A number of new furo and thienoquinolinones carrying an electron-withdrawing function or unsubstituted at the position 3 were synthesized in order to obtain new potential photochemotherapeutic agents with increased antiproliferative activity and decreased toxic side effects. Our interest in studying the SAR of these derivatives also prompted us to investigate the influence of N-methylation on biological activity, by preparing N-methyl derivatives. The antiproliferative activity of all the newly synthesized compounds was evaluated and compared to 8-methoxypsoralen (8-MOP), the drug widely used in PUVA-therapy. The 3-unsubstituted thienoquinolinones were generally the most potent derivatives, followed by the furo-analogues. In particular, the unsubstituted thieno[2,3-h]quinoline-2(1H)one showed the highest activity in T2 bacteriophage, HeLa cells and Ehrlich cells tests. All the compounds, assayed on Escherichia coli WP2 TM9, showed a similar mutagenic activity, very close to that of 8-MOP. Except for 2-oxo-1,2-dihydrothieno[2,3-h]quinoline-3-carboxylic acid, which appeared to be very effective, all compounds generated singlet oxygen to slightly larger amounts when compared to 8-MOP. The N-methyl analogues only induced moderate skin erythemas on albino guinea pigs, while all other derivatives appeared to be entirely inactive. On the basis of these results, the unsubstituted thieno[2,3h]quinoline 2(1H)one seems to be the most interesting potential drug for PUVA photochemotherapy and photopheresis.