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As cancer immunotherapy has been emerged as a new pillar of cancer therapy, many strategies modulating host immunity were suggested for cancer treatment. Stimulator of interferon genes (STING) is a promising target for anticancer immunotherapy. However, dysregulated STING expression, or poor pharmacokinetic profiles of STING agonists pose major challenges. Recently, modulating target protein levels via the ubiquitin-proteasome system, such as proteolysis-targeting chimera (PROTAC), has broadened the scope of pharmacological inventions. Herein, we propose UPPRIS (upregulation of target proteins by protein-protein interaction strategy) to overcome these limitations. We discovered the small molecule SB24011 that inhibits STING-E3 ligase interaction, thereby induces the blockade of E3 ligase-mediated STING degradation. As a result, SB24011 enhanced the STING agonist-mediated immune responses by upregulating cellular STING protein level. Thus, co-administration of SB24011 markedly improved the immuno-oncological efficacy of STING agonist cGAMP and anti-PD-1 therapy for tumor regression and robust systemic antitumor response. Taken together, we successfully demonstrated that targeted STING protein upregulation is a promising strategy for cancer immunotherapy. Citation Format: Wansang Cho, Solchan Won, Yoona Choi, Jong Beom Park, Jun-Gyu Park, Sihyeong Yi, Caroline E. Kim, Chintam Narayana, Dong-Sup Lee, Seung Bum Park. Targeted protein upregulation strategy potentiates STING agonist immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2585.

Introduction: STimulator of INterferon Genes (STING) has been reported to be essential for antitumor immunity by inducing type I IFN expression, resulting in activation of both innate and adaptive immunity. Thus, pharmacological activation of STING pathway is actively being tested in many clinical studies. However, the clinical efficacy was not that clear, probably due to the limited expression of STING protein, which is tightly regulated by TRIM29 (E3 ligase)-mediated protein degradation pathway. Here, we present a novel small-molecule drug candidate with strong antitumor response in murine syngeneic model by upregulating cellular levels of STING through the inhibition of protein-protein interaction. Methods: The inhibitors of STING-TRIM29 interaction were identified using luciferase-complementation high-throughput screening system. Target protein of the inhibitor was identified in cellular thermostability shift assay. Their immunological activity was tested in Raw264.7 and A431 cells as well as CT26 murine syngeneic model. Results: The lead compound, SB24011, bound to STING and upregulated cellular levels of STING by inhibiting ubiquitin-mediated degradation. Together with STING agonist, SB24011 synergistically enhanced the inflammatory cytokine expression in cell lines as well as in CT26 syngeneic animal model. Intratumoral injection of SB24011 showed strong abscopal activity as single agent as well as synergistic antitumor responses with anti-PD1 antibody in CT26 syngeneic animal model. Conclusions: We identified novel small-molecule drug candidate, SB24011, which prevents the degradation of both murine and human STING proteins, resulting in enhanced immune responses together with STING agonist, cGAMP. SB24011 showed strong abscopal antitumor activity as single agent and synergistic antitumor activity as combination agent with other IO agents in CT26 syngeneic models. Citation Format: Seung Bum Park, Wansang Cho, Won Sol Chan, young il choi, Sungoh Ahn, Dong-Sup Lee. Inhibition of protein-protein interaction between STING and TRIM29 is a new approach to enhance anti-tumor immune response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 667.

Fluorogenic bioorthogonal probes are ideal for fluorescent imaging in live cell conditions. By taking advantage of the dual functionality of tetrazine (Tz), as a bioorthogonal reaction unit as well as a fluorescence quencher, a fluorophore–Tz conjugate (FLTz) has been utilized for fluorescent live cell imaging via inverse electron-demand Diels–Alder (iEDDA) type bioorthogonal reactions. However, most FLTz strategies rely on a donor–acceptor-type energy transfer mechanism, which limits red-shifting of probes’ emission wavelength without deterioration of the fluorescent turn-on/off ratio. To address this constraint, herein we present a monochromophoric design strategy for making a series of FLTzs spanning a broad range of emission colors. For the systematic comparison of design strategies with minimized structural differences, we selected indolizine-based emission-tunable Seoul-Fluor (SF) as a model fluorophore system. As a result, by inducing strong electronic coupling between Tz and π-conjugation systems ...

Selective bioactive compounds have emerged as major players in chemical biology for their potential in disrupting diverse biological pathways with minimal adverse effects. Using phenotypic screening, we identified an anti-cancer agent, SB2001, with a highly specific cytotoxicity toward HeLa human cervical cancer cells. The subsequent mechanistic study revealed that SB2001 induced apoptotic cell death through restoring p53 function and suppressed the human papillomavirus (HPV)-mediated oncoprotein signaling pathway via oxidative damage in HeLa cells. SB2001 also selectively induced HeLa-specific tumor regression without any adverse effects in an in vivo tumor xenograft model, demonstrating its potential as a promising chemical probe.

A series of air-stable boron complexes 1-5 were prepared by using N-aryl iminopyrrolide ligands. Designed as minimalist structural mimics of the privileged BODIPY motif, these new BOIMPY (BOron complexes of IMinoPYrrolide ligands) fluorophores feature low molecular symmetry that promotes emission from CT-type excited states with large Stokes shifts and little self-quenching. Through comparative studies on the homologous set of compounds 1-4, we have confirmed that a delicate interplay between conformational twisting and donor-acceptor interaction dictates the mechanism of de-excitation, which responds sensitively to solvent polarity as well as protonation states. Over a wide visible spectral range, the structure-dependent light-emitting properties of BOIMPY molecules are well manifested, even in the solid-state. In order to exploit the environment-sensitive nature of CT-type emission, the BOIMPY motif was elaborated further into a bioprobe molecule 5. Live-cell fluorescence imaging studies have established that 5 is localized exclusively at lipid droplets to produce well-resolved staining patterns without affecting cell viability. These findings promise future elaboration of BOIMPY-based functional molecules for applications in biological imaging, chemical sensing, and molecular switching.

Sepsis is one of the major causes of death worldwide when associated with multiple organ failure. However, there is a critical lack of adequate sepsis therapies because of its diverse patterns of pathogenesis. The pro-inflammatory cytokine cascade mediates sepsis pathogenesis, and high mobility group box proteins (HMGBs) play an important role as late-stage cytokines. We previously reported the small-molecule modulator, inflachromene (1d), which inhibits the release of HMGBs and, thereby, reduces the production of pro-inflammatory cytokines. In this context, we intraperitoneally administered 1d to a cecal ligation and puncture (CLP)-induced mouse model of sepsis and confirmed that it successfully ameliorated sepsis pathogenesis. On the basis of a structure-activity relationship study, we discovered new candidate compounds, 2j and 2l, with improved therapeutic efficacy in vivo. Therefore, our study clearly demonstrates that the regulation of HMGB1 release using small molecules is a promising strategy for the treatment of sepsis.

Although stage I melanoma is highly curable, the prognosis of late stage melanoma is poor and treatment options are limited to immune checkpoint modulators (ICMs). In this study, we synthesized a novel small molecule, SB1703, and identified high-mobility group box (HMGB) proteins as the direct binding target proteins. SB1703 inhibited melanoma growth in B16F10 syngeneic mouse model. To elucidate the tumor growth inhibitory mechanism, we determined the changes in myeloid-derived suppressor cell (MDSC) number during drug treatment. MDSCs expand in disease states such as cancer and have immune suppressive functions. Serum HMGB1 and interleukin (IL)-6 levels were increased in vehicle administered mice and decreased in SB1703 administered mice. We examined the possibility of combinatorial treatment with SB1703 and ICMs. SB1703 showed synergistic effect with ICMs. In conclusion, SB1703 exhibited therapeutic effects against melanoma in both single and combination treatments with ICMs, which might be due to modulation of myeloid cells such as MDSCs Citation Format: Hyo Young Kim, Hee-Jung Kim, Jae Hoon Jung, So Young Jo, Ju Hee Kim, Wansang Cho, Mingi Kim, Junhyeong Yim, Seung Bum Park. SB1703 has therapeutic effect on syngeneic mouse model as an immune modulator in TME [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2816.

Diversity-oriented synthesis (DOS) can provide a collection of diverse and complex drug-like small molecules, which is critical in the development of new chemical probes for biological research of undruggable targets. However, the design and synthesis of small-molecule libraries with improved biological relevance as well as maximized molecular diversity represent a key challenge. Herein, we employ functional group-pairing strategy for the DOS of a chemical library containing privileged substructures, pyrimidodiazepine or pyrimidine moieties, as chemical navigators towards unexplored bioactive chemical space. To validate the utility of this DOS library, we identify a new small-molecule inhibitor of leucyl-tRNA synthetase–RagD protein–protein interaction, which regulates the amino acid-dependent activation of mechanistic target of rapamycin complex 1 signalling pathway. This work highlights that privileged substructure-based DOS strategy can be a powerful research tool for the construction of drug-like compounds to address challenging biological targets., Diversity-oriented synthesis is useful for generating complex molecular structures occupying diverse molecular space. Here the authors report a strategy to access libraries of privileged heterocyclic structures, and furthermore identify an inhibitor of LRS–RagD protein–protein interaction.

Neuroinflammation is a key process for many neurodegenerative diseases. Activated microglia and astrocytes play an essential role in neuroinflammation by producing nitric oxide (NO), inflammatory cytokines, chemokines, and neurotoxins. Therefore, targeting glia-mediated neuroinflammation using small-molecules is a potential therapeutic strategy. In this study, we performed a phenotypic screen using microglia cell-based assay to identify a hit compound containing N-carbamoylated urethane moiety (SNU-BP), which inhibits lipopolysaccharide (LPS)-induced NO production in microglia. SNU-BP inhibited pro-inflammatory cytokines and inducible nitric oxide synthase in LPS-stimulated microglia, and potentiated interleukin-4-induced arginase-1 expression. PPAR-γ was identified as a molecular target of SNU-BP. The PPAR response element reporter assay revealed that SNU-BP specifically activated PPAR-γ, but not PPAR-δ or -α, confirming that PPAR-γ is the target protein of SNU-BP. The anti-inflammatory effect of SNU-BP was attenuated by genetic and pharmacological inhibition of PPAR-γ. In addition, SNU-BP induced an anti-inflammatory phenotype in astrocytes as well, by inhibiting pro-inflammatory NO and TNF-α, while increasing anti-inflammatory genes, such as arginase-1 and Ym-1. Finally, SNU-BP exhibited an anti-inflammatory effect in the LPS-injected mouse brain, demonstrating a protective potential for neuroinflammatory diseases.