Components of cannabis, including THC, may decelerate peripheral monocyte processes that are implicated in HIV-associated neuroinflammation.

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New approach methodologies (NAMs) are increasingly being used for regulatory decision making by agencies worldwide because of their potential to reliably and efficiently produce information that is fit for purpose while reducing animal use. This article summarizes the ability to use NAMs for the assessment of human health effects of industrial chemicals and pesticides within the United States, Canada, and European Union regulatory frameworks. While all regulations include some flexibility to allow for the use of NAMs, the implementation of this flexibility varies across product type and regulatory scheme. This article provides an overview of various agencies’ guidelines and strategic plans on the use of NAMs, and specific examples of the successful application of NAMs to meet regulatory requirements. It also summarizes intra- and inter-agency collaborations that strengthen scientific, regulatory, and public confidence in NAMs, thereby fostering their global use as reliable and relevant tools for toxicological evaluations. Ultimately, understanding the current regulatory landscape helps inform the scientific community on the steps needed to further advance timely uptake of approaches that best protect human health and the environment.

The aryl hydrocarbon receptor (AHR) is a cytosolic ligand-activated transcription factor involved in xenobiotic sensing, cell cycle regulation, and cell development. In humans, the activation of AHR by 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a high affinity AHR-ligand, impairs the secretion of immunoglobulin M (IgM) to suppress humoral immunity. However, the mechanisms bridging the activation of AHR and the impairment of IgM secretion by human primary B cells remain poorly understood. Recent transcriptomic analysis revealed upregulation of lymphocyte-specific protein tyrosine kinase (LCK) in AHR-activated human primary B cells. LCK is a well-characterized tyrosine kinase that phosphorylates critical signaling proteins involved in activation and cytokine production in T cells. Conversely, the role of LCK in human primary B cells is not well understood. In the current studies, we have verified the transcriptomic finding by detecting AHR-mediated upregulation of LCK protein in human primary B cells. We also confirmed the role of AHR in the upregulation of LCK by using a specific AHR antagonist, which abolished the AHR-mediated increase of LCK. Furthermore, we have confirmed the role of LCK in the AHR-mediated suppression of IgM by using LCK specific inhibitors, which restored the IgM secretion by human B cells in the presence of TCDD. Collectively, the current studies demonstrate a novel role of LCK in IgM response and provide new insights into the mechanism for AHR-mediated impairment of immunoglobulin secretion by human primary B cells.

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Plasmacytoid dendritic cells (pDC) play a crucial role in host anti-viral immune response through the secretion of type I interferon. Interferon alpha (IFNα), a type I IFN, is critical for mounting the initial response to viral pathogens. A consequence of Human Immunodeficiency Virus-1 (HIV) infection is a decrease in both pDC number and function, but prolonged pDC activity has been linked with progression from HIV infection to the development of AIDS. HIV patients in the US routinely use cannabinoid-based therapies to combat the side effects of HIV infection and antiretroviral therapy (ART). However, cannabinoids, including Δ9-tetrahydrocannabinol (THC), are well-characterized immunosuppressants. Here we report that THC suppressed secretion of IFNα by pDC from both healthy and HIV+ donors through a mechanism involving impaired phosphorylation of interferon regulatory factor 7 (IRF-7). These results suggest that THC can suppress pDC function during the early host antiviral by dampening pDC activation.

Skin sensitization testing is a regulatory requirement for safety evaluations of pesticides in multiple countries. Globally harmonized test guidelines that include in chemico and in vitro methods reduce animal use, but no single assay is recommended as a complete replacement for animal tests. Defined approaches (DAs) that integrate data from multiple non-animal methods are accepted; however, the methods that comprise them have been evaluated using monoconstituent substances rather than mixtures or formulations. To address this data gap, we tested 27 agrochemical formulations in the direct peptide reactivity assay (DPRA), the KeratinoSens™ assay, and the human cell line activation test (h-CLAT). These data were used as inputs to evaluate three DAs for hazard classification of skin sensitization potential and two DAs for potency categorization. When compared to historical animal results, balanced accuracy for the DAs for predicting in vivo skin sensitization hazard (i.e., sensitizer vs. nonsensitizer) ranged from 56 to 78%. The best performing DA was the “2 out of 3 (2o3)” DA, in which the hazard classification was based on two concordant results from the DPRA, KeratinoSens, or h-CLAT. The KE 3/1 sequential testing strategy (STS), which uses h-CLAT and DPRA results, and the integrated testing strategy (ITSv2), which uses h-CLAT, DPRA, and an in silico hazard prediction from OECD QSAR Toolbox, had balanced accuracies of 56–57% for hazard classification. Of the individual test methods, KeratinoSens had the best performance for predicting in vivo hazard outcomes. Its balanced accuracy of 81% was similar to that of the 2o3 DA (78%). For predicting potency categories defined by the United Nations Globally Harmonized System of Classification and Labelling of Chemicals (GHS), the correct classification rate of the STS was 52% and that of the ITSv2 was 43%. These results demonstrate that non-animal test methods have utility for evaluating the skin sensitization potential of agrochemical formulations as compared to animal reference data. While additional data generation is needed, testing strategies such as DAs anchored to human biology and mechanistic information provide a promising approach for agrochemical formulation testing.

Plasmacytoid dendritic cells (pDC) compose 0.2-0.5% of circulating leukocytes but play a significant role in mounting host immune responses. Elevated and chronic activation of pDC are implicated in autoimmune disease like systemic lupus erythematosus and rheumatoid arthritis. Δ 9tetrahydrocannabinol (THC) is a well characterized cannabinoid with potent anti-inflammatory activity, but acceptance of THC as a treatment for autoimmune disorders has been hindered due to psychotropic activity. The psychotropic effects of THC are mediated through cannabinoid receptor 1 ( CB1) expressed in the central nervous system while the immunomodulatory effects of THC result from THC binding to CB1 and CB2 on immune cells. Synthetic CB2-selective agonists have been developed to explore immune modulation by cannabinoids in the absence of psychotropic effects. The goal of these studies was to determine if the CB2-selective agonists, JWH-015 and JWH-133, have comparable efficacy to THC in modulating IFNα and TNFα responses by primary human pDC. Treatment with JWH-133 and JWH-015 inhibited CpG-induced IFNα and TNFα responses by pDC. Further, the phosphorylation of IRF7, TBK1, NFκB, and IKKγ, key events in pDC activation, were suppressed by THC, JWH-133, and JWH-015. Likewise, the phosphorylation of AKT at the S473 and T308 residues were differentially modulated by treatment with THC and both JWH compounds. Collectively, these results demonstrate the potential for CB2 targeted therapeutics for treatment of inflammatory conditions involving aberrant pDC activity.