ALDH and cancer stem cells: Pathways, challenges, and future directions in targeted therapy.

Human ALDH comprise 19 subfamilies in which ALDH1A1, ALDH1A3, ALDH3A1, ALDH5A1, ALDH7A1, and ALDH18A1 are implicated in CSC. Studies have shown that ALDH can also be involved in drug resistance and standard chemotherapy regimens are ineffective in treating patients at the stage of disease recurrence. Existing chemotherapeutic drugs eliminate the bulk of tumors but are usually not effective against CSC which express ALDH+ population. Henceforth, targeting ALDH is convincing to treat the patient's post-relapse. Combination therapies that interlink signaling mechanisms seem promising to increase the overall disease-free survival rate. Therefore, targeting ALDH through ALDH inhibitors along with immunotherapies may create a novel platform for translational research. This review aims to fill in the gap between ALDH1 family members in relation to its cell signaling mechanisms, highlighting their potential as molecular targets to sensitize recurrent tumors and bring forward the future development concerning the current progress and draw backs. This review summarizes the role of cancer stem cells and their upregulation by maintaining the tumor microenvironment in which ALDH is specifically highlighted. It discusses the regulation of ALDH family proteins and the crosstalk between ALDH and CSC in relation to cancer metabolism. Furthermore, it establishes the correlation between ALDH involved signaling mechanisms and their specific targeted inhibitors, as well as their functional modularity, bioavailability, and mechanistic role in various cancers.

Targeting the retinoic acid signaling pathway as a modern precision therapy against cancers.

Retinoic acid (RA) is a vital metabolite derived from vitamin A. RA plays a prominent role during development, which helps in embryological advancement and cellular differentiation. Mechanistically, RA binds to its definite nuclear receptors including the retinoic acid receptor and retinoid X receptor, thus triggering gene transcription and further consequences in gene regulation. This functional heterodimer activation later results in gene activation/inactivation. Several reports have been published related to the detailed embryonic and developmental role of retinoic acids and as an anti-cancer drug for specific cancers, including acute promyelocytic leukemia, breast cancer, and prostate cancer. Nonetheless, the other side of all-trans retinoic acid (ATRA) has not been explored widely yet. In this review, we focused on the role of the RA pathway and its downstream gene activation in relation to cancer progression. Furthermore, we explored the ways of targeting the retinoic acid pathway by focusing on the dual role of aldehyde dehydrogenase (ALDH) family enzymes. Combination strategies by combining RA targets with ALDH-specific targets make the tumor cells sensitive to the treatment and improve the progression-free survival of the patients. In addition to the genomic effects of ATRA, we also highlighted the role of ATRA in non-canonical mechanisms as an immune checkpoint inhibitor, thus targeting the immune oncological perspective of cancer treatments in the current era. The role of ATRA in activating independent mechanisms is also explained in this review. This review also highlights the current clinical trials of ATRA in combination with other chemotherapeutic drugs and explains the future directional insights related to ATRA usage.

ALDH1A1 promotes PARP inhibitor resistance by enhancing retinoic acid receptor-mediated DNA polymerase θ expression.

Poly (ADP-ribose) Polymerase (PARP) inhibitors (PARPi) have been approved for both frontline and recurrent setting in ovarian cancer with homologous recombination (HR) repair deficiency. However, more than 40% of BRCA1/2-mutated ovarian cancer lack the initial response to PARPi treatment, and the majority of those that initially respond eventually develop resistance. Our previous study has demonstrated that increased expression of aldehyde dehydrogenase 1A1 (ALDH1A1) contributes to PARPi resistance in BRCA2-mutated ovarian cancer cells by enhancing microhomology-mediated end joining (MMEJ) but the mechanism remains unknown. Here, we find that ALDH1A1 enhances the expression of DNA polymerase θ (Polθ, encoded by the POLQ gene) in ovarian cancer cells. Furthermore, we demonstrate that the retinoic acid (RA) pathway is involved in the transcription activation of the POLQ gene. The RA receptor (RAR) can bind to the retinoic acid response element (RARE) located in the promoter of the POLQ gene, promoting transcription activation-related histone modification in the presence of RA. Given that ALDH1A1 catalyzes the biosynthesis of RA, we conclude that ALDH1A1 promotes POLQ expression via the activation of the RA signaling pathway. Finally, using a clinically-relevant patient-derived organoid (PDO) model, we find that ALDH1A1 inhibition by the pharmacological inhibitor NCT-505 in combination with the PARP inhibitor olaparib synergistically reduce the cell viability of PDOs carrying BRCA1/2 mutation and positive ALDH1A1 expression. In summary, our study elucidates a new mechanism contributing to PARPi resistance in HR-deficient ovarian cancer and shows the therapeutic potential of combining PARPi and ALDH1A1 inhibition in treating these patients.

ERK inactivation enhances stemness of NSCLC cells via promoting Slug-mediated epithelial-to-mesenchymal transition.

Rationale: The mitogen-activated protein kinase pathway (MAPK) is one of the major cancer-driving pathways found in non-small cell lung cancer (NSCLC) patients. ERK inhibitors (ERKi) have been shown to be effective in NSCLC patients with MAPK pathway mutations. However, like other MAPK inhibitors, ERKi rarely confers complete and durable responses. The mechanism of tumor relapse after ERKi treatment is yet defined. Methods: To best study the mechanism of tumor relapse after ERK inhibitor treatment in NSCLC patients, we treated various NSCLC cell lines and patient-derived xenograft (PDX) with ERK inhibitors and evaluated the enrichment of cancer stem cell (CSC) population. We then performed a Next-generation sequencing (NGS) to identify potential pathways that are responsible for the CSC enrichment. Further, the involvement of specific pathways was examined using molecular and cellular methods. Finally, we investigated the therapeutic benefits of ERKi treatment combined with JAK/STAT pathway inhibitor using cellular and xenograft NSCLC models. Results: We found that ERKi treatment expands the CSC population in NSCLC cells through enhanced epithelial-to-mesenchymal transition (EMT)-mediated cancer cell dedifferentiation. Mechanistically, ERK inactivation induces EMT via pSTAT3-mediated upregulation of Slug, in which, upregulation of miR-204 and downregulation of SPDEF, a transcription repressor of Slug, are involved. Finally, the JAK/STAT pathway inhibitor Ruxolitinib blocks the ERK inactivation-induced EMT and CSC expansion, as well as the tumor progression in xenograft models after ERKi treatment. Conclusions: This study revealed a potential tumor relapse mechanism of NSCLC after ERK inhibition through the unintended activation of the EMT program, ascertained the pSTAT-miR-204-SPDEF-Slug axis, and provided a promising combination inhibitor approach to prevent tumor relapse in patients.

2-Dimensional in vitro culture assessment of ovarian cancer cell line using cost effective silver nanoparticles from Macrotyloma uniflorum seed extracts.

Our research focused on generating AgNPs using Macrotyloma uniflorum (MU) seed extracts and studied their efficacy in combating tumor growth using the 2-Dimensional method for ovarian cancer cell line-PA-1. Characterization studies including a UV-visible spectrophotometer confirmed the surface plasmon resonance peak of 436 nm. Particle size determination data validated the nanoparticle diameter of 91.8 nm. Synthesized AgNPs possess a negative charge of -28.0 mV, which was confirmed through the zeta potential study. Structural characterization studies including XRD determined the crystal phase of AgNPs at four distant peaks at 2θ (38.17, 44.36, 64.52, and 77.46) and were assigned to 111, 200, 220, and 311 planes of the FCC. FTIR studies have confirmed the presence of O-H, N-H, C=O, ethers, C-Br, and C-I groups in AgNPs respectively. DPPH study has confirmed the presence of free radicles and we observed that at 500 µg/ml concentration, 76.08% of free radicles were formed which shows their efficiency. MTT assay shows the efficacy of MU-AgNPs in reducing the cell viability. At lower concentrations of MU-AgNP, 66% viability was observed and 9% of viability was observed at higher dose. ROS production (21%) was observed using MU-AgNPs with respect to 0.45% in controls, which affirms the capacity to induce DNA damage via apoptosis. Standard drug camptothecin generated 26% of ROS production which confirms higher potential of AgNPs in inducing DNA damage in tumor cells without causing lethality to the healthy cells. Further, the Fluorescence-activated cell sorting (FACS) study using a standard Caspase-3 marker confirms the generation of apoptotic bodies using two different concentrations of MU-AgNPs. At 40 µg, 64% of apoptotic cell death was observed, whereas, using 20 µg, 23% of apoptosis was recorded via fluorescent intensity. Propidium iodide-based Cell cycle study has shown a significant decrease in G0/G1 phase compared to control (88.8%), which further confirmed the apoptotic induction. Matrix metalloproteinases (MMP) studies using JC-1 dye, showed a significant increase in green fluorescence owing to lowered membrane potential, thus ensuring the breakdown of mitochondrial potential compared to untreated and standard drugs. With the obtained results, we are concluding that MU-AgNPs has a tremendous capacity to suppress the ovarian cancer cell proliferation in vitro by inducing DNA damage and apoptosis.

A Novel Estrogen Receptor ß Agonist Diminishes Ovarian Cancer Stem Cells via Suppressing the Epithelial-to-Mesenchymal Transition.

Epithelial ovarian cancer is the most lethal malignancy of the female reproductive tract. A healthy ovary expresses both Estrogen Receptor α (ERα) and ß (ERß). Given that ERα is generally considered to promote cell survival and proliferation, thereby, enhancing tumor growth, while ERß shows a protective effect against the development and progression of tumors, the activation of ERß by its agonists could be therapeutically beneficial for ovarian cancer. Here, we demonstrate that the activation of ERß using a newly developed ERß agonist, OSU-ERb-12, can impede ovarian cancer cell expansion and tumor growth in an ERα-independent manner. More interestingly, we found that OSU-ERb-12 also reduces the cancer stem cell (CSC) population in ovarian cancer by compromising non-CSC-to-CSC conversion. Mechanistically, we revealed that OSU-ERb-12 decreased the expression of Snail, a master regulator of the epithelial-to-mesenchymal transition (EMT), which is associated with de novo CSC generation. Given that ERα can mediate EMT and facilitate maintenance of the CSC subpopulation and that OSU-ERb-12 can block the transactivity of ERα, we conclude that OSU-ERb-12 reduces the CSC subpopulation by inhibiting EMT in an ERα-dependent manner. Taken together, our data indicate that the ERß agonist OSU-ERb-12 could be used to hinder tumor progression and limit the CSC subpopulation with the potential to prevent tumor relapse and metastasis in patients with ovarian cancer.

A bioinformatic approach of targeting SARS-CoV-2 replication by silencing a conserved alternative reserve of the orf8 gene using host miRNAs.

The causative agent of the COVID-19 pandemic, the SARS-CoV-2 virus has yielded multiple relevant mutations, many of which have branched into major variants. The Omicron variant has a huge similarity with the original viral strain (first COVID-19 strain from Wuhan). Among different genes, the highly variable orf8 gene is responsible for crucial host interactions and has undergone multiple mutations and indels. The sequence of the orf8 gene of the Omicron variant is, however, identical with the gene sequence of the wild type. orf8 modulates the host immunity making it easier for the virus to conceal itself and remain undetected. Variants seem to be deleting this gene without affecting the viral replication. While analyzing, we came across the conserved orf7a gene in the viral genome which exhibits a partial sequence homology as well as functional similarity with the SARS-CoV-2 orf8. Hence, we have proposed here in our hypothesis that, orf7a might be an alternative reserve of orf8 present in the virus which was compensating for the lost gene. A computational approach was adopted where we screened various miRNAs targeted against the orf8 gene. These miRNAs were then docked onto the orf8 mRNA sequences. The same set of miRNAs was then used to check for their binding affinity with the orf7a reference mRNA. Results showed that miRNAs targeting the orf8 had favorable shape complementarity and successfully docked with the orf7a gene as well. These findings provide a basis for developing new therapeutic approaches where both orf8 and orf7a can be targeted simultaneously.

Cytotoxic Potential of Biogenic Zinc Oxide Nanoparticles Synthesized From Swertia chirayita Leaf Extract on Colorectal Cancer Cells.

Chemotherapy side effects, medication resistance, and tumor metastasis impede the advancement of cancer treatments, resulting in a poor prognosis for cancer patients. In the last decade, nanoparticles (NPs) have emerged as a promising drug delivery system. Swertia chirayita has long been used as a treatment option to treat a variety of ailments. Zinc oxide nanoparticles (ZnO-NPs) were synthesized from ethanolic and methanolic extract of S. chirayita leaves. ZnO-NPs were characterized using UV-visible spectroscopy, Fourier transform infrared spectroscopy (FTIR), scanning electron Microscopy (SEM), high-resolution transmission electron microscopy (HRTEM), and X-ray diffraction (XRD). Its anti-cancer activities were analyzed using cytotoxicity assays [MTT assay and acridine orange (AO) staining] and quantitative real-time PCR (qRT-PCR) using colorectal cancer (CRC) cells (HCT-116 and Caco-2) and control cells (HEK-293). The ZnO-NPs synthesized from the ethanolic extract of S. chirayita have an average size of 24.67 nm, whereas those from methanolic extract have an average size of 22.95 nm with a spherical shape. MTT assay showed NPs' cytotoxic potential on cancer cells (HCT-116 and Caco-2) when compared to control cells (HEK-293). The IC50 values of ethanolic and methanolic extract ZnO-NPs for HCT-116, Caco-2, and HEK-293 were 34.356 ± 2.71 and 32.856 ± 2.99 µg/ml, 52.15 ± 8.23 and 63.1 ± 12.09 µg/ml, and 582.84 ± 5.26 and 615.35 ± 4.74 µg/ml, respectively. Acridine orange staining confirmed the ability of ZnO-NPs to induce apoptosis. qRT-PCR analysis revealed significantly enhanced expression of E-cadherin whereas a reduced expression of vimentin and CDK-1. Altogether, these results suggested anti-cancer properties of synthesized ZnO-NPs in CRC.