Biology subjects

Yu, W. A.

Publications and source records attributed to Yu, W. A..

3 recordsLinked to original sources

bioRxiv2021

Drug Conjugates of Antagonistic RSPO4 Mutant For Simultaneous Targeting of LGR4/5/6 for Cancer Treatment

LGR4-6 (Leucine-rich repeating containing, G-protein-coupled receptors 4, 5, and 6) are three related receptors with distinct roles in organ development and stem cell survival. All three receptors are upregulated in gastrointestinal cancers to different levels, and LGR5 has been shown to be enriched in cancer stem cells. Antibody-drug conjugates (ADCs) targeting LGR5 showed robust antitumor effect in vivo but could not eradicate tumors due to plasticity of LGR5-positive cancer cells. As LGR5-negative cells often express LGR4 or LGR6 or both, we reasoned that simultaneous targeting of all three LGRs may provide a more effective approach. R-spondins (RSPOs) bind to LGR4-6 with high affinity and potentiate Wnt signaling. We identified an RSPO4 mutant (Q65R) that retains potent LGR binding but no longer potentiates Wnt signaling. The RSPO4 mutant was fused to the N-terminus of human IgG1-Fc to create a peptibody which was then conjugated with cytotoxins monomethyl auristatin or duocarmycin by site-specific conjugation. The resulting peptibody drug conjugates (PDCs) showed potent cytotoxic effects on cancer cell lines expressing any LGR in vitro and suppressed tumor growth in vivo without inducing intestinal enlargement or other adverse effects. These results suggest that RSPO-derived PDCs may provide a novel approach to the treatment of cancers with high LGR expression.

pharmacology and toxicology↗

bioRxiv2020

Anti-GPR56 monoclonal antibody potentiates GPR56-mediated Src-Fak signaling to modulate cell adhesion

GPR56, also known as ADGRG1, is a member of the adhesion G protein-coupled receptor (aGPCR) family shown to play important roles in cell adhesion, brain development, immune system function, and tumorigenesis. GPR56 is upregulated in colorectal cancer and high expression correlates with poor prognosis. Several studies have shown that GPR56 couples to the G12/13 class of heterotrimeric G-proteins to promote RhoA activation. However, due to its structural complexity and lack of a high affinity receptor-specific ligand, the complete GPR56 signaling mechanism still remains largely unknown. To delineate the activation mechanism and intracellular signaling functions of GPR56, we generated a monoclonal antibody (mAb) which binds GPR56 with high affinity and specificity to the extracellular domain (ECD). We show that overexpression of GPR56 in 293T cells lead to increased phosphorylation of Src, Fak, and paxillin adhesion proteins and activation of the G12/13-RhoA-mediated serum response factor (SRF) pathway. Treatment of cells with anti-GPR56 mAb potentiated Src-Fak phosphorylation, RhoA-SRF signaling, and cell adhesion. Consistently, knockdown of GPR56 in colorectal cancer cells decreased Src-Fak pathway phosphorylation and cell adhesion. Interestingly, GPR56-mediated activation of Src-Fak phosphorylation occurred independent of RhoA, yet mAb-induced potentiation of RhoA-SRF signaling was Src-dependent. Furthermore, we show that the Serine-Threonine-Proline-rich (STP) region of the ECD of GPR56 was not essential for mAb binding, yet was required for activation of Src-Fak signaling. These data support a new ECD-dependent mechanism by which GPR56 functions to regulate cell adhesion through activation of Src-Fak signaling.

molecular biology↗

bioRxiv2019

The most common RNF43 mutant G659Vfs41 is fully functional in inhibiting Wnt signaling and unlikely to play a role in tumorigenesis

RNF43 is an E3 ligase that inhibits Wnt signaling by ubiquitinating Wnt receptors for degradation. It is mutated in various cancer types with the most recurrent mutation being the frameshift G659Vfs41 with frequencies of ~5-8% in colon, stomach and endometrial cancers. The mutation has always been assumed to be loss-of-function that would lead to increased Wnt signaling and tumorigenesis, yet no functional characterization has been reported. We analyzed the distribution of the G659Vfs41 mutation and its association with other cancer gene mutations, and found that the mutation occurred nearly exclusively in tumors with low expression of the DNA mismatch repair gene MLH1. Mutant RNF43-G659Vfs41 was no different from wild type RNF43 in expression, localization, R-spondin binding, promotion of Wnt receptor degradation, inhibition of Wnt signaling. We also found that colon tumors with RNF43-G659Vfs41 had low expression of Wnt/{beta}-catenin signaling markers and were frequently mutated in BRAF. A colon cancer cell line with RNF43-G659Vfs41 and BRAF-V600E mutations was uniquely sensitively to activation of Wnt/{beta}-catenin signaling. These findings indicate that RNF43-G659Vfs41 is likely a passenger mutation in MLH1-down regulated tumors, not a loss-of-function driver mutation that facilitates tumorigenesis. Tumors with RNF43-G659Vfs41 and BRAF-V600E may be susceptible to activators of Wnt signaling.

cancer biology↗