Abstract 114: Identification of the actionable target, LILRB4, through genetic linkage analysis of Diversity Outbred (DO) F1 mice expressing HER2(neu)
Authors: Jacob Jennifer B. , Wei Wei-Zen , Kidder Benjamin L. , Adeyelu Tolulope , Elliott Andrew , Bepler Gerold , Reyes Joyce D. 
Published: March 2024

Our goal is to identify, in situ, the regulatory genes that dictate the onset age and growth rate of spontaneously arising tumors to implement new intervention strategies. For genetic linkage analysis, we crossed HER2(neu) Tg mice with Diversity Outbred (DO) mice, an outbred population comprised of 8 founding strains (A/J, C57BL/6J, 129S1/SvlmJ, NOD/ShiLtJ, NZO/HILtJ, CAST/EiJ, PWK/PhJ, WSB/EiJ) maintained by non-sibling crossing to ensure each mouse is genetically unique. Using R/QTL package, we linked Quantitative Trait Loci (QTL) in Chr 1 and X with tumor onset age, and a Chr 10 QTL with tumor growth rate. The Chr1 (=human Chr2) QTL was linked to human breast cancer diagnosis in 11 Genome-Wide Association Studies (GWAS; the human GWAS catalog, https://www.ebi.ac.uk/gwas/). Because human GWAS data is deficient in Chr X mapping and also does not assess tumor growth rate as a trait, the QTL loci identified in mouse Chr X and 10 were discoveries beyond the capacity of human GWAS. In total, we identified 26 candidate genes across the 3 QTL. For clinical validation, the genes were analyzed with CodeAI (Caris LS) which associated cancer patient clinical data with candidate gene expression. We found 21/26 genes significantly associated with the survival of patients with primary (n=3,533) and/or metastatic (n=4,870) breast cancers and 17/26 were associated with lung cancer (n=11,334) survival, with 13 overlapping genes between lung and breast cancer, indicating broad applicability of these candidate genes.

We interrogated the immune status in BALB NeuT and (BALBxPWK)F1 NeuT mice because the F1 mice developed tumors around 9 wk, much earlier than the 14 wk observed in BALB NeuT mice. Surprisingly, (BALBxPWK)F1 mice responded to cancer vaccines more vigorously. Single cell RNA (scRNA) libraries from (BALBxPWK)F1 NeuT tumors showed an expanded lymphocyte cluster (12%), compared with BALB NeuT (2%) and an enrichment of T cell activation genes. These findings may reflect a deficiency in tumor cell recognition by the activated anti-tumor effectors and a window of opportunity for correction.

Of the 21 candidate genes, LILRB4, a myeloid cell checkpoint molecule, was of particular interest. Expression of LILRB4 is predictive of patient survival in both breast and lung cancer. Using scRNA sequencing, we found this transmembrane receptor expressed primarily by tumor infiltrating myeloid cells. In a reported scRNA sequencing dataset (GSE161529), LILRB4 was expressed by infiltrating macrophages in human breast cancers, and is also correlated with reduced survival. In a PAN Cancer ATACseq dataset (Corces et al., 2018), promoter accessibility of LILRB4 was associated with reduced survival. Taken together, there are multiple immune regulatory mechanisms of tumor progression and LILRB4 is an important new actionable target. CA76340 (WZW), KCI TBM program (JBJ) and CCSG P30 CA022453 (GB).

Citation Format: Jennifer B. Jacob, Wei-Zen Wei, Benjamin L. Kidder, Tolulope Adeyelu, Andrew Elliott, Gerold Bepler, Joyce D. Reyes. Identification of the actionable target, LILRB4, through genetic linkage analysis of Diversity Outbred (DO) F1 mice expressing HER2(neu) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 114.