Two-polarized roles of transcription factor FOSB in lung cancer progression and prognosis: dependent on p53 status.

BACKGROUND: Activator protein-1 (AP-1) represents a transcription factor family that has garnered growing attention for its extensive involvement in tumor biology. However, the roles of the AP-1 family in the evolution of lung cancer remain poorly characterized. FBJ Murine Osteosarcoma Viral Oncogene Homolog B (FOSB), a classic AP-1 family member, was previously reported to play bewilderingly two-polarized roles in non-small cell lung cancer (NSCLC) as an enigmatic double-edged sword, for which the reasons and significance warrant further elucidation. METHODS AND RESULTS: Based on the bioinformatics analysis of a large NSCLC cohort from the TCGA database, our current work found the well-known tumor suppressor gene TP53 served as a key code to decipher the two sides of FOSB - its expression indicated a positive prognosis in NSCLC patients harboring wild-type TP53 while a negative one in those harboring mutant TP53. By constructing a panel of syngeneically derived NSCLC cells expressing p53 in different statuses, the radically opposite prognostic effects of FOSB expression in NSCLC population were validated, with the TP53-R248Q mutation site emerging as particularly meaningful. Transcriptome sequencing showed that FOSB overexpression elicited diversifying transcriptomic landscapes across NSCLC cells with varying genetic backgrounds of TP53 and, combined with the validation by RT-qPCR, PREX1 (TP53-Null), IGFBP5 (TP53-WT), AKR1C3, and ALDH3A1 (TP53-R248Q) were respectively identified as p53-dependent transcriptional targets of FOSB. Subsequently, the heterogenous impacts of FOSB on the tumor biology in NSCLC cells via the above selective transcriptional targets were confirmed in vitro and in vivo. Mechanistic investigations revealed that wild-type or mutant p53 might guide FOSB to recognize and bind to distinct promoter sequences via protein-protein interactions to transcriptionally activate specific target genes, thereby creating disparate influences on the progression and prognosis in NSCLC. CONCLUSIONS: FOSB expression holds promise as a novel prognostic biomarker for NSCLC in combination with a given genetic background of TP53, and the unique interactions between FOSB and p53 may serve as underlying intervention targets for NSCLC.

Chronic restraint stress promotes oral squamous cell carcinoma development by inhibiting ALDH3A1 via stress response hormone.

BACKGROUND: Chronic restraint stress (CRS) has iteratively been reported to be possibly implicated in the development of numerous cancer types. However, its role in oral squamous cell carcinoma (OSCC) has not been well elucidated. Here we intended to evaluate the role and mechanism. METHODS: The effects of CRS were investigated in xenograft models of OSCC by using transcriptome sequencing, LC-MS, ELISA and RT-PCR. Moreover, the role of CRS and ALDH3A1 on OSCC cells was researched by using Trans-well, flow cytometry, western blotting, immunofluorescence, ATP activity and OCR assay. Furthermore, immunohistochemical staining was employed to observe the cell proliferation and invasion of OSCC in xenotransplantation models. RESULTS: CRS promoted the progression of OSCC in xenograft models, stimulated the secretion of norepinephrine and the expression of ADRB2, but decreased the expression of ALDH3A1. Moreover, CRS changed energy metabolism and increased mitochondrial metabolism markers. However, ALDH3A1 overexpression suppressed proliferation, EMT and mitochondrial metabolism of OSCC cells. CONCLUSION: Inhibition of ALDH3A1 expression plays a pivotal role in CRS promoting tumorigenic potential of OSCC cells, and the regulatory of ALDH3A1 on mitochondrial metabolism may be involved in this process.

Transcriptomics and molecular docking reveal the potential mechanism of lycorine against pancreatic cancer.

BACKGROUND: Pancreatic cancer is an extremely malignant digestive tumor, however, owing to its high drug resistance of pancreatic cancer, the search for more effective anti-pancreatic cancer drugs is urgently needed. Lycorine, an alkaloid of natural plant origin, exerts antitumor effects on a variety of tumors. PURPOSE: This study aimed to investigate the therapeutic effect of lycorine on pancreatic cancer and elucidate its potential molecular mechanism. METHODS: Two pancreatic cancer cell lines, PANC-1 and BxPC-3, were used to investigate the therapeutic effects of lycorine on pancreatic cancer in vitro using the CCK8 assay, colony formation assay, 5-Ethynyl-2'- deoxyuridine (EdU) incorporation assay, flow cytometry, and western blotting. Transcriptome sequencing and gene set enrichment analysis (GSEA) were used to analyze the differentially expressed genes and pathways after lycorine treatment. Molecular docking, quantitative real-time PCR (qRT-PCR), oil red O staining, small interfering RNA (siRNA) transfection, and other experiments were performed to further validate the differentially expressed genes and pathways. In vivo experiments were conducted to investigate lycorine's inhibitory effects and toxicity on pancreatic cancer using a tumor-bearing mouse model. RESULTS: Lycorine inhibited the proliferation of pancreatic cancer cells, caused G2/M phase cycle arrest and induced apoptosis. Transcriptome sequencing and GSEA showed that lycorine inhibition of pancreatic cancer was associated with fatty acid metabolism, and aldehyde dehydrogenase 3A1 (ALDH3A1) was a significantly enriched target in the fatty acid metabolism process. ALDH3A1 expression was significantly upregulated in pancreatic cancer and was closely associated with prognosis. Molecular docking showed that lycorine binds strongly to ALDH3A1. Further studies revealed that lycorine inhibited the fatty acid oxidation (FAO) process in pancreatic cancer cells and induced cell growth inhibition and apoptosis through ALDH3A1. Lycorine also showed significant suppressive effects in tumor-bearing mice. Importantly, it did not result in significant toxicity to liver and kidney of mice, demonstrating its therapeutic potential as a safe antitumor agent. CONCLUSION: Lycorine inhibited pancreatic cancer cell proliferation, blocked the cell cycle, and induced apoptosis by targeting ALDH3A1. FAO inhibition was identified for the first time as a possible mechanism for the anticancer effects of lycorine. These findings enrich the theory of targeted therapy for pancreatic cancer, expand our understanding of the pharmacological targets of lycorine, and provide a reference for exploring its natural components.

Identification of genetic variants in two families with Keratoconus.

BACKGROUND: This research investigated the genetic characteristic of two Chinese families with keratoconus (KC). METHODS: For all people in the two families with KC, their history, clinical data, and peripheral blood were collected. One hundred healthy participants without KC and 112 sporadic KC patients were recruited as the controls. Whole exome sequencing of the genomic DNA and polymerase chain reaction were conducted for all the controls and family members to verify the variants. Functional analyses of the variants was performed using the software programs. RESULTS: A missense tuberous sclerosis 1 (TSC1) variant g.135797247A > G (c.622A > G, p.Ser208Gly) was detected in family 1. A single nucleotide polymorphism (SNP) rs761232139 (p.Gly235Arg) in aldehyde dehydrogenase 3 family member A1 (ALDH3A1) gene was detected in family 2. The variant c.622A > G in TSC1 and the SNP rs761232139 in ALDH3A1 were predicted as being probably damaging. CONCLUSIONS: Novel variant c.622A > G in TSC1 and SNP rs761232139 in ALDH3A1 have been detected in families with KC. These two findings may play a role in the pathogenesis of KC.

BNTA attenuates temporomandibular joint osteoarthritis progression by directly targeting ALDH3A1: An in vivo and in vitro study.

BNTA is known to have a therapeutic effect on knee osteoarthritis and inflammatory osteoclastogenesis. However, the protective effect of BNTA regarding temporomandibular mandibular joint osteoarthritis (TMJOA) and its underlying mechanism and physiological target remains unclear. In the present study, BNTA ameliorated cartilage degradation and inflammation responses in monosodium iodoacetate (MIA)-induced TMJOA in vivo. In IL-1ß-induced condylar chondrocytes, BNTA prevents oxidative stress, inflammatory responses and increasing synthesis of cartilage extracellular matrix through activating nuclear factor-E2-related factor 2 (NRF2) signaling. Suppression of NRF2 signaling abolishes the protective effect of BNTA in TMJOA. Notably, BNTA may bind directly to ALDH3A1 and act as a stabilizer, as evidenced by drug affinity responsive target stability assay (DARTS), cellular thermal shift assay (CETSA) and molecular docking results. Further investigation of the underlying molecular and cellular mechanism infers a positive correlation of ALDH3A1 regulating NRF2 signaling. In conclusion, BNTA may attenuate TMJOA progression via the ALDH3A1/NRF2 axis, inferring that BNTA is a therapeutic target for treating temporomandibular mandibular joint osteoarthritis.

Identification of a peptide ligand for human ALDH3A1 through peptide phage display: Prediction and characterization of protein interaction sites and inhibition of ALDH3A1 enzymatic activity.

Aldehyde dehydrogenase 3A1 (ALDH3A1) by oxidizing medium chain aldehydes to their corresponding carboxylic acids, is involved in the detoxification of toxic byproducts and is considered to play an important role in antioxidant cellular defense. ALDH3A1 has been implicated in various other functions such as cell proliferation, cell cycle regulation, and DNA damage response. Recently, it has been identified as a putative biomarker of prostate, gastric, and lung cancer stem cell phenotype. Although ALDH3A1 has multifaceted functions in both normal and cancer homeostasis, its modes of action are currently unknown. To this end, we utilized a random 12-mer peptide phage display library to identify efficiently human ALDH3A1-interacting peptides. One prevailing peptide (P1) was systematically demonstrated to interact with the protein of interest, which was further validated in vitro by peptide ELISA. Bioinformatic analysis indicated two putative P1 binding sites on the protein surface implying biomedical potential and potent inhibitory activity of the P1 peptide on hALDH3A1 activity was demonstrated by enzymatic studies. Furthermore, in search of potential hALDH3A1 interacting players, a BLASTp search demonstrated that no protein in the database includes the full-length amino acid sequence of P1, but identified a list of proteins containing parts of the P1 sequence, which may prove potential hALDH3A1 interacting partners. Among them, Protein Kinase C Binding Protein 1 and General Transcription Factor II-I are candidates of high interest due to their cellular localization and function. To conclude, this study identifies a novel peptide with potential biomedical applications and further suggests a list of protein candidates be explored as possible hALDH3A1-interacting partners in future studies.

ALDH3A1 overexpression in OSCC inhibits inflammation via phospho-Ser727 at STAT3 in tumor-associated macrophages.

OBJECTIVES: Cancer-related inflammation (CRI) significantly increases the difficulty of treating oral squamous cell carcinoma (OSCC) and remains a major treatment challenge. Our objective was to determine whether tumor ALDH3A1 could attenuate OSCC tumorigenesis by inhibiting tumor-associated macrophages (TAMs) that promoted CRI. MATERIALS AND METHODS: ALDH3A1 in Cal27 cells was overexpressed, and the tumor-conditioned medium (TCM) was collected. We induced THP-1 cells with TCM and recombinant human IL-6. The phosphorylation of STAT3 and the TLR4/TRAF6/TBK1 cascade reaction in TAMs was analyzed using Western blotting, and mitochondrial ROS (mtROS) production was measured using a MitoSox kit. A tumorigenicity assay was performed to examine the tumor volume and weight, and the expression of CD68, CD11b, IL-6, Ki67, and CD31 was analyzed via immunohistochemistry. RESULTS: ALDH3A1 attenuated STAT3 phosphorylation at Ser727 rapidly and mtROS production earlier in TAMs via inhibiting TLR4/TRAF6/TBK1 cascade reaction. MtROS reduction inhibited IL-1ß and IL-8 secretions by NLRP3/caspase-1/IL-1ß/IL-8 pathway. Meanwhile, the inhibition of pro-tumor phenotypes of TAMs, tumor proliferation, and tumor angiogenesis during the process was proved in vivo. CONCLUSION: ALDH3A1 was associated closely with CRI and inhibited CRI regulated by TAMs. This finding may achieve clinical transformation and open new therapeutic options for targeting CRI regulated by TAMs.

Profiling of Aldehyde Dehydrogenase Isoforms in In Vitro Formed Tumorspheres.

BACKGROUND/AIM: Aldehyde dehydrogenases (ALDHs) are considered as markers for normal and cancer stem cells (CSC) and are involved in cell metabolism, proliferation, differentiation, stemness, and retinoic acid (RA) biosynthesis. The aim of the present study was to identify the ALDH isoforms that are associated with the CSC phenotype in non-small cell lung and hepatocellular carcinomas. MATERIALS AND METHODS: We utilized lung (A549) and hepatocellular (HepG2) cancer cells and generated tumor spheres to isolate the CSC sub-population. RESULTS: The CSC enrichment was confirmed by the up-regulation of various CSC-related genes. Comparative qPCR analysis indicated the up-regulation of several ALDH isoforms in A549 and HepG2 spheres. Interestingly, cyclin D1 and Akt, down-stream targets of the RA signaling pathway, were also shown to be significantly up-regulated in both sphere populations. CONCLUSION: Specific ALDH isoforms appear to be important mediators for the acquisition of an CSC phenotype and thus, are potential promising targets for CSC-based therapeutic approaches in lung and hepatocellular carcinomas.

Design, Synthesis, Biological Evaluation and In Silico Study of Benzyloxybenzaldehyde Derivatives as Selective ALDH1A3 Inhibitors.

Aldehyde dehydrogenase 1A3 (ALDH1A3) has recently gained attention from researchers in the cancer field. Several studies have reported ALDH1A3 overexpression in different cancer types, which has been found to correlate with poor treatment recovery. Therefore, finding selective inhibitors against ALDH1A3 could result in new treatment options for cancer treatment. In this study, ALDH1A3-selective candidates were designed based on the physiological substrate resemblance, synthesized and investigated for ALDH1A1, ALDH1A3 and ALDH3A1 selectivity and cytotoxicity using ALDH-positive A549 and ALDH-negative H1299 cells. Two compounds (ABMM-15 and ABMM-16), with a benzyloxybenzaldehyde scaffold, were found to be the most potent and selective inhibitors for ALDH1A3, with IC50 values of 0.23 and 1.29 µM, respectively. The results also show no significant cytotoxicity for ABMM-15 and ABMM-16 on either cell line. However, a few other candidates (ABMM-6, ABMM-24, ABMM-32) showed considerable cytotoxicity on H1299 cells, when compared to A549 cells, with IC50 values of 14.0, 13.7 and 13.0 µM, respectively. The computational study supported the experimental results and suggested a good binding for ABMM-15 and ABMM-16 to the ALDH1A3 isoform. From the obtained results, it can be concluded that benzyloxybenzaldehyde might be considered a promising scaffold for further drug discovery aimed at exploiting ALDH1A3 for therapeutic intervention.

ALDH3A1 driving tumor metastasis is mediated by p53/BAG1 in lung adenocarcinoma.

Lung adenocarcinoma (LUAD) is a lethal malignancy with metastasis, a major tumor feature that predominantly correlated with progression, but the molecules that mediated tumor metastasis remain elusive. To declare the critical regulatory genes, RNA sequencing data in LUAD patients was acquired from The Cancer Genome Atlas (TCGA) and found that ALDH3A1 was distinctly highly expressed in LUAD patients with metastasis (M1) compared with those without metastasis (M0), linked to the property of cancer stem cell and epithelial-mesenchymal transition (EMT). Besides, high ALDH3A1 expression predicted a poor prognosis. Knockdown of ALDH3A1 showed decreased proliferation, migration, and invasion in A549 cell line. Furthermore, BAG1 was regulated by ALDH3A1 through p53, enhanced cell proliferation, and predicted clinical prognosis. Our findings collectively uncovered a novel mechanism that orchestrates tumor cells' metastasis, and decreasing ALDH3A1 represented a potential therapeutic target for reprogramming metastasis.

Association with Corneal Remodeling Related Genes, ALDH3A1, LOX, and SPARC Genes Variations in Korean Keratoconus Patients.

PURPOSE: To determine whether the cornea remodeling-related genes aldehyde dehydrogenase 3A1 (ALDH3A1), lysyl oxidase (LOX), and secreted protein acidic and rich in cysteine (SPARC) were potential susceptibility candidate genes for keratoconus in Korean patients, we investigated the associations of single nucleotide polymorphisms (SNPs) in these three genes in Korean patients with keratoconus. METHODS: Genomic DNA was extracted from blood samples of unrelated patients with keratoconus and healthy control individuals. For screening of genetic variations, all exons from the entire coding regions of the ALDH3A1, LOX, and SPARC genes were directly sequenced to determine the presence of mutations. Control individuals were selected from the general population without keratoconus. RESULTS: In this study, we detected nine SNPs in ALDH3A1, four SNPs in LOX, and 18 SNPs in SPARC. rs116992290, IVS3-62c>t, rs116962241, and rs2228100 in ALDH3A1 and rs2956540 and rs1800449 in LOX were significantly different between patient and control groups. In the SPARC gene, the distribution of the *G allele of EX10+225 T>G (p = 0.018; odds ratio, 1.869) was strongly associated with the risk of keratoconus in the Korean population. In haplotype analysis, C-G of rs2956540-rs2288393 in LOX(p = 0.046) and C-C-G and G-G-G of rs60610024-rs2228100-rs57555435 (p = 0.021 and p < 0.001), G-A of IVS3-62 a>g - rs116962241 in ALDH3A1(p = 0.048) predisposed significantly to keratoconus. After cross-validation consistency and permutation tests, two locus model was the best SNP variations interaction pattern. CONCLUSIONS: Our results suggested that genetic variations in ALDH3A1, LOX, and SPARC genes were associated with a predisposition for keratoconus in Korean individuals. Moreover, variations in ALDH3A1 and LOX may serve as strong biomarkers for keratoconus.

Shikonin enhances the antitumor effect of cabazitaxel in prostate cancer stem cells and reverses cabazitaxel resistance by inhibiting ABCG2 and ALDH3A1.

Cancer stem cells (CSCs) are a small population among cancer cells, defined as capable of self-renewal, and driving tumor growth, metastasis, and therapeutic relapse. The development of therapeutic strategies to target CSCs is of great importance to prevent tumor metastasis and relapse. Increasing evidence shows that shikonin has inhibiting effects on CSCs. This study was to determine the effect of shikonin on prostate CSCs, and on drug resistant cells. Sphere formation assay was used to enrich prostate CSCs. The effect of shikonin on viability, proliferation, migration, and invasion was studied. Typical CSCs markers were analyzed by flow cytometry and RT-qPCR. The cytotoxic mechanism of shikonin was analyzed by staining for annexin V, reactive oxygen species (ROS) and mitochondrial membrane potential. To study the effect of shikonin on drug-resistant cells a cabazitaxel resistant cell line was established. Shikonin inhibited the viability, proliferation, migration, and invasion of prostate CSCs. Shikonin enhanced the antitumor effect of cabazitaxel, which is a second-line chemotherapeutic drug in advanced prostate cancer. Shikonin induced apoptosis through generating ROS and disrupting the mitochondrial membrane potential. Furthermore, shikonin suppressed the expression of ALDH3A1 and ABCG2 in prostate CSCs, which are two markers related to drug-resistance. When inhibiting the expression of ABCG2 and ALDH3A1, the cabazitaxel resistant cells acquired more sensibility to cabazitaxel. Shikonin enhances the cytotoxic activity of cabazitaxel in prostate CSCs and reverses the cabazitaxel-resistant state.

Elevated 4-hydroxynonenal induces hyperglycaemia via Aldh3a1 loss in zebrafish and associates with diabetes progression in humans.

Increased methylglyoxal (MG) formation is associated with diabetes and its complications. In zebrafish, knockout of the main MG detoxifying system Glyoxalase 1, led to limited MG elevation but significantly elevated aldehyde dehydrogenases (ALDH) activity and aldh3a1 expression, suggesting the compensatory role of Aldh3a1 in diabetes. To evaluate the function of Aldh3a1 in glucose homeostasis and diabetes, aldh3a1-/- zebrafish mutants were generated using CRISPR-Cas9. Vasculature and pancreas morphology were analysed by zebrafish transgenic reporter lines. Corresponding reactive carbonyl species (RCS), glucose, transcriptome and metabolomics screenings were performed and ALDH activity was measured for further verification. Aldh3a1-/- zebrafish larvae displayed retinal vasodilatory alterations, impaired glucose homeostasis, which can be aggravated via pdx1 silencing induced hyperglycaemia. Unexpectedly, MG was not altered, but 4-hydroxynonenal (4-HNE), another prominent lipid peroxidation RCS exhibited high affinity with Aldh3a1, was increased in aldh3a1 mutants. 4-HNE was responsible for the retinal phenotype via pancreas disruption induced hyperglycaemia and can be rescued via l-Carnosine treatment. Furthermore, in type 2 diabetic patients, serum 4-HNE was increased and correlated with disease progression. Thus, our data suggest impaired 4-HNE detoxification and elevated 4-HNE concentration as biomarkers but also the possible inducers for diabetes, from genetic susceptibility to the pathological progression.

ALDH3A1 acts as a prognostic biomarker and inhibits the epithelial mesenchymal transition of oral squamous cell carcinoma through IL-6/STAT3 signaling pathway.

Objectives: Aldehyde dehydrogenase 3A1 (ALDH3A1) is a member of the ALDH superfamily and its relationship with oral squamous cell carcinoma (OSCC) still unknown. In our subject, we aimed to reveal the expression pattern and clinical value of ALDH3A1 in OSCC and its biological function in OSCC cell lines. Materials and methods: The expression level of ALDH3A1 in paired OSCC tissues and adjacent noncancerous tissues were detected by quantitative real-time PCR, Western blot and immunohistochemistry. The relationship between ALDH3A1 expression and clinical characteristics was analyzed. Besides, cell-counting kit 8, colony formation, wound healing, transwell invasion, apoptosis and cell cycle assays were employed to assess the role of ALDH3A1 in OSCC cells. To explore the influence of ALDH3A1 on OSCC epithelial-to-mesenchymal transition (EMT), the expression of EMT markers (E-cadherin, vimentin, snail, MMP3) on OSCC cells were detected, and possible mechanisms were analyzed. Results: In OSCC tissues, ALDH3A1 was significantly decreased compared to the adjacent normal tissues. Lower ALDH3A1 expression in OSCC tissues was associated with a higher incidence of lymph node metastasis (LNM). Moreover, the overall survival of OSCC with low ALDH3A1 expression was significantly worse compared to that of OSCC with high ALDH3A1 expression. Restored expression of ALDH3A1 suppressed cell proliferation, migration and invasion in OSCC cells. Further experiments showed that ALDH3A1 might inhibit EMT in OSCC via a regulation of the IL-6/STAT3 signal pathway. Conclusion: These data indicate that ALDH3A1 may serve as a biomarker and may be developed into a novel treatment for OSCC.

Aldehyde dehydrogenase 3A1 confers oxidative stress resistance accompanied by altered DNA damage response in human corneal epithelial cells.

Aldehyde dehydrogenase 3A1 is constitutively expressed in a taxon-specific manner in the cornea, where, due to its high abundance, it has been characterized as a corneal crystallin. ALDH3A1 has been proposed to be a multifaceted protein that protects cellular homeostasis through several modes of action. The present study examines the mechanisms by which ALDH3A1 exerts its cytoprotective role under conditions of oxidative stress. To this end, we have utilized an isogenic HCE-2 (human corneal epithelium) cell line pair differing in the expression of ALDH3A1. Single cell gel electrophoresis assay and H2DCFDA analysis revealed that the expression of ALDH3A1 protected HCE-2 cells from H2O2-, tert-butyl peroxide- and etoposide-induced oxidative and genotoxic effects. Furthermore, comparative qPCR analysis revealed that a panel of cell cycle (Cyclins B1, B2, D, E), apoptosis (p53, BAX, BCL-2, BCL-XL) and DNA damage response (DNA-PK, NBS1) genes were up-regulated in the ALDH3A1 expressing HCE-2 cells. Moreover, the expression profile of a variety of DNA damage signaling (DDS)-related genes, was investigated (under normal and oxidative stress conditions) by utilizing the RT2 profiler™ PCR array in both isogenic HCE-2 cell lines. Our results demonstrated that several genes associated with ATM/ATR signaling, cell cycle regulation, apoptosis and DNA damage repair were differentially expressed under all conditions tested. In conclusion, this study suggests that ALDH3A1 significantly contributes to the antioxidant defense of corneal homeostasis by maintaining DNA integrity possibly through altering the expression of specific DDS-related genes. Further studies will shed light on the precise role(s) of this multifunctional protein.

Linking endotypes to omics profiles in difficult-to-control asthma using the diagnostic Chinese medicine syndrome differentiation algorithm.

Objective: Patients with difficult-to-control asthma have difficulty breathing almost all of the time, even leading to life-threatening asthma attacks. However, only few diagnostic markers for this disease have been identified. We aimed to take advantage of unique Chinese medicine theories for phenotypic classification and to explore molecular signatures in difficult-to-control asthma. Methods: The Chinese medicine syndrome differentiation algorithm (CMSDA) is a syndrome-scoring classification method based on the Chinese medicine overall observation theory. Patients with difficult-to-control asthma were classified into Cold- and Hot-pattern groups according to the CMSDA. DNA methylation and metabolomic profiles were obtained using Infinium Human Methylation 450 BeadChip and gas chromatography-mass spectrometer. Subsequently, an integrated bioinformatics analysis was performed to compare those two patterns and identify Cold/Hot-associated candidates, followed by functional validation studies. Results: A total of 20 patients with difficult-to-control asthma were enrolled in the study. Ten were grouped as Cold and 10 as Hot according to the CMSDA. We identified distinct whole-genome DNA methylation and metabolomic profiles between Cold- and Hot-pattern groups. ALDH3A1 gene exhibited variations in the DNA methylation probe cg10791966, while two metabolic pathways were associated with those two patterns. Conclusions: Our study introduced a novel diagnostic classification approach, the CMSDA, for difficult-to-control asthma. This is an alternative way to categorize diverse syndromes and link endotypes with omics profiles of this disease. ALDH3A1 might be a potential biomarker for precision diagnosis of difficult-to-control asthma.

ALDH3A1 Overexpression in Melanoma and Lung Tumors Drives Cancer Stem Cell Expansion, Impairing Immune Surveillance through Enhanced PD-L1 Output.

Melanoma and non-small-cell lung carcinoma (NSCLC) cell lines are characterized by an intrinsic population of cancer stem-like cells (CSC), and high expression of detoxifying isozymes, the aldehyde dehydrogenases (ALDHs), regulating the redox state. In this study, using melanoma and NSCLC cells, we demonstrate that ALDH3A1 isozyme overexpression and activity is closely associated with a highly aggressive mesenchymal and immunosuppressive profile. The contribution of ALDH3A1 to the stemness and immunogenic status of melanoma and NSCLC cells was evaluated by their ability to grow in 3D forming tumorspheres, and by the expression of markers for stemness, epithelial to mesenchymal transition (EMT), and inflammation. Furthermore, in specimens from melanoma and NSCLC patients, we investigated the expression of ALDH3A1, PD-L1, and cyclooxygenase-2 (COX-2) by immunohistochemistry. We show that cells engineered to overexpress the ALDH3A1 enzyme enriched the CSCs population in melanoma and NSCLC cultures, changing their transcriptome. In fact, we found increased expression of EMT markers, such as vimentin, fibronectin, and Zeb1, and of pro-inflammatory and immunosuppressive mediators, such as NFkB, prostaglandin E2, and interleukin-6 and -13. ALDH3A1 overexpression enhanced PD-L1 output in tumor cells and resulted in reduced proliferation of peripheral blood mononuclear cells when co-cultured with tumor cells. Furthermore, in tumor specimens from melanoma and NSCLC patients, ALDH3A1 expression was invariably correlated with PD-L1 and the pro-inflammatory marker COX-2. These findings link ALDH3A1 expression to tumor stemness, EMT and PD-L1 expression, and suggest that aldehyde detoxification is a redox metabolic pathway that tunes the immunological output of tumors.

Severe and prolonged cyclophosphamide-induced hepatotoxicity in a breast cancer patient carrying a CYP2B6*7 variant.

As a widely used alkylating agent, cyclophosphamide often leads to various toxicities. Severe hepatotoxicity has been rarely reported in breast cancer patients receiving chemotherapy containing cyclophosphamide. Differences in cyclophosphamide metabolism may contribute to variability in adverse events of patients. Here, we report on a case of a 68-year-old Chinese female with breast cancer who experienced severe and prolonged hepatotoxicity induced by cyclophosphamide. Pharmacogenetic tests showed that she was a carrier of CYP2B6*7 allele and this is the first case of a CYP2B6*7 variant in the Han Chinese population so far reported. In addition, the patient was also a carrier of an ALDH3A1*2 variant potentially contributing to the occurrence of hepatotoxicity. CYP2B6 and ALDH3A1 genotyping may play a role in guiding cyclophosphamide therapy.

Using an in-vivo syngeneic spontaneous metastasis model identifies ID2 as a promoter of breast cancer colonisation in the brain.

BACKGROUND: Dissemination of breast cancers to the brain is associated with poor patient outcome and limited therapeutic options. In this study we sought to identify novel regulators of brain metastasis by profiling mouse mammary carcinoma cells spontaneously metastasising from the primary tumour in an immunocompetent syngeneic host. METHODS: 4T1 mouse mammary carcinoma sublines derived from primary tumours and spontaneous brain and lung metastases in BALB/c mice were subject to genome-wide expression profiling. Two differentially expressed genes, Id2 and Aldh3a1, were validated in in-vivo models using mouse and human cancer cell lines. Clinical relevance was investigated in datasets of breast cancer patients with regards to distant metastasis-free survival and brain metastasis relapse-free survival. The role of bone morphogenetic protein (BMP)7 in regulating Id2 expression and promoting cell survival was investigated in two-dimensional and three-dimensional in-vitro assays. RESULTS: In the spontaneous metastasis model, expression of Id2 and Aldh3a1 was significantly higher in 4T1 brain-derived sublines compared with sublines from lung metastases or primary tumour. Downregulation of expression impairs the ability of cells to colonise the brain parenchyma whereas ectopic expression in 4T1 and human MDA-MB-231 cells promotes dissemination to the brain following intracardiac inoculation but has no impact on the efficiency of lung colonisation. Both genes are highly expressed in oestrogen receptor (ER)-negative breast cancers and, within this poor prognosis sub-group, increased expression correlates with reduced distant metastasis-free survival. ID2 expression also associates with reduced brain metastasis relapse-free survival. Mechanistically, BMP7, which is present at significantly higher levels in brain tissue compared with the lungs, upregulates ID2 expression and, after BMP7 withdrawal, this elevated expression is retained. Finally, we demonstrate that either ectopic expression of ID2 or BMP7-induced ID2 expression protects tumour cells from anoikis. CONCLUSIONS: This study identifies ID2 as a key regulator of breast cancer metastasis to the brain. Our data support a model in which breast cancer cells that have disseminated to the brain upregulate ID2 expression in response to astrocyte-secreted BMP7 and this serves to support metastatic expansion. Moreover, elevated ID2 expression identifies breast cancer patients at increased risk of developing metastatic relapse in the brain.

Mutation of the gene encoding the circadian clock component PERIOD2 in oncogenic cells confers chemoresistance by up-regulating the Aldh3a1 gene.

Disruption of circadian rhythms has been implicated in an increased risk for cancer development. The Period2 (Per2) gene encodes one of the major components of the mammalian circadian clock, which plays a key role in controlling the circadian rhythms in physiology and behavior. PER2 has also been reported to suppress the malignant transformation of cells, but its role in the regulation of cancer susceptibility to chemotherapeutic drugs remains unclear. In this study, we found that oncogene-transformed embryonic fibroblasts prepared from Per2-mutant (Per2m/m ) mice, which are susceptible to both spontaneous and radiation-induced tumorigenesis, were resistant against common chemotherapeutic drugs and that this resistance is associated with up-regulation of the aldehyde dehydrogenase 3a1 (Aldh3a1) gene. Co-expression of the oncogenes H-rasV12 and SV40 large T-antigen induced malignant transformation of both WT and Per2m/m cells, but the cytotoxic effects of the chemotherapeutic agents methotrexate, gemcitabine, etoposide, vincristine, and oxaliplatin were significantly alleviated in the oncogene-transformed Per2m/m cells. Although introduction of the two oncogenes increased the expression of Aldh3a1 in both WT and Per2m/m cells, the ALDH3A1 protein levels in the Per2m/m cells were ∼7-fold higher than in WT cells. The elevated ALDH3A1 levels in the oncogene-transformed Per2m/m cells were sufficient to prevent chemotherapeutic drug-induced accumulation of reactive oxygen species. Consequently, shRNA-mediated suppression of Aldh3a1 expression relieved the chemoresistance of the Per2m/m cells. These results suggest a role for mutated PER2 in the development of multiple drug resistance and may inform therapeutic strategies for cancer management.