In tumour development, the degradation of heparan sulfate (HS) by heparanase (HPSE) is associated with cell-surface and extracellular matrix remodelling and as well as the release of HSbound signalling molecules, allowing cancer cell migration, invasion and angiogenesis. Because of their structural similarity with HS, sulfated polysaccharides are considered a promising source of molecules to control these activities. In this study, we used a depolymerisation method for producing λ-carrageenan oligosaccharides (λ-CO), with progressive desulfation over time. This These were then used allowed to investigating investigate the influence of the polymeric chain length and degree of sulfation (DS), on their anti-HPSE activity. The effects of these two features on λ-CO anticoagulant properties were also investigated to eliminate a potential limitation on the use of a candidate λ-CO as a chemotherapeutic agent. HPSE inhibition was mainly related to the DS of λ-CO, but this correlation was not complete. On the other hand, both the chain length and the DS modulated the λ-CO activities activity for factor Xa and thrombin IIa inhibition, two enzymes involved in the coagulation cascade, and different mechanisms of inhibition were observed. A λcarrageenan oligosaccharide of 5.9 KDa was identified as a suitable anticancer candidate because it displayed one of the lowest anticoagulant properties among the λ-CO produced, while showing a remarkable inhibitory effect on MDA-MB-231 breast cancer cell migration.