Introduction The process of cancer treatment presents significant challenges for both patients and their caregivers. Caregivers often neglect their own health and quality of life while trying to meet the needs of patients. Aims This study aimed to investigate the relationship between caregiver burden and the levels of anxiety and spiritual well-being among caregivers of individuals with cancer. Methods: This descriptive and correlational study included 218 caregivers of individuals undergoing cancer treatment. Research data were collected with the personal information form, caregiver burden scale, generalised anxiety disorder scale and three-factor spiritual well-being scale. Results According to the research findings, caregivers experienced a moderate level of caregiving burden (34.45±19.11) and moderate anxiety (7.62±5.65), while their spiritual well-being levels were high (112.93±17.56). A weak positive relationship was found between caregiving burden and anxiety levels, whereas a weak negative relationship emerged between caregiving burden and spiritual well-being. Regression analysis revealed that anxiety and spiritual well-being significantly predicted caregiving burden, explaining 20% of the variance (adjusted R²= 0.203, p<0.001). Conclusion The impact of anxiety and spiritual well-being on caregiving burden clearly highlights the importance of psychological and spiritual support interventions to alleviate caregivers’ stress.