Chronic exposure to tertiary butyl alcohol (TBA) in drinking water (DW) is associated with low but increased incidence (15% vs 3% in controls) of thyroid follicular cell adenomas in female mice. The U.S. Environmental Protection Agency evaluated this data in the context of an anti-thyroid (AT) mode of action (MOA) and identified data gaps while noting evidence for thyroid adenomas as “suggestive”. A study was designed to address data gaps in the MOA for TBA thyroid tumors. Female mice were exposed for 5 days to TBA by oral gavage (500 or 1000 mg/kg) or via DW (5-40 mg/mL) for up to 28 days. TBA activated upstream key events (KEs) within the AT MOA framework including CAR/PXR signaling and upregulation of UGT and SULT transcripts. However, increased UGT and SULT activity was not observed. Downstream events (decreased serum TSH and thyroid histopathology prior to 2-years) were not demonstrated. The inability to demonstrate downstream KEs is likely a consequence of a weak tumor response with chronic exposure to TBA. With previously published data, an AT MOA is a plausible explanation for a weak TBA tumor response, with analysis highlighting important consequence of conducting risk assessments on weak, suggestive tumor data.