In Vivo Models of irAE for Safety and Toxicity Screening of Immunotherapies

Currently Available HuGEMM Models for Safety and Toxicity Evaluation

HuGEMM models are immunocompetent chimeric mouse models, engineered to express humanized drug targets (instead of their murine counterparts) such as genes encoding for immune checkpoint proteins.

Single Knock-in	Double Knock-in	Triple Knock-in
B7H3	TIGIT/PVR	CD47/Sirpα/PD-1
BTLA	PD-L1/TIM-3	CD47/Sirpα/PD-L1
CCR2	PD-L1/TIGIT	PD-1/PD-L1/CD137
CCR8	PD-L1/OX40	PD-1/PD-L1/CTLA4
CD137	PD-L1/LAG3	PD-1/PD-L1/IDO-1
CD27	PD-L1/CTLA4	PD-1/PD-L1/LAG-3
CD28	PD-L1/CD47	PD-1/PD-L1/OX40
CD38	PD-L1/CD40	PD-1/PD-L1/TIGIT
CD39	PD-L1/CD27	PD-1/PD-L1/TIM-3
CD3E	PD-L1/CD137	PD-1/TIM-3/TIGIT
CD40	PD-1/Tim3
CD47	PD-1/TIGIT
CD73	PD-1/Sirpα
CTLA4	PD-1/PD-L1
GITR	PD-1/OX40
IL-1b	PD-1/LAG3
LAG3	PD-1/GITR
OX40	PD-1/CTLA4
OX40L	PD-1/CD47
PD-1	PD-1/CD40
PD-L1	PD-1/CD28
SIGLEC15	PD-1/CD27
Sirpα	PD-1/CD137
STING	PD-1/BTLA
TIGIT	OX40/CD137
TIM-3	NKG2A/CD94
TNFR2	IL2RA/IL2
VEGFR2	CTLA4/Tim3
	CTLA4/OX40
	CTLA4/LAG3
	CTLA4/CD137
	CD47/Sirpα
	CD40/CD137
	CD27/CD137

Safety and Toxicity Readouts Available

• Cytokine analysis
• Liver function tests (ALT, AST)
• Histopathological evaluation of organ toxicity
• Immune cell infiltration in target organs by IHC, Flow cytometry
• Complete blood count (CBC)
• Clinical observations to identify potential adverse effects related to cytokine storm/CRS

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Fig 1. The tumor growth inhibition of urelumab and chimeric urelumab on murine colon tumor CT26.WT in CD137 HuGEMM model. Tumor growth inhibition (TGI) was calculated as: TGI% = (1-Ti/Vi)*100; Ti as the mean tumor volume of the treatment group on the measurement day; Vi as the mean tumor volume of control group at the measurement day.

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Fig 2. Measurement of liver toxicity based on ALT (alanine aminotransferase) and AST (aspartate aminotransferase) levels in CD137 HuGEMM animals. Fasting serum level of ALT (alanine aminotransferase) and AST (aspartate aminotransferase) were measured 2 and 7 days after final dose. One way ANOVA *, **, and *** refer to p<0.05, p<0.01, and p<0.001, respectively.

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Fig 3. Histological assessment of liver inflammation following urelumab and chimeric urelumab treatment in CD137 HuGEMM in vivo model. H&E staining of liver tissue. Liver inflammation was evaluated on a scale of 0-3: 0 = none; 1 = mild; 2 = moderate; 3 = severe.

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Fig 4. Infiltration of CD45+ immune cells in the liver following urelumab and chimeric urelumab treatment in CD137 HuGEMM in vivo model, as indicated by IHC staining - CD45+ cell density were measured by HALO v3.0.311.363.