PARP14 has been implicated by genetic knockout studies to promote protumor macrophage polarization and suppress the antitumor inflammatory response due to its role in modulating interleukin-4 (IL-4) ...and interferon-γ signaling pathways. Here, we describe structure-based design efforts leading to the discovery of a potent and highly selective PARP14 chemical probe. RBN012759 inhibits PARP14 with a biochemical half-maximal inhibitory concentration of 0.003 μM, exhibits >300-fold selectivity over all PARP family members, and its profile enables further study of PARP14 biology and disease association both in vitro and in vivo. Inhibition of PARP14 with RBN012759 reverses IL-4-driven protumor gene expression in macrophages and induces an inflammatory mRNA signature similar to that induced by immune checkpoint inhibitor therapy in primary human tumor explants. These data support an immune suppressive role of PARP14 in tumors and suggest potential utility of PARP14 inhibitors in the treatment of cancer.
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•The first highly potent and selective inhibitor of PARP14, RBN012759, is described•The profile of RBN012759 enables its use as an in vitro and in vivo chemical probe•RBN012759 reversed IL-4-driven protumor gene expression•An inflammatory gene signature was activated with RBN012759 treatment
Schenkel et al. disclose the first highly potent and selective inhibitor of PARP14, RBN012759, an in vitro and in vivo chemical probe. RBN012759 reverses IL-4-driven protumor gene expression in macrophages and induces an inflammatory mRNA signature in primary human tumor explants, suggesting utility of PARP14 inhibitors for treating cancer.
Placental abnormalities have been sporadically implicated as a source of developmental heart defects. Yet it remains unknown how often the placenta is at the root of congenital heart defects (CHDs), ...and what the cellular mechanisms are that underpin this connection. Here, we selected three mouse mutant lines, Atp11a, Smg9 and Ssr2, that presented with placental and heart defects in a recent phenotyping screen, resulting in embryonic lethality. To dissect phenotype causality, we generated embryo- and trophoblast-specific conditional knockouts for each of these lines. This was facilitated by the establishment of a new transgenic mouse, Sox2-Flp, that enables the efficient generation of trophoblast-specific conditional knockouts. We demonstrate a strictly trophoblast-driven cause of the CHD and embryonic lethality in one of the three lines (Atp11a) and a significant contribution of the placenta to the embryonic phenotypes in another line (Smg9). Importantly, our data reveal defects in the maternal blood-facing syncytiotrophoblast layer as a shared pathology in placentally induced CHD models. This study highlights the placenta as a significant source of developmental heart disorders, insights that will transform our understanding of the vast number of unexplained congenital heart defects.
PARP14 is an interferon‐stimulated gene that is overexpressed in multiple tumor types, influencing pro‐tumor macrophage polarization as well as suppressing the antitumor inflammation response by ...modulating IFN‐γ and IL‐4 signaling. PARP14 is a 203 kDa protein that possesses a catalytic domain responsible for the transfer of mono‐ADP‐ribose to its substrates. PARP14 also contains three macrodomains and a WWE domain which are binding modules for mono‐ADP‐ribose and poly‐ADP‐ribose, respectively, in addition to two RNA recognition motifs. Catalytic inhibitors of PARP14 have been shown to reverse IL‐4 driven pro‐tumor gene expression in macrophages, however it is not clear what roles the non‐enzymatic biomolecular recognition motifs play in PARP14‐driven immunology and inflammation. To further understand this, we have discovered a heterobifunctional small molecule designed based on a catalytic inhibitor of PARP14 that binds in the enzyme's NAD+‐binding site and recruits cereblon to ubiquitinate it and selectively target it for degradation.
A tool to dissect PARP14: A heterobifunctional small molecule that selectively degrades PARP14 is described. It can be used in studies to unravel the biological function of this large protein's mono‐ADP‐ribosyltransferase catalytic domain and its macrodomains, WWE domain and RNA recognition motifs in immunity and inflammation.
PARP7 is a monoPARP that catalyzes the transfer of single units of ADP-ribose onto substrates to change their function. Here, we identify PARP7 as a negative regulator of nucleic acid sensing in ...tumor cells. Inhibition of PARP7 restores type I interferon (IFN) signaling responses to nucleic acids in tumor models. Restored signaling can directly inhibit cell proliferation and activate the immune system, both of which contribute to tumor regression. Oral dosing of the PARP7 small-molecule inhibitor, RBN-2397, results in complete tumor regression in a lung cancer xenograft and induces tumor-specific adaptive immune memory in an immunocompetent mouse cancer model, dependent on inducing type I IFN signaling in tumor cells. PARP7 is a therapeutic target whose inhibition induces both cancer cell-autonomous and immune stimulatory effects via enhanced IFN signaling. These data support the targeting of a monoPARP in cancer and introduce a potent and selective PARP7 inhibitor to enter clinical development.
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•PARP7 acts as a brake in cytosolic nucleic acid sensing in a TBK1-dependent manner•RBN-2397 is a potent and selective PARP7 inhibitor targeting this cancer vulnerability•RBN-2397 restores the type I interferon (IFN) response in tumor cells•RBN-2397 induces antitumor immunity dependent on type I IFN and CD8 T cells
Engaging cytosolic nucleic acid sensing and the type I interferon (IFN) response is an emerging therapeutic strategy in oncology. Gozgit et al. find that PARP7 is a cancer-specific brake on this pathway and that inhibition of PARP7 by a selective inhibitor, RBN-2397, induces both cancer cell-autonomous effects and antitumor immunity via enhanced IFN signaling.
Mutations in RNA-binding proteins can lead to pleiotropic phenotypes including craniofacial, skeletal, limb, and neurological symptoms. Heterogeneous nuclear ribonucleoproteins (hnRNPs) are involved ...in nucleic acid binding, transcription, and splicing through direct binding to DNA and RNA, or through interaction with other proteins in the spliceosome. We show a developmental role for Hnrnpul1 in zebrafish, resulting in reduced body and fin growth and missing bones. Defects in craniofacial tendon growth and adult-onset caudal scoliosis are also seen. We demonstrate a role for Hnrnpul1 in alternative splicing and transcriptional regulation using RNA-sequencing, particularly of genes involved in translation, ubiquitination, and DNA damage. Given its cross-species conservation and role in splicing, it would not be surprising if it had a role in human development. Whole-exome sequencing detected a homozygous frameshift variant in HNRNPUL1 in 2 siblings with congenital limb malformations, which is a candidate gene for their limb malformations. Zebrafish Hnrnpul1 mutants suggest an important developmental role of hnRNPUL1 and provide motivation for exploring the potential conservation of ancient regulatory circuits involving hnRNPUL1 in human development.
The EZH2 small-molecule inhibitor tazemetostat (EPZ-6438) is currently being evaluated in phase II clinical trials for the treatment of non-Hodgkin lymphoma (NHL). We have previously shown that EZH2 ...inhibitors display an antiproliferative effect in multiple preclinical models of NHL, and that models bearing gain-of-function mutations in
were consistently more sensitive to EZH2 inhibition than lymphomas with wild-type (WT)
Here, we demonstrate that cell lines bearing
mutations show a cytotoxic response, while cell lines with WT-
show a cytostatic response and only tumor growth inhibition without regression in a xenograft model. Previous work has demonstrated that cotreatment with tazemetostat and glucocorticoid receptor agonists lead to a synergistic antiproliferative effect in both mutant and wild-type backgrounds, which may provide clues to the mechanism of action of EZH2 inhibition in WT-
models. Multiple agents that inhibit the B-cell receptor pathway (e.g., ibrutinib) were found to have synergistic benefit when combined with tazemetostat in both mutant and WT-
backgrounds of diffuse large B-cell lymphomas (DLBCL). The relationship between B-cell activation and EZH2 inhibition is consistent with the proposed role of EZH2 in B-cell maturation. To further support this, we observe that cell lines treated with tazemetostat show an increase in the B-cell maturation regulator,
/BLIMP1, and gene signatures corresponding to more advanced stages of maturation. These findings suggest that EZH2 inhibition in both mutant and wild-type backgrounds leads to increased B-cell maturation and a greater dependence on B-cell activation signaling.
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Skeletal muscle stem cells (MuSC) are crucial for tissue homoeostasis and repair after injury. Following activation, they proliferate to generate differentiating myoblasts. A proportion of cells ...self-renew, re-enter the MuSC niche under the basal lamina outside the myofiber and become quiescent. Quiescent MuSC have a primary cilium, which is disassembled upon cell cycle entry. Ex vivo experiments suggest cilia are important for MuSC self-renewal, however, their requirement for muscle regeneration in vivo remains poorly understood. Talpid3 (TA
) is essential for primary cilia formation and Hedgehog (Hh) signalling. Here we use tamoxifen-inducible conditional deletion of TA
in MuSC (iSC-KO) and show that regeneration is impaired in response to cytotoxic injury. Depletion of MuSC after regeneration suggests impaired self-renewal, also consistent with an exacerbated phenotype in TA
mice after repeat injury. Single cell transcriptomics of MuSC progeny isolated from myofibers identifies components of several signalling pathways, which are deregulated in absence of TA
, including Hh and Wnt. Pharmacological activation of Wnt restores muscle regeneration, while purmorphamine, an activator of the Smoothened (Smo) co-receptor in the Hh pathway, has no effect. Together, our data show that TA
and primary cilia are important for MuSC self-renewal and pharmacological treatment can efficiently restore muscle regeneration.
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