Carbon nanotubes (CNTs) consist of carbon atoms arranged in sheets of graphene rolled up into cylindrical shapes. This class of nanomaterials has attracted attentionbecause of their extraordinary properties, such as high electrical and thermal conductivity. In addition, development in CNT functionalization chemistry has led to an enhanced dispersibility in aqueous physiological media which indeed broadens the spectrum for their potential biological applications including gene delivery. The aim of this study is to determine the capability of different cationic polymergrafted multiwalled carbon nanotubes (MWNTs) (polymer- g -MWNTs) to efﬁ ciently complex and transfer plasmid DNA (pCMV-β Gal) in vitro without promoting cytotoxicity. Carboxylated MWNT is chemically conjugated to the cationic polymers polyethylenimine (PEI), polyallylamine (PAA), or a mixture of the two polymers. In order to explore the potential of these polymer-g-MWNTs as gene delivery systems, we ﬁ rst study their capacity to complex plasmid DNA (pDNA) using agarose gel electrophoresis. Gel migration studies conﬁrm pDNA binding to polymer-g-MWNT with different afﬁnities, highest for PEI-g-MWNT and PEI/PAA-g-CNT constructs. β-galactosidase expression is assessed in human lung epithelial (A549) cells, and the cytotoxicity is determined by modiﬁ ed LDH assay after 24 h incubation period. Additionally, PEI- g -MWNT and/or PEI/PAA- g -MWNT reveal an improvement in gene expression when compared to the naked pDNA or to the equivalent amounts of PEI polymer alone. Mechanistically, pDNA was delivered by the polymer- g -MWNT constructs via a different pathway compared to those used by polyplexes. In conclusion, polymer- g -MWNTs may be considered in the future as a versatile tool for efﬁ cient gene transfer in cancer cells in vitro, provided their toxicological proﬁle is established.