A series of 24 benzothiadiazine derivatives with structural novelty were designed, synthesized and biologically evaluated as PI3Kδ-selective inhibitors. As a consequence of the structure-activity relationship (SAR) study, compounds 63 and 71 were identified with single-digit nanomolar IC50 values against PI3Kδ and submicromolar GI50 values against human malignant B-cell line SU-DHL-6. Furthermore, chiral resolution of the key amine intermediate of these two compounds was performed to achieve corresponding enantiomers. In subsequent biological evaluation, S-63 (IC50: 4.6 nM) and S-71 (IC50: below 0.32 nM) demonstrated comparable and superior PI3Kδ inhibitory activity, respectively, to that of idelalisib. Additionally, both S-63 (GI50: 33.2 nM) and S-71 (GI50: 15.9 nM) exerted enhanced anti-proliferative activity against the SU-DHL-6 cell line than that of idelalisib. Moreover, both S-63 and S-71 exhibited excellent PI3Kδ selectivity. In the further in vivo pharmacokinetic (PK) study, S-63 displayed a good plasma exposure and an acceptable oral bioavailability of 29.2%. By virtue of its biological performance, S-63 merits further development as a potential therapeutic agent for battling B-cell-mediated malignancies.