Isolation, Structural Modification, and HIV Inhibition of Pentacyclic Lupane-Type Triterpenoids from Cassine xylocarpa and Maytenus cuzcoina Oliver Callies, † Luis M. Bedoya, ‡,§ Manuela Beltra ́ n, ‡ Alejandro Muñ oz, ‡ Patricia Obregó n Calderó n, ‡ Alex A. Osorio, † Ignacio A. Jime ́ nez, † Jose ́ Alcamí, ‡ and Isabel L. Bazzocchi* ,† † Instituto Universitario de Bio-Orga ́ nica “Antonio Gonza ́ lez”, Universidad de La Laguna, Avenida Astrofísico Francisco Sa ́ nchez 2, 38206 La Laguna, Tenerife, Spain ‡ Unidad de Inmunopatología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Carretera Pozuelo Km.2, 28220-Majadahonda, Madrid, Spain § Departamento de Farmacología, Facultad de Farmacia, Universidad Complutense de Madrid, Ramó n y Cajal s/n, 28040, Madrid, Spain * S Supporting Information ABSTRACT: As a part of our investigation into new anti-HIV agents, we report herein the isolation, structure elucidation, and biological activity of six new (1-6) and 20 known (7-26) pentacyclic lupane-type triterpenoids from the stem of Cassine xylocarpa and root bark of Maytenus cuzcoina. Their stereo- structures were elucidated on the basis of spectroscopic and spectrometric methods, including 1D and 2D NMR techniques. To gain a more complete understanding of the structural requirements for anti-HIV activity, derivatives 27-48 were prepared by chemical modification of the main secondary metabolites. Sixteen compounds from this series displayed inhibitory effects of human immunodeficiency virus type 1 replication with IC 50 values in the micromolar range, highlighting compounds 12, 38, and 42 (IC 50 4.08, 4.18, and 1.70 μM, respectively) as the most promising anti-HIV agents. T hree decades into the acquired immunodeficiency syndrome (AIDS) epidemic, 35 million people are living with this disease worldwide and, up to now, 39 million people have died of AIDS-related illness. Currently, antiretroviral therapy has dramatically decreased morbidity and mortality and is capable of suppressing viral loads to undetectable levels, making AIDS a chronic disease. However, despite these advances, toxicity problems, patient adherence, adverse effects, and, particularly, the emergence of drug-resistant strains of human immunodeficiency virus (HIV), along with restricted accessibility in resource-poor areas, emphasize the urgent need for new anti-HIV drugs with simpler treatment regimens, acceptable toxicity, and novel mechanisms of action. 1 Natural compounds play an important role in overcoming the current urgency in anti-HIV/AIDS therapies. In fact, various natural compounds are currently in preclinical or clinical studies for the treatment of HIV infections. 2 In particular, lupane-type pentacyclic triterpenoids have been extensively studied as anti-HIV agents, providing a versatile structural platform for the discovery of new biologically active compounds. 3 Over the last two decades, many derivatives of the well-known lupane-type triterpenoid betulinic acid (3β- hydroxylup-20(29)-en-28-oic acid) have been reported to inhibit HIV replication, 4-6 including bevirimat [3-O-(3′,3′- dimethylsuccinyl)betulinic acid], a first-in-class HIV-1 matura- tion inhibitor. 7,8 In our previous search for anti-HIV agents from plants, pentacyclic triterpenoids from the oleanane series isolated from Celastraceae species have demonstrated potent anti-HIV activity. 9 In continuing research toward the discovery of naturally occurring antiretroviral agents, the current study reports the isolation, structural modification, and anti-HIV evaluation of a series of pentacyclic lupane-type triterpenoids with unusual substitution pattern on the triterpene scaffold. This series included 26 compounds (1-26), isolated from Cassine xylocarpa and Maytenus cuzcoina, and 22 analogues (27-48), 13 of which are reported for the first time. Their structures have been elucidated on the basis of spectrocopic analysis, including 1D and 2D NMR techniques (COSY, HSQC, HMBC, and ROESY), and spectrometric methods. The compounds have been tested for their effects on HIV type 1 replication, and the most effective inhibitors, those with a percentage of HIV-1 inhibition higher than 50%, were selected for further IC 50 calculations to compare potencies. The structure-activity relationship revealed that the regiosubstitu- Received: December 18, 2014 Article pubs.acs.org/jnp © XXXX American Chemical Society and American Society of Pharmacognosy A DOI: 10.1021/np501025r J. Nat. Prod. XXXX, XXX, XXX-XXX