# Time for arginine methylation: PRMT5 inhibition to advance cholangiocarcinoma treatment

Romain Désert, Lipika Goyal, Thomas F Baumert

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Romain Désert, Lipika Goyal, Thomas F Baumert. Time for arginine methylation: PRMT5 inhibition to advance cholangiocarcinoma treatment. Gut, 2024, Online ahead of print. 10.1136/gutjnl-2024-333632. [https://doi.org/10.1136/gutjnl-2024-333632](https://doi.org/10.1136/gutjnl-2024-333632). [hal-04902649](https://HAL.inhal-04902649)

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# Time for arginine methylation: PRMT5 inhibition to advance cholangiocarcinoma treatment

Romain Désert,<fnref n="1" /> Lipika Goyal,<fnref n="2" /> Thomas F Baumert ID <fnref n="1" />,<fnref n="3" />

Cholangiocarcinoma (CCA) is a highly aggressive adenocarcinoma of the biliary tract system with unsatisfactory therapeutic options.<fnref n="1" /> Standard frontline treatment for unresectable or metastatic CCA consisting of cisplatin and gemcitabine combined with checkpoint inhibitors targeting programmed cell death ligand 1 or programmed cell death 1 offers objective response rates of less than 30% and a median survival of approximately a year.<fnref n="1" /> Targeted therapies against FGFR2 fusions and IDH1 mutations have gained regulatory approval in CCA, but these are applicable only in a minority of patients.<fnref n="1" /> Disease-agnostic approvals of therapies targeting HER2 overexpression, NTRK fusions, RET fusions and microsatellite-unstable tumours also benefit patients with CCA, but again, only a small minority. Therefore, novel strategies to treat CCA are urgently needed.

Molecular heterogeneity stands as a major barrier to improving outcomes in CCA. Genetic alterations in DNA only explain a part of this heterogeneity. A rising number of studies suggest a major role for epigenetic perturbations in controlling CCA fate.<fnref n="2" /> Indeed, epigenetic vulnerabilities including histone modifications have been suggested as novel CCA targets.<fnref n="1" /> An example of a histone regulator is the protein arginine methyltransferase 5 (PRMT5). PRMT5 forms a homotetramer that associates with methylosome protein 50 (MEP50) in a highly active complex that exhibits high affinity for arginine residues. Via histone methylation, PRMT5 functions as a transcriptional co-repressor supporting gene expression of oncogenic signalling via regulation of genes such as p53, NFκB or p21.<fnref n="2" /> In addition, PRMT5 regulates splicing via its role as the enzymatic component of the methylosome, a multi-subunit complex containing

MEP50, facilitating small nuclear ribonucleoprotein assembly (figure 1). PRMT5 and MEP50 functions have been shown to be important in regulating genome stability and DNA repair.<fnref n="3" /> Preclinical studies of PRMT5 inhibitors have shown antitumour activity across several tumour types including lymphoma, breast cancer, head and neck or oesophageal cancer,<fnref n="2" /> but the potential of PRMT5 as a therapeutic target in CCA has been largely unexplored to date.

Elurbide *et al*⁴ investigated the potential of PRMT5 as a novel therapeutic target in CCA by evaluating expression of the target in patient tissues and inhibition of the target in preclinical models. First, using publicly available transcriptomic datasets from human CCA tissues, they showed overexpression of PRMT5 and MEP50 in intrahepatic CCA but not extrahepatic CCA. However, by immunohistochemistry, Elurbide *et al* showed overexpression of both proteins in both anatomic subtypes. Using several preclinical models, the authors convincingly showed proof of concept for PRMT5 as a key mediator of DNA stability in CCA. They further showed that the PRMT5 inhibitors, GSK3326595 and JNJ64619178, significantly reduced both the proliferation of CCA cell lines and organoid growth and showed synergistic effects with cisplatin and gemcitabine. PRMT5 inhibition induced apoptosis and led to decreased expression of genes involved in chromatin remodelling and DNA repair. Notably, in two immunocompetent mouse models including an orthotopic model based on the hydrodynamic tail vein injection technique (HDTV), PRMT5 inhibition significantly reduced tumour burden without significant adverse effects. PRMT5 inhibition also led to increased expression of genes involved in antigen presentation and immune response, shown by increased infiltration of CD4+ and CD8+ T cells, suggesting a potential effect in promoting antitumour immunity as seen in preclinical models of other tumour types.⁵ The use of two complementary immunocompetent mouse models in the study provides robust in vivo proof of concept.

Future studies may evaluate the functional impact on the tumour microenvironment in other HDTV-based models mimicking the immune variability found in patients⁶ and investigate the combination of PRMT5 inhibitors with checkpoint inhibitors. Collectively, the study of Elurbide *et al*⁴ provides evidence for the important role of PRMT5 in maintaining genomic stability in CCA, extending similar findings in other cancer types,<fnref n="3" /> and identifies PRMT5 as a promising therapeutic target in CCA.

A clinical application of this work includes the opportunity to treat patients with tumours bearing methylthioadenosine phosphorylase (MTAP) deletion. MTAP cleaves methylthioadenosine (MTA) to generate precursor substrates for methionine and adenine salvage pathways. Hence, MTAP deletion causes MTA to accumulate leading to PRMT5 inhibition, creating a state whereby MTAP-deleted cells are particularly vulnerable to further inhibition of PRMT5⁷ (figure 1). This was functionally confirmed for CCA by Elurbide *et al*⁴ by showing that MTAP-deleted cell lines were far more sensitive to PRMT5 inhibitors than cell lines with preserved MTAP expression. MTAP is often co-deleted with CDKN2A, a tumour suppressor gene, due to their close proximity on chromosome 9p21.3.⁸ Homozygous deletions in this region, encompassing both CDKN2A and MTAP, are frequently observed in various cancers (40% of glioblastomas; 25% of melanomas, urothelial carcinomas and pancreatic adenocarcinomas; and 15% of non-small cell lung carcinomas).⁸ For CCA, Elurbide *et al*⁴ showed that MTAP protein expression was undetectable in ~25% of intrahepatic CCA suggesting that MTAP deletion is also a frequent event in CCA. By genomic sequencing, the frequency of MTAP deletion was found to be 15% in a large cohort of patients with intrahepatic CCA.⁹ Overall, these studies identify an expanded population of patients that could benefit from PRMT5 inhibitors.<fnref n="1" /> Notably, multiple tumour profiling companies report CDKN2A loss but not MTAP loss, so improved reporting or further education of the oncology community will be important as this class of drugs emerges in the clinic.

Recent advances in small molecule-based epigenetic therapies have sparked significant hope in the field of cancer treatment. A variety of inhibitors, including those targeting histone deacetylase, DNA N-methyltransferase, enhancer of zeste homolog 2, bromodomain and extra-terminal domain and notably

<footnote n="1">¹University of Strasbourg, Inserm, Institute for Translational Medicine and Liver Disease (ITM), UMR S1110, Strasbourg, France</footnote>

<footnote n="2">²Division of Oncology, Department of Medicine, Stanford School of Medicine, Palo Alto, California, USA</footnote>

<footnote n="3">³Gastroenterologie-Hepatology Service, Strasbourg University Hospitals, Strasbourg, France</footnote>

<footnote>**Correspondence to** Prof. Thomas F Baumert; Thomas.Baumert@unistra.fr</footnote>
<figure><img src="image_2.png" /><figcaption>**Figure 1** Mechanism of action model for PRMT5 as a therapeutic target in CCA. In CCA, PRMT5 and MEP50 are overexpressed and play an important role in regulating genome stability as part of the methylosome complex, which controls the assembly and stability of splicing complexes. Pharmacological PRMT5 inhibition induces DNA damage and subsequent apoptosis in CCA cell lines with inhibition of tumour growth as shown in in vivo models by Elurbide *et al*.⁴ Drug-induced PRMT5 inhibition is further enhanced in cancer cells carrying an MTAP deletion by modulating MTA levels. MTAP loss occurs in 25% of intrahepatic CCAs as shown by Elurbide *et al* in CCA tissues.⁴ MTA accumulation potently and selectively inhibits PRMT5, acting as 'double-edged sword' in the context of PRMT5 inhibition. CCA, cholangiocarcinoma; MEP50, methylosome protein 50; MTAP, methylthioadenosine phosphorylase; MTA, methyl-thioadenosine; PRMT5, protein arginine methyltransferase 5.</figcaption></figure>

PRMT5, have entered clinical trials.¹⁰
PRMT5 inhibitors have shown encour-
aging antitumour activity across a variety
of MTAP-deleted solid tumours including
CCA and gallbladder cancer,¹¹ but the full
potential of this therapy remains yet to
be established. Although these inhibitors
can cause GI side effects and cytopenias,
their adverse event profile is considered
manageable.² ¹⁰ If their full therapeutic
potential, either as monotherapy or in
combination with immune checkpoint
inhibitors, can be realised, they stand to
significantly improve patient outcomes.
The study by Elurbide et al⁴ offers compel-
ling preclinical evidence for pursuing
the evaluation of PRMT5 inhibitors in
patients with CCA, one of the most lethal
and challenging cancers to treat.

**Acknowledgements** TFB is supported by the European Union (ERC-AdG-2020-FIBCAN #101021417, HORIZON-HLTH-2021-DISEASE-04-07

D-SOLVE #10105791), the US National Institute of Health (R01CA233794), the University of Strasbourg Foundation, the Marigny Award of the Alsatian Cancer Ligue, French state funds managed within the 'Plan Investissements d'Avenir' and ANR (references ANR-10-IAHU-02 and ANR-10-LABX-0028), along with French state funds managed by the ANR within the France 2030 program (reference ANR-21-RHUS-0001). This work of the Interdisciplinary Thematic Institute IMCBio, as part of the ITI 2021-2028 program of the University of Strasbourg, CNRS and Inserm, was supported by IdEx Unistra (ANR-10-IDEX-0002), the SFRI-STRAT'US project (ANR 20-SFRI-0012) and by EUR IMCBio (ANR-17-EURE-0023) under the framework of the French Investments for the Future Program.

**Contributors** RD wrote the first draft of manuscript and assembled the figures. LG revised and edited the manuscript. TB led the project and participated to the writing and revisions.

**Funding** The authors have not declared a specific grant for this research from any funding agency in the public, commercial or not-for-profit sectors.

**Competing interests** TB is a founder, shareholder and advisor of Alentis Therapeutics developing Claudin-1-specific antibodies to treat fibrosis and cancer. TB is

also the inventor of a patent application on the use
of Claudin-1-specific antibodies to treat CCA, filed by
the University of Strasbourg, Inserm, the Strasbourg
University Hospitals and Alentis Therapeutics. TB is an
advisor for Pureos Bioventures and Novo Holding. LG is
an advisor/consultant for AbbVie, Agenus, AstraZeneca
(DSMB), Boehringer Ingelheim, Compass Therapeutics,
Exelixis, Kinnate Biopharma, Merck, Relay Therapeutics,
Servier, Surface Oncology, Taiho, TransThera Biosciences,
Tyra Biosciences and receives research funding (to
institution) from Alyssum Therapeutics, Boehringer
Ingelheim, Genentech and AstraZeneca.

**Patient consent for publication** Not applicable.

**Ethics approval** Not applicable.

**Provenance and peer review** Commissioned; internally peer reviewed.

© Author(s) (or their employer(s)) 2024. No commercial
re-use. See rights and permissions. Published by BMJ.

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**To cite** Désert R, Goyal L, Baumert TF. *Gut* Epub ahead of print: [*please include* Day Month Year]. doi:10.1136/gutjnl-2024-333632
Received 29 October 2024
Accepted 13 November 2024

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► http://dx.doi.org/10.1136/gutjnl-2024-332998

Gut 2024;0:1–3.

doi:10.1136/gutjnl-2024-333632

ORCID iD

Thomas F Baumert http://orcid.org/0000-0002-8864-2168

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