Innovative silica nanomaterials have made the significant advancements in curative therapy against cancers with multidrug resistance (MDR). The study on different‐nanostructured mesoporous silica nanoparticles (MSNs) with discrepant pore sizes affecting biomacromolecules in resisting cancer MDR hasn't been reported yet. In this study, a systematic comparison of 6 nm‐pore sized hollow‐structured MSNs (HMSNs) and 10 nm‐pore sized dendrimers‐structured MSNs (LMSNs) for delivering Bcl‐2‐functional converting peptide (N9) or doxorubicin (DOX) to overcome cancer MDR was comprehensively carried out both in in vitro and in vivo resistant tumor models. Our results showed that both LMSNs and HMSNs exerted no significant difference in delivering DOX to treat drug‐resistant cancers. However, compared with N9@HMSNs, N9@LMSNs displayed the increased loading efficiency, the improved cell‐penetrative capability, the higher cancer cell apoptosis effect, the enhanced tumor accumulation and retention efficiency, and the final elevated tumor inhibition efficiency. Unexpectedly, naked LMSNs without surface modification especially at high dosage produced relatively more serious toxicity than HMSNs whatever in cells, zebrafish embryo or mice models. Collectively, our data provided the sufficient theoretical evidence that LMSNs might be a better choice for delivering biomacromolecules to treat resistant cancers after appropriate surface functionalization such as with PEGylation to weaken its intrinsic toxicity.This article is protected by copyright. All rights reserved