Our Research
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At the heart of our research projects are Wnt proteins and their signaling pathways. Wnt genes encode a family of secreted lipid-modified growth factors (there are 19 Wnt genes in the mammalian genome) that regulate embryonic development and tissue homeostasis. Deregulated Wnt signaling impacts a large number of human diseases, including neurodegeneration and cancer. In the Willert lab, we incorporate biochemical, genetic and cell biological approaches to study these proteins and their signaling cascades.
Selective Activation of WNT Signaling
In 2003, we published the first purification of a WNT protein, Wnt3a, which has enabled countless studies, including the dissection of WNT signal transduction and the manipulation of stem cell populations, to name a few. However, WNT proteins are extremely difficult to work with. First, because of a covalently attached lipid, WNTs are extremely hydrophobic, necessitating high concentrations of detergents to maintain their solubility in aqueous and biologically compatible buffers. Second, with their low expression levels (the best expression systems yield about 100 micrograms per liter of culture medium) and an extremely cumbersome and lengthy purification scheme (our current purification protocol involves 4 sequential chromatography runs), it has been challenging to produce adequate amounts to support ongoing research activities. Finally, some WNTs are quite promiscuous in their interactions with their receptors, making it difficult to selectively activate WNT pathways in complex biological systems where multiple WNT receptors may be expressed. To overcome these challenges, we along with several other groups have engineered bi-specific antibodies that, like WNTs, heterodimerize WNT receptors (Frizzled, FZD) with co-receptors (LRP5/6), and thereby activate downstream signaling. In our lab, we developed a FZD7 and LRP6 bispecific antibody, which we refer to as a WNT mimetic (other labs refer to such WNT agonists as WNT surrogates or FLAgs [Fzd-Lrp Agonist]). We are currently using these tools to selectively activate WNT signaling in pluripotent stem cells, organoids and in mice.
FZD7, a WNT Receptor, in Mammary Tumors
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Our studies on the role of WNT signaling in human embryonic stem cells led to the observation that the WNT receptor encoded by the FRIZZLED7 (FZD7) gene is critically important in the maintenance of the pluripotent stem cells. Furthermore, FZD7 is expressed in a variety of adult stem cell populations, including the intestinal crypt, the mammary gland and the hair follicle. Importantly, FZD7 expression is significantly upregulated in a variety of solid tumors, including ovarian and breast cancer. To further study the role of FZD7 in cancer, we have used a genetically modified mouse model for breast cancer (MMTV-Wnt1) and demonstrated that, upon transplantation into mouse mammary gland, FZD7-expressing cancer cells rapidly expand into tumors. Treatment of these mammary tumors with a FZD7-specific antibody drug conjugate (ADC) blocked tumor growth. We are currently exploring the viability of this ADC to treat human cancers.
Mice transplanted with mammary tumors were treated with a FZD7-specific antibody drug conjugate (F7-ADC) or with saline (PBS). Tumor growth is significantly reduced in F7-ADC treated mice.
WNT and Genomic Instability
WNT signaling is a major regulator of developmental processes and its deregulation contributes to many types of cancers. Mutations in β-catenin, Axin and APC, which encode critical components of WNT signal transduction, are observed in cancers, and the prevailing model is that these mutations lead to changes in expression of WNT target genes, such as cMyc and CyclinD1, which in turn contribute to cancer development and progression. However, mutations in these WNT signaling components have also been associated with an increase in chromosomal instability (CIN), raising the possibility that mis-regulated WNT signaling may also promote tumorigenesis by destabilizing the genome.
In our lab, we observed that an excess of WNT activity resulted in aneuploidy during reprogramming, consistent with findings that mutations in WNT pathway components contribute to chromosomal instability. On the other hand, our collaborator Professor Sergio Acebrón (University of Heidelberg, Germany) found that inhibition of autocrine WNT signaling in pluripotent stem cells triggers DNA replication stress and damage in S-phase, as well as chromosome mis-segregation in ensuing mitoses. To reconcile our seemingly disparate observations, we hypothesize that WNT signaling adheres to a “Goldilocks principle”, in which a fine-tuned level of activity is required to ensure faithful DNA replication and segregation of the genetic blueprints during stem cell maintenance and differentiation.