The IMiD target CRBN determines HSP90 activity toward transmembrane proteins essential in multiple myeloma.

The complex architecture of transmembrane proteins requires quality control (QC) of folding, membrane positioning, and trafficking as prerequisites for cellular homeostasis and intercellular communication. However, it has remained unclear whether transmembrane protein-specific QC hubs exist. Here we identify cereblon (CRBN), the target of immunomodulatory drugs (IMiDs), as a co-chaperone that specifically determines chaperone activity of HSP90 toward transmembrane proteins by means of counteracting AHA1. This function is abrogated by IMiDs, which disrupt the interaction of CRBN with HSP90. Among the multiple transmembrane protein clients of CRBN-AHA1-HSP90 revealed by cell surface proteomics, we identify the amino acid transporter LAT1/CD98hc as a determinant of IMiD activity in multiple myeloma (MM) and present an Anticalin-based CD98hc radiopharmaceutical for MM radio-theranostics. These data establish the CRBN-AHA1-HSP90 axis in the biogenesis of transmembrane proteins, link IMiD activity to tumor metabolism, and nominate CD98hc and LAT1 as attractive diagnostic and therapeutic targets in MM.

A druggable cascade links methionine metabolism to epigenomic reprogramming in squamous cell carcinoma.

Upper aerodigestive squamous cell carcinoma (UASCC) is a common and aggressive malignancy with few effective therapeutic options. Here, we investigate amino acid metabolism in this cancer, surprisingly noting that UASCC exhibits the highest methionine level across all human cancers, driven by its transporter LAT1. We show that LAT1 is also expressed at the highest level in UASCC, transcriptionally activated by UASCC-specific promoter and enhancers, which are directly coregulated by SCC master regulators TP63/KLF5/SREBF1. Unexpectedly, unbiased bioinformatic screen identifies EZH2 as the most significant target downstream of the LAT1-methionine pathway, directly linking methionine metabolism to epigenomic reprogramming. Importantly, this cascade is indispensable for the survival and proliferation of UASCC patient-derived tumor organoids. In addition, LAT1 expression is closely associated with cellular sensitivity to inhibition of the LAT1-methionine-EZH2 axis. Notably, this unique LAT1-methionine-EZH2 cascade can be targeted effectively by either pharmacological approaches or dietary intervention in vivo. In summary, this work maps a unique mechanistic cross talk between epigenomic reprogramming with methionine metabolism, establishes its biological significance in the biology of UASCC, and identifies a unique tumor-specific vulnerability which can be exploited both pharmacologically and dietarily.

L-type amino acid transporter 1 inhibitor JPH203 prevents the growth of cabazitaxel-resistant prostate cancer by inhibiting cyclin-dependent kinase activity.

L-type amino acid transporter 1 (LAT1, SLC7A5) is an amino acid transporter expressed in various carcinomas, and it is postulated to play an important role in the proliferation of cancer cells through the uptake of essential amino acids. Cabazitaxel is a widely used anticancer drug for treating castration-resistant prostate cancer (CRPC); however, its effectiveness is lost when cancer cells acquire drug resistance. In this study, we investigated the expression of LAT1 and the effects of a LAT1-specific inhibitor, JPH203, in cabazitaxel-resistant prostate cancer cells. LAT1 was more highly expressed in the cabazitaxel-resistant strains than in the normal strains. Administration of JPH203 inhibited the growth, migration, and invasive ability of cabazitaxel-resistant strains in vitro. Phosphoproteomics using liquid chromatography-mass spectrometry to comprehensively investigate changes in phosphorylation due to JPH203 administration revealed that cell cycle-related pathways were affected by JPH203, and that JPH203 significantly reduced the kinase activity of cyclin-dependent kinases 1 and 2. Moreover, JPH203 inhibited the proliferation of cabazitaxel-resistant cells in vivo. Taken together, the present study results suggest that LAT1 might be a valuable therapeutic target in cabazitaxel-resistant prostate cancer.

The LAT1 inhibitor JPH203 suppresses the growth of castration-resistant prostate cancer through a CD24-mediated mechanism.

L-type amino acid transporter 1 (LAT1) is specifically expressed in many malignancies, contributes to the transport of essential amino acids, such as leucine, and regulates the mammalian target of rapamycin (mTOR) signaling pathway. We investigated the expression profile and functional role of LAT1 in prostate cancer using JPH203, a specific inhibitor of LAT1. LAT1 was highly expressed in castration-resistant prostate cancer (CRPC) cells, including C4-2 and PC-3 cells, but its expression level was low in castration-sensitive LNCaP cells. JPH203 significantly inhibited [14C] leucine uptake in CRPC cells but had no effect in LNCaP cells. JPH203 inhibited the proliferation, migration, and invasion of CRPC cells but not of LNCaP cells. In C4-2 cells, Cluster of differentiation (CD) 24 was identified by RNA sequencing as a novel downstream target of JPH203. CD24 was downregulated in a JPH203 concentration-dependent manner and suppressed activation of the Wnt/ß-catenin signaling pathway. Furthermore, an in vivo study showed that JPH203 inhibited the proliferation of C4-2 cells in a castration environment. The results of this study indicate that JPH203 may exert its antitumor effect in CRPC cells via mTOR and CD24.

L-type Amino Acid Transporter 1 as a Therapeutic Target in Pancreatic Cancer.

Metabolic rewiring is a key feature of cancer cells to support the demands of growth and proliferation. The metabolism of amino acids is altered in many cancers, including pancreatic cancer. The cellular uptake of amino acids is regulated by amino acid transporters, such as L-type amino acid transporter 1 (LAT1). Accumulating evidence suggests that LAT1 is overexpressed in pancreatic cancer and confers a poor prognosis. Here we discuss the prospects of utilizing LAT1 as a novel target for pancreatic cancer therapy.

Fluorine-18 labeling PEGylated 6-boronotryptophan for PET scanning of mice for assessing the pharmacokinetics for boron neutron capture therapy of brain tumors.

Two tryptophan compound classes 5- and 6-borono PEGylated boronotryptophan derivatives have been prepared for assessing their aqueous solubility as formulation of injections for boron neutron capture therapy (BNCT). The PEGylation has improved their aqueous solubility thereby increasing their test concentration in 1 mM without suffering from toxicity. In-vitro uptake assay of PEGylated 5- and 6-boronotryptophan showed that the B-10 concentration can reach 15-50 ppm in U87 cell whereas the uptake in LN229 cell varies. Shorter PEG compound 6-boronotryptophanPEG200[18F] was obtained in 1.7 % radiochemical yield and the PET-derived radioradioactivity percentage in 18 % was taken up by U87 tumor at the limb of xenograft mouse. As high as tumor to normal uptake ratio in 170 (T/N) was obtained while an inferior radioactivity uptake of 3 % and T/N of 8 was observed in LN229 xenografted mouse.

Identification of tumor-suppressive miRNAs that target amino acid transporter LAT1 and exhibit anti-proliferative effects on cholangiocarcinoma cells.

Amino acid transporter LAT1 is highly upregulated in various cancer types, including cholangiocarcinoma (CHOL), and contributes to the rapid proliferation of cancer cells and disease progression. However, the molecular mechanisms underlying the pathological upregulation of LAT1 remain largely unknown. This study pursued the possibility of miRNA-mediated regulation of the LAT1 expression in CHOL cells. Using online target prediction methods, we extracted five candidate miRNAs commonly predicted to regulate the LAT1 expression. Three of them, miR-194-5p, miR-122-5p, and miR-126-3p, were significantly downregulated in CHOL cancer compared to normal tissues. Correlation analysis revealed weak-to-moderate negative correlations between the expression of these miRNAs and LAT1 mRNA in CHOL cancer tissues. We selected miR-194-5p and miR-122-5p for further analyses and found that both miRNAs functionally target 3'UTR of LAT1 mRNA by a luciferase-based reporter assay. Transfection of the miRNA mimics significantly suppressed the LAT1 expression at mRNA and protein levels and inhibited the proliferation of CHOL cells, with a trend of affecting intracellular amino acids and amino acid-related signaling pathways. This study indicates that the decreased expression of these LAT1-targeting tumor-suppressive miRNAs contributes to the upregulation of LAT1 and the proliferation of CHOL cells, highlighting their potential for developing novel cancer therapeutics and diagnostics.

Upregulation of ATF4 mediates the cellular adaptation to pharmacologic inhibition of amino acid transporter LAT1 in pancreatic ductal adenocarcinoma cells.

L-type amino acid transporter 1 (LAT1) is recognized as a promising target for cancer therapy; however, the cellular adaptive response to its pharmacological inhibition remains largely unexplored. This study examined the adaptive response to LAT1 inhibition using nanvuranlat, a high-affinity LAT1 inhibitor. Proteomic analysis revealed the activation of a stress-induced transcription factor ATF4 following LAT1 inhibition, aligning with the known cellular responses to amino acid deprivation. This activation was linked to the GCN2-eIF2α pathway which regulates translation initiation. Our results show that ATF4 upregulation counteracts the suppressive effect of nanvuranlat on cell proliferation in pancreatic ductal adenocarcinoma cell lines, suggesting a role for ATF4 in cellular adaptation to LAT1 inhibition. Importantly, dual targeting of LAT1 and ATF4 exhibited more substantial anti-proliferative effects in vitro than individual treatments. This study underscores the potential of combining LAT1 and ATF4 inhibition as a therapeutic strategy in cancer treatment.

Inhibition of amino acid transporter LAT1 in cancer cells suppresses G0/G1-S transition by downregulating cyclin D1 via p38 MAPK activation.

L-type amino acid transporter 1 (LAT1, SLC7A5) is upregulated in various cancers and associated with disease progression. Nanvuranlat (Nanv; JPH203, KYT-0353), a selective LAT1 inhibitor, suppresses the uptake of large neutral amino acids required for rapid growth and proliferation of cancer cells. Previous studies have suggested that the inhibition of LAT1 by Nanv induces the cell cycle arrest at G0/G1 phase, although the underlying mechanisms remain unclear. Using pancreatic cancer cells arrested at the restriction check point (R) by serum deprivation, we found that the Nanv drastically suppresses the G0/G1-S transition after release. This blockade of the cell cycle progression was accompanied by a sustained activation of p38 mitogen-activated protein kinase (MAPK) and subsequent phosphorylation-dependent proteasomal degradation of cyclin D1. Isoform-specific knockdown of p38 MAPK revealed the predominant contribution of p38α. Proteasome inhibitors restored the cyclin D1 amount and released the cell cycle arrest caused by Nanv. The increased phosphorylation of p38 MAPK and the decrease of cyclin D1 were recapitulated in xenograft tumor models treated with Nanv. This study contributes to delineating the pharmacological activities of LAT1 inhibitors as anti-cancer agents and provides significant insights into the molecular basis of the amino acid-dependent cell cycle checkpoint at G0/G1 phase.

Prolactin upregulates amino acids uptake in dairy cow mammary epithelial cells via LAT1.

The uptake of AA in mammary tissues is affected by prolactin (PRL). To investigate whether PRL-induced AA uptake is involved in L-type AA transporter 1 (LAT1), we analyzed the changes of AA in the medium of dairy cow mammary epithelial cells in the presence of PRL or PRL plus BCH, an inhibitor of LAT1. Then Western blot and luciferase assay were used to detect the regulation mechanism of PRL on LAT1 expression and function. Our results showed that Thr, Val, Met, Ile, Leu, Tyr, Lys, Phe, and His are LAT1 substrates and could be transported into mammary epithelial cells via LAT1. PRL stimulation increased the uptake of most AA into mammary epithelial cells of dairy cows, however, inhibition of LAT1 transport activity reduced PRL-induced AA uptake, suggesting that the effect of PRL on AA transport depends on LAT1 expression and function. PRL stimulation upregulated LAT1 expression and plasma membrane location not only in dairy cow mammary epithelial cells, but also in mouse mammary epithelial cell line HC11. Western blot showed that PI3K-AKT-mTOR signaling could be activated in PRL-stimulated mammary epithelial cells. Treatment of cells with LY294002 decreased PI3K-AKT-mTOR activation, as well LAT1 expression, that in turn decreased milk protein synthesis. Luciferase assay showed PRL treatment increased the promoter activity of LAT1 promoter fragment -419∼-86 bp. Treatment of cells with LY294002, an inhibitor of PI3K, or SC79, an activator of AKT abolished or promoted the transcriptional activity of this promoter fragment in the presence of PRL. These results suggested that the -419∼-86 bp fragment of LAT1 promoter mediates the action of PI3K-AKT-mTOR signaling on LAT1 transcription in mammary epithelial cells of dairy cows, which in turn increased LAT1 expression and AA uptake.

Invariant NKT cells metabolically adapt to the acute myeloid leukaemia environment.

Acute myeloid leukaemia (AML) creates an immunosuppressive environment to conventional T cells through Arginase 2 (ARG2)-induced arginine depletion. We identify that AML blasts release the acute phase protein serum amyloid A (SAA), which acts in an autocrine manner to upregulate ARG2 expression and activity, and promote AML blast viability. Following in vitro cross-talk invariant natural killer T (iNKT) cells become activated, upregulate mitochondrial capacity, and release IFN-γ. iNKT retain their ability to proliferate and be activated despite the low arginine AML environment, due to the upregulation of Large Neutral Amino Acid Transporter-1 (LAT-1) and Argininosuccinate Synthetase 1 (ASS)-dependent amino acid pathways, resulting in AML cell death. T cell proliferation is restored in vitro and in vivo. The capacity of iNKT cells to restore antigen-specific T cell immunity was similarly demonstrated against myeloid-derived suppressor cells (MDSCs) in wild-type and Jα18-/- syngeneic lymphoma-bearing models in vivo. Thus, stimulation of iNKT cell activity has the potential as an immunotherapy against AML or as an adjunct to boost antigen-specific T cell immunotherapies in haematological or solid cancers.

Combination effects of amino acid transporter LAT1 inhibitor nanvuranlat and cytotoxic anticancer drug gemcitabine on pancreatic and biliary tract cancer cells.

BACKGROUND: Cytotoxic anticancer drugs widely used in cancer chemotherapy have some limitations, such as the development of side effects and drug resistance. Furthermore, monotherapy is often less effective against heterogeneous cancer tissues. Combination therapies of cytotoxic anticancer drugs with molecularly targeted drugs have been pursued to solve such fundamental problems. Nanvuranlat (JPH203 or KYT-0353), an inhibitor for L-type amino acid transporter 1 (LAT1; SLC7A5), has novel mechanisms of action to suppress the cancer cell proliferation and tumor growth by inhibiting the transport of large neutral amino acids into cancer cells. This study investigated the potential of the combined use of nanvuranlat and cytotoxic anticancer drugs. METHODS: The combination effects of cytotoxic anticancer drugs and nanvuranlat on cell growth were examined by a water-soluble tetrazolium salt assay in two-dimensional cultures of pancreatic and biliary tract cancer cell lines. To elucidate the pharmacological mechanisms underlying the combination of gemcitabine and nanvuranlat, we investigated apoptotic cell death and cell cycle by flow cytometry. The phosphorylation levels of amino acid-related signaling pathways were analyzed by Western blot. Furthermore, growth inhibition was examined in cancer cell spheroids. RESULTS: All the tested seven types of cytotoxic anticancer drugs combined with nanvuranlat significantly inhibited the cell growth of pancreatic cancer MIA PaCa-2 cells compared to their single treatment. Among them, the combined effects of gemcitabine and nanvuranlat were relatively high and confirmed in multiple pancreatic and biliary tract cell lines in two-dimensional cultures. The growth inhibitory effects were suggested to be additive but not synergistic under the tested conditions. Gemcitabine generally induced cell cycle arrest at the S phase and apoptotic cell death, while nanvuranlat induced cell cycle arrest at the G0/G1 phase and affected amino acid-related mTORC1 and GAAC signaling pathways. In combination, each anticancer drug basically exerted its own pharmacological activities, although gemcitabine more strongly influenced the cell cycle than nanvuranlat. The combination effects of growth inhibition were also verified in cancer cell spheroids. CONCLUSIONS: Our study demonstrates the potential of first-in-class LAT1 inhibitor nanvuranlat as a concomitant drug with cytotoxic anticancer drugs, especially gemcitabine, on pancreatic and biliary tract cancers.

Overexpression of the LAT1 in primary human trophoblast cells increases the uptake of essential amino acids and activates mTOR signaling.

The System L amino acid transporter, particularly the isoform Large Neutral Amino Acid Transporter Small Subunit 1 (LAT1) encoded by SLC7A5, is believed to mediate the transfer of essential amino acids in the human placenta. Placental System L amino acid transporter expression and activity is decreased in pregnancies complicated by IUGR and increased in fetal overgrowth. However, it remains unknown if changes in the expression of LAT1 are mechanistically linked to System L amino acid transport activity. Here, we combined overexpression approaches with protein analysis and functional studies in cultured primary human trophoblast (PHT) cells to test the hypothesis that SLC7A5 overexpression increases the uptake of essential amino acids and activates mTOR signaling in PHT cells. Overexpression of SLC7A5 resulted in a marked increase in protein expression of LAT1 in the PHT cells microvillous plasma membrane and System L amino acid transporter activity. Moreover, mTOR signaling was activated, and System A amino acid transporter activity increased following SLC7A5 overexpression, suggesting coordination of trophoblast amino transporter expression and activity to ensure balanced nutrient flux to the fetus. This is the first report showing that overexpression of LAT1 is sufficient to increase the uptake of essential amino acids in PHT cells, which activates mTOR, a master regulator of placental function. The decreased placental System L activity in human IUGR and the increased placental activity of this transporter system in some cases of fetal overgrowth may directly contribute to changes in fetal amino acid availability and altered fetal growth in these pregnancy complications.

Sodium-Dependent Neutral Amino Acid Transporter 2 Can Serve as a Tertiary Carrier for l-Type Amino Acid Transporter 1-Utilizing Prodrugs.

Membrane transporters are the key determinants of the homeostasis of endogenous compounds in the cells and their exposure to drugs. However, the substrate specificities of distinct transporters can overlap. In the present study, the interactions of l-type amino acid transporter 1 (LAT1)-utilizing prodrugs with sodium-coupled neutral amino acid transporter 2 (SNAT2) were explored. The results showed that the cellular uptake of LAT1-utilizing prodrugs into a human breast cancer cell line, MCF-7 cells, was mediated via SNATs as the uptake was increased at higher pH (8.5), decreased in the absence of sodium, and inhibited in the presence of unselective SNAT-inhibitor, (α-(methylamino)isobutyric acid, MeAIB). Moreover, docking the compounds to a SNAT2 homology model (inward-open conformation) and further molecular dynamics simulations and the subsequent trajectory and principal component analyses confirmed the chemical features supporting the interactions of the studied compounds with SNAT2, which was found to be the main SNAT expressed in MCF-7 cells.

Structural Features Affecting the Interactions and Transportability of LAT1-Targeted Phenylalanine Drug Conjugates.

L-type amino acid transporter 1 (LAT1) transfers essential amino acids across cell membranes. Owing to its predominant expression in the blood-brain barrier and tumor cells, LAT1 has been exploited for drug delivery and targeting to the central nervous system (CNS) and various cancers. Although the interactions of amino acids and their mimicking compounds with LAT1 have been extensively investigated, the specific structural features for an optimal drug scaffold have not yet been determined. Here, we evaluated a series of LAT1-targeted drug-phenylalanine conjugates (ligands) by determining their uptake rates by in vitro studies and investigating their interaction with LAT1 via induced-fit docking. Combining the experimental and computational data, we concluded that although LAT1 can accommodate various types of structures, smaller compounds are preferred. As the ligand size increased, its flexibility became more crucial in determining the compound's transportability and interactions. Compounds with linear or planar structures exhibited reduced uptake; those with rigid lipophilic structures lacked interactions and likely utilized other transport mechanisms for cellular entry. Introducing polar groups between aromatic structures enhanced interactions. Interestingly, compounds with a carbamate bond in the aromatic ring's para-position displayed very good transport efficiencies for the larger compounds. Compared to the ester bond, the corresponding amide bond had superior hydrogen bond acceptor properties and increased interactions. A reverse amide bond was less favorable than a direct amide bond for interactions with LAT1. The present information can be applied broadly to design appropriate CNS or antineoplastic drug candidates with a prodrug strategy and to discover novel LAT1 inhibitors used either as direct or adjuvant cancer therapy.

Contribution of the L-Type Amino Acid Transporter Family in the Diagnosis and Treatment of Prostate Cancer.

The L-type amino acid transporter (LAT) family contains four members, LAT1~4, which are important amino acid transporters. They mainly transport specific amino acids through cell membranes, provide nutrients to cells, and are involved in a variety of metabolic pathways. They regulate the mTOR signaling pathway which has been found to be strongly linked to cancer in recent years. However, in the field of prostate cancer (PCa), the LAT family is still in the nascent stage of research, and the importance of LATs in the diagnosis and treatment of prostate cancer is still unknown. Therefore, this article aims to report the role of LATs in prostate cancer and their clinical significance and application. LATs promote the progression of prostate cancer by increasing amino acid uptake, activating the mammalian target of rapamycin (mTOR) pathway and downstream signals, mediating castration-resistance, promoting tumor angiogenesis, and enhancing chemotherapy resistance. The importance of LATs as diagnostic and therapeutic targets for prostate cancer was emphasized and the latest research results were introduced. In addition, we introduced selective LAT1 inhibitors, including JPH203 and OKY034, which showed excellent inhibitory effects on the proliferation of various tumor cells. This is the future direction of amino acid transporter targeting therapy drugs.

Genetic Ablation of LAT1 Inhibits Growth of Liver Cancer Cells and Downregulates mTORC1 Signaling.

Through a comprehensive analysis of the gene expression and dependency in HCC patients and cell lines, LAT1 was identified as the top amino acid transporter candidate supporting HCC tumorigenesis. To assess the suitability of LAT1 as a HCC therapeutic target, we used CRISPR/Cas9 to knockout (KO) LAT1 in the epithelial HCC cell line, Huh7. Knockout of LAT1 diminished its branched chain amino acid (BCAA) transport activity and significantly reduced cell proliferation in Huh7. Consistent with in vitro studies, LAT1 ablation led to suppression of tumor growth in a xenograft model. To elucidate the mechanism underlying the observed inhibition of cell proliferation upon LAT1 KO, we performed RNA-sequencing analysis and investigated the changes in the mTORC1 signaling pathway. LAT1 ablation resulted in a notable reduction in phosphorylation of p70S6K, a downstream target of mTORC1, as well as its substrate S6RP. This reduced cell proliferation and mTORC1 activity were rescued when LAT1 was overexpressed. These findings imply an essential role of LAT1 for maintenance of tumor cell growth and additional therapeutic angles against liver cancer.

mTORC2-AKT-LAT1 signalling participates in methionine-induced ß-CASEIN expression in mammary epithelial cells of dairy cows.

This study investigated the role of the mammalian target of rapamycin complex 2 (mTORC2)-protein kinase B (AKT) signalling in methionine (Met)-induced L-type amino acid transporter 1 (LAT1) expression and milk protein production. Primary mammary epithelial cells (MECs) from mammary parenchymal tissues of three lactating cows and MAC-T bovine MECs were cultured with or without 0.6 mM Met. Rapamycin-insensitive companion of mTOR (RICTOR) siRNA, the mTORC1 inhibitor rapamycin and the AKT activator SC79 were used to evaluate the effects of mTORC2-AKT signalling on Met-induced LAT1 expression and function. Each experiment was performed three times. Data were analysed with a two-sided unpaired t test or ANOVA with the Bonferroni multiple-comparison test. Western blotting showed that Met stimulation increased RICTOR expression (~244.67%; p < 0.05; control, 0.15 ± 0.026; Met, 0.517 ± 0.109) and AKT-S473 levels (~281.42%; p < 0.01; control, 0.253 ± 0.067; Met, 0.965 ± 0.019) in both primary MECs and MAC-T cells. Rapamycin-induced mTORC1 signalling inhibition decreased only Met-induced ß-CASEIN expression by ~21.24% (p < 0.01; Met, 0.777 ± 0.01; Met and rapamycin, 0.612 ± 0.04) and did not affect Met-stimulated AKT-S473 levels, suggesting that mTORC2-AKT activation upon Met stimulation also contributes to milk protein synthesis. LAT1 participates in Met-induced ß-CASEIN expression. In dairy cow MECs, mTORC2 inhibition by RICTOR siRNA decreased LAT1 levels on the plasma membrane by ~45.13% (p < 0.01; control, 0.359 ± 0.006; siRICTOR, 0.197 ± 0.004). However, SC79-induced AKT activation had the opposite effect (p < 0.01). In primary MECs and MAC-T cells, Met stimulation increased cytosolic and plasma membrane LAT1 expression respectively (MECs, 113.98% and 58.43%; MAC-T, 165.85% and 396.39%; p < 0.05). However, RICTOR siRNA significantly reduced Met-induced plasma membrane LAT1 expression (~76.48%; Met, 0.539 ± 0.05; Met and siRICTOR, 0.127 ± 0.012; p < 0.05). Thus, Met increased LAT1 expression and function via mTORC2-AKT signalling, upregulating milk protein synthesis in dairy cow MECs.

Pharmacologic inhibition of LAT1 predominantly suppresses transport of large neutral amino acids and downregulates global translation in cancer cells.

L-type amino acid transporter 1 (LAT1; SLC7A5), which preferentially transports large neutral amino acids, is highly upregulated in various cancers. LAT1 supplies cancer cells with amino acids as substrates for enhanced biosynthetic and bioenergetic reactions and stimulates signalling networks involved in the regulation of survival, growth and proliferation. LAT1 inhibitors show anti-cancer effects and a representative compound, JPH203, is under clinical evaluation. However, pharmacological impacts of LAT1 inhibition on the cellular amino acid transport and the translational activity in cancer cells that are conceptually pivotal for its anti-proliferative effect have not been elucidated yet. Here, we demonstrated that JPH203 drastically inhibits the transport of all the large neutral amino acids in pancreatic ductal adenocarcinoma cells. The inhibitory effects of JPH203 were observed even in competition with high concentrations of amino acids in a cell culture medium. The analyses of the nutrient-sensing mTORC1 and GAAC pathways and the protein synthesis activity revealed that JPH203 downregulates the global translation. This study demonstrates a predominant contribution of LAT1 to the transport of large neutral amino acids in cancer cells and the suppression of protein synthesis by JPH203 supposed to underly its broad anti-proliferative effects across various types of cancer cells.

Inactivation of Lats1 and Lats2 highlights the role of hippo pathway effector YAP in larynx and vocal fold epithelium morphogenesis.

Proliferation and differentiation of vocal fold epithelial cells during embryonic development is poorly understood. We examined the role of Hippo signaling, a vital pathway known for regulating organ size, in murine laryngeal development. Conditional inactivation of the Hippo kinase genes Lats1 and Lats2, specifically in vocal fold epithelial cells, resulted in severe morphogenetic defects. Deletion of Lats1 and Lats2 caused abnormalities in epithelial differentiation, epithelial lamina separation, cellular adhesion, basement membrane organization with secondary failed cartilage, and laryngeal muscle development. Further, Lats1 and Lats2 inactivation led to failure in differentiation of p63+ basal progenitors. Our results reveal novel roles of Hippo-Lats-YAP signaling in proper regulation of VF epithelial fate and larynx morphogenesis.