Human NAD(P)H: quinone oxidoreductase 1 (hNQO1), a proteinase that engages in detoxification of quinones and capable of activating anti-tumor drugs, has drawn increasing attention as tumor biomarker and drug target. To date, the detection of hNQO1 primarily uses stimulus-responsive probes, involving metabolization of synthetic quinone-functionalized substrates, which however, remain challenging to improve the sensing signal-to-noise ratio, and are lack of sufficient stability. Herein, we report a facile but general way for hNQO1 detection and associated drug screening as well by ECL sensing of the metabolic H2O2 enabled by futile redox cycle reaction. Taking advantage of the intrinsic circulatory amplification and the luminol-modified nickel foam electrode, the sensing system exhibited a record-level performance in electrochemiluminescent detection of hNQO1. The same strategy was also successfully applied to rapidly screening hNQO1-directed anti-tumor candidate drugs. The proposed new principle for hNQO1 detection would stimulate ECL as a promising tool that combines diagnostic and drug screening functions for the popularization of proteinases in cancer management.