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Many species belonging to the genus <i>Echinops</i> are widely used in traditional medicine to treat infectious diseases and cancers. The present study aimed to evaluate the antibacterial and antiproliferative properties of <i>Echinops lanceolatus</i> Mattf. (Asteraceae). The activity of the methanolic extract and subsequent partition fractions was investigated against drug-resistant bacteria (Gram-negative and Gram-positive) and human tumor cell lines using broth microdilution and sulforhodamine B (SRB) assay, respectively. Our findings revealed weak to moderate antibacterial activities of tested extracts, with the recorded minimal inhibitory concentrations ranging from 256 to 1024 µg/mL. The ethyl acetate fraction (EL-EA) was found to be the most effective. Likewise, that fraction displayed strong antiproliferative potential with recorded IC₅₀ of 8.27 µg/mL and 28.27 µg/mL on A549 and HeLa cells, respectively. An analysis based on the ultra-performance liquid chromatography–electrospray ionization tandem mass spectrometry (UPLC–ESI–MS/MS) of the EL-EA fraction allowed the identification of 32 compounds, of which quinic acid and derivatives, cinnamic acid derivatives, dihydrokaempferol, naringenin-7-<i>O</i>-glucoside, apigenin-7-<i>O</i>-<span style="font-variant: small-caps;">d</span>-glucoside, naringin, apigenin, rhoifolin, coniferyl aldehyde, and secoisolariciresinol are well-known compounds of biological importance. This study is first to report on the biological activity and phytochemical profile of <i>E. lanceolatus</i>. We provide a baseline to consider <i>E. lanceolatus</i> as a valuable source of anti-infective and antiproliferative agents.