Endoplasmic Reticulum Targeting Research Articles

Endoplasmic Reticulum-Targeted Polymer-Manganese Nanocomplexes for Tumor Immunotherapy.

Manganese ions (Mn2+) are an immune activator that enhances the activation of both cGAS and STING proteins. The STING signaling activation and subsequential immune responses are predominantly associated with endoplasmic reticulum (ER). Therefore, ER targeting of Mn2+ in the subcellular compartments would promote the activation of STING signaling pathways. Herein, we report the design of ER-targeted manganese-based nanocomplexes (NCs) by complexation of Mn2+ with a zwitterionic polymer, poly[2-(N-oxide-N,N-dimethylamino) ethyl methacrylate] (OPDMA). The Mn/OPDMA nanocomplexes (Mn/OPDMA NCs) keep a long blood circulation for tumor accumulation and trigger adsorption-mediated transcytosis for extravasation and deep tumor penetration. Notably, in the tumor-associated macrophages, the Mn/OPDMA NCs can preferentially translocate to their ERs, significantly enhancing cGAS-STING pathway activation for tumor-associated macrophage polarization and IFN-β secretion. In mouse colon and hepatocellular cancer models, the intravenously administrated Mn/OPDMA NCs efficiently remodel tumor immune microenvironment, greatly retard tumor growths by 2.4- to 5-fold, and prolong the mouse survivals compared to free Mn2+-treated mice. This study provides the ER-targeted delivery of Mn2+ that achieves robust STING activation and, thus, potent systemic tumor inhibition without the toxicity of free Mn2+.

Photo-Facilitated Nitric Oxide-Triggered Turn-on Photodynamic Therapy for Precise Antitumor Application.

Photodynamic therapy (PDT) is a powerful strategy for tumor therapy with noninvasiveness and desirable efficacy. However, the phototoxicity of photosensitizer after the post-PDT is the major obstacle limiting the clinic applications. Herein, a nitric oxide (NO)-activatable photosensitizer is reported with turn-on PDT behavior and endoplasmic reticulum (ER) targeting ability for precise tumor therapy. Four o-thiophenediamine derivatives with reaction-tunable donor/acceptor push-pull electronic effect are established, and the systematic structure and property relationship observation reveals the following features: 1) the reactivity against NO can be improved by enhancing the electron density and further facilitated upon photo-irradiation. 2) the reactivity with NO enables the improved intramolecular charge transfer process with the evoking of photosensitizing effect. 3) only o-thiophenediamine derivative with ER enrichment behavior exhibited cancer cell ablation effect compared to photosensitizers localized in lysosome and lipid droplet. Thus, the efficient inhibition of cancer cells both in vitro and in vivo is realized based on the photo-controlled PDT strategy. This work provides more insights into developing promising activatable photosensitizers for advanced therapy based on tumor microenvironment trigger.

Tröger's Base as a Potential Bridge to Type-I Photosensitizers: Mechanism and Antitumor Applications.

In contrast to Type-II photodynamic therapy (PDT), Type-I PDT with less oxygen consumption has shown great potential against tumor hypoxia. However, there are limited strategies available for designing Type-I photosensitizers (PSs). Herein, we present a promising strategy for synthesizing Type-I PSs (TBC-1-TBC-4) using Tröger's base (TB) framework. The TB framework can promote intersystem crossing efficiency and create an electron-rich environment, making it the most likely site for electron transfer to O2 to generate Type-I ROS. As anticipated, TBC-1-TBC-4 demonstrates Type-I ROS generation capability and their impressive visible light-harvesting ability significantly enhances this capability. Among them, TBC-1 demonstrates outstanding biocompatibility and PDT efficiency in vitro under both normoxia and hypoxia. Furthermore, TBC-1 effectively inhibits tumor growth in vivo, with negligible side effects. This is attributed to TBC-1's efficient generation of Type-I ROS and endoplasmic reticulum targeting ability. This study thus offers useful insights into developing Type-I PSs.

Endoplasmic reticulum targeting photodynamic oxidizer to boost anti-tumor immunity by intensifying immunogenic cell death in conjunction with IDO1 inhibition

Endoplasmic reticulum targeting photodynamic oxidizer to boost anti-tumor immunity by intensifying immunogenic cell death in conjunction with IDO1 inhibition

Theranostic Rhenium(I)-Based ER-Phagy Retardant Promotes Immunogenic Cell Death.

ER-phagy is a double-edged sword in the occurrence, development, and treatment of cancer; especially, its functions in immunotherapy are still unknown. In this work, we designed a theranostic Re complex (Re1) containing a BODIPY-derived ligand and a β-carboline ligand to target the endoplasmic reticulum (ER) and block ER-phagy at the late stages. Interestingly, as validated both in vitro and in vivo, ER-phagy blockage greatly enhances the capability of Re1 to induce immunogenic cell death (ICD). In summary, we dexterously fused two molecular modules for ER targeting and ER-phagy blockage into a coordination complex to afford a highly effective ICD inducer, which provides clues for designing new cancer immunotherapeutics.

NIR-715 photodynamic therapy induces immunogenic cancer cell death by enhancing the endoplasmic reticulum stress response

Effectively interfering with endoplasmic reticulum (ER) function in tumor cells and simultaneously activating an anti-tumor immune microenvironment to attack the tumor cells are promising strategies for cancer treatment. However, precise ER-stress induction is still a huge challenge. In this study, we synthesized a near-infrared (NIR) probe, NIR-715, which induces tumor cell death and inhibits tumor growth without causing apparent side effects. NIR-715 triggers severe ER stress and immunogenic cell death (ICD) after visible light exposure. NIR-715 induced ICD-associated HMGB1 release in vitro and anti-tumor immune responses, including increased cytotoxic T lymphocyte (GZMB+ CD8+ T cell) infiltration and decreased numbers of exhausted T lymphocytes (PD-L1+ CD8+ T cell). These findings suggest that NIR-715 may be a novel agent for “cold” tumor photodynamic therapy (PDT).Schematic illustration of NIR-715 photodynamic therapy for visible light-triggered, endoplasmic reticulum-targeting antitumor therapy.

Research Progress of Endoplasmic Reticulum Targeting Metal Complexes in Cancer Therapy.

The development of anticancer drugs that target different organelles has received extensive attention due to the characteristics of cancer recurrence, metastasis, and drug resistance. The endoplasmic reticulum (ER) is an important structure within the cell that is primarily responsible for protein synthesis, folding, modification, and transport and plays a crucial role in cell function and health. ER stress activation induces cancer cell apoptosis. New anticancer drugs with different anticancer mechanisms and selectivity can be designed because of redox activity, composition diversity, and metal complexes structure regulation. Over the past few decades, dozens of metal complexes have killed cancer cells through ER stress, showing powerful tumor-suppressive effects. This review summarizes the progress of research on anticancer metallic drugs that induce ER stress over the past few years, which is expected to bring more breakthroughs in the field of medicine and life science.

Near-Infrared-II-Activated Transition Metal(II)-Coordinated Ligand Radical Primes Robust Anticancer Immunity.

Photoactivatable metallodrugs combining tumor cell eradication and immune stimulation hold immense promise for targeted cancer therapy. However, limitations such as oxygen dependence, narrow visible light responsiveness, and poor immunogenicity hinder their efficacy in deep solid tumors with hypoxic and immunosuppressive microenvironments. Herein, we present a novel design strategy for transition metal(II)-coordinated ligand radicals exhibiting intense near-infrared-II (NIR-II) absorption, unique endoplasmic reticulum-targeting capability, and oxygen-independent photothermal performance, effectively addressing these constraints. Proof-of-concept results demonstrate the potent efficacy of our cobalt(II)-coordinated ligand radical (BPDP-Co) in inducing highly immunogenic pyroptosis in tumor cells under both normoxic and severe hypoxic conditions upon 1064 nm laser irradiation. This NIR-II activation triggers the release of damage-associated molecular patterns (DAMPs) and proinflammatory cytokines, fueling a robust antitumor immune response. In vivo studies demonstrate that treatment with BPDP-Co/NIR-II significantly inhibited 4T1 tumor growth in BALB/c mice with a high inhibitory rate of 85.7%, highlighting its therapeutic potential in tumor immunotherapy.

Monitoring Glutathione Content of the Endoplasmic Reticulum under Scrap Leather-Induced Endoplasmic Reticulum Stress via an Endoplasmic Reticulum-Targeted Two-Photon Fluorescent Probe.

Maintaining tissue homeostasis necessitates the coordinated efforts of various cell types to regulate inflammation. Endoplasmic reticulum (ER) stress, a hallmark of inflammation, exacerbates tissue pathology in various human diseases. Glutathione (GSH), a pivotal regulator of cellular redox balance, controls disulfide bond formation in the ER, thereby shielding cells from oxidative stress. In this study, we developed a two-photon fluorescent probe, ER-GSH, with specific ER targeting and demonstrated its high sensitivity and rapid response to GSH. Experiments conducted on BV2 cells and a mice model of neuroinflammation induced by scrap leather revealed that inflammatory reactions led to ER stress and a substantial reduction in GSH levels. Notably, the anti-inflammatory drug NS-398 effectively inhibited cell inflammation and ER stress by maintaining GSH levels. These findings underscore the potential therapeutic significance of modulating GSH levels to alleviate the impact of neuroinflammation.

Coumarin-Quinazolinone based photosensitizers: Mitochondria and endoplasmic reticulum targeting for enhanced phototherapy via different cell death pathways

Organelle-targeted photosensitizers (PSs) offer valuable tools for improving photodynamic therapy (PDT), yet systematic studies on how different organelles influence phototherapeutic outcomes are limited. In particular, the connection between organelle targeting and various modes of programmed cell death remains unclear. In this study, we developed a series of PSs using the Coumarin-Quinazolinone (CQ) scaffold, each designed to target different organelles, including the mitochondria, endoplasmic reticulum (ER), lysosome, and nucleolus. Our results show that their PDT performance is highly dependent on their localization, with phototoxic index (PI) ranging from 2 to 245. Notably, the mitochondria-targeted CQ-Mito and ER-targeted CQ-ER exhibited profound phototherapeutic performances, with PI of 167 and 245 respectively. Our further study reveals that CQ-Mito causes cell death by both apoptosis and ferroptosis, while CQ-ER primarily triggers ferroptosis. This study not only provides new agents for PDT but also offers insights into how organelle targeting influences cell death mechanisms, which can shed light on the design of PSs for controlled cell death.

Exploration of glyoxals’ level changes in endoplasmic reticulum during ferroptosis by a photocaged fluorescent probe

Both ferroptosis and elevated glyoxals (e.g. methylglyoxal and glyoxal), are associated with endoplasmic reticulum stress, but their correlations, in particular, how level of glyoxals changes in endoplasmic reticulum during ferroptosis remains unclear. In this work, a new photoactivatable fluorescent probe Photo-ER-GOS was developed to study level changes of glyoxals in endoplasmic reticulum during ferroptosis via in situ photodecaging and fluorescence detection, avoiding potential interference from glyoxals in cytosol. The probe contains a photolabile nitrobenzyloxymethyl group functioned both as the reactivity blocking group and the fluorescence quenching group, which linked to the guanidino group of NAP-DCP-3, the active endoplasmic reticulum-targeting fluorescent probe for glyoxals previously developed in our lab. It was found that the level of glyoxals in endoplasmic reticulum increases significantly during erastin-induced ferroptosis in RAW 264.7 cells via comparison of relative fluorescence intensity increases. The endoplasmic reticulum glyoxal level increase was also confirmed by the Ultra Performance Liquid Chromatography-Mass Spectrum studies. In contrast, no significant change of level of glyoxals in endoplasmic reticulum was found in staurosporine-induced apoptosis, suggesting that upregulated glyoxals in endoplasmic reticulum may be a previously overlooked characteristic of ferroptosis. Erastin-induced level increase of glyoxals was also found in zebrafish. The caged probe Photo-ER-GOS thus provides a useful chemical tool to study GOS levels changes in endoplasmic reticulum.

Strategically engineered Au(I) complexes for orchestrated tumor eradication via chemo-phototherapy and induced immunogenic cell death

Cancer is a significant cause of death around the world, and for many varieties, treatment is not successful. Therefore, there is a need for the development of innovative, efficacious, and precisely targeted treatments. Here, we develop a series of Au(I) complexes (1-4) through rational manipulation of ligand structures, thereby achieving tumor cell specific targeting and orchestrated tumor eradication via chemo-phototherapy and induced immunogenic cell death. A comprehensive exploration based on in vitro and in vivo female mice experimentation shows that complex 4 exhibits proficiency in specific tumor imaging, endoplasmic reticulum targeting, and has robust therapeutic capabilities. Mechanistic elucidation indicates that the anticancer effect derives from the synergistic actions of thioredoxin reductase inhibition, highly efficient reactive oxygen species production and immunogenic cell death. This work presents a report on a robust Au(I) complex integrating three therapeutic modalities within a singular system. The strategy presented in this work provides a valuable reference for the development of high-performance therapeutic agents.

Extremely Sensitive Genetically Encoded Temperature Indicator Enabling Measurement at the Organelle Level.

Intracellular temperature is a fundamental parameter in biochemical reactions. Genetically encoded fluorescent temperature indicators (GETIs) have been developed to visualize intracellular thermogenesis; however, the temperature sensitivity or localization capability in specific organelles should have been further improved to clearly capture when and where intracellular temperature changes at the subcellular level occur. Here, we developed a new GETI, gMELT, composed of donor and acceptor subunits, in which cyan and yellow fluorescent proteins, respectively, as a Förster resonance energy transfer (FRET) pair were fused with temperature-sensitive domains. The donor and acceptor subunits associated and dissociated in response to temperature changes, altering the FRET efficiency. Consequently, gMELT functioned as a fluorescence ratiometric indicator. Untagged gMELT was expressed in the cytoplasm, whereas versions fused with specific localization signals were targeted to the endoplasmic reticulum (ER) or mitochondria. All gMELT variations enabled more sensitive temperature measurements in cellular compartments than those in previous GETIs. The gMELTs, tagged with ER or mitochondrial targeting sequences, were used to detect thermogenesis in organelles stimulated chemically, a method previously known to induce thermogenesis. The observed temperature changes were comparable to previous reports, assuming that the fluorescence readout changes were exclusively due to temperature variations. Furthermore, we demonstrated how macromolecular crowding influences gMELT fluorescence given that this factor can subtly affect the fluorescence readout. Investigating thermogenesis with gMELT, accounting for factors such as macromolecular crowding, will enhance our understanding of intracellular thermogenesis phenomena.

A novel theranostic strategy for myocardial infarction through neutralization of endogenous SO2 using an endoplasmic reticulum-targeted fluorescent probe

Myocardial infarction (MI), one of the leading causes of death worldwide, urgently needs further understanding of the pathological process and effective therapies. SO2 in endoplasmic reticulum in several cardiovascular diseases has been reported to be particularly important. However, the role of endogenous SO2 in endoplasmic reticulum in treating myocardial infarction is still ambiguous and needs to be elucidated. Herein, we developed TPA-HI-SO2 as the first endoplasmic reticulum-targeting fluorescent agent for specific imaging and detection of sulfur dioxide derivatives both in vitro and in vivo. TPA-HI-SO2 shows a highly sensitive and selective response to SO2 derivatives over other anions in aqueous solution with a satisfactory response time and detection limit. Furthermore, TPA-HI-SO2 decreased the SO2 concentration in H9C2 cells treated with H2O2 and in an MI mouse model. Most importantly, TPA-HI-SO2 protects H9C2 cells from H2O2-induced apoptosis and obviously protects against myocardial infarction in vivo through neutralization of endogenous SO2. Taken together, we developed the first ER-targeting ratiometric fluorescent probe for endogenous SO2 with excellent biocompatibility, high selectivity and sensitivity in this paper. More importantly, we demonstrated an obvious increase of the endogenous SO2 concentration in a myocardial infarction mouse model for the first time, which suggests that neutralization of endogenous SO2 in endoplasmic reticulum could be a promising therapeutic strategy for myocardial infarction.

Ru(II)-Photoactive Agents for Targeting ER Stress and Immunogenic Cell Death.

Immunotherapy has emerged as a promising avenue for cancer treatment by bolstering the immune system's ability to recognize and attack cancer cells. Photodynamic therapy shows potential in enhancing antitumor immunity, though the mechanisms behind its success are not fully understood. In this manuscript, we investigate two previously reported green light activated PCT/PDT agents where compound 2 - [Ru(tpy)(Me2bpy)( 3 )] 2+ , (tpy = 2,2':6',2''- terpyridine, Me2bpy = 6,6'-dimethyl-2,2'-bipyridine, 3 = pyridyl-BODIPY-I2,) - shows remarkable photoselectivity in assays containing both 2D cancer cells and 3D cocultures containing BALB/c macrophages and 4T1 murine breast cancer cells. Through flow cytometry and protein analysis, we found complex 2 displays superior evidence of induced endoplasmic reticulum (ER) stress markers and indicators of immunogenic cell death (ICD) compared to its ligand 3 , despite its weaker photoselectivity. Most importantly, these results were supported by in vivo studies where 2 produced anti-tumor immunity against the 4T1 tumor model in BALB/c mice. Complete tumor elimination was achieved in 2/8 mice, and these mice were both protected against a subsequent contralateral rechallenge and showed increased ex vivo peripheral tumor antigen-specific recall, suggesting memory T cells are induced by 2 . Signatures of M1 macrophage polarization were also evident in tumor tissue from the remaining 6/8 mice treated with 2 compared to untreated tumors. These findings demonstrate Ru(II) complexation plays a critical role in ER targeting which triggers ICD, highlighting the potential of Ru(II) agents as future in situ tumor vaccines.

In Situ Visualizing Carboxylesterase Activity in Type 2 Diabetes Mellitus Using an Activatable Endoplasmic Reticulum Targetable Proximity Labeling Far-Red Fluorescent Probe.

Carboxylesterase (CE), an enzyme widely present in organisms, is involved in various physiological and pathological processes. Changes in the levels of CEs in the liver may predict the presence of type 2 diabetes mellitus (T2DM). Here, a novel dicyanoisophorone (DCI)-based proximity-labeled far-red fluorescent probe DCI2F-Ac with endoplasmic reticulum targeting was proposed for real-time monitoring and imaging of the CEs activity. DCI2F-Ac featured very low cytotoxicity and biotoxicity and was highly selective and sensitive for CEs. Compared with traditional CEs probes, DCI2F-Ac was covalently anchored directly to CEs, thus effectively reducing the loss of in situ fluorescent signals due to diffusion. Through the "on-off" fluorescence signal readout, DCI2F-Ac was able to distinguish cell lines and screen for CEs inhibitors. In terms of endoplasmic reticulum (ER) stress, it was found that thapsigargin (Tg) induced upregulation of CEs levels but not tunicamycin (Tm), which was related to the calcium homeostasis of the ER. DCI2F-Ac could efficiently detect downregulated CEs in the livers of T2DM, and the therapeutic efficacy of metformin, acarbose, and a combination of these two drugs was assessed by tracking the fluctuation of CEs levels. The results showed that combining metformin and acarbose could restore CEs levels to near-normal levels with the best antidiabetic effect. Thus, the DCI2F-Ac probe provides a great opportunity to explore the untapped potential of CEs in liver metabolic disorders and drug efficacy assessment.

Nanoparticle-mediated celastrol ER targeting delivery amplify immunogenic cell death in melanoma

IntroductionChemoimmunotherapy, which benefits from the combination of chemotherapy and immunotherapy, has emerged as a promising strategy in cancer treatment. However, effectively inducing a robust immune response remains challenging due to the limited responsiveness across patients. Endoplasmic reticulum (ER) stress is essential for activating intracellular signaling pathways associated with immunogenic cell death (ICD), targeting drugs to ER might enhance ER stress and improve ICD-related immunotherapy. ObjectivesTo improve the immune response of Chemoimmunotherapy. MethodsER targeting nanoparticles TSE-CEL/NP were constructed to enhance immunogenic cancer cell death. Flow cytometry, confocal microscope, TEM and immunofluorescence were used to evaluate the ER targeting effect and immunogenic tumor cell death in vitro on B16F10 tumor cells. Unilateral and bilateral tumor models were constructed to investigate the efficacy of anti-tumor and immunotherapy in vivo. Lung metastasis B16F10 melanoma tumor-bearing mice were used to assess the anti-metastasis efficacy. ResultsTSE-CEL/NP could specially accumulate in ER, thereby induce ER stress. High ER stress trigger the exposure of CRT, the extracellular release of HMGB1 and ATP. These danger signals subsequently promote the recruitment and maturation of dendritic cells (DCs), which in turn increase the proliferation of cytotoxic T lymphocytes (CD8+ T cells), ultimately resulted in an improved immunotherapy efficacy against melanoma. Invivo experiments showed that TSE-CEL/NP exhibits excellent antitumor efficacy and triggers a strong immune response. ConclusionOur findings demonstrated that celastrol ER targeting delivery could amplify immunogenic cell death in melanoma, which provide experimental basis for melanoma immunotherapy.

Rational construction of a novel fluorescent probe for imaging peroxynitrite in the endoplasmic reticulum during drug-induced liver injury

Drug-induced liver injury (DILI) not only poses a serious health threat to patients, but is also a major obstacle to the development of new drugs. The liver toxicity of many drugs is not detected until late in the development process or even after marketing, which significantly increases the cost of new drug development. This may be limited by the lack of tools to detect DILI. Studies have shown that abnormal changes in peroxynitrite (ONOO-) levels in the endoplasmic reticulum of liver cells may be an early manifestation of DILI. Therefore, the development of fluorescent probes to detect ONOO- in the endoplasmic reticulum may provide assistance in the detection of DILI and the screening of new drugs. Herein, we report a fluorescent probe, ER-N, capable of detecting ONOO- in the endoplasmic reticulum with high selectivity and sensitivity. The probe selects di(trifluoromethyl)benzene as the targeting group of the endoplasmic reticulum, which possesses excellent targeting to the endoplasmic reticulum, and provides a new design method for the establishment of other endoplasmic reticulum-targeting probes. In addition, the probe ER-N successfully realized the detection of endogenous and exogenous ONOO- in cells and zebrafish. More importantly, the probe ER-N was able to be used for tracing changes in the levels of ONOO- in the endoplasmic reticulum of cells and tissues under drug stimulation. Together, these results suggest that the probe ER-N has the potential to be a tool for DILI detection and new drug screening.

Mutations in the non-catalytic polyproline motif destabilize TREX1 and amplify cGAS-STING signaling.

The cGAS-STING pathway detects cytosolic DNA and activates a signaling cascade that results in a type I interferon (IFN) response. The endoplasmic reticulum (ER)-associated exonuclease TREX1 suppresses cGAS-STING by eliminating DNA from the cytosol. Mutations that compromise TREX1 function are linked to autoinflammatory disorders, including systemic lupus erythematosus (SLE) and Aicardi-Goutières syndrome (AGS). Despite key roles in regulating cGAS-STING and suppressing excessive inflammation, the impact of many disease-associated TREX1 mutations-particularly those outside of the core catalytic domains-remains poorly understood. Here, we characterize a recessive AGS-linked TREX1 P61Q mutation occurring within the poorly characterized polyproline helix (PPII) motif. In keeping with its position outside of the catalytic core or ER targeting motifs, neither the P61Q mutation, nor aggregate proline-to-alanine PPII mutation, disrupts TREX1 exonuclease activity, subcellular localization, or cGAS-STING regulation in overexpression systems. Introducing targeted mutations into the endogenous TREX1 locus revealed that PPII mutations destabilize the protein, resulting in impaired exonuclease activity and unrestrained cGAS-STING activation. Overall, these results demonstrate that TREX1 PPII mutations, including P61Q, impair proper immune regulation and lead to autoimmune disease through TREX1 destabilization.

Biomaterials for Targeting Endoplasmic Reticulum in Cancer.

Endoplasmic reticulum (ER) is one of the most important sub-cellular organelles which controls myriads of biological functions including protein biosynthesis with proper functional folded form, protein misfolding, protein transport into Golgi body for secretion, Ca2+ homeostasis and so on. Subsequently, dysregulation in ER function leads to ER stress followed by disease pathology like cancer. Hence, targeting ER in the cancer cells emerged as one of the futuristic strategies for cancer treatment. However, the major challenge is to selectively and specifically target ER in the sub-cellular milieu in the cancer tissues, due to the lack of ER targeting chemical moieties to recognize the ER markers. To address this, in the last decade, numerous biomaterials were explored to selectively impair and image ER in cancer cells to induce ER stress. This review outlines those biomaterials which consists of carbon and silicon materials, lipid nanoparticles (liposomes and micelles), supramolecular self-assembled nanostructures, cell membrane-coated nanoparticles and metallic nanoparticles. Moreover, we also discuss the challenges and possible solutions of this promising field to usher the readers towards next-generation ER targeted cancer therapy.