Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. They are associated with alcohol and tobacco consumption, as well as infection with human papillomaviruses (HPV). Therapeutic options include radiochemotherapy, surgery or chemotherapy. Nanoparticles are becoming more and more important in medicine. They can be used diagnostically, but also therapeutically. In order to provide therapeutic alternatives in the treatment of HNSCC, the effect of citrate-coated superparamagnetic iron oxide nanoparticles (Citrate-SPIONs) and gold-coated superparamagnetic iron oxide nanoparticles (Au-SPIONs) in combination with ionizing irradiation (IR) on two HPV positive and two HPV negative HNSCC and healthy fibroblasts and keratinocytes cell lines were tested. Effects on apoptosis and necrosis were analyzed by using flow cytometry. Cell survival studies were performed with a colony formation assay. To better understand where the SPIONs interact, light microscopy images and immunofluorescence studies were performed. The HNSCC and healthy cell lines showed different responses to the investigated SPIONs. The cytotoxic effects of SPIONs, in combination with IR, are dependent on the type of SPIONs, the dose administered and the cell type treated. They are independent of HPV status. Reasons for the different cytotoxic effect are probably the different compositions of the SPIONs and the related different interaction of the SPIONs intracellularly and paramembranously, which lead to different strong formations of double strand breaks.