BACKGROUND Malaria is responsible for 429,000 deaths per year worldwide, and more than 200 million cases were reported in 2015. Increasing parasite resistance has imposed restrictions to the currently available antimalarial drugs. Thus, the search for new, effective and safe antimalarial drugs is crucial. Heterocyclic compounds, such as dihydropyrimidinones (DHPM), synthesised via the Biginelli multicomponent reaction, as well as bicyclic compounds synthesised from DHPMs, have emerged as potential antimalarial candidates in the last few years. METHODS Thirty compounds were synthesised employing the Biginelli multicomponent reaction and subsequent one-pot substitution/cyclisation protocol; the compounds were then evaluated in vitro against chloroquine-resistant Plasmodium falciparum parasites (W2 strain). Drug cytotoxicity in baseline kidney African Green Monkey cells (BGM) was also evaluated. The most active in vitro compounds were evaluated against P. berghei parasites in mice. Additionally, we performed an in silico target fishing approach with the most active compounds, aiming to shed some light into the mechanism at a molecular level. RESULTS The synthetic route chosen was effective, leading to products with high purity and yields ranging from 10-84%. Three out of the 30 compounds tested were identified as active against the parasite and presented low toxicity. The in silico study suggested that among all the molecular targets identified by our target fishing approach, Protein Kinase 3 (PK5) and Glycogen Synthase Kinase 3β (GSK-3β) are the most likely molecular targets for the synthesised compounds. CONCLUSIONS We were able to easily obtain a collection of heterocyclic compounds with in vitro anti-P. falciparum activity that can be used as scaffolds for the design and development of new antiplasmodial drugs.