Differential Transcriptomic Signatures of Small Airway Cell Cultures Derived from IPF and COVID-19-Induced Exacerbation of Interstitial Lung Disease.

Idiopathic pulmonary fibrosis (IPF) is a pathological condition wherein lung injury precipitates the deposition of scar tissue, ultimately leading to a decline in pulmonary function. Existing research indicates a notable exacerbation in the clinical prognosis of IPF patients following infection with COVID-19. This investigation employed bulk RNA-sequencing methodologies to describe the transcriptomic profiles of small airway cell cultures derived from IPF and post-COVID fibrosis patients. Differential gene expression analysis unveiled heightened activation of pathways associated with microtubule assembly and interferon signaling in IPF cell cultures. Conversely, post-COVID fibrosis cell cultures exhibited distinctive characteristics, including the upregulation of pathways linked to extracellular matrix remodeling, immune system response, and TGF-ß1 signaling. Notably, BMP signaling levels were elevated in cell cultures derived from IPF patients compared to non-IPF control and post-COVID fibrosis samples. These findings underscore the molecular distinctions between IPF and post-COVID fibrosis, particularly in the context of signaling pathways associated with each condition. A better understanding of the underlying molecular mechanisms holds the promise of identifying potential therapeutic targets for future interventions in these diseases.

Detection of viral infection in cell lines using ViralCellDetector.

Cell lines are commonly used in research to study biology, including gene expression regulation, cancer progression, and drug responses. However, cross-contaminations with bacteria, mycoplasma, and viruses are common issues in cell line experiments. Detection of bacteria and mycoplasma infections in cell lines is relatively easy but identifying viral infections in cell lines is difficult. Currently, there are no established methods or tools available for detecting viral infections in cell lines. To address this challenge, we developed a tool called ViralCellDetector that detects viruses through mapping RNA-seq data to a library of virus genome. Using this tool, we observed that around 10% of experiments with the MCF7 cell line were likely infected with viruses. Furthermore, to facilitate the detection of samples with unknown sources of viral infection, we identified the differentially expressed genes involved in viral infection from two different cell lines and used these genes in a machine learning approach to classify infected samples based on the host response gene expression biomarkers. Our model reclassifies the infected and non-infected samples with an AUC of 0.91 and an accuracy of 0.93. Overall, our mapping- and marker-based approaches can detect viral infections in any cell line simply based on readily accessible RNA-seq data, allowing researchers to avoid the use of unintentionally infected cell lines in their studies.

Dihydroxy-Metabolites of Dihomo-γ-linolenic Acid Drive Ferroptosis-Mediated Neurodegeneration.

Even after decades of research, the mechanism of neurodegeneration remains understudied, hindering the discovery of effective treatments for neurodegenerative diseases. Recent reports suggest that ferroptosis could be a novel therapeutic target for neurodegenerative diseases. While polyunsaturated fatty acid (PUFA) plays an important role in neurodegeneration and ferroptosis, how PUFAs may trigger these processes remains largely unknown. PUFA metabolites from cytochrome P450 and epoxide hydrolase metabolic pathways may modulate neurodegeneration. Here, we test the hypothesis that specific PUFAs regulate neurodegeneration through the action of their downstream metabolites by affecting ferroptosis. We find that the PUFA dihomo-γ-linolenic acid (DGLA) specifically induces ferroptosis-mediated neurodegeneration in dopaminergic neurons. Using synthetic chemical probes, targeted metabolomics, and genetic mutants, we show that DGLA triggers neurodegeneration upon conversion to dihydroxyeicosadienoic acid through the action of CYP-EH (CYP, cytochrome P450; EH, epoxide hydrolase), representing a new class of lipid metabolites that induce neurodegeneration via ferroptosis.

Targeting ATP12A, a Nongastric Proton Pump α Subunit, for Idiopathic Pulmonary Fibrosis Treatment.

Idiopathic pulmonary fibrosis (IPF) is a pathological condition of unknown etiology that results from injury to the lung and an ensuing fibrotic response that leads to the thickening of the alveolar walls and obliteration of the alveolar space. The pathogenesis is not clear, and there are currently no effective therapies for IPF. Small airway disease and mucus accumulation are prominent features in IPF lungs, similar to cystic fibrosis lung disease. The ATP12A gene encodes the α-subunit of the nongastric H+, K+-ATPase, which functions to acidify the airway surface fluid and impairs mucociliary transport function in patients with cystic fibrosis. It is hypothesized that the ATP12A protein may play a role in the pathogenesis of IPF. The authors' studies demonstrate that ATP12A protein is overexpressed in distal small airways from the lungs of patients with IPF compared with normal human lungs. In addition, overexpression of the ATP12A protein in mouse lungs worsened bleomycin induced experimental pulmonary fibrosis. This was prevented by a potassium competitive proton pump blocker, vonoprazan. These data support the concept that the ATP12A protein plays an important role in the pathogenesis of lung fibrosis. Inhibition of the ATP12A protein has potential as a novel therapeutic strategy in IPF treatment.

Dihydroxy-Metabolites of Dihomo-gamma-linolenic Acid Drive Ferroptosis-Mediated Neurodegeneration.

Even after decades of research, the mechanism of neurodegeneration remains understudied, hindering the discovery of effective treatments for neurodegenerative diseases. Recent reports suggest that ferroptosis could be a novel therapeutic target for neurodegenerative diseases. While polyunsaturated fatty acid (PUFA) plays an important role in neurodegeneration and ferroptosis, how PUFAs may trigger these processes remains largely unknown. PUFA metabolites from cytochrome P450 and epoxide hydrolase metabolic pathways may modulate neurodegeneration. Here, we test the hypothesis that specific PUFAs regulate neurodegeneration through the action of their downstream metabolites by affecting ferroptosis. We find that the PUFA, dihomo gamma linolenic acid (DGLA), specifically induces ferroptosis-mediated neurodegeneration in dopaminergic neurons. Using synthetic chemical probes, targeted metabolomics, and genetic mutants, we show that DGLA triggers neurodegeneration upon conversion to dihydroxyeicosadienoic acid through the action of CYP-EH, representing a new class of lipid metabolite that induces neurodegeneration via ferroptosis.

BGLM: big data-guided LOINC mapping with multi-language support.

Motivation: Mapping internal, locally used lab test codes to standardized logical observation identifiers names and codes (LOINC) terminology has become an essential step in harmonizing electronic health record (EHR) data across different institutions. However, most existing LOINC code mappers are based on text-mining technology and do not provide robust multi-language support. Materials and methods: We introduce a simple, yet effective tool called big data-guided LOINC code mapper (BGLM), which leverages the large amount of patient data stored in EHR systems to perform LOINC coding mapping. Distinguishing from existing methods, BGLM conducts mapping based on distributional similarity. Results: We validated the performance of BGLM with real-world datasets and showed that high mapping precision could be achieved under proper false discovery rate control. In addition, we showed that the mapping results of BGLM could be used to boost the performance of Regenstrief LOINC Mapping Assistant (RELMA), one of the most widely used LOINC code mappers. Conclusions: BGLM paves a new way for LOINC code mapping and therefore could be applied to EHR systems without the restriction of languages. BGLM is freely available at https://github.com/Bin-Chen-Lab/BGLM.

Reversal of cancer gene expression identifies repurposed drugs for diffuse intrinsic pontine glioma.

Diffuse intrinsic pontine glioma (DIPG) is an aggressive incurable brainstem tumor that targets young children. Complete resection is not possible, and chemotherapy and radiotherapy are currently only palliative. This study aimed to identify potential therapeutic agents using a computational pipeline to perform an in silico screen for novel drugs. We then tested the identified drugs against a panel of patient-derived DIPG cell lines. Using a systematic computational approach with publicly available databases of gene signature in DIPG patients and cancer cell lines treated with a library of clinically available drugs, we identified drug hits with the ability to reverse a DIPG gene signature to one that matches normal tissue background. The biological and molecular effects of drug treatment was analyzed by cell viability assay and RNA sequence. In vivo DIPG mouse model survival studies were also conducted. As a result, two of three identified drugs showed potency against the DIPG cell lines Triptolide and mycophenolate mofetil (MMF) demonstrated significant inhibition of cell viability in DIPG cell lines. Guanosine rescued reduced cell viability induced by MMF. In vivo, MMF treatment significantly inhibited tumor growth in subcutaneous xenograft mice models. In conclusion, we identified clinically available drugs with the ability to reverse DIPG gene signatures and anti-DIPG activity in vitro and in vivo. This novel approach can repurpose drugs and significantly decrease the cost and time normally required in drug discovery.

Deciphering COVID-19 host transcriptomic complexity and variations for therapeutic discovery against new variants.

The molecular manifestations of host cells responding to SARS-CoV-2 and its evolving variants of infection are vastly different across the studied models and conditions, imposing challenges for host-based antiviral drug discovery. Based on the postulation that antiviral drugs tend to reverse the global host gene expression induced by viral infection, we retrospectively evaluated hundreds of signatures derived from 1,700 published host transcriptomic profiles of SARS/MERS/SARS-CoV-2 infection using an iterative data-driven approach. A few of these signatures could be reversed by known anti-SARS-CoV-2 inhibitors, suggesting the potential of extrapolating the biology for new variant research. We discovered IMD-0354 as a promising candidate to reverse the signatures globally with nanomolar IC50 against SARS-CoV-2 and its five variants. IMD-0354 stimulated type I interferon antiviral response, inhibited viral entry, and down-regulated hijacked proteins. This study demonstrates that the conserved coronavirus signatures and the transcriptomic reversal approach that leverages polypharmacological effects could guide new variant therapeutic discovery.

Ibrutinib Blocks YAP1 Activation and Reverses BRAF Inhibitor Resistance in Melanoma Cells.

Most B-Raf proto-oncogene (BRAF)-mutant melanoma tumors respond initially to BRAF inhibitor (BRAFi)/mitogen-activated protein kinase kinase 1 inhibitor (MEKi) therapy, although few patients have durable long-term responses to these agents. The goal of this study was to use an unbiased computational approach to identify inhibitors that reverse an experimentally derived BRAFi resistance gene expression signature. Using this approach, we found that ibrutinib effectively reverses this signature, and we demonstrate experimentally that ibrutinib resensitizes a subset of BRAFi-resistant melanoma cells to vemurafenib. Ibrutinib is used clinically as an inhibitor of the Src family kinase Bruton tyrosine kinase (BTK); however, neither BTK deletion nor treatment with acalabrutinib, another BTK inhibitor with reduced off-target activity, resensitized cells to vemurafenib. These data suggest that ibrutinib acts through a BTK-independent mechanism in vemurafenib resensitization. To better understand this mechanism, we analyzed the transcriptional profile of ibrutinib-treated BRAFi-resistant melanoma cells and found that the transcriptional profile of ibrutinib was highly similar to that of multiple Src proto-oncogene kinase inhibitors. Since ibrutinib, but not acalabrutinib, has appreciable off-target activity against multiple Src family kinases, it suggests that ibrutinib may be acting through this mechanism. Furthermore, genes that are differentially expressed in ibrutinib-treated cells are enriched in Yes1-associated transcriptional regulator (YAP1) target genes, and we showed that ibrutinib, but not acalabrutinib, reduces YAP1 activity in BRAFi-resistant melanoma cells. Taken together, these data suggest that ibrutinib, or other Src family kinase inhibitors, may be useful for treating some BRAFi/MEKi-refractory melanoma tumors. SIGNIFICANCE STATEMENT: MAPK-targeted therapies provide dramatic initial responses, but resistance develops rapidly; a subset of these tumors may be rendered sensitive again by treatment with an approved Src family kinase inhibitor-ibrutinub-potentially providing improved clinical outcomes.

Published anti-SARS-CoV-2 in vitro hits share common mechanisms of action that synergize with antivirals.

The global efforts in the past year have led to the discovery of nearly 200 drug repurposing candidates for COVID-19. Gaining more insights into their mechanisms of action could facilitate a better understanding of infection and the development of therapeutics. Leveraging large-scale drug-induced gene expression profiles, we found 36% of the active compounds regulate genes related to cholesterol homeostasis and microtubule cytoskeleton organization. Following bioinformatics analyses revealed that the expression of these genes is associated with COVID-19 patient severity and has predictive power on anti-SARS-CoV-2 efficacy in vitro. Monensin, a top new compound that regulates these genes, was further confirmed as an inhibitor of SARS-CoV-2 replication in Vero-E6 cells. Interestingly, drugs co-targeting cholesterol homeostasis and microtubule cytoskeleton organization processes more likely present a synergistic effect with antivirals. Therefore, potential therapeutics could be centered around combinations of targeting these processes and viral proteins.

Published Anti-SARS-CoV-2 In Vitro Hits Share Common Mechanisms of Action that Synergize with Antivirals.

The global efforts in the past few months have led to the discovery of around 200 drug repurposing candidates for COVID-19. Although most of them only exhibited moderate anti- SARS-CoV-2 activity, gaining more insights into their mechanisms of action could facilitate a better understanding of infection and the development of therapeutics. Leveraging large-scale drug-induced gene expression profiles, we found 36% of the active compounds regulate genes related to cholesterol homeostasis and microtubule cytoskeleton organization. The expression change upon drug treatment was further experimentally confirmed in human lung primary small airway. Following bioinformatics analysis on COVID-19 patient data revealed that these genes are associated with COVID-19 patient severity. The expression level of these genes also has predicted power on anti-SARS-CoV-2 efficacy in vitro, which led to the discovery of monensin as an inhibitor of SARS-CoV-2 replication in Vero-E6 cells. The final survey of recent drug- combination data indicated that drugs co-targeting cholesterol homeostasis and microtubule cytoskeleton organization processes more likely present a synergistic effect with antivirals. Therefore, potential therapeutics should be centered around combinations of targeting these processes and viral proteins.

Sex differences in viral entry protein expression and host transcript responses to SARS-CoV-2.

Epidemiological studies suggest that men exhibit a higher mortality rate to COVID-19 than women, yet the underlying biology is largely unknown. Here, we seek to delineate sex differences in the gene expression of viral entry proteins ACE2 and TMPRSS2, and host transcriptional responses to SARS-CoV-2 through large-scale analysis of genomic and clinical data. We first compiled 220,000 human gene expression profiles from three databases and completed the meta-information through machine learning and manual annotation. Large scale analysis of these profiles indicated that male samples show higher expression levels of ACE2 and TMPRSS2 than female samples, especially in the older group (>60 years) and in the kidney. Subsequent analysis of 6,031 COVID-19 patients at Mount Sinai Health System revealed that men have significantly higher creatinine levels, an indicator of impaired kidney function. Further analysis of 782 COVID-19 patient gene expression profiles taken from upper airway and blood suggested men and women present distinct expression changes. Computational deconvolution analysis of these profiles revealed male COVID-19 patients have enriched kidney-specific mesangial cells in blood compared to healthy patients. Together, this study suggests biological differences in the kidney between sexes may contribute to sex disparity in COVID-19.

The Relationship between Leptin, the Leptin Receptor and FGFR1 in Primary Human Breast Tumors.

Obesity is associated with increased breast cancer risk and poorer cancer outcomes; however, the precise etiology of these observations has not been fully identified. Our previous research suggests that adipose tissue-derived fibroblast growth factor-2 (FGF2) promotes the malignant transformation of epithelial cells through the activation of fibroblast growth factor receptor-1 (FGFR1). FGF2 is increased in the context of obesity, and increased sera levels have been associated with endocrine-resistant breast cancer. Leptin is a marker of obesity and promotes breast carcinogenesis through several mechanisms. In this study, we leverage public gene expression datasets to evaluate the associations between FGFR1, leptin, and the leptin receptor (LepR) in breast cancer. We show a positive association between FGFR1 and leptin protein copy number in primary breast tumors. These observations coincided with a positive association between Janus kinase 2 (Jak2) mRNA with both leptin receptor (LepR) mRNA and FGFR1 mRNA. Moreover, two separate Jak2 inhibitors attenuated both leptin+FGF2-stimulated and mouse adipose tissue-stimulated MCF-10A transformation. These results demonstrate how elevated sera FGF2 and leptin in obese patients may promote cancer progression in tumors that express elevated FGFR1 and LepR through Jak2 signaling. Therefore, Jak2 is a potential therapeutic target for FGFR1 amplified breast cancer, especially in the context of obesity.

Analysis of Infected Host Gene Expression Reveals Repurposed Drug Candidates and Time-Dependent Host Response Dynamics for COVID-19.

The repurposing of existing drugs offers the potential to expedite therapeutic discovery against the current COVID-19 pandemic caused by the SARS-CoV-2 virus. We have developed an integrative approach to predict repurposed drug candidates that can reverse SARS-CoV-2-induced gene expression in host cells, and evaluate their efficacy against SARS-CoV-2 infection in vitro. We found that 13 virus-induced gene expression signatures computed from various viral preclinical models could be reversed by compounds previously identified to be effective against SARS- or MERS-CoV, as well as drug candidates recently reported to be efficacious against SARS-CoV-2. Based on the ability of candidate drugs to reverse these 13 infection signatures, as well as other clinical criteria, we identified 10 novel candidates. The four drugs bortezomib, dactolisib, alvocidib, and methotrexate inhibited SARS-CoV-2 infection-induced cytopathic effect in Vero E6 cells at < 1 µM, but only methotrexate did not exhibit unfavorable cytotoxicity. Although further improvement of cytotoxicity prediction and bench testing is required, our computational approach has the potential to rapidly and rationally identify repurposed drug candidates against SARS-CoV-2. The analysis of signature genes induced by SARS-CoV-2 also revealed interesting time-dependent host response dynamics and critical pathways for therapeutic interventions (e.g. Rho GTPase activation and cytokine signaling suppression).

Evaluating cell lines as models for metastatic breast cancer through integrative analysis of genomic data.

Cell lines are widely-used models to study metastatic cancer although the extent to which they recapitulate the disease in patients remains unknown. The recent accumulation of genomic data provides an unprecedented opportunity to evaluate the utility of them for metastatic cancer research. Here, we reveal substantial genomic differences between breast cancer cell lines and metastatic breast cancer patient samples. We also identify cell lines that more closely resemble the different subtypes of metastatic breast cancer seen in the clinic and show that surprisingly, MDA-MB-231 cells bear little genomic similarities to basal-like metastatic breast cancer patient samples. Further comparison suggests that organoids more closely resemble the transcriptome of metastatic breast cancer samples compared to cell lines. Our work provides a guide for cell line selection in the context of breast cancer metastasis and highlights the potential of organoids in these studies.