The physicochemical properties of therapeutic particles, including size, shape, surface chemistry, stability, and biodegradability, have been widely investigated how they influence their biological behaviors and improve the efficacy for therapeutic delivery. Recent report have indicated that stiffness is a key parameter for governing bio-nano interactions. However, stiffness is an easily overlooked parameter due to the difficulty associated with tuning the stiffness of many particles (e.g., inorganic particles) and the challenge associated with stiffness characterization. The flexibility of fabrication techniques in precise control over polymer particles has facilitated their application in cancer theranostics. Soft polymer particles have shown significant advantages in overcoming biological barriers and increasing drug delivery efficacy. This review focus on (i) characterization methods for investigating the stiffness of polymer particles, (ii) strategies for the fabrication of soft polymer particles, and (iii) advances in the biological behavior of soft polymer particles. The engineering of soft polymer particles with tunable stiffness will revolutionize our understanding of their bio-nano interactions, which is vital to provide key steps for developing the next generation of cancer theranostics that minimise side-effects while maximising therapeutic efficacy.