated HSV1-TK and the at least one pro-apoptotic agent.
Cumulative retroviral vector dose in TVF units
At least 1×10^5 TVF of retroviral vector is administered cumulatively to the patient.
Overall, the claim set requires a mutated HSV1-TK with an NLS containing at least two mutated amino acids and a pro-apoptotic agent, with an expressed increase in cell kill activity relative to wild-type HSV1-TK. Dependent claims further narrow the NLS residue positions, optionally add a nuclear export signal sequence, specify p53 or Rb, and extend into retroviral vector delivery and cumulative TVF dosing.
Stated Advantages
Increases cell kill activity relative to a wild-type HSV1-TK.
Documented Applications
Phase 1 clinical trial of Reximmune-C2 (HSV-TK + GM-CSF), including dose escalation, imaging-based surrogate expression assessment using [18F]FHBG PET, and tumor response assessment criteria (irRC and RECIST 1.1).
Therapeutic treatment using a retroviral delivery vector quantified in TVP/TVF units, including intra-arterial infusion approaches such as hepatic arterial infusion and systemic administration schedules with therapeutic course and rest periods.
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