SRPK1 expression was estimated by IHC on paraffin-embedded tissue sections using a SRPK1-specific monoclonal antibody. In normal testicular parenchyma, we observed high SRPK1 expression () as anticipated [11
]. Positive nuclear and cytoplasmic staining of SRPK1 was found in spermatogonia, spermatocytes, and spermatids (), whereas no staining was observed in mature spermatozoa, Sertoli cells, or blood vasculature. In carcinoma in situ
(the precursor of testicular GCTs), we also found SRPK1 (). Strong SRPK1 staining was found in the majority of randomly selected and standard chemotherapy-sensitive NS (). In contrast, in most tumors from patients with treatment-refractory disease and patients with advanced disease responding to high-dose chemotherapy only, SRPK1 expression was considerably lower or even absent. A large number of the treatment-refractory DOD cases were negative or stained only weakly for SRPK1 (, and ). We did not see obvious differences in staining between embryonal carcinoma, yolk sac tumor, teratoma, and choriocarcinoma components.
Figure 1 Representative examples of immunohistochemical staining for SRPK1. (A) Normal spermatogenesis (left) and carcinoma in situ (right). (a) Spermatogonium; (b) spermatocyte; (c) spermatid; (d) Sertoli cell; and (e) carcinoma in situ cell. (B) Chemotherapy-sensitive (more ...)
Regardless of their histological subclassification, for several samples examined, a more or less heterogeneous SRPK1 staining was noticed among the tumor cells. Therefore, a total of five randomly chosen microscopic tumor fields per IHC slide was scored in order to get a representative estimate of SRPK1 expression. The semiquantitative score of (-) to (+++) was assigned the value 1, 2, 3, 4, or 5, respectively, and the cumulative scores of five different tumor fields were calculated. Although most randomly selected and standard chemotherapy-sensitive GCTs showed overall scores > 15 corresponding to strong or very strong SRPK1 staining, the majority of treatment-refractory and high-dose-responsive GCTs displayed scores ≤ 15 for negative, weak, or intermediate staining (). The median cumulative IHC scores for the randomly selected, standard chemotherapy-responsive, refractory, and high-dose-responsive NS were 20.5 (range 10–25), 19 (11–25), 7.5 (5–20), and 15 (5–18), respectively (). SRPK1 expression in the randomly selected versus refractory and high-dose-responsive groups was significantly different at P < .0001 and P < .0001 (two-sided Wilcoxon rank sum test), respectively. The relative expression in the standard chemotherapy-responsive versus the refractory and high-dose-responsive group was significantly different at P < .0001 and P < .001 (two-sided Wilcoxon rank sum test), respectively. We conclude that high SRPK1 expression might be an important prognostic indicator for the responsiveness of nonseminomatous GCTs toward platinum-containing chemotherapy, whereas its absence or low expression might predict resistance.
Figure 2 Relative SRPK1 expression was compared between randomly selected germ cell tumors, chemorefractory germ cell tumors (DOD), poorprognosis high-dose-responsive, and standard chemotherapy-responsive germ cell tumors. (A) Pie chart showing the percentages (more ...)