STOCK Ctla4^tm1All Pdcd1^tm1.1Shr/J

Stock number: 037076
Product name: Ctla4 KO ; Pdcd1 KO
Research areas: Cancer Research, Cardiovascular Research, Diabetes and Obesity Research, Immunology, Inflammation and Autoimmunity Research, Internal/Organ Research

Description

Mice incorporating knock-out alleles of both the Ctla4 (cytotoxic T-lymphocyte-associated protein 4) and Pdcd1 (programmed cell death 1) genes on mouse Chromosome 1 are a preclinical model of immune checkpoint inhibitors (ICI)‐associated myocarditis. Organ-specific autoimmunity leading to premature death occurs in approximately half of mice heterozygous for the Ctla4 mutation and homozygous for the Pdcd1 mutation (median survival ~100 days).

Detailed description

Immune checkpoint inhibitors (ICI) targeting CTLA4 (cytotoxic T-lymphocyte-associated protein 4) or PDCD1 (programmed cell death 1; also called PD-1/PD-L1) have shown remarkable efficacy across multiple tumor types but are associated with immune-related adverse events, including myocarditis. ICI-associated myocarditis is characterized by myocardial T-cell and macrophage infiltration and significant ECG disturbances.

Mice that are heterozygous for a Ctla4 knock-out allele and homozygous for a Pdcd1 knock-out allele (Ctla4^+/- Pdcd1^-/-) provide a robust preclinical mouse model of ICI-associated myocarditis. Ctla4^+/- Pdcd1^-/- mice develop organ-specific autoimmunity leading to premature death in approximately half of mice (median survival ~100 days). Premature death results from myocardial infiltration by T cells and macrophages and severe ECG abnormalities, closely recapitulating the clinical and pathologic hallmarks of ICI-associated myocarditis observed in patients. The frequency of mortality is reported to be partially sex-dependent, with female mice dying at higher frequency than males, consistent with the characteristics of ICI-associated myocarditis in patients. Mice that succumb to premature death may exhibit some clinical signs prior to death (e.g., reduced body weight gain, reduced movement, and dyspnea as early as 1 month of age), but most mice appear normal. Approximately 25% of mice survive beyond 1 year.

Mice homozygous for both the Ctla4 and Pdcd1 knock-out alleles are embryonic/perinatal lethal, and 100% of mice homozygous for the Ctla4 knock-out alone succumb to a massive lymphoproliferative disease at ~1 month.

Development

Mice carrying the Ctla4^tm1All allele on an C57BL/6 genetic background (The Jackson Laboratory offers an NOD background line, Stock No. 005144) were intercrossed with mice carrying the Pdcd1^tm1.1Shr allele (Stock No. 021157) that had been backcrossed once to C57BL/6J. F1 hybrid mice were intercrossed to recombine the mutant alleles onto the same member of the Chromosome 1 pair (cis) at a calculated distance of 17.03 cM.

Ctla4^tm1All mutation: A neomycin cassette was introduced to exon 3 of the mouse Ctla4 gene via homologous recombination in R1 ((129X1/SvJ x 129S1/Sv)F1-Kitl⁺) embryonic stem cells (ES cells). Resultant mice were backcrossed to C57BL/6.

Pdcd1^tm1.1Shr mutation: A loxP-flanked neomycin resistance (neo) cassette was introduced to intron 1 and a single loxP site was placed in intron 3 of the mouse Pdcd1 gene via homologous recombination in B6.Cg-derived Bruce 4 embryonic stem (ES) cells. Correctly targeted ES cells were transiently transfected with a Cre expression plasmid to delete the neo cassette and exons 2-3. Resultant chimeric mice were bred with C57BL/6J mice for at least 15 generations, but SNP results for Stock No. 021157 show 2 of 27 tested autosomal markers are segregating in an incomplete backcross.

Upon arrival at The Jackson Laboratory, sperm was cryopreserved. Cryorecovery/live colony development will involve C57BL/6J (Stock No. 000664).

Allele

Symbol: Ctla4^tm1All
Name: targeted mutation 1, James P Allison
MGI accession ID: MGI:2180670
Generation method: Targeted
Attributes: Null/Knockout
Synonyms: CTLA-4 KO; CTLA-4^-; Ctla4^o; Ctla4o
Marker symbol: Ctla4
Marker name: cytotoxic T-lymphocyte-associated protein 4
Marker synonyms: ALPS5; CD; Cd152; CELIAC3; CTLA-4; GRD4; GSE; IDDM12; Ly-56; sCTLA4
Chromosome: 1
Strain of origin: (129X1/SvJ x 129S1/Sv)F1-Kitl⁺
Molecular note: The insertion of neomycin selection cassette under the control of a PGK promoter disrupted exon 3.

Allele

Symbol: Pdcd1^tm1.1Shr
Name: targeted mutation 1.1, Arlene H Sharpe
MGI accession ID: MGI:4397682
Generation method: Targeted
Attributes: Null/Knockout
Synonyms: PD-1^-
Marker symbol: Pdcd1
Marker name: programmed cell death 1
Marker synonyms: ADMIO4; AIMTBS; CD279; hPD-1; hPD-l; hSLE1; PD1; PD-1; Pdc1; SLEB2
Chromosome: 1
Strain of origin: B6.Cg-Thy1^a
Molecular note: A floxed neo cassette was inserted upstream of exon 2 and an additional loxP site was inserted downstream of exon 3. Transient cre expression in ES cells removed exons 2 and 3 along with the neo cassette.