Death Response Research Articles

Mechanisms regulating host cell death during Leishmania infection.

Parasites from the Leishmania genus are the causative agents of leishmaniasis and primarily reside within macrophages during mammalian infection. Their ability to establish intracellular infection provides a secure niche for proliferation while evading detection. However, successful multiplication within mammalian cells requires the orchestration of multiple mechanisms that control host cell viability. In contrast, innate immune cells, such as macrophages, can undergo different forms of cell death in response to pathogenic intracellular microbes. Thus, modulation of these different forms of host cell death is crucial for Leishmaniasis development. The regulation of host cell apoptosis, a form of programmed cell death, is crucial for sustaining parasites within viable host cells. Accordingly, several studies have demonstrated evasion of apoptosis induced by dermotropic and viscerotropic Leishmania species. Conversely, the prevention of pyroptosis, an inflammatory form of cell death, ensures the establishment of infection by silencing the release of mediators that could trigger massive proinflammatory responses. This manuscript explores how Leishmania regulates various host cell death pathways and overviews seminal studies on regulating host cell apoptosis by different Leishmania species.

DDDR-27. BMP SIGNALING MODULATES APOPTOTIC AND NON-APOPTOTIC CELL DEATH IN GLIOBLASTOMA

Abstract Recurrence occurs in approximately 90% of glioblastoma (GBM) patients, in part due to the tumors becoming resistant to cell death in response to existing therapies. These therapies often trigger apoptosis; however, apoptosis is only one of several different forms of cell death. We hypothesize that activating non-canonical death mechanisms could provide a back door to killing treatment-resistant GBM cells. To test this hypothesis, we first developed a new imaging-based high-throughput method to directly quantify cell death in 3D models. Using this method, we find patient-derived GBM spheroids to be resistant to the induction of cell death in response to various standard-of-care and experimental therapeutic compounds. To further investigate mechanisms underlying this resistance, we screened a panel of extracellular protein ligands for the modulation of cell death response. We find that bone morphogenic protein (BMP) signaling increases resistance to several apoptosis-inducing compounds, including temozolomide and tyrosine kinase inhibitors. Intriguingly, BMP signaling simultaneously enhanced sensitivity to a novel non-apoptotic cell death pathway triggered by the small molecule CIL56. Efforts are ongoing to further understand how BMP signaling governs apoptotic and non-apoptotic cell death mechanisms. Nevertheless, our results suggest a novel approach to treating GBM, centered upon the modulation of BMP signaling combined with the use of agents that induce non-apoptotic cell death.

Integrating transcriptomic data and digital pathology for NRG-based prediction of prognosis and therapy response in gastric cancer

Background Cancer is characterized by its ability to resist cell death, and emerging evidence suggests a potential correlation between non-apoptotic regulated cell death (RCD), tumor progression, and therapy response. However, the prognostic significance of non-apoptotic RCD-related genes (NRGs) and their relationships with immune response in gastric cancer (GC) remain unclear. Methods In this study, RNA-seq gene expression and clinical information of GC patients were acquired from The Cancer Genome Atlas and the Gene Expression Omnibus databases. Cox and LASSO regression analyses were used to construct the NRG signature. Moreover, we developed a deep learning model based on ResNet50 to predict the NRG signature from digital pathology slides. The expression of signature hub genes was validated using real-time quantitative PCR and single-cell RNA sequencing data. Results We identified 13 NRGs as signature genes for predicting the prognosis of patients with GC. The high-risk group, characterized by higher NRG scores, demonstrated a shorter overall survival rate, increased immunosuppressive cell infiltration, and immune dysfunction. Moreover, associations were observed between the NRG signature and chemotherapeutic drug responsiveness, as well as immunotherapy effectiveness in GC patients. Furthermore, the deep learning model effectively stratified GC patients based on the NRG signature by leveraging morphological variances, showing promising results for the classification of GC patients. Validation experiments demonstrated that the expression level of SERPINE1 was significantly upregulated in GC, while the expression levels of GPX3 and APOD were significantly downregulated. Conclusion The NRG signature and its deep learning model have significant clinical implications, highlighting the importance of individualized treatment strategies based on GC subtyping. These findings provide valuable insights for guiding clinical decision-making and treatment approaches for GC.

Maternal and perinatal death surveillance and response in Bangladesh: A case study on measuring impact through health information systems

The health sector in Bangladesh is thriving, and the Maternal and Perinatal Death Surveillance and Response (MPDSR) program has been in place to investigate the causes of maternal and perinatal death. This study aimed to investigate the availability of information systems for measuring the status of an MPDSR program and the planning process and actions developed based on real-time information. This study delved into innovative information systems that can be used to monitor and measure the impact of health programs. The study included both the primary and secondary data. A total of six key informant interviews were conducted to explore the enablers and strengths of information systems. Secondary data were collected from government websites and MPDSR action plans. This study highlights the potential and challenges of governmental health information systems in notifying maternal and neonatal deaths. While the current notification rates are not up to national standards, it is encouraging to see health managers develop action plans and address this issue. The MPDSR information system is a valuable tool to help health managers measure changes in maternal and perinatal mortality rates and design locally targeted interventions. However, several obstacles must be addressed, such as insufficient data, lack of monitoring, and irregularities in MPDSR review meetings. By finding ways to overcome these challenges, we can unlock the full potential of MPDSR and improve maternal and neonatal health outcomes. The success of an MPDSR program relies on how the information is utilized to take action and measure the outcome. Advanced information systems, such as health dashboards, scorecards, and administrative data, can play a vital role in measuring the progress and impact of a program. The primary challenge lies in capturing all deaths, including their causes, and having the capacity to analyze the data effectively to develop action plans for health managers.

Using normalisation process theory (NPT) to explore implementation of the maternal perinatal death surveillance and response (MPDSR) policy in Uganda: a reflection

BackgroundThe implementation of the maternal perinatal death surveillance and response (MPDSR) policy is among the envisaged strategies to reduce the high global burden of maternal and perinatal mortality and morbidity. However, implementation of this policy across various contexts is inconsistent. Theoretically informed approaches to process evaluation can support assessment the implementation of policy interventions such as MPDSR, particularly in understanding what the actors involved actually do. In this article, we reflect on how the normalisation process theory (NPT) was used to explore implementation of the MPDSR policy in Uganda. NPT is a sociological theory concerned with the social organisation of the work (implementation) of making practices routine elements of everyday life (embedding) and of sustaining embedded practices in their social contexts (integration).MethodsThis qualitative multiple case study conducted across eight districts in Uganda and among 10 health facilities (cases) representing four out of the seven levels of the Uganda health care system. NPT was utilised in several ways including informing the study design, structuring the data collection tools (semi-structured interview guides), providing an organising framework for analysis, interpreting and reporting of study findings as well as making recommendations. Study participants were purposely selected to reflect the range of actors involved in the policy implementation process. This included direct care providers located at each of the cases, the Ministry of Health and from agencies and professional associations. Data were collected using semi-structured, in-depth interviews and were inductively and deductively analysed using NPT constructs and subconstructs.Results and conclusionNPT served useful for process evaluation, particularly in identifying factors that contribute to variations in policy implementation. Considering the NPT focus on the agency of people involved in implementation, additional efforts are required to understand how recipients of the policy intervention influence how the intervention becomes embedded within the various contexts.

Loss of interferon regulatory factor-1 prevents lung fibrosis by upregulation of pon1 expression

BackgroundInterferon regulatory factor-1 (IRF1) is a transcription factor that plays a significant role in various biological processes, including inflammatory injury, viral infection, cell death, and immune responses, and it has been extensively studied in the context of different lung diseases. However, the mechanism underlying its involvement in lung fibrosis remains largely unknown.MethodsWild type (WT) mice, IRF1 global-null mice (Irf1−/−) were subjected to a bleomycin-induced lung fibrosis model to enable examination of the role of IRF1 in lung fibrosis. Proteomic analysis of lung tissue from WT and Irf1−/− mice treated with saline or bleomycin was performed to explore the mechanism of IRF1 in regulating lung fibrosis.ResultsIn the bleomycin-induced fibrosis mouse model, increased expression of IRF1 was observed. Irf1 knockout mice displayed decreased lung fibrosis relative to WT mice following treatment with bleomycin. The protein expression of fibronectin, as assessed by the Western blot analysis of lung tissues, was downregulated in Irf1−/− mice. We observed a similar reduction in collagen content using hydroxyproline detection. Histologically, there was less collagen deposition in the lungs of Irf1−/− mice compared with WT mice. Proteomics data revealed that IRF1 may be involved in lung fibrosis via the regulation of ferroptosis. We determined that paraoxonase 1(PON1), a poorly characterized protein in lung fibrosis, was upregulated in Irf1−/− mice following exposure to bleomycin. In vitro experiments revealed that IRF1 could regulate the level of GSH and MDA through PON1. We also determined that PON1 levels were lower in the plasma of IPF patients compared with healthy controls.ConclusionOur data highlight the importance of IRF1 in the fibrotic process, and PON1 may be a potential mediator of IRF1 in the progression of lung fibrosis.

Telaglenastat as an alternative to cisplatin as a radiosensitizer in the treatment of head and neck squamous cell carcinoma

The efficacy of radiation treatment (RT) of head and neck squamous cell carcinoma (HNSCC) is limited by radioresistance and the toxicity of FDA approved radiosensitizers. In extension to our previous research where we demonstrated that telaglenastat (CB839) increased efficacy of RT in in vitro and in vivo HNSCC models, here, we examine the radiosensitizing effects of telaglenastat in comparison to cisplatin's, as cisplatin is currently the standard of care for concurrent therapy. Combination of telaglenastat with RT reduced tumor volume in a HNSCC patient derived xenograft mouse model. The efficacy of telaglenastat with RT in reducing cell survival and increasing apoptosis was similar if not greater than that of cisplatin with RT in Cal27 and HN5 HNSCC cells. The addition of telaglenastat increased reactive oxygen species and reduced the antioxidant glutathione in both Cal27 and HN5 cells. Reverse Phase Protein Array analyses revealed alterations in cell death and DNA damage response proteins. This study provides the scientific underpinnings for the use of telaglenastat as a radiosensitizer in the treatment of HNSCC either as an alternative to cisplatin or in cisplatin-ineligible patients.

Necroptosis as a consequence of photodynamic therapy in tumor cells.

Photodynamic therapy (PDT) is an alternative to cancer treatment, demonstrating selectivity and significant cytotoxicity on malignant tissues. Such therapy involves two nontoxic components: photosensitizer (PS) and non-ionizing radiation. In optimal dosage combinations, PDT causes cellular and tissue effects by oxygen-dependent processes, leading tumor cells to regulated cell death pathways. Regulated necrosis, called necroptosis, can be triggered by PDT and is characterized by caspase-8 inhibition and RIPK1, RIPK3, and MLKL activities, leading to plasma membrane pores formation with subsequent cellular content release into the extracellular space. For this review, studies accessed by PubMed describing the relation between necroptosis and PDT were summarized. The results showed that PDT can trigger necroptosis mechanisms in different tumor cells. Moreover, a mix of different cell death types can co-occur. It is also important to highlight that necroptosis triggered by PDT is related to damage-associated molecular patterns (DAMPs) release, involving immunogenic cell death and vaccination. The cell death response is directly related to the photosensitizer chemical characteristics, concentration, incubation time, cellular location, and irradiation parameters. The synergism among all cell death types is an excellent advantage for avowing tumor resistance mechanisms and developing new solutions.

Understanding the molecular regulatory mechanisms of autophagy in lung disease pathogenesis.

Lung disease development involves multiple cellular processes, including inflammation, cell death, and proliferation. Research increasingly indicates that autophagy and its regulatory proteins can influence inflammation, programmed cell death, cell proliferation, and innate immune responses. Autophagy plays a vital role in the maintenance of homeostasis and the adaptation of eukaryotic cells to stress by enabling the chelation, transport, and degradation of subcellular components, including proteins and organelles. This process is essential for sustaining cellular balance and ensuring the health of the mitochondrial population. Recent studies have begun to explore the connection between autophagy and the development of different lung diseases. This article reviews the latest findings on the molecular regulatory mechanisms of autophagy in lung diseases, with an emphasis on potential targeted therapies for autophagy.

Stepwise phosphorylation and SUMOylation of PIDD1 drive PIDDosome assembly in response to DNA repair failure

SUMOylation regulates numerous cellular stress responses, yet targets in the apoptotic machinery remain elusive. We show that a single, DNA damage-induced monoSUMOylation event controls PIDDosome (PIDD1/RAIDD/caspase-2) formation and apoptotic death in response to unresolved DNA interstrand crosslinks (ICLs). SUMO-1 conjugation occurs on conserved K879 in the PIDD1 death domain (DD); is catalyzed by PIAS1 and countered by SENP3; and is triggered by ATR phosphorylation of neighboring T788 in the PIDD1 DD, which enables PIAS1 docking. Phospho/SUMO-PIDD1 proteins are captured by nucleolar RAIDD monomers via a SUMO-interacting motif (SIM) in the RAIDD DD, thus compartmentalizing nascent PIDDosomes for caspase-2 recruitment. Denying SUMOylation or the SUMO-SIM interaction spares the onset of PIDDosome assembly but blocks its completion, thus eliminating the apoptotic response to ICL repair failure. Conversely, removal of SENP3 forces apoptosis, even in cells with tolerable ICL levels. SUMO-mediated PIDDosome control is also seen in response to DNA breaks but not supernumerary centrosomes. These results illuminate PIDDosome formation in space and time and identify a direct role for SUMOylation in the assembly of a major pro-apoptotic device.

3D Neurovascular Unit Tissue Model to Assess Responses to Traumatic Brain Injury.

The neurovascular unit (NVU) is a critical interface in the central nervous system that links vascular interactions with glial and neural tissue. Disruption of the NVU has been linked to the onset and progression of neurodegenerative diseases. Despite its significance the NVU remains challenging to study in a physiologically relevant manner. Here, a 3D cell triculture model of the NVU is developed that incorporates human primary brain microvascular endothelial cells, astrocytes, and pericytes into a tissue system that can be sustained invitro for several weeks. This tissue model helps recapitulate the complexity of the NVU and can be used to interrogate the mechanisms of disease and cell-cell interactions. The NVU tissue model displays elevated cell death and inflammatory responses following mechanical damage, to emulate traumatic brain injury (TBI) under controlled laboratory conditions, including lactate dehydrogenase (LDH) release, elevated inflammatory markers TNF-α and monocyte chemoattractant cytokines MCP-2 and MCP-3 and reduced expression of the tight junction marker ZO-1. This 3D tissue model serves as a tool for deciphering mechanisms of TBIs and immune responses associated with the NVU.

Insights into ubiquitinome dynamics in the host‒pathogen interplay during Francisella novicida infection

Ubiquitination functions as an important posttranslational modification for orchestrating inflammatory immune responses and cell death during pathogenic infection. The ubiquitination machinery is a major target hijacked by pathogenic bacteria to promote their survival and proliferation. Type I interferon (IFN-I) plays detrimental roles in host defense against Francisella novicida (F. novicida) infection. The effects of IFN-I on the ubiquitination of host proteins during F. novicida infection remain unclear. Herein, we delineate the dynamic ubiquitinome alterations in both wild-type (WT) and interferon-alpha receptor-deficient (Ifnar–/–) primary bone marrow-derived macrophages (BMDMs) during F. novicida infection. Using diGly proteomics and stable isotope labeling (SILAC), we quantified ubiquitination sites in proteins from primary WT and Ifnar–/– BMDMs with and without F. novicida infection. Our mass spectrometry analysis identified 2,491 ubiquitination sites in 1,077 endogenous proteins. Our study revealed that F. novicida infection induces dynamic changes in the ubiquitination of proteins involved in the cell death, phagocytosis, and inflammatory response pathways. IFN-I signaling is essential for both the increase and reduction in ubiquitination in response to F. novicida infection. We identified IFN-I-dependent ubiquitination in proteins involved in glycolysis and vesicle transport processes and highlighted key hub proteins modified by ubiquitination within cell death pathways. These findings underscore the significant influence of IFN-I signaling on modulating ubiquitination during F. novicida infection and provide valuable insights into the complex interplay between the host and F. novicida.

AIBP Protects Müller Glial Cells Against Oxidative Stress-Induced Mitochondrial Dysfunction and Reduces Retinal Neuroinflammation.

Glaucoma, an optic neuropathy with the loss of retinal ganglion cells (RGCs), is a leading cause of irreversible vision loss. Oxidative stress and mitochondrial dysfunction have a significant role in triggering glia-driven neuroinflammation and subsequent glaucomatous RGC degeneration in the context of glaucoma. It has previously been shown that apolipoprotein A-I binding protein (APOA1BP or AIBP) has an anti-inflammatory function. Moreover, Apoa1bp-/- mice are characterized by retinal neuroinflammation and RGC loss. In this study, we found that AIBP deficiency exacerbated the oxidative stress-induced disruption of mitochondrial dynamics and function in the retina, leading to a further decline in visual function. Mechanistically, AIBP deficiency-induced oxidative stress triggered a reduction in glycogen synthase kinase 3β and dynamin-related protein 1 phosphorylation, optic atrophy type 1 and mitofusin 1 and 2 expression, and oxidative phosphorylation, as well as the activation of mitogen-activated protein kinase (MAPK) in Müller glia dysfunction, leading to cell death and inflammatory responses. In vivo, the administration of recombinant AIBP (rAIBP) effectively protected the structural and functional integrity of retinal mitochondria under oxidative stress conditions and prevented vision loss. In vitro, incubation with rAIBP safeguarded the structural integrity and bioenergetic performance of mitochondria and concurrently suppressed MAPK activation, apoptotic cell death, and inflammatory response in Müller glia. These findings support the possibility that AIBP promotes RGC survival and restores visual function in glaucomatous mice by ameliorating glia-driven mitochondrial dysfunction and neuroinflammation.

IMPACT OF REAL-LIFE ENVIRONMENTAL EXPOSURES ON REPRODUCTION: Systemic and ovarian impacts of heat stress in the porcine model.

This review describes how heat stress causes systemic endocrine and metabolic alterations that contribute to intracellular ovarian perturbations, resulting in female infertility. Heat stress (HS) in mammals results from an imbalance in heat accumulation and dissipation. Fertility impairments consequential to HS have been recognized for decades in production animals, and more recently, observations have been extended to other species, including women. There are several systemic impacts of HS that can independently affect reproduction, including metabolic endotoxemia, reduced plane of nutrition, and endocrine disruption. At the level of the ovary, molecular pathways are altered by HS, such as inflammation, JAK-STAT, PI3K, oxidative stress, cell death, and heat shock response. Taken together, impaired ovarian function contributes to seasonal infertility that results from HS. This review paper describes the physiological and endocrine systemic impacts of HS that may independently and collaboratively impair fertility in the porcine model. The review then details ovarian intracellular events that are altered during HS and finally determines future needs in this area of research.

Membrane Ruffles: Composition, Function, Formation and Visualization

Membrane ruffles are cell actin-based membrane protrusions that have distinct structural characteristics. Linear ruffles with columnar spike-like and veil-like structures assemble at the leading edge of cell membranes. Circular dorsal ruffles (CDRs) have no supporting columnar structures but their veil-like structures, connecting from end to end, present an enclosed ring-shaped circular outline. Membrane ruffles are involved in multiple cell functions such as cell motility, macropinocytosis, receptor internalization, fluid viscosity sensing in a two-dimensional culture environment, and protecting cells from death in response to physiologically compressive loads. Herein, we review the state-of-the-art knowledge on membrane ruffle structure and function, the growth factor-induced membrane ruffling process, and the growth factor-independent ruffling mode triggered by calcium and other stimulating factors, together with the respective underlying mechanisms. We also summarize the inhibitors used in ruffle formation studies and their specificity. In the last part, an overview is given of the various techniques in which the membrane ruffles have been visualized up to now.

Ventral Hernia Repair With a Hybrid Absorbable-permanent Preperitoneal Mesh.

To analyze device safety and clinical outcomes of ventral hernia repair with the GORE SYNECOR Preperitoneal Biomaterial (PRE device), a permanent high-strength mesh with bioabsorbable web scaffold technology. This multicenter retrospective review analyzed device/procedure endpoints and patient-reported outcomes in patients treated for hernia repair ≥1 year from study enrollment. Included in this analysis were 148 patients with a mean age of 56 years; 66.2% met the Ventral Hernia Working Group grade 2 classification. Median hernia size was 30.0cm 2 and 58.8% of patients had an incisional hernia. Repairs were primarily a robotic (53.4%) or open approach (41.9%). All meshes were placed extraperitoneal. Procedure-related adverse events within 30 days occurred in 13 (8.8%) patients and included 7 (4.8%) patients with surgical site infection, 2 (1.4%) with surgical site occurrence (SSO), 4 (2.7%) requiring readmission, and 3 (2.0%) who had reoperation. The rate of SSO events requiring procedural intervention was 2.7% (4 patients) through 30 days and 3.4% (5 patients) at 12 months. The rate of procedure-related surgical site infection remained at 4.8% through 12 months (no further reports after 30d) and 3.4% for SSO (2 reports after 30d). There were no site-reported clinically diagnosed hernia recurrences throughout the study. Median patient follow-up including in-person visit, physical examination, reported adverse event, explant, death, and questionnaire response was 28 months (n = 148). Median patient follow-up with patient questionnaire was 36 months (n = 88). Use of the PRE device, which incorporates the proven advantages of both an absorbable synthetic mesh and the long-term durability of a permanent macroporous mesh, is safe and effective in complex ventral hernia repairs. When used in the retromuscular space, the combination of these 2 materials had lower wound complications and recurrence rates than either type of material alone.

Protective effect of recombinant interleukin-10 on newborn rat lungs exposed to short-term sublethal hyperoxia.

Lung injury imposed by hyperoxia is the main cause of bronchopulmonary dysplasia in newborns. These injuries are generated from the early stage of hyperoxia through the main biologic effects of cell death and inflammatory response. Interleukin (IL)-10 is a potent anti-inflammatory cytokine that may have the inhibitory effects on these biologic actions induced by hyperoxia. Based on our former in vitro studies investigating the effect of recombinant IL-10 (rIL-10) on protecting cultured alveolar type II cells exposed to short-term hyperoxia, we performed the in vivo study to investigate the effect of rIL-10 in newborn rats aged P4 exposed to hyperoxia. Rats were classified into 3 groups; the control group exposed to normoxia for 24 hours; the hyperoxia group exposed to 65% hyperoxia for 24 hours; and the IL10 group treated with intratracheal instillation of rIL-10 prior to exposure to 65% hyperoxia for 24 hours. Following each treatment, the rats were euthanized. Individual lobes of the right lung were prepared for hematoxyling and eosin (H&E) staining and immunohistochemical staining for thyroid transcription factor-1 (TTF1). Bronchoalveolar lavage (BAL) was performed in the left lung to analyze cell counts and cytokines. The IL10 group showed preserved air spaces similar to the control group, with decreased cellularity compared to the hyperoxia group, whereas the hyperoxia group showed markedly reduced air spaces with increased cellularity compared to the IL10 group. And, the IL10 group showed more TTF1-positive cells, which represented alveolar type II cells, compared to the hyperoxia group. Inflammatory cells, such as neutrophils and lymphocytes and proinflammatory cytokines of tumor necrosis factor-α, IL-1α, IL-8, and macrophage inflammatory protein-1α were significantly lower in BAL fluid of the IL10 group compared to the hyperoxia group. These results indicate that rIL-10 may be a promising pharmaceutical measure for protecting newborn lungs from injury induced at the early stage of hyper oxia.

An array of Zymoseptoria tritici effectors suppress plant immune responses.

Zymoseptoria tritici is the most economically significant fungal pathogen of wheat in Europe. However, despite the importance of this pathogen, the molecular interactions between pathogen and host during infection are not well understood. Herein, we describe the use of two libraries of cloned Z. tritici effectors that were screened to identify effector candidates with putative pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI)-suppressing activity. The effectors from each library were transiently expressed in Nicotiana benthamiana, and expressing leaves were treated with bacterial or fungal PAMPs to assess the effectors' ability to suppress reactive oxygen species (ROS) production. From these screens, numerous effectors were identified with PTI-suppressing activity. In addition, some effectors were able to suppress cell death responses induced by other Z. tritici secreted proteins. We used structural prediction tools to predict the putative structures of all of the Z. tritici effectors and used these predictions to examine whether there was enrichment of specific structural signatures among the PTI-suppressing effectors. From among the libraries, multiple members of the killer protein-like 4 (KP4) and killer protein-like 6 (KP6) effector families were identified as PTI suppressors. This observation is intriguing, as these protein families were previously associated with antimicrobial activity rather than virulence or host manipulation. This data provides mechanistic insight into immune suppression by Z. tritici during infection and suggests that, similar to biotrophic pathogens, this fungus relies on a battery of secreted effectors to suppress host immunity during early phases of colonization.

Targeting caspase-8/c-FLIPL heterodimer in complex II promotes DL-mediated cell death.

Death receptor (DR) networks are controlled by the assembly of the Death-Inducing Signaling Complex (DISC) and complex II. The family of small molecules FLIPins (FLIP interactors) were developed to target the caspase-8/c-FLIPL heterodimer. FLIPin compounds were shown to promote apoptosis and caspase-8 activation at the DISC upon stimulation with death ligands (DLs) such as CD95L and TRAIL. To further investigate the role of FLIPin compounds in the DL-mediated cell death response, we analyzed their effects in combination with DLs and SMAC mimetics treatment. FLIPins were found to enhance cell viability loss and cell death induced by DL and SMAC mimetics in acute myeloid leukemia (AML), colon and pancreatic cancer cells. FLIPins enhanced both DL/BV6-induced apoptosis and DL/BV6/zVAD-fmk-induced necroptosis via an increase in complex II formation. Our results indicate that targeting the caspase-8/c-FLIPL heterodimer plays a prominent role in enhancing cell death induced by co-stimulation of DL/SMAC mimetics and opens new therapeutic strategies for targeting DR networks.

Modulating the unfolded protein response with ISRIB mitigates cisplatin ototoxicity

Cisplatin is a commonly used chemotherapy agent with a nearly universal side effect of sensorineural hearing loss. The cellular mechanisms underlying cisplatin ototoxicity are poorly understood. Efforts in drug development to prevent or reverse cisplatin ototoxicity have largely focused on pathways of oxidative stress and apoptosis. An effective treatment for cisplatin ototoxicity, sodium thiosulfate (STS), while beneficial when used in standard risk hepatoblastoma, is associated with reduced survival in disseminated pediatric malignancy, highlighting the need for more specific drugs without potential tumor protective effects. The unfolded protein response (UPR) and endoplasmic reticulum (ER) stress pathways have been shown to be involved in the pathogenesis of noise-induced hearing loss and cochlear synaptopathy in vivo, and these pathways have been implicated broadly in cisplatin cytotoxicity. This study sought to determine whether the UPR can be targeted to prevent cisplatin ototoxicity. Neonatal cochlear cultures and HEK cells were exposed to cisplatin, and UPR marker gene expression and cell death measured. Treatment with ISRIB (Integrated Stress Response InhIBitor), a drug that activates eif2B and downregulates the pro-apoptotic PERK/CHOP pathway of the UPR, was tested for its ability to reduce apoptosis in HEK cells, hair-cell death in cochlear cultures, and hearing loss using an in vivo mouse model of cisplatin ototoxicity. Finally, to evaluate whether ISRIB might interfere with cisplatin chemoeffectiveness, we tested it in head and neck squamous cell carcinoma (HNSCC) cell-based assays of cisplatin cytotoxicity. Cisplatin exhibited a biphasic, non-linear dose–response of cell death and apoptosis that correlated with different patterns of UPR marker gene expression in HEK cells and cochlear cultures. ISRIB treatment protected against cisplatin-induced hearing loss and hair-cell death, but did not impact cisplatin’s cytotoxic effects on HNSCC cell viability, unlike STS. These findings demonstrate that targeting the pro-apoptotic PERK/CHOP pathway with ISRIB can mitigate cisplatin ototoxicity without reducing anti-cancer cell effects, suggesting that this may be a viable strategy for drug development.