Figure 6
From: Maximizing the potential of aggressive mouse tumor models in preclinical drug testing
Ispinesib is efficacious against a second model of pediatric embryonal brain tumor. (a) KIF11 (target of ispinesib) was expressed on our PDOX models of medulloblastoma (MB), another type of embryonal brain tumor. MB is the most common malignant brain tumor in children. Immunohistochemical staining of several of our MB PDOX models with KIF11 antibody demonstrated KIF11 expression co-localized with mitotic figures, shown here in Model ICb-1572. KIF11 was expressed on different cell populations within PDOX tumor (shown here in brown staining), and not restricted to mitoses, indicating ispinesib targeted not only mitoses (actively dividing cells) but also other tumor cell populations within the tumor. Cytoplasmic staining of KIF11 was observed on 5–10% of cells, including mitotic figures (red arrows), ×400. Normal brain parenchyma was devoid of KIF11 expression, interfacing with tumor core densely packed with tumor cells. Actively dividing cells were marked by KIF11+ mitotic figures—a good target for ispinesib. (b) Pediatric MB is molecularly classified into 4 molecular subtypes: sonic-hedgehog (SHH)-activated MB, Wingless (WNT)-activated, and less characterized Group 3 and 4 subtypes20,21. Group 3 and 4 MBs are the most aggressive subtypes and Group 3 MBs have the worst survival outcome22,23. PDOX Model ICb-1572 is Group 3 subtype24. (c) Hematoxylin & eosin stains showing a large xenograft MB tumor of Group 3 subtype (Model ICb-1572), occupying almost the entire mouse cerebellum. Histological appearance (40X) of xenograft tumor was consistent with densely packed, small round blue cell typical of MB. Our panel of xenograft tumors recapitulate morphologic, histological and immunohistochemical features of matched patient original tumors1. Animal survival was monitored up to Passage IV-IX for Model ICb-1572. Log rank analysis of animal survival times of ICb-1572 MB during serial sub-transplantation of retrieved xenograft cells from cryopreservation (Passage III to VIII is shown here using Passage I as reference). Survival days were consistent and maintained within and across passages in mice. (d) Patient characteristics of Model ICb1572. (e) Tumor proliferation activity—ispinesib against in-vitro 3D-Neurosphere cultures of Model ICb1572. Cancer stem cell enriched neurospheres derived from aggressive Group 3 subtype (Model ICb-1572) demonstrated susceptibility to 7-day in-vitro treatment with ispinesib. Monolayer tumor cells of Model ICb-1572 demonstrated similar susceptibility. Area under curve (AUC) > 0.7 was the significant cut-off. (f) Top view of cranial cavity of mice brains (Treatment Group = 7, Control Group = 6). We employed the clinical staging system to evaluate animal health fitness and determine their suitability to continue ispinesib therapy. Brains, particularly the tumor-bearing cerebellum (yellow arrows), from ispinesib-treated animals (right panel) were structurally more normal in appearance, in contrast to untreated control animals (left panel) where the cerebellums were distorted by tumor growth, soft and necrotic in appearance. (g) Overall survival and progression survival (Stage 3) of animals was significantly improved with ispinesib treatment (p = 0.0003, p = 0.03 respectively, Log-rank test).
