Immunodetection of phosphohistone H3 as a surrogate of mitotic figure count and clinical outcome in cutaneous melanoma

In the American Joint Committee on Cancer (AJCC)-TNM (2009) staging system, the key prognostic factor in cutaneous melanoma is the depth of dermal invasion (Breslow thickness) with further refinement according to the presence of epidermal ulceration or dermal mitoses. Immunodetection of phosphohistone H3 has been shown to facilitate the identification of mitotic figures in various neoplasms. We selected 120 cases of primary cutaneous melanoma with completely annotated histopathologic parameters and clinical outcomes and performed double immunohistochemical staining for MLANA (Mart-1/Melan-A) and phosphohistone H3. One hundred and thirteen cases were amenable to antiphosphohistone H3 staining from 66 men and 47 women, with mean age of 64 years (9-93), including 61 superficial spreading type, 24 nodular, 6 lentigo maligna, 8 acral lentiginous, and 14 unclassified. The mean Breslow thickness was 2.53 mm (0.20-25), ulceration was present in 25/113 (22%) and the mean mitotic count was 3.2/mm(2) (<1-29/mm())(2). In 27/113 (24%) of the cases, antiphosphohistone H3 failed to highlight mitotic figures anywhere in the tissue (normal or tumor cell), whereas in 86/113 (76%) antiphosphohistone H3 detected at least one mitotic figure. Among the latter, antiphosphohistone H3 did not detect mitotic figures in dermal tumor cells in 37/86 cases (43%), whereas anti-PHH3 identified at least one melanocytic mitotic figure in the other 49/86 cases (57%; range: 1-66/mm(2)). The relationship between phosphohistone H3 and manual mitotic count was statistically significant (Pearson correlation = 0.59, P<0.0001). Logistic regression analyses demonstrated an association between the development of subsequent metastatic disease and the following variables: mitotic figures (odds ratio (OR) = 5.7; P=0.0001); phosphohistone H3-positive mitotic figures (OR = 3.0; P=0.008); Breslow thickness (OR = 4.0 per mm; P=0.0002); ulceration (OR = 3.94; P=0.008). The application of phosphohistone H3 immunohistochemistry to the description of primary cutaneous melanoma is useful in identifying mitotic figures, improves upon the specificity of this designation when used together with MLANA, and correlates with an increased risk for metastasis in univariate analyses.