Palm hearts that do not meet export quality standards are typically used as animal feed, while sambo seeds are rarely consumed and often discarded. Both are promising raw materials for the development of high value-added food products. This study aimed to develop gluten-free cookies by partially replacing oat flour and dairy fat with palm heart flour and sambo seed paste, respectively. Cookies were produced under good manufacturing practices using an all-in mixing method. The formulation included oat and palm heart flour (29.7%), sunflower and sambo seed paste (17.7%), erythritol (17%), egg (16.0%), vanilla extract (0.6%), cocoa powder (4.2%), and chocolate chips (14.8%). Response surface methodology (RSM) with a 2² factorial central composite design was employed to quantify the individual and interactive effects of the substitutes on texture (fracture strength, hardness, and gumminess), moisture, and color. Proximate composition and sensory acceptability were also determined using standardized methods. The optimized formulation achieved an overall desirability of 0.70, incorporating 19.47% palm heart flour (relative to total flours) and 40.70% sambo seed paste (relative to total fat). The product provided 14.0% protein, 30.0% fat, 6.4% crude fiber, 3.2% moisture, and 2.0% ash and exhibited a balanced texture profile (fracture strength: 0.3 N; hardness: 80 N; gumminess: 11). Sensory acceptability was moderately high (7.17/9): flavor and aroma received the highest scores, whereas color was penalized due to the greenish hue imparted by the sambo paste. A significant interaction between ingredients strongly influenced textural attributes, underscoring the importance of formulation optimization to maximize overall acceptance. These results demonstrate that agro-industrial by-products such as palm heart flour and sambo seed paste can be effectively leveraged to formulate functional, sustainable, gluten-free cookies. Further studies should address amino acid and fatty acid profiles, mineral content, and advanced sensory analysis to enhance product characteristics and increase consumer acceptance.