Cannabis research has historically focused on the most prevalent cannabinoids. However, extracts with a broad spectrum of secondary metabolites may have increased efficacy and decreased adverse effects compared to cannabinoids in isolation. Cannabis's complexity contributes to the length and breadth of its historical usage, including the individual application of the leaves, stem barks, and roots, for which modern research has not fully developed its therapeutic potential. This study is the first attempt to profile secondary metabolites groups in individual plant parts comprehensively. We profiled 14 cannabinoids, 47 terpenoids (29 monoterpenoids, 15 sesquiterpenoids, and 3 triterpenoids), 3 sterols, and 7 flavonoids in cannabis flowers, leaves, stem barks, and roots in three chemovars available. Cannabis inflorescence was characterized by cannabinoids (15.77-20.37%), terpenoids (1.28-2.14%), and flavonoids (0.07-0.14%); the leaf by cannabinoids (1.10-2.10%), terpenoids (0.13-0.28%), and flavonoids (0.34-0.44%); stem barks by sterols (0.07-0.08%) and triterpenoids (0.05-0.15%); roots by sterols (0.06-0.09%) and triterpenoids (0.13-0.24%). This comprehensive profile of bioactive compounds can form a baseline of reference values useful for research and clinical studies to understand the "entourage effect" of cannabis as a whole, and also to rediscover therapeutic potential for each part of cannabis from their traditional use by applying modern scientific methodologies. Cannabis is a complex herbal medicine containing several classes of secondary metabolites, including at least 104 cannabinoids, 120 terpenoids (including 61 monoterpenes, 52 sesquiterpenoids, and 5 triterpenoids), 26 flavonoids, and 11 steroids among 545 identified compounds 1-6. The postulated biosynthetic pathways for these metabolite groups 7,8 are outlined in Fig. 1. Cannabis has attracted a new wave of interest for its broad medicinal applications as 1) an analgesic, potentially as an adjunct to or substitute for opiates in the treatment of chronic pain 9 , and 2) an appetite stimulant and digestive aid 10 , among others. Since the 1960s, the research has focussed mainly on cannabinoids, ∆ 9-tetrahydrocannabinol (∆ 9-THC), and cannabidiol (CBD) in particular 11-28. The major psychoactive content expressed as total THC decreases in the order of inflorescences (10-12%), leaves (1-2%), stems (0.1-0.3%), roots (<0.03%), and seeds (generally absent) 29. As such, female flower tops are harvested while other parts are often discarded by growers 29. This is a potentially unnecessary waste. As an ancient medicine in various cultures, each part of the cannabis plant has been historically indicated with a wide range of applications relating mostly to painkilling, inflammation releasing, and mental illness treatment 30-33. Compounds other than ∆ 9-THC and CBD may contribute to the therapeutic effects of each plant part in their traditional uses. Minor cannabinoids, such as cannabinol (CBN), cannabigerol (CBG), cannabichromene (CBC), also have broad therapeu...

Cdc42 plays an evolutionarily conserved role in promoting cell polarity and is indispensable during epithelial morphogenesis. To further investigate the role of Cdc42, we have used a three-dimensional matrigel model, in which single Caco-2 cells develop to form polarized cysts. Using this system, we previously reported that Cdc42 controls mitotic spindle orientation during cell division to correctly position the apical surface in a growing epithelial structure. In the present study, we have investigated the specific downstream effectors through which Cdc42 controls this process. Here, we report that Par6B and its binding partner, atypical protein kinase C (aPKC), are required to regulate Caco-2 morphogenesis. Depletion or inhibition of Par6B or aPKC phenocopies the loss of Cdc42, inducing misorientation of the mitotic spindle, mispositioning of the nascent apical surface, and ultimately, the formation of aberrant cysts with multiple lumens. Mechanistically, Par6B and aPKC function interdependently in this context. Par6B localizes to the apical surface of Caco-2 cysts and is required to recruit aPKC to this compartment. Conversely, aPKC protects Par6B from proteasomal degradation, in a kinase-independent manner. In addition, we report that depletion or inhibition of aPKC induces robust apoptotic cell death in Caco-2 cells, significantly reducing both cyst size and number. Cell survival and apical positioning depend upon different thresholds of aPKC expression, suggesting that they are controlled by distinct downstream pathways. We conclude that Par6B and aPKC control mitotic spindle orientation in polarized epithelia and, furthermore, that aPKC coordinately regulates multiple processes to promote morphogenesis.Cdc42 is a small GTPase that plays a fundamental role in promoting cell polarity across evolution and in a wide range of biological contexts (1). For instance, Cdc42 is required for bud emergence in Saccharomyces cerevisiae (2), asymmetric cell division in Caenorhabditis elegans (3) and directed cell migration in astrocytes (4). In epithelial cells, Cdc42 is indispensable during morphogenesis, controlling tight junction formation (5-7), the delivery of basolateral proteins (8), and apical surface formation through the regulated trafficking of vacuolar apical components (9). To further investigate the role of Cdc42, we have exploited a three-dimensional model of epithelial morphogenesis, in which single Caco-2 cells are cultured in matrigel to form polarized cysts (Fig. 1A). Using this system, we previously reported that Cdc42 controls mitotic spindle orientation to correctly position the nascent apical surface in a growing cyst (10). These findings highlight an intriguing relationship between epithelial mitosis and morphogenesis, which couples cell proliferation to tissue architecture.A major challenge now is to determine the specific upstream regulators and downstream effectors through which Cdc42 controls epithelial morphogenesis. Recent work has focused on the identification of specific guanine nucleotid...

A systematic screen of Cdc42 targets was carried out in human bronchial epithelial cells. Two kinases, PAK4 and Par6B/aPKC, were identified and are required for maturation of primordial junctions into apical junctions. PAK4 recruitment to primordial junctions is Cdc42-dependent, but maintenance at junctions during maturation is Par6B-dependent.

Background BCMA-specific chimeric antigen receptor-T cells (CAR-Ts) have exhibited remarkable efficacy in refractory or relapsed multiple myeloma (RRMM); however, primary resistance and relapse exist with single-target immunotherapy. Bispecific CARs are proposed to mitigate these limitations. Methods We constructed a humanized bispecific BM38 CAR targeting BCMA and CD38 and tested the antimyeloma activity of BM38 CAR-Ts in vitro and in vivo. Twenty-three patients with RRMM received infusions of BM38 CAR-Ts in a phase I trial. Results BM38 CAR-Ts showed stronger in vitro cytotoxicity to heterogeneous MM cells than did T cells expressing an individual BCMA or CD38 CAR. BM38 CAR-Ts also exhibited potent antimyeloma activity in xenograft mouse models. In the phase I trial, cytokine release syndrome occurred in 20 patients (87%) and was mostly grade 1–2 (65%). Neurotoxicity was not observed. Hematologic toxicities were common, including neutropenia in 96% of the patients, leukopenia in 87%, anemia in 43% and thrombocytopenia in 61%. At a median follow-up of 9.0 months (range 0.5 to 18.5), 20 patients (87%) attained a clinical response and minimal residual disease-negativity (≤ 10–4 nucleated cells), with 12 (52%) achieving a stringent complete response. Extramedullary plasmacytoma was eliminated completely in 56% and partially in 33% and of 9 patients. The median progression-free survival was 17.2 months. Two relapsed patients maintained BCMA and CD38 expression on MM cells. Notably, BM38 CAR-Ts cells were detectable in 77.8% of evaluable patients at 9 months and 62.2% at 12 months. Conclusion Bispecific BM38 CAR-Ts were feasible, safe and significantly effective in patient with RRMM. Trial registration: Chictr.org.cn ChiCTR1800018143.

Although researchers have established that DNA methylation and active demethylation are dynamically regulated in plant cells, the molecular mechanism for the regulation of active DNA demethylation is not well understood. By using an Arabidopsis (Arabidopsis thaliana) line expressing the Promoter RESPONSIVE TO DEHYDRATION 29A:LUCIFERASE (ProRD29A:LUC) and Promoter cauliflower mosaic virus 35S:NEOMYCIN PHOSPHOTRANSFERASE II (Pro35S:NPTII) transgenes, we isolated an mbd7 (for methyl-CpG-binding domain protein7) mutant. The mbd7 mutation causes an inactivation of the Pro35S:NPTII transgene but does not affect the expression of the ProRD29A:LUC transgene. The silencing of the Pro35S:NPTII reporter gene is associated with DNA hypermethylation of the reporter gene. MBD7 interacts physically with REPRESSOR OF SILENCING5/INCREASED DNA METHYLATION2, a protein in the small heat shock protein family. MBD7 prefers to target the genomic loci with high densities of DNA methylation around chromocenters. The Gypsy-type long terminal repeat retrotransposons mainly distributed around chromocenters are most affected by mbd7 in all transposons. Our results suggest that MBD7 is required for active DNA demethylation and antisilencing of the genomic loci with high densities of DNA methylation in Arabidopsis.