Downregulation of TRIB3 enhances the sensitivity of lung cancer cells to amino acid deprivation by suppressing AKT activation.

Tribbles pseudokinase 3 (TRIB3), a member of the mammalian Tribbles family, is implicated in multiple biological processes. This study aimed to investigate the biological functions of TRIB3 in lung cancer and its effect on amino acid-deprived lung cancer cells. TRIB3 mRNA expression was elevated in lung cancer tissues and cell lines compared to normal lung tissues and cells. TRIB3 knockdown markedly reduced the viability and proliferation of H1299 lung cancer cells. Deprivation of amino acids, particularly arginine, glutamine, lysine, or methionine, strongly increased TRIB3 expression via ATF4 activation in H1299 lung cancer cells. Knockdown of TRIB3 led to transcriptional downregulation of ATF4 and reduced AKT activation induced by amino acid deprivation, ultimately increasing the sensitivity of H1299 lung cancer cells to amino acid deprivation. Additionally, TRIB3 knockdown enhanced the sensitivity of H1299 cells to V-9302, a competitive antagonist of transmembrane glutamine flux. These results suggest that TRIB3 is a pro-survival regulator of cell viability in amino acid-deficient tumor microenvironments and a promising therapeutic target for lung cancer treatment.

Duloxetine enhances the sensitivity of non-small cell lung cancer cells to EGFR inhibitors by REDD1-induced mTORC1/S6K1 suppression.

Although epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) have been effective targeted therapies for non-small cell lung cancer (NSCLC), most advanced NSCLC inevitably develop resistance to these therapies. Combination therapies emerge as valuable approach to preventing, delaying, or overcoming disease progression. Duloxetine, an antidepressant known as a serotonin-noradrenaline reuptake inhibitor, is commonly prescribed for the treatment of chemotherapy-induced peripheral neuropathy. In the present study, we investigated the combined effects of duloxetine and EGFR-TKIs and their possible mechanism in NSCLC cells. Compared with either monotherapy, the combination of duloxetine and EGFR-TKIs leads to synergistic cell death. Mechanistically, duloxetine suppresses 70-kDa ribosomal protein S6 kinase 1 (p70S6K1) activity through mechanistic target of rapamycin complex 1 (mTORC1), and this effect is associated with the synergistic induction of cell death of duloxetine combined with EGFR-TKIs. More importantly, activating transcription factor 4 (ATF4)-induced regulated in development and DNA damage response 1 (REDD1) is responsible for the suppression of mTORC1/S6K1 activation. Additionally, we found that the combination effect was significantly attenuated in REDD1 knockout NSCLC cells. Taken together, our findings reveal that the ATF4/REDD1/mTORC1/S6K1 signaling axis, as a novel mechanism, is responsible for the synergistic therapeutic effect of duloxetine with EGFR-TKIs. These results suggest that combining EGFR-TKIs with duloxetine appears to be a promising way to improve EGFR-TKI efficacy against NSCLC.

Amino acid deprivation induces TXNIP expression by NRF2 downregulation.

Thioredoxin-interacting protein (TXNIP) is sensitive to oxidative stress and is involved in the pathogenesis of various metabolic, cardiovascular, and neurodegenerative disorders. Therefore, several studies have suggested that TXNIP is a promising therapeutic target for several diseases, particularly cancer and diabetes. However, the regulation of TXNIP expression under amino acid (AA)-restricted conditions is not well understood. In the present study, we demonstrated that TXNIP expression was promoted by the deprivation of AAs, especially arginine, glutamine, lysine, and methionine, in non-small cell lung cancer (NSCLC) cells. Interestingly, we determined that increased TXNIP expression induced by AA deprivation was associated with nuclear factor erythroid 2-related factor 2 (NRF2) downregulation, but not with activating transcription factor 4 (ATF4) activation. Furthermore, N-acetyl-l-cysteine (NAC), a scavenger of reactive oxygen species (ROS), suppressed TXNIP expression in NSCLC cells deprived of AA. Collectively, the induction of TXNIP expression by AA deprivation was mediated by ROS production, potentially through NRF2 downregulation. Our findings suggest that TXNIP expression may be associated with the redox homeostasis of AA metabolism and provide a possible rationale for a therapeutic strategy to treat cancer with AA restriction.

Identification of a Splice Variant (c.5074+3A>C) of BRCA1 by RNA Sequencing and TOPO Cloning.

Grading the pathogenicity of BRCA1/2 variants has great clinical importance in patient treatment as well as in the prevention and screening of hereditary breast and ovarian cancer (HBOC). For accurate evaluation, confirming the splicing effect of a possible splice site variant is crucial. We report a significant splicing variant (c.5074+3A>C) in BRCA1 in a patient with recurrent ovarian cancer. Next-generation sequencing (NGS) of BRCA1/2 from patient's peripheral blood identified the variant, which was strongly suspected of being a splicing mutation based on in silico predictions. Direct RNA analysis yielded multiple transcripts, and TOPO cloning of the complementary DNA (cDNA) and Sanger sequencing revealed an aberrant transcript with an insertion of the first 153 bp of intron 17, and another transcript with the 153 bp insertion along with an exon 18 deletion. A premature termination codon was presumed to be formed by the 153 bp partial intron retention common to the two transcripts. Therefore, BRCA1 c.5074+3A>C was classified as a likely pathogenic variant. Our findings show that active use of functional studies of variants suspected of altered splicing are of great help in classifying them.

tRNA-Derived Small RNAs: Novel Epigenetic Regulators.

An epigenetic change is a heritable genetic alteration that does not involve any nucleotide changes. While the methylation of specific DNA regions such as CpG islands or histone modifications, including acetylation or methylation, have been investigated in detail, the role of small RNAs in epigenetic regulation is largely unknown. Among the many types of small RNAs, tRNA-derived small RNAs (tsRNAs) represent a class of noncoding small RNAs with multiple roles in diverse physiological processes, including neovascularization, sperm maturation, immune modulation, and stress response. Regarding these roles, several pioneering studies have revealed that dysregulated tsRNAs are associated with human diseases, such as systemic lupus, neurological disorder, metabolic disorder, and cancer. Moreover, recent findings suggest that tsRNAs regulate the expression of critical genes linked with these diseases by a variety of mechanisms, including epigenetic regulation. In this review, we will describe different classes of tsRNAs based on their biogenesis and will focus on their role in epigenetic regulation.