marine drugs Article Isomalabaricane Triterpenes from the Marine Sponge Rhabdastrella sp. Kuei-Hung Lai 1,2,3,*, Zheng-Hao Huang 4,5, Mohamed El-Shazly 6,7, Bo-Rong Peng 4,5, Wen-Chi Wei 8,* and Jui-Hsin Su 4,5,* 1 PhD Program in Clinical Drug Development of Herbal Medicine, College of Pharmacy, Taipei Medical University, Taipei 11031, Taiwan 2 Graduate Institute of Pharmacognosy, College of Pharmacy, Taipei Medical University, Taipei 11031, Taiwan 3 Traditional Herbal Medicine Research Center, Taipei Medical University Hospital, Taipei 11031, Taiwan 4 National Museum of Marine Biology & Aquarium, Pingtung 94450, Taiwan; [email protected] (Z.-H.H.); [email protected] (B.-R.P.) 5 Graduate Institute of Marine Biology, National Dong Hwa University, Pingtung 94450, Taiwan 6 Department of Pharmaceutical Biology, German University in Cairo, Cairo 11432, Egypt; [email protected] 7 Department of Pharmacognosy and Natural Products Chemistry, Faculty of Pharmacy, Ain-Shams University, Organization of African Unity Street, Abassia, Cairo 11566, Egypt 8 Division of Basic Chinese Medicine, National Research Institute of Chinese Medicine, Taipei 11221, Taiwan * Correspondence: [email protected] (K.-H.L.); [email protected] (W.-C.W.); [email protected] (J.-H.S.); Tel.: +886-2-2736-1661 (ext. 6157) (K.-H.L.); +886-2-2820-1999 (ext. 3561) (W.-C.W.); +886-8-882-5001 (ext. 1326) (J.-H.S.) Abstract: The marine sponge of the genus Geodia, Jaspis, Rhabdastrella, and Stelletta are characterized chemically by a variety of isomalabaricane triterpenes. This class of compounds drew spotlights in marine lead discovery due to their profound anti-proliferative properties. Further research on Citation: Lai, K.-H.; Huang, Z.-H.; exploring its chemical diversity led to the identifications of two new isomalabaricane-type triterpenes El-Shazly, M.; Peng, B.-R.; Wei, W.-C.; rhabdastin H (1) and rhabdastin I (2). Their structures were unraveled using a series of spectroscopic Su, J.-H. Isomalabaricane Triterpenes approaches. These isolates were found to exhibit unique structural features with the only reported from the Marine Sponge Rhabdastrella tetrahydrofuran functionality among all marine-derived isomalabaricanes. Both compounds 1 and 2 sp. Mar. Drugs 2021, 19, 206. https:// showed activities against K562 (IC50 11.7 and 9.8 µM) and Molt4 (IC50 16.5 and 11.0 µM) leukemic doi.org/10.3390/md19040206 cells in MTT cell proliferative assay. Academic Editor: Keywords: isomalabaricane; tetrahydrofuran moiety; anti-leukemia Orazio Taglialatela-Scafati Received: 18 March 2021 Accepted: 3 April 2021 1. Introduction Published: 6 April 2021 Marine sponges continue to act as a fruitful source of bioactive and unusual metabo- Publisher’s Note: MDPI stays neutral lites. Among these metabolites are the sponge-derived isomalabaricanes. They were with regard to jurisdictional claims in reported to exhibit potent anti-proliferative properties against a series of cancer cells, in- published maps and institutional affil- cluding colorectal carcinoma [1], ovarian carcinoma [2], acute promyelocytic leukemia [3], iations. acute lymphoblastic leukemia [4], prostate carcinoma [3], gastric adenocarcinoma [3], duc- tal carcinoma [3], hepatocellular carcinoma [3], cervix carcinoma [3], papillomavirus-related endocervical adenocarcinoma [5], and malignant melanoma [4]. The isomalabaricane-type triterpenoids are characterized by an a-methyl group at C-8, owning to their trans-syn-trans tricyclic ring junction instead of trans-anti-trans of their isomers, malabaricanes [1]. Isomal- Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. abaricanes are only found in sponges and are considered as chemotaxonomic markers of This article is an open access article Rhabdastrella sp. [1] Despite the significant difficulties in the isolation and characterization distributed under the terms and of isomalabaricanes due to light induced 13-E/Z isomerization [6], their significant cyto- conditions of the Creative Commons toxic activity prompted extensive research on this class of compounds aiming to find new Attribution (CC BY) license (https:// drug leads. creativecommons.org/licenses/by/ The current study aimed to explore novel isomalabaricanes from marine sources. We 4.0/). isolated two unusual compounds with tetrahydrofuran functional group, the moiety that Mar. Drugs 2021, 19, 206. https://doi.org/10.3390/md19040206 https://www.mdpi.com/journal/marinedrugs Mar. Drugs 2021, 19, x FOR PEER REVIEW 2 of 8 Mar. Drugs 2021, 19, 206 2 of 8 The current study aimed to explore novel isomalabaricanes from marine sources. We isolated two unusual compounds with tetrahydrofuran functional group, the moiety that has been reported previously in malabaricane-type triterpenoids but without any cyto- has been reported previously in malabaricane-type triterpenoids but without any cytotoxic toxic assessments [7,8]. Then, the isolated compounds were evaluated against several can- assessments [7,8]. Then, the isolated compounds were evaluated against several cancer cell cer cell lines using MTT assay to further interpret their anti-proliferative properties. lines using MTT assay to further interpret their anti-proliferative properties. 2.2. Results TheThe freeze-driedfreeze-dried specimenspecimen ofof thethe wild-typewild-type spongesponge RhabdastrellaRhabdastrella sp.sp. (Figure(Figure1 1)) was was extracted with a 1:1 mixture of methanol (MeOH) and dichloromethane (CH2Cl2) (1:1) to extracted with a 1:1 mixture of methanol (MeOH) and dichloromethane (CH2Cl2) (1:1) to provideprovide thethe crude crude extract. extract. The The obtained obtained crude crude extract extract was was further further fractionated fractionated and and purified puri- usingfied using normal normal and reversed-phaseand reversed-phase column column chromatography chromatography yielding yielding two isomalabaricanes, two isomalabar- rhabdastinicanes, rhabdastin H (1) and H (1 rhabdastin) and rhabdastin I (2). TheseI (2). These isolates isolates demonstrated demonstrated unique unique structural struc- featurestural features with with the presence the presence of the of the first first identified identified tetrahydrofuran tetrahydrofuran moiety moiety in in this this classclass ofof compounds.compounds. FigureFigure 1.1. ((A)) AquaticAquatic ecology of the sponge sponge RhabdastrellaRhabdastrella sp.sp. and and ( (BB)) the the isolated isolated isomalabaricane isomalabaricane triterpenes.triterpenes. 1313 TheThe molecularmolecular formulaformula ofof 11was was suggestedsuggested as as C C3232HH5050OO55 basedbased onon CC NMRNMR andand HRESIMSHRESIMS data that that showed showed a a molecular molecular ion ion peak peak at atm/mz 537.3539/z 537.3539 [M + [M Na] ++ Na] implying+ implying eight eightdegrees degrees of unsaturation. of unsaturation. The IR The spectrum IR spectrum of 1 revealed of 1 revealed the presence the presence of carbonyl of carbonyl groups − groupsfrom absorptions from absorptions at 1742, at 1726, 1742, an 1726,d 1703 and cm 1703−1. Its cm 13C1. NMR Its 13C spectrum NMR spectrum of 1 (Table of 1 (1),Table meas-1 ), measuredured in CDCl in CDCl3, showed3, showed the presence the presence of thirty of thirty-two-two carbon carbon signals, signals, which which were were assigned assigned by bythe theassistance assistance of DEPT of DEPT data datato nine to methyl nine methyl groups, groups, eight sp eight3 methylenes, sp3 methylenes, six sp3 methines six sp3 3 2 methines(including (including two oxymathin two oxymathines),es), four sp3 quaternary four sp quaternary carbons, carbons,one sp2 methine one sp methineand four and sp2 2 fourquaternary sp quaternary carbons (including carbons (including two ketone two carbonyl). ketone The carbonyl). NMR signals The NMR at δC signals169.8 (C) at anddC −1 169.820.9 (CH (C) and3) and 20.9 δH (CH2.123 )(3H, and s)dH and2.12 the (3H, IR s)absorption and the IR at absorption 1742 cm−1 atsuggested 1742 cm thesuggested presence theof an presence acetoxy ofgr anoup. acetoxy Carbon group. signals Carbon of the eight signals methyl of the groups eight methyl(δC 31.8, groups 29.3, 26.0, (dC 31.8,24.5, 29.3,24.0, 26.0,19.3, 24.5,19.0 and 24.0, 12.1), 19.3, one 19.0 oxygen and 12.1),-bearing one oxygen-bearingmethine carbon ( methineδC 79.8), carbonand one ( doxygen-C 79.8), andated one quaternary oxygenated carbon quaternary (δC 86.2), carbon one trisubstituted (dC 86.2), one carbon trisubstituted-carbon double carbon-carbon bonds (δ doubleC 116.9, bondsCH; 142.6, (dC 116.9, C), two CH; ketones 142.6, ( C),δC 220.3 two ketonesand 207.4) (dC were220.3 also and assigned. 207.4) were The also resonances assigned. of Theone d J resonancesolefinic proton of one (δH olefinic 5.11, d, protonJ = 10.0 ( Hz)H 5.11, and d,two= oxygenated 10.0 Hz) and methines two oxygenated (δH 6.09, methinesd, J = 10.0 (d 6.09, d, J = 10.0 Hz; d 3.76, ddd, J = 14.0, 8.5, 4.5 Hz) were observed from the 1H NMR Hz;H δH 3.76, ddd, J = 14.0,H 8.5, 4.5 Hz) were observed from the 1H NMR spectroscopic data spectroscopicof 1 (Table 1). data of 1 (Table1). 1 1 BasedBased on the the above above results results and and with with the the assistance assistance of 1H of–1HH– COSYH COSY and HMBC and HMBC exper- experimentsiments (Figure (Figure 2), the2), planar the planar structure structure of 1 of was1 was determined. determined. To establish To establish the theproton proton se- 1 1 1 sequencesquences in in 1, the, the 1HH––1HH COSY COSY spectrum spectrum analysis analysis est establishedablished five five proton sequences fromfrom H2-1 to H2-2, H-5 to H2-7, and H2-11 to H2-12, H2-15 to H-21, and H-23 to H-24. These H2-1 to H2-2, H-5 to H2-7, and H2-11 to H2-12, H2-15 to H-21, and H-23 to H-24. These data, data, together with the HMBC correlations (Figure2) from H 2-1 and H2-2 to C-3, H2-11 together with the HMBC correlations (Figure 2) from H2-1 and H2-2 to C-3, H2-11 and H2- and H2-12 to C-9, H2-12 to C-8 and C-13, H3-19 to C-1, C-5, C-9 and C-10, H3-28 and H3-29 to C-3, C-4 and C-5 and H3-30 to C-7, C-8, C-9 and C-13 established the connectivity within the 6-membered (A), 6-membered (B), and 5-membered (C) rings.