A novel cycloartane triterpenoid alkaloid, kleinhospitine E (1), six new cycloartane triterpenoids (2–7), three known cycloartane triterpenoids (8–10), and taraxerone (11) were isolated from a methanol extract of Kleinhovia hospita. Their structures were elucidated by 1D- and 2D-NMR spectroscopy as well as HRMS analysis. The absolute configurations of all isolated compounds were determined from their ECD spectra by comparison with theoretical values. Kleinhospitine E (1) is the first cycloartane alkaloid possessing an unusual γ-lactam with an oxopropylidene side chain. Compounds 2, 3, and 6were assigned as cycloartane triterpenoids with a 9α,10α-cyclopropyl ring, which is found rarely among naturally occurring compounds, while 4and 5were established as isomers of compound 3containing a 21,23-diacetal side chain. Biological evaluation revealed that compounds 4and 9exhibited more potent antiproliferative activities against a multidrug-resistant tumor cell line compared with its parent chemosensitive cell line. Furthermore, compound 6exhibited submicromolar anti-HIV activity.