Human African trypanosomiasis (HAT), commonly known as sleeping sickness has remained a serious health problem in many African countries with thousands of new infected cases annually. Chemotherapy, which is the main form of control against HAT has been characterized lately by the viewpoints of toxicity and drug resistance issues. Recently, there have been a lot of emphases on the use of medicinal plants world-wide. Morinda lucida Benth. is one of the most popular medicinal plants widely distributed in Africa and several groups have reported on its anti-protozoa activities. In this study, we have isolated one novel tetracyclic iridoid, named as molucidin, from the CHCl3 fraction of the M. lucida leaves by bioassay-guided fractionation and purification. Molucidin was structurally elucidated by (1)H and (13)C NMR including HMQC, HMBC, H-H COSY and NOESY resulting in tetracyclic iridoid skeleton, and its absolute configuration was determined. We have further demonstrated that molucidin presented a strong anti-trypanosomal activity, indicating an IC50 value of 1.27 μM. The cytotoxicity study using human normal and cancer cell lines indicated that molucidin exhibited selectivity index (SI) against two normal fibroblasts greater than 4.73. Furthermore, structure-activity relationship (SAR) study was undertaken with molucidin and oregonin, which is identical to anti-trypanosomal active components of Alnus japonica. Overlapping analysis of the lowest energy conformation of molucidin with oregonin suggested a certain similarities of aromatic rings of both oregonin and molucidin. These results contribute to the future drug design studies for HAT.

Trypanosoma brucei parasites are kinetoplastid protozoa that devastate the health and economic well-being of millions of people in Africa through the disease human African trypanosomiasis (HAT). New chemotherapy has been eagerly awaited due to severe side effects and the drug resistance issues plaguing current drugs. Recently, there has been an emphasis on the use of medicinal plants worldwide. Morinda lucida Benth. is a popular medicinal plant widely distributed in Africa, and several research groups have reported on the antiprotozoal activities of this plant. In this study, we identified three novel tetracyclic iridoids, molucidin, ML-2-3, and ML-F52, from the CHCl3 fraction of M. lucida leaves, which possess activity against the GUTat 3.1 strain of T. brucei brucei. The 50% inhibitory concentrations (IC50) of molucidin, ML-2-3, and ML-F52 were 1.27 μM, 3.75 μM, and 0.43 μM, respectively. ML-2-3 and ML-F52 suppressed the expression of paraflagellum rod protein subunit 2, PFR-2, and caused cell cycle alteration, which preceded apoptosis induction in the bloodstream form of Trypanosoma parasites. Novel tetracyclic iridoids may be promising lead compounds for the development of new chemotherapies for African trypanosomal infections in humans and animals.

Macropinocytosis is a ubiquitous cellular uptake mechanism of peptide‐based intracellular delivery. This entry pathway shows promise as a route for the intracellular uptake of biomacromolecules and nanoparticles. In this work, we obtained the 8‐residue analogue P4A bearing higher macropinocytosis induction ability. P4A contains vital cysteine residues in its sequence, which immediately reacts with cystine in culture medium to convert into its oxidized forms, including the intramolecularly oxidized form (oxP4A) as the dominant and active species. The conjugate of oxP4A and the membrane lytic peptide LK15 delivered bioactive proteins into cells; notably, this peptide delivered functional proteins fused with a negatively charged protein tag at a significantly reduced amount (up to nanomolar range) without compromising the delivery efficiency and the cellular activities of delivered proteins.

A critical part of the development of CXCR4 modulators is to have a simple and sensitive assay system to complement the search by screening and evaluating the binding affinity. Herein, a NanoBRET assay system was developed, and its feasibility as a high-throughput screening tool for potent CXCR4 ligands was ascertained. TAMRA-Ac-TZ14011, a fluorescent-labeled CXCR4 antagonist, was adopted as a fluorescent acceptor of bioluminescent energy from N-terminally fused NanoLuc-CXCR4 stably expressed in CHO cells. The ratio of fluorescence at 620 nm to the luminescence at 460 nm represents the interaction between test compounds and CXCR4. We have demonstrated in the present study that the NanoBRET assay system is applicable for the evaluation of CXCR4 ligands using the combination of TAMRA-Ac-TZ14011 as an acceptor and NanoLuc tagged to CXCR4 as a bioluminescent donor expressed in living cells. IC50 values of known CXCR4 ligands were determined and found to be compatible with the values obtained by other existing and sensitive methods, such as SDF-1:3.2 nM, Ac-TZ14011:15.3 nM, and FC131:4.5 nM, which confirmed the feasibility of our system (Z′ values ≥0.5). The introduction of an IL-6 secretory signaling peptide (secNluc-CXCR4) further enhanced the expression and trafficking of the tagged receptor, which, in turn, increased the dynamic range of the NanoBRET assay system. Thus, we have successfully developed a NanoBRET system in living cells, which is simple, homogeneous, and useful in multiwell plate screening of potent CXCR4 ligands.

Despite remarkable advances in combination antiretroviral therapy (cART), human immunodeficiency virus type 1 (HIV-1) infection remains incurable due to the incomplete elimination of the replication-competent virus, which persists in latent reservoirs. Strategies for targeting HIV reservoirs for eradication that involves reactivation of latent proviruses while protecting uninfected cells by cART are urgently needed for cure of HIV infection. We screened medicinal plant extracts for compounds that could reactivate the latent HIV-1 provirus and identified a procyanidin trimer C1 derived from Theobroma cacao as a potent activator of the provirus in human T cells latently infected with HIV-1. This reactivation largely depends on the NF-κB and MAPK signaling pathways because either overexpression of a super-repressor form of IκBα or pretreatment with a MEK inhibitor U0126 diminished provirus reactivation by C1. A pan-PKC inhibitor significantly blocked the phorbol ester-induced but not the C1-induced HIV-1 reactivation. Although C1-induced viral gene expression persisted for as long as 48 h post-stimulation, NF-κB-dependent transcription peaked at 12 h post-stimulation and then quickly declined, suggesting Tat-mediated self-sustainment of HIV-1 expression. These results suggest that procyanidin C1 trimer is a potential compound for reactivation of latent HIV-1 reservoirs.