*Article* **Mapping HPV 16 Sub-Lineages in Anal Cancer and Implications for Disease Outcomes**

**Daniel Guerendiain 1,2,\* , Laila Sara Arroyo Mühr <sup>3</sup> , Raluca Grigorescu <sup>4</sup> , Matthew T. G. Holden <sup>2</sup> and Kate Cuschieri <sup>1</sup>**


**Abstract:** The incidence of anal cancer is rising worldwide. As identified in cervical cancer management, an improvement in the early detection and management of anal pre-cancer is essential. In other cancers associated with human papillomavirus (HPV), HPV 16 sub-lineages have been shown to be associated with disease status and prognosis. However, in anal cancer, they have been under-explored. A total of 119 HPV 16-positive anal cancer lesions diagnosed between 2009 and 2018 in Scotland and 134 HPV 16-positive residual rectal swabs from asymptomatic men collected in 2016/7 were whole genome sequenced. The association of HPV 16 sub-lineages with underlying disease status (cancer vs. asymptomatic) and overall survival in anal cancer samples was assessed (comparing A1 vs non-A1 sub-lineages). A1 was the dominant sub-lineage present in the anal cancer (76.5%) and the asymptomatic (76.1%) cohorts. A2 was the second most dominant sub-lineage in both groups (16.8% and 17.2%, respectively). We did not observe significant associations of sub-lineage with demographics, clinical variables or survival (A1 vs. non-A1 sub-lineages (HR 0.83, 0.28–2.46 *p* = 0.743)). HPV 16 sub-lineages do to not appear to cluster with disease vs asymptomatic carriage or be independently associated with outcomes in anal cancer patients. Further international studies on anal HPV sub-lineage mapping will help to determine whether this is a consistent observation.

**Keywords:** human papillomavirus; HPV 16 sub-lineages; anal cancer
