N-methyl-D-aspartate (NMDA) receptors are highly expressed in the CNS, mediate the slow component of excitatory transmission and play key roles in synaptic plasticity and excitotoxicity. These ligand-gated ion channels are heteromultimers composed of NR1 and NR2 subunits activated by glycine and glutamate. In this study, patch-clamp recordings were used to study the temperature sensitivity of recombinant NR1/NR2B receptors expressed in human embryonic kidney (HEK) 293 cells. Rate constants were assessed by fitting a six-state kinetic scheme to time courses of transient macroscopic currents induced by glutamate at 21.9-46.5 degrees C. Arrhenius transformation of the rate constants characterizing NMDA receptor channel activity indicates that the most sensitive were the rate constants of desensitization (temperature coefficient Q(10)=10.3), resensitization (Q(10)=4.6) and unbinding (Q(10)=3.6). Other rate constants and the amplitude of single-channel currents were less temperature sensitive. Deactivation of responses mediated by NR1/NR2B receptors after a brief application of glutamate was best fit by a double exponential function (tau(fast): Q(10)=3.7; tau(slow): Q(10)=2.7). From these data, we conclude that desensitization/resensitization of the NMDA receptor and glutamate unbinding are especially temperature sensitive and imply that at physiological temperatures the channel kinetics play an important role in determining amplitude and time course of NMDA receptor-mediated postsynaptic currents and these receptors mediated synaptic plasticity.

• MeSH
• Algorithms MeSH
• Cell Line MeSH
• Electrophysiology MeSH
• Financing, Organized MeSH
• Ion Channel Gating physiology   MeSH
• Kinetics MeSH
• Glutamic Acid metabolism   MeSH
• Kidney cytology   metabolism   MeSH
• Humans MeSH
• Patch-Clamp Techniques MeSH
• Receptors, N-Methyl-D-Aspartate physiology   genetics   drug effects   MeSH
• Recombinant Proteins MeSH
• Models, Statistical MeSH
• In Vitro Techniques MeSH
• Temperature MeSH
• Transfection MeSH
• Check Tag
• Humans MeSH

P2X receptors (P2XRs) are ATP-gated cationic channels that are allosterically modulated by numerous compounds, including steroids and neurosteroids. These compounds may both inhibit and potentiate the activity of P2XRs, but sex steroids such as 17β-estradiol or progesterone are reported to be inactive. Here, we tested a hypothesis that testosterone, another sex hormone, modulates activity of P2XRs. We examined actions of native testosterone and a series of testosterone derivatives on the gating of recombinant P2X2R, P2X4R and P2X7R and native channels expressed in pituitary cells and hypothalamic neurons. The 17β-ester derivatives of testosterone rapidly and positively modulate the 1 µM ATP-evoked currents in P2X2R- and P2X4R-expressing cells, but not agonist-evoked currents in P2X7R-expressing cells. In general, most of the tested testosterone derivatives are more potent modulators than endogenous testosterone. The comparison of chemical structures and whole-cell recordings revealed that their interactions with P2XRs depend on the lipophilicity and length of the alkyl chain at position C-17. Pre-treatment with testosterone butyrate or valerate increases the sensitivity of P2X2R and P2X4R to ATP by several fold, reduces the rate of P2X4R desensitization, accelerates resensitization, and enhances ethidium uptake by P2X4R. Native channels are also potentiated by testosterone derivatives, while endogenously expressed GABA receptors type A are inhibited. The effect of ivermectin, a P2X4R-specific allosteric modulator, on deactivation is antagonized by testosterone derivatives in a concentration-dependent manner. Together, our results provide evidence for potentiation of particular subtypes of P2XRs by testosterone derivatives and suggest a potential role of ivermectin binding site for steroid-induced modulation. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.

• MeSH
• Ion Channel Gating drug effects   physiology   MeSH
• HEK293 Cells MeSH
• Rats MeSH
• Humans MeSH
• Rats, Wistar MeSH
• Receptors, Purinergic P2X2 metabolism   MeSH
• Receptors, Purinergic P2X4 metabolism   MeSH
• Testosterone pharmacology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

A growing body of evidence supports the notion that cancer resistance is driven by a small subset of cancer stem cells (CSC), responsible for tumor initiation, growth, and metastasis. Both CSC and chemoresistant cancer cells may share common qualities to activate a series of self-defense mechanisms against chemotherapeutic drugs. Here, we aimed to identify proteins in chemoresistant triple-negative breast cancer (TNBC) cells and corresponding CSC-like spheroid cells that may contribute to their resistance. We have identified several candidate proteins representing the subfamilies of DNA damage response (DDR) system, the ATP-binding cassette, and the 26S proteasome degradation machinery. We have also demonstrated that both cell types exhibit enhanced DDR when compared to corresponding parental counterparts, and identified RAD50 as one of the major contributors in the resistance phenotype. Finally, we have provided evidence that depleting or blocking RAD50 within the Mre11-Rad50-NBS1 (MRN) complex resensitizes CSC and chemoresistant TNBC cells to chemotherapeutic drugs.

• MeSH
• Drug Resistance, Neoplasm drug effects   genetics   MeSH
• Cisplatin administration & dosage   MeSH
• Cyclophosphamide administration & dosage   MeSH
• DNA-Binding Proteins genetics   MeSH
• Doxorubicin administration & dosage   MeSH
• DNA Repair Enzymes genetics   MeSH
• MRE11 Homologue Protein genetics   MeSH
• Acid Anhydride Hydrolases genetics   MeSH
• Nuclear Proteins genetics   MeSH
• Humans MeSH
• Neoplastic Stem Cells drug effects   metabolism   MeSH
• DNA Damage drug effects   MeSH
• Disease-Free Survival MeSH
• Cell Cycle Proteins genetics   MeSH
• Triple Negative Breast Neoplasms drug therapy   genetics   MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

While the contribution of specific tumor suppressor networks to cancer development has been the subject of considerable recent study, it remains unclear how alterations in these networks are integrated to influence the response of tumors to anti-cancer treatments. Here, we show that mechanisms commonly used by tumors to bypass early neoplastic checkpoints ultimately determine chemotherapeutic response and generate tumor-specific vulnerabilities that can be exploited with targeted therapies. Specifically, evaluation of the combined status of ATM and p53, two commonly mutated tumor suppressor genes, can help to predict the clinical response to genotoxic chemotherapies. We show that in p53-deficient settings, suppression of ATM dramatically sensitizes tumors to DNA-damaging chemotherapy, whereas, conversely, in the presence of functional p53, suppression of ATM or its downstream target Chk2 actually protects tumors from being killed by genotoxic agents. Furthermore, ATM-deficient cancer cells display strong nononcogene addiction to DNA-PKcs for survival after DNA damage, such that suppression of DNA-PKcs in vivo resensitizes inherently chemoresistant ATM-deficient tumors to genotoxic chemotherapy. Thus, the specific set of alterations induced during tumor development plays a dominant role in determining both the tumor response to conventional chemotherapy and specific susceptibilities to targeted therapies in a given malignancy.

• MeSH
• Survival Analysis MeSH
• Apoptosis drug effects   MeSH
• Ataxia Telangiectasia Mutated Proteins MeSH
• NIH 3T3 Cells MeSH
• Checkpoint Kinase 2 MeSH
• Drug Resistance, Neoplasm MeSH
• DNA-Binding Proteins * deficiency   metabolism   MeSH
• Humans MeSH
• Mice, Nude MeSH
• Mice MeSH
• Cell Line, Tumor MeSH
• Tumor Suppressor Proteins * deficiency   metabolism   MeSH
• Tumor Suppressor Protein p53 * metabolism   MeSH
• Breast Neoplasms drug therapy   physiopathology   MeSH
• Neoplasms * drug therapy   physiopathology   MeSH
• Protein Serine-Threonine Kinases * deficiency   metabolism   MeSH
• Cell Cycle Proteins * metabolism   MeSH
• Antineoplastic Agents * pharmacology   therapeutic use   MeSH
• Signal Transduction MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH

Interferon (IFN)-related DNA damage resistant signature (IRDS) genes are a subgroup of interferon-stimulated genes (ISGs) found upregulated in different cancer types, which promotes resistance to DNA damaging chemotherapy and radiotherapy. Along with briefly discussing IFNs and signalling in this review, we highlighted how different IRDS genes are affected by viruses. On the contrary, different strategies adopted to suppress a set of IRDS genes (STAT1, IRF7, OAS family, and BST2) to induce (chemo- and radiotherapy) sensitivity were deliberated. Significant biological pathways that comprise these genes were classified, along with their frequently associated genes (IFIT1/3, IFITM1, IRF7, ISG15, MX1/2 and OAS1/3/L). Major upstream regulators from the IRDS genes were identified, and different IFN types regulating these genes were outlined. Functional interfaces of IRDS proteins with DNA/RNA/ATP/GTP/NADP biomolecules featured a well-defined pharmacophore model for STAT1/IRF7-dsDNA and OAS1/OAS3/IFIH1-dsRNA complexes, as well as for the genes binding to GDP or NADP+. The Lys amino acid was found commonly interacting with the ATP phosphate group from OAS1/EIF2AK2/IFIH1 genes. Considering the premise that targeting IRDS genes mediated resistance offers an efficient strategy to resensitize tumour cells and enhances the outcome of anti-cancer treatment, this review can add some novel insights to the field.

• MeSH
• Adaptor Proteins, Signal Transducing physiology   MeSH
• Transcriptional Activation MeSH
• Drug Resistance, Neoplasm genetics   physiology   MeSH
• RNA, Double-Stranded MeSH
• Interferon Regulatory Factor-7 MeSH
• Interferons metabolism   physiology   MeSH
• Intracellular Signaling Peptides and Proteins MeSH
• Humans MeSH
• Cell Line, Tumor MeSH
• DNA Damage genetics   physiology   MeSH
• RNA-Binding Proteins MeSH
• Signal Transduction MeSH
• STAT1 Transcription Factor MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

Chemoresistance remains the major challenge for successful treatment of acute myeloid leukemia (AML). Although recent mouse studies suggest that treatment response of genetically and immunophenotypically indistinguishable AML can be influenced by their different cells of origin, corresponding evidence in human disease is still largely lacking. By combining prospective disease modeling using highly purified human hematopoietic stem or progenitor cells with retrospective deconvolution study of leukemia stem cells (LSCs) from primary patient samples, we identified human hematopoietic stem cells (HSCs) and common myeloid progenitors (CMPs) as two distinctive origins of human AML driven by Mixed Lineage Leukemia (MLL) gene fusions (MLL-AML). Despite LSCs from either MLL-rearranged HSCs or MLL-rearranged CMPs having a mature CD34-/lo/CD38+ immunophenotype in both a humanized mouse model and primary patient samples, the resulting AML cells exhibited contrasting responses to chemotherapy. HSC-derived MLL-AML was highly resistant to chemotherapy and expressed elevated amounts of the multispecific anion transporter ABCC3. Inhibition of ABCC3 by shRNA-mediated knockdown or with small-molecule inhibitor fidaxomicin, currently used for diarrhea associated with Clostridium difficile infection, effectively resensitized HSC-derived MLL-AML toward standard chemotherapeutic drugs. This study not only functionally established two distinctive origins of human LSCs for MLL-AML and their role in mediating chemoresistance but also identified a potential therapeutic avenue for stem cell-associated treatment resistance by repurposing a well-tolerated antidiarrhea drug already used in the clinic.

• MeSH
• Leukemia, Myeloid, Acute * drug therapy   genetics   MeSH
• Hematopoietic Stem Cells MeSH
• Humans MeSH
• Mice MeSH
• Prospective Studies MeSH
• Myeloid-Lymphoid Leukemia Protein * genetics   MeSH
• Retrospective Studies MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH