Flexile strain sensors hold vast potential for applications in monitoring human motion, enabling human-machine interaction, and facilitating information transfer, etc. However, the available materials and manufacturing techniques for fabricating flexible strain sensors with high sensitivity and extended sensing range still face significant challenges. By utilizing solution casting and twin-screw extrusion techniques, this study prepared high-performance thermoplastic polyurethane/multi-walled carbon nanotube (TPU/MWCNT) nanocomposite 3D printable filaments, and employed material extrusion 3D printing technology to facilely fabricate flexible strain sensors. The 1-pyrene carboxylic acid (PCA)-modified TPU/MWCNT(3 wt%) nanocomposite exhibited outstanding mechanical properties, with a tensile strength of 24.3 ± 0.5 MPa and a fracture elongation of 625.8 ± 12.3 %. The 3D printed TPU/MWCNT(3 wt%)/PCA flexible strain sensors demonstrated amazing sensitivity (GFmax = 10279.95) within a wide strain range (0–300 % strain). The addition of PCA brings the benefits of improved dispersion of MWCNTs in the TPU composite, as well as enhanced thermal stability, and upgraded mechanical and electrical properties under different tensile strains. Meanwhile, the 3D printed samples displayed remarkable durability and reproducibility. Finally, the flexible strain sensors fabricated from this nanocomposite material could accurately perceive human motion, providing great prospects for wearable device applications.