Health literacy has a direct impact on the health of populations. It is related to education, capacity for self-care, and management of health resources. The Health Literacy Survey Questionnaire HLS-Q12 is one of the reference instruments but has not yet been adapted to Spanish. The aims of the study were to cross-culturally adapt and evaluate the psychometric properties of the Spanish version of the HLS-Q12. Data was collected from June 2020 to March 2022. The sample consisted of 60 patients who initiated cancer treatment for the first time within a clinical trial. Double direct translation, back-translation, cognitive debriefing with a 10-patient sample, and an expert committee were used for cross-cultural adaptation. For validation of the HLS-Q12, a psychometric analysis was performed to assess feasibility, reliability, sensitivity to change and construct validity with other measures such as health-related quality of life, empowerment, and health needs. The HLS-Q12 is equivalent at the semantic, conceptual, and content level to the original version and its psychometric properties demonstrated good internal consistency with a Cronbach's alpha of 0.88 and a McDonald´s omega of 0.91, a high degree of fit for the confirmatory factor analysis, and a statistically significant sensitivity to change (p = 0.025). Based on robust psychometric values, the Spanish version of HLS-Q12 was found to be a good cross-culturally adapted tool for collecting correct information on health literacy in cancer patients regardless of tumour type or stage. Although more studies are needed, this version of HLS-Q12 could be used in research for collecting data on the health literacy needs of Spanish-speaking patients.