Post-Hyaluronic Acid Filler Reaction Treated With Abrocitinib: A Case Report.

Post-hyaluronic acid filler nodules are uncommon, unpredictable complications that present a challenge to clinical therapy. We report a case of a female in her fifties who developed edema and nodules 6 weeks after hyaluronic acid (HA) filler injection. After minimal improvement with oral steroids and intralesional hyaluronidase, a trial of oral abrocitinib was initiated, which yielded significant clinical improvement. Thus, abrocitinib may be a novel therapeutic option for delayed-onset nodules following injection of hyaluronic acid. J Drugs Dermatol. 2024;23(1):1355-1356.   doi:10.36849/JDD.7271.

Talquetamab-Induced Grover's Disease.

Kresch M, Guénin S, Mubasher A, et al. Talquetamab-induced Grover’s disease. J Drugs Dermatol. 2023;22(8):828-829. doi:10.36849/JDD.7170.

Rethinking the Inflammatory Balance in Psoriasis and Atherosclerosis.

Guénin S, Kazemi A, Cline A, et al. Rethinking the inflammatory balance in psoriasis and atherosclerosis. J Drugs Dermatol. 2023;22(8):832-833. doi:10.36849/JDD.7082.

Characteristics, outcome, and response to therapy of multirefractory chronic immune thrombocytopenia.

Refractory immune thrombocytopenia (ITP) was previously defined as lack of a minimum response to splenectomy and the requirement for long-term treatment to reduce the risk of significant bleeding events. In this multicenter study, we included 37 patients with multirefractory ITP, defined as no response to splenectomy, rituximab, romiplostim, and eltrombopag. As compared with a historical cohort of 183 ITP patients, matched on the calendar year of ITP diagnosis with a 5:1 ratio, patients with multirefractory ITP were more likely to have secondary ITP (odds ratio [OR], 4.84; 95% confidence interval [CI], 1.31-17.86; P = .018) and monoclonal gammopathy of undetermined significance (OR, 5.94; 95% CI, 1.08-32.48; P = .04). The median duration of ITP before being recognized as multirefractory was 78 months (range, 6-450). The patients showed failure of a median of 10.5 prior treatment lines for ITP (range, 6-15). At the end of follow-up (median, 84 months; range, 12-455), only 1/14 patients achieved response with immunosuppressant therapy alone. By contrast, 7/10 patients achieved response with a combination of immunosuppressant therapy and thrombopoietin-receptor agonists that lasted for a median of 15 months (range, 6-32). Throughout the course of ITP, 5/37 patients died, 3 with ITP (bleeding, n = 2; sepsis n = 1); 15 (40%) had at least 1 bacterial infection and 9 (24%) at least 1 episode of thrombosis. In conclusion, multirefractory ITP was associated with high morbidity and mortality. Combining an immunosuppressant therapy with thrombopoietin-receptor agonists may be a good strategy for management for these patients with severe disease.

Hamster Melatonin Receptors: Cloning and Binding Characterization of MT1 and Attempt to Clone MT2.

For many years, it was of interest to identify the sequences encoding the two melatonin receptors (MT1 and MT2) from various species. After publishing the basic molecular characterization of the human, rat, mouse, sheep, and platypus MT1, MT2, or Mel1c receptors, we began cloning the genes from other animals, such as birds, bats, and vipers. The goal was to advance the receptor crystallization, which could greatly contribute the understanding of the sequence/stability relationship. European hamster MT1 receptor was cloned for the first time from this gender, was expressed in stable form in cells, and its binding characterized with a sample of 19 melatonin ligands. Siberian hamster (Phodopus sungorus) expresses a non-functional MT2. We observed that unlike this hamster, the European hamster (Cricetus cricetus) does not have a stop codon in the MT2 sequence. Thus, we undertook the tedious task of cloning the MT2 receptor. We partially succeeded, sequencing the complete exon 2 and a fragment of exon 1 (from putative amino acids 12 to 38 and 77 to 323), after several years of efforts. In order to show that the protein parts we cloned were capable to sustain some binding capacities, we designed a chimeric MT2 receptor using a consensus sequence to replace the unknown amino acids, based on other small rodent MT2 sequences. This chimeric construct could bind melatonin in the nanomolar range. This work is meant to be the basis for attempts from other laboratories of the community to determine the complete natural sequence of the European hamster MT2 receptor. The present work is the first to show that, among the hamsters, if the Siberian is a natural knockout for MT2, the European one is not.

Why Are We Still Cloning Melatonin Receptors? A Commentary.

Cloning may seem to be a view from the past. The time before software, computers and AI were invented. It seems to us worth discussing these points in view of our favorite target: the melatoninergic system. In a few stances, it might be important to point out that even in the new era of dry science, there is still a need to experiment and to prove at the bench that our in silico assertions are right. Most of the living animals express to some extend the melatonin receptors. Some of these animal genomes were completely or partially sequenced, and it is tempting to extract from this huge information the sequence(s) of our favorite genes (MLT receptors). Then, why bother cloning, as opposed to simply built the gene and express it in a host cell? Because the genetic boundaries of the expressed sequence(s) are not 100% sure. Because the melatonin receptor gene(s) comprise a first exon 25,000 base pair far from the second one and the limits between this Ex1 and In1-as between In1 and Ex2-are subject to changes that might have a huge impact on the biochemical properties of the receptor, once expressed. Because a receptor is a biochemical entity with characteristics that are important for the functioning of this particular pathway, and more generally, for the functioning of life.

Approach to the specificity and selectivity between D2 and D3 receptors by mutagenesis and binding experiments part I: Expression and characterization of D2 and D3 receptor mutants.

D3/D2 sub-specificity is a complex problem to solve. Indeed, in the absence of easy structural biology of the G-protein coupled receptors, and despite key progresses in this area, the systematic knowledge of the ligand/receptor relationship is difficult to obtain. Due to these structural biology limitations concerning membrane proteins, we favored the use of directed mutagenesis to document a rational towards the discovery of markedly specific D3 ligands over D2 ligands together with basic binding experiments. Using our methodology of stable expression of receptors in HEK cells, we constructed the gene encoding for 24 mutants and 4 chimeras of either D2 or D3 receptors and expressed them stably. Those cell lines, expressing a single copy of one receptor mutant each, were stably constructed, selected, amplified and the membranes from them were prepared. Binding data at those receptors were obtained using standard binding conditions for D2 and D3 dopamine receptors. We generated 26 new molecules derived from D2 or D3 ligands. Using 8 reference compounds and those 26 molecules, we characterized their binding at those mutants and chimeras, exemplifying an approach to better understand the difference at the molecular level of the D2 and D3 receptors. Although all the individual results are presented and could be used for minute analyses, the present report does not discuss the differences between D2 and D3 data. It simply shows the feasibility of the approach and its potential.

Biochemistry, structure, and cellular internalization of a four nanobody-bearing Fc dimer.

VHH stands for the variable regions of heavy chain only of camelid IgGs. The VHH family forms a set of interesting proteins derived from antibodies that maintain their capacity to recognize the antigen, despite their relatively small molecular weight (in the 12,000 Da range). Continuing our exploration of the possibilities of those molecules, we chose to design alternative molecules with maintained antigen recognition, but enhanced capacity, by fusing four VHH with one Fc, the fragment crystallizable region of antibodies. In doing so, we aimed at having a molecule with superior quantitative antigen recognition (×4) while maintaining its size below the 110 kDa. In the present paper, we described the building of those molecules that we coined VHH2 -Fc-VHH2 . The structure of VHH2 -Fc-VHH2 in complex with HER2 antigen was determined using electronic microscopy and modeling. The molecule is shown to bind four HER2 proteins at the end of its flexible arms. VHH2 -Fc-VHH2 also shows an internalization capacity via HER2 receptor superior to the reference anti-HER2 monoclonal antibody, Herceptin®, and to a simple fusion of two VHH with one Fc (VHH2 -Fc). This new type of molecules, VHH2 -Fc-VHH2 , could be an interesting addition to the therapeutic arsenal with multiple applications, from diagnostic to therapy.

Studies of the melatonin binding site location onto quinone reductase 2 by directed mutagenesis.

Melatonin is a neurohormone implicated in both biorhythm synchronization and neuroprotection from oxidative stress. Its functions are mediated by two G-protein-coupled-receptors (MT1 and MT2) and MT3, which corresponds to quinone oxidoreductase 2 (QR2). To determine the binding site of QR2 for melatonin, point mutations of residues crucial for the enzymatic activity of hQR2 were performed. The substitution of the hydrophobic residues Phe126, Ile128 and Phe178 by tyrosines at the active site significantly increased enzymatic activity and decreased the affinity of a structural analog of melatonin, the 2[(125)I]iodo-MCANAT. The mutation of residues implicated in zinc chelating (His(173); His(177)) had no effect on radioligand binding. Destabilisation of the cofactor FAD by mutation N18E showed that 2[(125)I]iodo-MCANAT binding was closely linked to the conformational integrity of human QR2. Surprisingly, the mutations C222F and N161A, which are distant from the determined binding site of the ligand, increased the affinity of 2[(125)I]iodo-MCANAT for hQR2. What seems to better explain the binding variations among the mutants are the activity recorded with BNAH and coenzyme Q1. Various hypotheses are discussed based on the various parameters used in the study: nature of the substrates and co-substrates and nature of the amino acid changes. This study, which constitutes the first structural analysis of hQR2, should enable to better understand the biological role of melatonin on this enzyme and particularly, the discrepancies between the pharmacologies of the melatonin binding site (MT3) and the QR2 catalytic activity.

Characterization of the Mel1c melatoninergic receptor in platypus (Ornithorhynchus anatinus).

Melatonin is a neurohormone produced in both animals and plants. It binds at least three G-protein-coupled receptors: MT1 and MT2, and Mel1cGPR. Mammalian GPR50 evolved from the reptilian/avian Mel1c and lost its capacity to bind melatonin in all the therian mammal species that have been tested. In order to determine if binding is lost in the oldest surviving mammalian lineage of monotremes we investigated whether the melatonin receptor has the ability to bind melatonin in the platypus (Ornithorhynchus anatinus), and evaluated its pharmacological profile. Sequence and phylogenetic analysis showed that platypus has in fact retained the ancestral Mel1c and has the capacity to bind melatonin similar to other mammalian melatonin receptors (MT1 and MT2), with an affinity in the 1 nM range. We also investigated the binding of a set of melatoninergic ligands used previously to characterize the molecular pharmacology of the melatonin receptors from sheep, rats, mice, and humans and found that the general profiles of these compounds make Mel1c resemble human MT1 more than MT2. This work shows that the loss of GPR50 binding evolved after the divergence of monotremes less than 190MYA in therian mammals.

Gene expression profiling during hibernation in the European hamster.

Hibernation is an exceptional physiological response to a hostile environment, characterized by a seasonal period of torpor cycles involving dramatic reductions of body temperature and metabolism, and arousal back to normothermia. As the mechanisms regulating hibernation are still poorly understood, here we analysed the expression of genes involved in energy homeostasis, torpor regulation, and daily or seasonal timing using digital droplet PCR in various central and peripheral tissues sampled at different stages of torpor/arousal cycles in the European hamster. During torpor, the hypothalamus exhibited strongly down-regulated gene expression, suggesting that hypothalamic functions were reduced during this period of low metabolic activity. During both torpor and arousal, many structures (notably the brown adipose tissue) exhibited altered expression of deiodinases, potentially leading to reduced tissular triiodothyronine availability. During the arousal phase, all analysed tissues showed increased expression of the core clock genes Per1 and Per2. Overall, our data indicated that the hypothalamus and brown adipose tissue were the tissues most affected during the torpor/arousal cycle, and that clock genes may play critical roles in resetting the body's clocks at the beginning of the active period.

Cellular knock-down of quinone reductase 2: a laborious road to successful inhibition by RNA interference.

NRH:quinone oxidoreductase 2 (QR2) is a long forgotten oxidoreductive enzyme that metabolizes quinones and binds melatonin. We used the potency of the RNA interference (RNAi)-mediated gene silencing to build a cellular model in which the role of QR2 could be studied. Because standard approaches were poorly successful, we successively used: (1) two chemically synthesized fluorescent small interfering RNA (siRNA) duplexes designed and tested for their gene silencing capacity leading to a maximal 40% QR2 gene silencing 48h post-transfection; (2) double transfection and cell-sorting of high fluorescent siRNA-transfected HT22 cells further enhancing QR2 RNAi silencing to 88%; (3) stable QR2 knock-down HT22 cell lines established with H1and U6 promoter driven QR2 short hairpin RNA (shRNA) encoding vectors, resulting in a 71-80% reduction of QR2 enzymatic activity in both QR2 shRNA HT22 cells. Finally, as a first step in the study of this cellular model, we observed a 42-48% reduction of menadione/BNAH-mediated toxicity in QR2 shRNA cells compared to the wild-type HT22 cells. Although becoming widespread and in some cases effective, siRNA-mediated cellular knock-down proves in the present work to be of marginal efficiency. Much development is required for this technique to be of general application.

Update on human alpha1-adrenoceptor subtype signaling and genomic organization.

Alpha1-adrenoceptors are G-protein-coupled receptors that bind catecholamines. Sixteen distinct human alpha1A-adrenoceptor isoforms have been identified from human tissues, including five full-length and 11 truncated versions. An updated scheme for the identification of alpha1A-adrenoceptor splice variants is proposed. Given the established roles of alpha1-adrenoceptors in benign prostatic hyperplasia, myocardial hypertrophy and other cardiovascular disorders, elucidation of the biological significance of the signaling diversity and potential pharmacological roles of alpha1A-adrenoceptor splice variants are important areas of future research.

Characterization of the melatoninergic MT3 binding site on the NRH:quinone oxidoreductase 2 enzyme.

Melatonin acts through a series of molecular targets: the G-protein coupled receptors, MT1 and MT2, and a third binding site, MT3, recently identified as the enzyme NRH:quinone oxydoreductase 2 (QR2). The relationship between the multiple physiological functions of melatonin and this enzyme remains unclear. Because of the relationship of QR2 with the redox status of cells, these studies could bring the first tools for a molecular rationale of the antioxidant effects of melatonin. In the present paper, we used a QR2-stably expressing cell line and hamster kidneys to compare the 2-[125I]-iodomelatonin and 2-[125I]-iodo-5-methoxycarbonylamino-N-acetyltryptamine binding data, and to characterize the MT3 binding site. We designed and tested compounds from two distinct chemicals series in a displacement assay of the two MT3 ligands, 2-[125I]-iodomelatonin and 2-[125I]-iodo-5-methoxycarbonylamino-N-acetyltryptamine from their cloned target. We also tested their ability to inhibit QR2 catalytic activity. These compounds were separated into two classes: those that bind within the catalytic site (and being inhibitors) and those that bind outside it (and therefore not being inhibitors). Compounds range from potent ligands (K(i) = 1 nM) to potent inhibitors (14 nM), and include one compound [NMDPEF: N-[2-(2-methoxy-6H-dipyrido[2,3-a:3,2-e]pyrrolizin-11-yl)ethyl]-2-furamide] active on both parameters in the low nanomolar range. To dissect the physio-pathological pathways in which QR2, MT3 and melatonin meet, one needs more compounds binding to MT3 and/or inhibitors of QR2 enzymatic activity. The compounds described in the present paper are new tools for such a task.