Malalignment of the lower limbs is the main biomechanical factor for knee osteoarthritis (KOA). The static hip-knee-ankle angle (S-HKAA) measured from radiograph is regarded as the “gold standard” of the malalignment. However, many evidences showed that the S-HKAA has no significant correlation with the knee dynamic-load distribution, unlike the dynamic HKAA (D-HKAA). The purpose of this study was to quantitatively analyze the D-HKAA and investigate the relationship between D-HKAA and S-HKAA for both KOA and healthy participants. In this paper, twenty-five healthy subjects and twenty-five medial compartment KOA (M-KOA) patients were recruited. Three-dimensional motion analysis and standing lower-limbs-full-length radiograph were utilized to obtain the D-HKAA and S-HKAA, respectively. The results showed that the mean D-HKAA was more varus than the S-HKAA (
p
<
0.05
). For the mean D-HKAA, larger varus angle was observed in swing phase than stance phase (
p
<
0.05
). Compared with healthy subjects, the M-KOA patients had remarkably smaller S-HKAA and D-HKAA during gait cycle (
p
<
0.01
). For the relationship between the S-HKAA and mean D-HKAA, no significant correlation was found for both healthy subjects and M-KOA patients (
r
<
0.357
,
n
=
25
,
p
>
0.05
, Spearman correlation analysis). In conclusion, the S-HKAA was limited to predict the D-HKAA for both M-KOA patients and healthy subjects. The D-HKAA should be given more attention to the orthopedist and the designer of knee brace and orthotics.