Biology subjects

Carmon, K. S.

Publications and source records attributed to Carmon, K. S..

10 recordsLinked to original sources

bioRxiv2026

Antibody-drug conjugate combination therapy targeting LGR5 and MET with different payloads enhances efficacy in preclinical colorectal cancer models

Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5) is a marker of cancer stem-like cells frequently upregulated in colorectal cancer (CRC) with lower expression in normal tissues, making it a favorable target for antibody-drug conjugates (ADCs). ADCs combine antibody specificity with potent cytotoxic payloads to enhance efficacy while minimizing systemic toxicity. LGR5-targeting ADCs incorporating different payloads demonstrate strong initial tumor inhibition, yet tumors eventually recur due in part to LGR5 downregulation, necessitating more effective strategies to prevent relapse. We show treatment with chemotherapies or an LGR5-targeting ADC coupled to a topoisomerase 1 inhibitor payload (8E11-CPT2) reduces LGR5 levels and increases MET receptor expression and/or activation in CRC cells, supporting a therapeutic approach to target LGR5 and MET simultaneously. Accordingly, we engineered a MET-targeting ADC (ABT-700-SG3199) via site-specific conjugation of the anti-MET antibody telisotuzumab (ABT-700) with the DNA-crosslinking pyrrolobenzodiazepine (PBD) dimer SG3199. ABT-700-SG3199 exhibited superior potency and efficacy in CRC models compared to the clinical-stage MET-targeting ADCs ABBV-399 and ABBV-400, which use the same antibody backbone conjugated to different payloads. Treatment with ABT-700-SG3199 increased LGR5 expression, and the combination of ABT-700-SG3199 with 8E11-CPT2 enhanced CRC cell-killing efficacy, reinforcing the rationale for a dual-targeting approach. In patient-derived xenografts, combined administration of 8E11-CPT2 and ABT-700-SG3199 markedly delayed tumor relapse and prolonged survival compared with single-agent treatment. Taken together, these findings reveal a reciprocal regulation between MET and LGR5 in response to LGR5- or MET-targeting ADCs in CRC models and support a LGR5/MET dual-targeting therapeutic strategy to enhance efficacy and potentially overcome resistance and relapse.

cancer biology↗

bioRxiv2025

Cetuximab increases LGR5 expression and augments LGR5-targeting antibody-drug conjugate efficacy in patient-derived colorectal cancer models

Colorectal cancer (CRC) remains the second-leading cause of cancer-associated deaths, indicating an urgent need for improved therapeutic options. We previously generated antibody-drug conjugates (ADCs) targeting the cancer stem-like cell marker leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5). However, tumor relapse due to LGR5 downregulation and suboptimal payload selection warranted strategies to improve ADC efficacy. Here we report cetuximab, an EGFR-targeting monoclonal antibody indicated for RASWT metastatic CRC, augments LGR5 expression independent of RAS/PIK3CA mutation status and promotes EGFR-LGR5 interactions. Furthermore, we describe the development of LGR5 ADCs incorporating a camptothecin-derived payload that is well-tolerated and significantly inhibits tumor growth. Importantly, cetuximab in combination with LGR5 ADCs results in enhanced tumor inhibition or regression versus single-agent treatment and extends survival in RASMUT patient-derived xenografts. These findings support growing evidence that ADC combination therapies may be more effective than monotherapies and suggests a broader clinical use for cetuximab in treating RASMUT CRC.

cancer biology↗

bioRxiv2024

Anti-tumor activity of camptothecin analog conjugate of a RSPO4-based peptibody targeting LGR4/5/6 in preclinical models of colorectal cancer.

Antibody-drug conjugates (ADCs) have emerged as a major modality of targeted cancer therapy, yet no ADC has been approved specifically for colorectal cancer (CRC). LGR4/5/6 (leucine-rich repeat containing, G protein-coupled receptor 4, 5, 6) are three related receptors that are expressed at high levels together or alternately in nearly all cases of CRC. ADCs targeting LGR5 have been shown to have robust anti-tumor potency, but not all CRC cells express LGR5 and LGR5-positive tumor cells may lose LGR5 expression due to cancer cell plasticity. R-spondin 4 (RSPO4) is a natural protein ligand of LGR4/5/6 with high affinity for all three receptors. We fused a mutant form of RSPO4 furin domain that retains high affinity binding to LGR4/5/6 to human IgG1 Fc to create a peptibody designated R462. Conjugation of R462 with a camptothecin analog designated CPT2 at eight drugs per peptibody led to the generation of R462-CPT2 that showed highly potent cytotoxic activity in vitro in CRC cell lines expressing any of LG4/5/6. In cell line xenograft and PDX models of CRC, R462-CPT2 demonstrated robust anti-tumor effect. Importantly, R462-CPT2 showed no major adverse effect at therapeutically effective dose levels. These results strongly support the use of RSPO ligand drug-conjugates that target LGR4/5/6 simultaneously for the treatment of CRC.

pharmacology and toxicology↗

bioRxiv2024

Antibody-drug conjugates targeting EGFR ligand Epiregulin inhibit colorectal tumor growth irrespective of RAS mutational status

As colorectal cancer (CRC) remains a leading cause of cancer-related death, identifying therapeutic targets and approaches is essential to improve patient outcomes. The EGFR ligand epiregulin (EREG) is highly expressed in RAS wildtype and mutant CRC with minimal expression in normal tissues, making it an attractive target for antibody-drug conjugate (ADC) development. In this study, we produced and purified an EREG monoclonal antibody (mAb), H231, that had high specificity and affinity for human and mouse EREG. H231 also internalized to lysosomes, which is important for ADC payload release. ImmunoPET and ex vivo biodistribution studies showed significant tumor uptake of 89Zr-labeled H231 with minimal uptake in normal tissues. H231 was conjugated to either cleavable dipeptide or tripeptide chemical linkers attached to the DNA-alkylating payload duocarmycin DM, and cytotoxicity of EREG ADCs was assessed in a panel of CRC cell lines. EREG ADCs incorporating tripeptide linkers demonstrated the highest potency in EREG-expressing CRC cells irrespective of RAS mutations. Preclinical safety and efficacy studies showed EREG ADCs were well-tolerated, neutralized EGFR pathway activity, caused significant tumor growth inhibition or regression, and increased survival in CRC cell line and patient-derived xenograft models. These data suggest EREG is a promising target for the development of ADCs for treating CRC and other cancer types that express high levels of EREG. While the efficacy of clinically approved anti-EGFR mAbs are largely limited by RAS mutational status, EREG ADCs may show promise for both RAS mutant and wildtype patients, thus improving existing treatment options. Significance: EREG-targeting antibody-drug conjugates demonstrate acceptable safety and robust therapeutic efficacy in RAS mutant and wildtype colorectal cancer, suggesting their potential as an alternative to EGFR-targeted therapy to benefit a broader patient population.

cancer biology↗

bioRxiv2022

An antibody-drug conjugate targeting GPR56 demonstrates efficacy in preclinical models of colorectal cancer

Antibody-drug conjugates (ADCs) have become an increasingly successful class of anticancer therapy, particularly within the past few years. Though ADCs are in clinical trials for colorectal cancer (CRC), a candidate has yet to be approved. CRC continues to be a leading cause of cancer-related death, emphasizing the need to identify novel target antigens for ADC development. GPR56, a member of the 7TM receptor family, is upregulated in colorectal tumors compared to normal tissues and located on the surface of CRC cells, making it a promising ADC target. Furthermore, high GPR56 expression occurs in tumors that are microsatellite stable, negative for the CpG methylator phenotype, and show chromosomal instability. We previously reported the generation of a high affinity GPR56-specific monoclonal antibody, 10C7, and we have now mapped the epitope to the C-terminal end of the extracellular domain, proximal to the GPCR proteolysis site. Here, we describe the development of a duocarmycin-conjugated 10C7 ADC. 10C7 co-internalized with GPR56 and trafficked to the lysosomes of CRC cells, which is critical for efficient ADC payload release. Evaluation of the ADC in a panel of CRC cell lines and tumor organoids with different levels of GPR56 expression showed the ADC selectively induced cytotoxicity at low nanomolar concentrations in a GPR56-dependent manner. A nontargeting control ADC showed minimal to no activity. Furthermore, GPR56 ADC exhibited significant antitumor efficacy against GPR56-expressing patient-derived xenograft models of CRC. This study provides rationale for the development of a GPR56-targeted ADC approach to potentially treat a large fraction of CRC patients.

cancer biology↗

bioRxiv2022

Loss of LGR5 through plasticity or gene ablation is associated with therapy resistance and enhanced MET-STAT3 signaling in colorectal cancer cells

Plasticity plays a significant role in colorectal tumor initiation, progression, and drug resistance. LGR5 is highly expressed in colorectal cancer (CRC) and marks functional cancer stem cells (CSCs). While LGR5+ CSCs are tumor-initiating, the majority of CRC cells that disseminate to seed metastases are LGR5-; however, reemergence of LGR5+ CSCs is required to drive metastatic outgrowth. LGR5+ CSCs have been shown to convert to LGR5- CRC cells in response to chemotherapies and this loss of LGR5 promotes a more drug-resistant phenotype. However, the molecular mechanisms that mediate plasticity remain elusive. In this study, we demonstrate conversion of LGR5+ CRC cells to an LGR5- state in response to chemotherapy, LGR5-targeted antibody-drug conjugates (ADCs), or LGR5 gene ablation, led to activation of STAT3. Further investigation revealed increased STAT3 activation occurred a result of increased MET activity. LGR5 overexpression decreased MET-STAT3 activity and sensitized CRC cells to therapy. STAT3 inhibition suppressed MET phosphorylation, while constitutively active STAT3 reduced LGR5 levels and increased MET activity, suggesting a potential feedback mechanism. Combination treatment of MET-STAT3 inhibitors with irinotecan or ADCs substantiated synergistic effects in vitro. In CRC xenografts, STAT3 inhibition combined with irinotecan enhanced tumor growth suppression and prolonged survival. These findings suggest a mechanism by which drug-resistant LGR5- CRC cells acquire a survival advantage through activation of MET-STAT3 and provide rationale for new treatment strategies that target CRC cell plasticity. SignificanceThis study reveals that transition of highly plastic LGR5+ CRC cells to a more drug-resistant LGR5- state involves activation of MET-STAT3 signaling and provides new insight into therapeutic strategies to combat plasticity.

cancer biology↗

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

Chromatin State Dynamics Confers Specific Therapeutic Strategies in Enhancer Subtypes of Colorectal Cancer

The extent and function of chromatin state aberrations during colorectal cancer (CRC) progression is not completely understood. Here, by comprehensive epigenomic characterization of 56 tumors, adenomas, and their matched normal tissues, we define the dynamics of chromatin states during the progression of colorectal cancer. H3K27ac-marked active enhancer state could distinguish between different stages of CRC progression. By epigenomic editing, we present evidence that gains of tumor-specific enhancers for crucial oncogenes, such as ASCL2 and FZD10, was crucial for excessive proliferation. Consistently, combination of MEK plus bromodomain (BET) inhibition was found to have synergistic effects in CRC patient-derived xenograft (PDX) models. Probing inter-tumor heterogeneity, we identified four distinct enhancer subtypes (EpiC), three of which correlate well with previously defined transcriptomic subtypes (CMSs). Importantly, CMS2 can be divided into two EpiC subgroups with significant survival differences. Leveraging such correlation, we devised a combinatorial therapeutic strategy of enhancer-blocking bromodomain inhibitors with pathway-specific inhibitors (PARPi, EGFRi, and TGF{beta}i) for three EPIC groups. Our data suggest that the dynamics of active enhancer underlies colorectal cancer progression and the patient-specific active enhancer patterns govern their unique gene expression patterns which can be leveraged for precision combination therapy.

cancer biology↗

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↗