Background: This study aimed to explore the differences in quantitative diffusion-weighted (DW) MRI parameters in oropharyngeal squamous cell carcinoma (OPC) based on Human Papillomavirus (HPV) status before and during radiotherapy (RT). Methods: Echo planar DW sequences acquired before and during (chemo)radiotherapy (CRT) of 178 patients with histologically proven OPC were prospectively analyzed. The volumetric region of interest (ROI) was manually drawn on the apparent diffusion coefficient (ADC) map, and 105 DW-MRI radiomic parameters were extracted. Change in ADC values (Δ ADC) was calculated as the difference between baseline and during RT at week 4, normalized by the baseline values. Results: Pre-treatment first-order 10th percentile ADC and Gray Level co-occurrence matrix (GLCM)-correlation were significantly lower in HPV-positive compared with HPV-negative tumors (82.4 × 10−5 mm2/s vs. 90.3 × 10−5 mm2/s, p = 0.03 and 0.18 vs. 0.30, p < 0.01). In the fourth week of RT, all first-order ADC values were significantly higher in HPV-positive tumors (p < 0.01). Δ ADC mean was significantly higher for the HPV-positive compared with the HPV-negative OPC group (95% vs. 55%, p < 0.01). A predictive model for HPV status based on smoking status, alcohol consumption, GLCM correlation, and mean ADC and 10th percentile ADC values yielded an area under the curve of 0.77 (95% CI 0.70–0.84). Conclusion: Our results highlight the potential of DW-MR imaging as a non-invasive biomarker for the prediction of HPV status, although its current role remains supplementary to pathological confirmation.