Examination of the roles of insulin receptors in estrogen receptor-positive breast cancer

Abstract

The insulin growth factor (IGF) system plays a crucial role in regulating cell proliferation, differentiation, and survival across multiple physiological contexts, including normal development and tumorigenesis. Dysregulations of the IGF signaling axis are strongly implicated in the pathogenesis of breast cancer. Activation of the type I IGF receptor (IGF1R) and insulin receptor (IR) stimulates downstream signaling cascades such as phosphoinositide 3-kinase (PI3K)/AKT and mitogen-activated protein kinase (MAPK) pathways, promoting oncogenic processes that support tumor initiation, progression, and resistance to endocrine therapies. Although considerable preclinical evidence supported targeting IGF1R in breast cancer, clinical translation has been limited, particularly in endocrine-resistant breast cancer where loss of IGF1R expression is frequently observed. Given that IR remains expressed in these resistant cells, increasing attention has turned to IR as a potential therapeutic target. Recent studies have demonstrated that modulating IR activity can suppress tumor growth and inhibit progression of endocrine-resistant breast cancer cells.In parallel, metabolic dysfunction characterized by obesity and hyperinsulinemia has been increasingly recognized as a modifying factor that further promotes breast cancer progression and therapeutic resistance. Obesity-associated hyperinsulinemia elevates circulating insulin levels, which can directly stimulate IR signaling in tumor cells. Furthermore, breast cancers that develop resistance to endocrine therapies often show downregulated IGF1R expression while shifting sensitivity toward IR, thereby complicating therapeutic strategies that target the IGF system. These observations underscore the need for innovative approaches capable of modulating insulin receptor activity while preserving metabolic homeostasis. Chapter 2 of my thesis focuses on the functional evaluation of AKS-130, a novel insulin-Fc fusion protein. In both parental estrogen receptor-positive (ER+) breast cancer cell lines and their tamoxifen-resistant (TamR) derivatives, AKS-130 demonstrated effective downregulation of IR expression and suppression of downstream insulin-mediated signaling. In vivo studies using xenograft models showed that AKS-130 modestly suppressed tumor growth and exhibited enhanced efficacy when combined with tamoxifen treatment. Importantly, these therapeutic effects were achieved without inducing significant metabolic side effects, supporting the feasibility of IR modulation as a complementary approach in the management of endocrine-resistant breast cancer. Chapter 3 explores the impact of housing and diet condition on mouse metabolism and breast tumor biology. Utilizing a refined xenograft model incorporating diet- and thermoneutral housing-induced obesity, we recreated a physiologically relevant metabolic environment characterized by elevated body mass, hyperinsulinemia, glucose intolerance. In this context, breast tumor xenografts exhibited modestly accelerated growth, reflecting the tumor-promoting influence of metabolic dysregulation. The effects of AKS-130 differed between obese and lean mice. These findings demonstrate the metabolic alterations that occur under varying environmental and dietary conditions and emphasize the complex interplay between systemic metabolic status and tumor responses to IR-targeted therapies. The IR exists in two isoforms, IR-A and IR-B. IR-A deletes a 12-amino-acid sequence encoded by exon 11. Chapter 4 investigates an alternative therapeutic strategy aimed at selectively targeting the IR-A isoform using the engineered T7 gene 2 protein (Gp2) protein scaffolds. Given that IR-A is often preferentially expressed in malignant tissues and mediates mitogenic signaling distinct from the metabolic functions of IR-B, isoform-selective inhibition presents an attractive approach to minimize systemic toxicity while disrupting tumor growth in endocrine-resistant breast cancer. A former colleague in the Yee lab successfully generated three lead Gp2 binders to IR using directed evolution, however, subsequent optimization efforts revealed significant challenges: improvements in binding affinity frequently compromised isoform selectivity due to the high structural similarity between IR-A and IR-B. Furthermore, Fc-fusion formatting of promising binders, which was intended to enhance stability and serum half-life, unexpectedly reduced binding capacity and abolished biological activity, likely due to conformational constraints or steric hindrance imposed by the fusion architecture. These observations highlight intrinsic limitations of current scaffold platforms for achieving robust, isoform-specific IR-A binder suitable for therapeutic development. In this thesis, we explored the complex interactions between insulin receptor signaling, metabolic dysfunction, and endocrine resistance in ER+ breast cancer using multiple complementary approaches. Our findings demonstrate that modulating the insulin receptor holds potential as a strategy to overcome endocrine therapy resistance. Collectively, these studies advance our understanding of how dysregulated IGF1R/IR signaling contributes to ER+ breast cancer progression and underscore both the promise and the challenges of targeting IGF1R/IR pathways for therapeutic gain.