Human pressures on marine ecosystems significantly increased during last decades. Among the intense anthropic activities, industrial fisheries have caused the alteration of habitats, the reduction of biodiversity and the main fish stocks. The aim of this research, carried out in the Adriatic Sea, was to test a repeatable Marine Spatial Planning framework aimed at enhancing fisheries sustainability through the application of Decision Support Tools and the composition of a catalog of possible measures. The use of these tools proved very useful to identify possible criticalities and facilitate an effective exchange with fisheries stakeholders, local authorities, and fishermen, whose involvement was an indispensable step in the process. Tool-based analyses allowed to assess the spatial footprint of a range of anthropogenic pressures from human activities (e.g., fisheries, maritime traffic, and aquaculture). Within this multi-pressure scenario, special attention was paid to fishing-related disturbances and potential conflicts across different fishing métier and with other sectors. Specifically, results highlighted the spatial features of the major fishing pressures (e.g., abrasion from trawling) affecting essential fish habitats, marine mammals and turtles in the study area. A portfolio of possible management measures is identified for the study area. It provides clear evidence that, in order to mitigate emerging conflicts and cumulative impacts, it is necessary to combine and integrate different types of measures: spatial measures modulated over time, monitoring and control, actions to fill knowledge gaps, concertation—involvement—co-management actions, improvement of governance systems, actions to support innovation in the sector, etc. Given the complex set of measures discussed, this work can provide a useful contribution to the management of fisheries both at local and regional level, fostering the transition to sustainable fisheries.