The Lupane-type Triterpene 30-Oxo-calenduladiol Is a CCR5 Antagonist with Anti-HIV-1 and Anti-chemotactic Activities

Abstract

The existence of drug-resistant human immunodeficiency virus (HIV) viruses in patients receiving antiretroviral treatment urgently requires the characterization and development of new antiretroviral drugs designed to inhibit resistant viruses and to complement the existing antiretroviral strategies against AIDS. We assayed several natural or semi-synthetic lupane-type pentacyclic triterpenes in their ability to inhibit HIV-1 infection in permissive cells. We observed that the 30-oxo-calenduladiol triterpene, compound 1, specifically impaired R5-tropic HIV-1 envelope-mediated viral infection and cell fusion in permissive cells, without affecting X4-tropic virus. This lupane derivative competed for the binding of a specific anti-CCR5 monoclonal antibody or the natural CCL5 chemokine to the CCR5 viral coreceptor with high affinity. 30-Oxo-calenduladiol seems not to interact with the CD4 antigen, the main HIV receptor, or the CXCR4 viral coreceptor. Our results suggest that compound 1 is a specific CCR5 antagonist, because it binds to the CCR5 receptor without triggering cell signaling or receptor internalization, and inhibits RANTES (regulated on activation normal T cell expressed and secreted)-mediated CCR5 internalization, intracellular calcium mobilization, and cell chemotaxis. Furthermore, compound 1 appeared not to interact with ␤-chemokine receptors CCR1, CCR2b, CCR3, or CCR4. Thereby, the 30-oxocalenduladiol-associated anti-HIV-1 activity against R5-tropic virus appears to rely on the selective occupancy of the CCR5 receptor to inhibit CCR5-mediated HIV-1 infection. Therefore, it is plausible that the chemical structure of 30-oxo-calenduladiol or other related dihydroxylated lupane-type triterpenes could represent a good model to develop more potent anti-HIV-1 molecules to inhibit viral infection by interfering with early fusion and entry steps in the HIV life cycle. The human immunodeficiency virus (HIV)7 pandemic is a medical challenge and represents the public health crisis of our time (1-5). Antiretroviral treatment achieves long-lasting viral suppression and, subsequently, reduces the morbidity and mortality of HIV-infected individuals. However, current drugs do not eradicate HIV infection and lifelong treatment might be needed (2).Emerging drug-resistant HIV viruses, in patients receiving high active antiretroviral treatment, urgently needs the development of new antiretroviral molecules designed to inhibit resistant viruses, because many patients treated during the past decades harbor viral strains with reduced susceptibilities to many if not all available drugs (2, 6). In this matter, pentacyclic triterpenes represent a varied class of natural products presenting antitumor and antiviral activities (7-9). A well studied pentacyclic lupane-type triterpene is the betulinic acid (3␤-hydroxy-lup-20(29)-en-28-oic acid), widely distributed throughout the plant kingdom, which presents anti-inflammatory, anti-malarial, and anti-HIV-1 effects in vitro (7, 9, 10). Although its mechanism of action has not been f...

Gelsolin activity controls efficient early HIV-1 infection

“…Cells were passaged every 3 days. The HeLa-P5 cells, constitutively expressing CXCR4 and stably transfected with human CD4 and CCR5 cDNAs and with an HIV-long terminal repeat-driven-β-galactosidase reporter gene [23,45,47], as well as, HeLa-243 and HeLa-ADA cells, co-expressing the Tat and X4- and R5-tropic HIV- 1-Env proteins, respectively, were provided by Dr. M. Alizon (Hôpital Cochin, Paris, France) and cultured as previously described [23,45,47]. The 293T cell line was similarly cultured in supplemented DMEM (Lonza).…”

“…A β-Galactosidase cell fusion assay was performed as previously described [23,45,47]. Briefly, HeLa-243 or HeLa ADA cells were mixed with previously treated HeLa-P5 cells ( overexpressing free EGFP, gelsolin-EGFP, free gelsolin, scrambled or siRNA(1 + 2)-GSN oligos ), in 96-well plates, in a 1:1 ratio (20,000 total cells), under each experimental condition.…”

HIV-1 requires Arf6-mediated membrane dynamics to efficiently enter and infect T lymphocytes

“…Cells were then washed by ice-cold PBS, fixed in PBS with 1% paraformaldehyde, and analyzed by flow cytometry (XL-MCL system; Beckman-Coulter, CA), as described (Valenzuela-Fernandez et al , 2005; Barrero-Villar et al , 2008, 2009; Barroso-Gonzalez et al , 2009a). …”

“…Permissive TZMbl cells, transiently expressing the different Arf6-HA or Arf6-EGFP constructs or fluorescent siRNA oligonucleotides (λ Em 680 nm) together with PH-ECFP and CD4-DsRed molecules, or Arf6-ECFP constructs with CD4-DsRed, were imaged with an inverted microscope Zeiss 200M (Zeiss, Jena, Germany) through a 1.45 NA objective (αFluar, 100×, Zeiss), as described (Barroso-Gonzalez et al , 2009a, 2009b). Experiments were performed and imaged with cells in a Krebs-HEPES buffer containing 2 mM Ca 2 + at 37°C.…”

Unifying Mechanisms of Action of the Anticancer Activities of Triterpenoids and Synthetic Analogs

“…A recent study investigated the effects of several lupane-derived compounds for their anti-HIV and anti-chemotactic activity by their interactions with chemokine receptors [95]. 30-Oxo-calenduladiol exhibited high CCR antagonist activity, suggesting a potential role for lupane-derived molecules as antiviral agents.…”