Sodium-coupled electrogenic transport of pyroglutamate (5-oxoproline) via SLC5A8, a monocarboxylate transporter

Pyroglutamate, also known as 5-oxoproline, is a structural analog of proline. This amino acid derivative is a byproduct of glutathione metabolism, and is reabsorbed efficiently in kidney by Na⁺-coupled transport mechanisms. Previous studies have focused on potential participation of amino acid transport systems in renal reabsorption of this compound. Here we show that it is not the amino acid transport systems but instead the Na⁺-coupled monocarboxylate transporter SLC5A8 that plays a predominant role in this reabsorptive process. Expression of cloned human and mouse SLC5A8 in mammalian cells induces Na⁺-dependent transport of pyroglutamate that is inhibitable by various SLC5A8 substrates. SLC5A8-mediated transport of pyroglutamate is saturable with a Michaelis constant of 0.36 ± 0.04 mM. Na⁺-activation of the transport process exhibits sigmoidal kinetics with a Hill coefficient of 1.8 ± 0.4, indicating involvement of more than one Na⁺ in the activation process. Expression of SLC5A8 in Xenopuslaevis oocytes induces Na⁺-dependent inward currents in the presence of pyroglutamate under voltage-clamp conditions. The concentration of pyroglutamate necessary for induction of half-maximal current is 0.19 ± 0.01 mM. The Na⁺-activation kinetics is sigmoidal with a Hill coefficient of 2.3 ± 0.2. Ibuprofen, a blocker of SLC5A8, suppressed pyroglutamate-induced currents in SLC5A8-expressing oocytes; the concentration of the blocker necessary for causing half-maximal inhibition is 14 ± 1 μM. The involvement of SLC5A8 can be demonstrated in rabbit renal brush border membrane vesicles by showing that the Na⁺-dependent uptake of pyroglutamate in these vesicles is inhibitable by known substrates of SLC5A8. The Na⁺ gradient-driven pyroglutamate uptake was stimulated by an inside-negative K⁺ diffusion potential induced by valinomycin, showing that the uptake process is electrogenic.     

Synthesis and pharmacological evaluation of pyrazoline derivatives as new anti-inflammatory and analgesic agents

A series of 3-(4-biphenyl)-5-substituted phenyl-2-pyrazolines (2a-h) and 1-benzoyl-3-(4-biphenyl)-5-substituted phenyl-2-pyrazolines (3a-h) were synthesized by condensation of chalcones with hydrazine hydrate in solvent system ethanol and DMF. The newly synthesized compounds were screened for their anti-inflammatory and analgesic activity, and were compared with standard drug. Among the compounds studied, compound 2e showed more potent anti-inflammatory and analgesic activity than the standard drug, along with minimum ulcerogenic index.     

Heat shock protein A12A activates migration of hepatocellular carcinoma cells in a monocarboxylate transporter 4–dependent manner

Metastasis is responsible for most of the hepatocellular carcinoma (HCC)–associated death. However, its underlying mechanism has yet to be fully elucidated. Glycolysis-derived lactate has been shown to be a powerful regulator of cancer metastasis. Heat shock protein A12A (HSPA12A) encodes a novel member of HSP70 family. We have recently demonstrated that heat shock protein A12A (HSPA12A) inhibited renal cancer cell migration by suppressing lactate output and glycolytic activity, which were mediated by unstabilizing CD147 and promoting its degradation. By striking contrast, here we demonstrated that HSPA12A promoted migration of human HCC cells. Extracellular acidification, lactate export, and glycolytic activity in HCC cells were also promoted following HSPA12A overexpression. Further analysis revealed that HSPA12A interacted with MCT4 and increased its membrane localization, thereby promoting export of lactate generated from glycolysis; this led, ultimately, to HCC cell migration. Our results revealed the opposite effect of HSPA12A on migration of renal cancer cells and that of HCC cells. Of note, in contrast to the inhibitory effect on CD147 expression in renal cancer cells, we found that HSPA12A increased CD147 expression in HCC cells, indicating that the expression of CD147 might exist heterogeneity in different cancer cell types. Taken together, we identified HSPA12A as an activator of HCC migration, a role opposite to that of renal cancer cells. Inhibiting HSPA12A might be a potential therapeutic intervention for HCC metastasis.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12192-021-01251-z.     

Synthesis and pharmacological evaluation of 4-(3,4-dichlorophenyl)-N-methyl-1,2,3,4-tetrahydronaphthalenyl amines as triple reuptake inhibitors

The present work describes a series of novel chiral amines that potently inhibit the in vitro reuptake of serotonin, norepinephrine and dopamine (triple reuptake inhibitors) and were active in vivo in a mouse model predictive of antidepressant like activity. The detailed synthesis and in vitro activity and ADME profile of compounds is described, which represent a previously undisclosed triple reuptake inhibitor chemotype.     

Calcium signaling and the therapeutic targeting of cancer cells

The calcium signal is implicated in a variety of processes important in tumor progression (e.g. proliferation and invasiveness). The calcium signal has also been shown to be important in other processes important in cancer progression including the development of resistance to current cancer therapies. In this review, we discuss how Ca²⁺ channels, pumps and exchangers may be drug targets in some cancer types. We consider what factors should be taken into account when considering an optimal Ca²⁺ channel, pump or exchanger as a candidate for further assessment as a novel drug target in cancer. We also present and summarize how some therapies for the treatment of cancer intersect with Ca²⁺ signaling and how pharmacological manipulation of the machinery of Ca²⁺ signaling could promote the effectiveness of some therapies. We also review new therapeutic opportunities for Ca²⁺ signal modulators in the context of the tumor microenvironment.     

Synthesis and pharmacological evaluation of a new class of peroxisome proliferator-activated receptor modulators

A series of 5-substituted 2-benzoylaminobenzoic acids has been synthesized and assayed for PPAR/γ activity. Both dual activators and selective PPARγ agonists have been identified. This class of compounds was shown to activate the PPARγ receptor through interaction with a novel binding site.     

Synthesis and evaluation of naphthalene-based thiosemicarbazone derivatives as new anticancer agents against LNCaP prostate cancer cells

Fourteen new naphthalene-based thiosemicarbazone derivatives were designed as anticancer agents against LNCaP human prostate cancer cells and synthesized. MTT assay indicated that compounds 6, 8 and 11 exhibited inhibitory effect on LNCaP cells. Among these compounds, 4-(naphthalen-1-yl)-1-[1-(4-hydroxyphenyl)ethylidene)thiosemicarbazide (6), which caused more than 50% death on LNCaP cells, was chosen for flow cytometric analysis of apoptosis. Flow cytometric analysis pointed out that compound 6 also showed apoptotic effect on LNCaP cells. Compound 6 can be considered as a promising anticancer agent against LNCaP cells owing to its potent cytotoxic activity and apoptotic effect.     

Hormonal regulation of lactate dehydrogenase-A through activation of protein kinase C pathways in MCF-7 breast cancer cells

Lactate dehydrogenase A (LDH-A) is hormonally regulated in rodents, and increased expression of LDH-A is observed during mammary gland tumorigenesis. The mechanisms of hormonal regulation of LDH-A were investigated using a series of deletion and mutant constructs derived from the rat LDH-A gene promoter. Results of these studies show that constructs containing the -92 to -37 region of the LDH-A promoter are important for basal and E2-induced transactivation, and mutation of the consensus CRE motif within this region results in significant loss of basal activity and hormone-responsiveness. Gel mobility shift assays using nuclear extracts from MCF-7 cells show that both CREB and ATF-1 interact with the CRE. Studies with kinase inhibitors show that E2-induced activation of this CRE is dependent on protein kinase C, and these data indicate that LDH-A is induced through a non-genomic pathway of estrogen action.     

NK cell-mediated targeting of human cancer and possibilities for new means of immunotherapy

Insights into the molecular basis for natural killer (NK) cell recognition of human cancer have been obtained in recent years. Here, we review current knowledge on the molecular specificity and function of human NK cells. Evidence for NK cell targeting of human tumors is provided and new strategies for NK cell-based immunotherapy against human cancer are discussed. Based on current knowledge, we foresee a development where more cancers may be subject to treatment with drugs or other immunomodulatory agents affecting NK cells, either directly or indirectly. We also envisage a possibility that certain forms of cancers may be subject to treatment with adoptively transferred NK cells, either alone or in combination with other therapeutic interventions.     

Synthesis and biological evaluation of indolyl chalcones as antitumor agents

A series of indolyl chalcones were synthesized and evaluated in vitro for their anticancer activity against three human cancer cell lines. Compounds 3b–d, 3h, 3j, 3l, 3m, 4g, and 4j showed significant cytotoxicity, particularly, indolyl chalcones 3l and 3m were identified as the most potent and selective anticancer agents with IC₅₀ values 0.03 and 0.09 μM, against PaCa-2 cell line, respectively.     

Engineered production of cancer targeting peptide (CTP)-containing C-1027 in Streptomyces globisporus and biological evaluation

Conjugation of cancer targeting peptides (CTPs) with small molecular therapeutics has emerged as a promising strategy to deliver potent (but typically nonspecific) cytotoxic agents selectively to cancer cells. Here we report the engineered production of a CTP (NGR)-containing C-1027 and evaluation of its activity against selected cancer cell lines. C-1027 is an enediyne chromoprotein produced by Streptomyces globisporus, consisting of an apo-protein (CagA) and an enediyne chromophore (C-1027). NGR is a CTP that targets CD13 in tumor vasculature. S. globisporus SB1026, a recombinant strain engineered to encode CagA with the NGR sequence fused at its C-terminus, directly produces the NGR-containing C-1027 that is equally active as the native C-1027. Our results demonstrate the feasibility to produce CTP-containing enediyne chromoproteins by metabolic pathway engineering and microbial fermentation and will inspire efforts to engineer other CTP-containing drug binding proteins for targeted delivery.     

Synthesis and evaluation of unsymmetrical polyamine derivatives as antitumor agents

A series of unsymmetrically substituted polyamine derivatives were prepared and their cytotoxicities in mouse leukemia L1210, melanoma B16, and HeLa cells were investigated. The in vitro cytotoxicity revealed that these conjugates could recognize the polyamine transporter, and the N-ethyl modified homospermidine moiety may be another efficient carrier as homospermidine even though the introduction of terminal alkyl groups led to reduced cytotoxicity in comparison with the un-substituted counterpart 1. The ornithine decarboxylase and topoisomerase II inhibition experiments indicated that ODC and TOPO II were potential, but not unique targets of these conjugates. Furthermore, the in vivo antitumor activities illustrated that the representative conjugate 2f and the homospermidine analogue 1 evidently inhibited the tumor growth and significantly increased the survival time of mice-bearing sarcoma 180 cells.     

A mechanistic kinetic description of lactate dehydrogenase elucidating cancer diagnosis and inhibitor evaluation

As a key enzyme for glycolysis, lactate dehydrogenase (LDH) remains as a topic of great interest in cancer study. Though a number of kinetic models have been applied to describe the dynamic behavior of LDH, few can reflect its actual mechanism, making it difficult to explain the observed substrate and competitor inhibitions at wide concentration ranges. A novel mechanistic kinetic model is developed based on the enzymatic processes and the interactive properties of LDH. Better kinetic simulation as well as new enzyme interactivity information and kinetic properties extracted from published articles via the novel model was presented. Case studies were presented to a comprehensive understanding of the effect of temperature, substrate, and inhibitor on LDH kinetic activities for promising application in cancer diagnosis, inhibitor evaluation, and adequate drug dosage prediction.     

Synthesis of sulfonamide-containing N-hydroxyindole-2-carboxylates as inhibitors of human lactate dehydrogenase-isoform 5

N-Hydroxyindole-2-carboxylates possessing sulfonamide-substituents at either position 5 or 6 were designed and synthesized. The inhibitory activities of these compounds against isoforms 1 and 5 of human lactate dehydrogenase were analysed, and K_i values of the most efficient inhibitors were determined by standard enzyme kinetic studies. Some of these compounds displayed state-of-the-art inhibitory potencies against isoform 5 (K_i values as low as 5.6 μM) and behaved as competitive inhibitors versus both the substrate and the cofactor.     

Synthesis and evaluation of new 5-fluorouracil antitumor cell differentiating derivatives

Three new antitumour drugs containing two 5-fluorouracil moieties at both ends of the structure and a two amide bond linker were synthesized. Appropriated bis-acetal were reacted with two equivalents of 5-FU to afford the desired compounds. These drugs were evaluated for their ability to induce myogenic maturation in vitro on human rhabdomyosarcoma cells in an experimental model. Compounds 5 and 6 induced morphological and phenotypical differentiation in rhabdomyosarcoma cells at 4.5 and 3.5 microM, respectively. These new cell differentiating agents could be used as an alternative to selective destruction of undifferentiated cells. A potential role of the differentiation therapy as an alternative approach to the treatment of rhabdomyosarcomas is suggested.     

Oxime-based inhibitors of glucose transporter 1 displaying antiproliferative effects in cancer cells

An analysis of the main pharmacophoric features present in the still limited number of inhibitors of glucose transporter GLUT1 led to the identification of new oxime-based inhibitors, which proved to be able to efficiently hinder glucose uptake and cell growth in H1299 lung cancer cells. The most important interactions of a representative inhibitor were indicated by a novel computational model of GLUT1, which was purposely developed to explain these results and to provide useful indications for the design and the development of new and more efficient GLUT1 inhibitors.     

Synthesis and biological evaluation of cyclopenta[c]thiophene related compounds as new antitumor agents

A series of 22 cyclopenta[c]thiophene related compounds was obtained by the pharmacomodulation of 6-amino-5,6-dihydro-4H-cyclopenta[c]thiophen-4-ones 1a-g. All compounds were evaluated for potential anticancer activity in the NCI's in vitro human disease-oriented tumor cell line screening panel that consisted of 60 human tumor cell lines arranged in nine subpanels, representing diverse histologies. Among these tested compounds, seven were found to be cytotoxic, especially against leukemia cell lines, allowing us to point out some structure-activity relationships. These derivatives were further evaluated for potential in vivo anticancer activity in the hollow fiber assay developed at the NCI, which selected two compounds, 1f and 3a for standard xenograft testing.     

Design,synthesis and biological evaluation of new inhibitors of Bax/Bcl-xL interaction in cancer cells

We describe the synthesis of a series of new molecules containing phenol and triazoles moieties, compounds which have been evaluated for their ability to inhibit Bax/Bcl-xL interactions in cancer cells, by using BRET assays, and to induce cell death. Several derivatives exhibit a very promising activity, being more potent than the reference compounds acylpyrogallol A and ABT-737. These preliminary results demonstrate that derivatives of this family can be attractive to develop new molecules with potent anticancer activity.