The microstrip patch antenna (MPA) with a thermoplastic substrate has been widely used in wearable sensors primarily due to its lightweight, flexibility, strength, and compactness. But up till now, little has been conveyed on the use of three-dimensional (3D) printed thermoplastic polyurethane (TPU) on cotton-lycra woven (CLW) fabric-based substrate as a wearable sensor. This study presents a novel and scalable approach for characterizing, simulating, and fabricating the substrate for wearable sensor applications. In this study 3D printing of TPU on CLW fabric (70% cotton (on the warp side)-30% lycra (on the weft side) using fused filament fabrication (FFF) process) has been reported with a tradeoff between flexibility and strength (of a substrate for wearable sensor applications). The rheological, mechanical, morphological, four-dimensional (4D), and resonance frequency (RF) characterization of the substrate has been established. As regards rheological properties, the melt flow index (MFI) of primary (1°) recycled TPU was observed as 29.758 g/(10min). Further, the optimized 3D printing settings of TPU were obtained based on the mechanical testing (minimum stiffness, maximum peak strength (PS), maximum Young's modulus (E), and maximum strain energy (SE)) which comes out to be 225°C nozzle temperature, 18 mm/s printing speed and 60% infill density with a stiffness of 3.311 N/mm and PS of 9.628 MPa. The mechanical properties of TPU printed on stretched and unstretched CLW fabric were tested for lower stiffness (more flexibility). To ascertain the mechanical properties, porosity analyses of samples printed on CLW fabric have been performed based on scanning electron microscopy (SEM). For dielectric properties, and RF characterization, the ring resonator (RR) test was performed based on a Vector network analyzer (VNA). Finally, 4D characterization of the substrate was performed by applying a load from 0 to 25 N, resulting in programable RF output (in the industry scientific and medicine (ISM) band).