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Jun Shinoda, Hiromu Yamada, Noboru Sakai, Takashi Ando, Toshifumi Hirata and Yoshiaki Miwa

✓ A sensitive enzyme-linked immunosorbent assay (ELISA) was used in a retrospective study of placental alkaline phosphatase (PLAP) levels in serum, cerebrospinal fluid (CSF), and intratumoral cyst fluid in primary intracranial germinoma. The ELISA showed no cross-reactivity with intestinal alkaline phosphatase except in very high concentrations, after samples had been heat-treated. Three patients with germinoma were studied for serum PLAP levels and in all the levels were elevated (3.78, 0.52, and 2.11 IU/liter). Two of the germinoma patients were studied for PLAP levels in the CSF, and both had elevated levels (0.83 and 9.83 IU/liter). The intratumoral cyst fluid in one case of germinoma was tested for PLAP and the level was found to be very high (603 IU/liter). These PLAP levels decreased concomitantly with the reduction in tumor size during irradiation. Serum PLAP levels were measured in 40 control adult male individuals and in the CSF of 20 nonpregnant patients with subarachnoid hemorrhage. The upper normal limits were 0.20 and 0.11 IU/liter in the serum and the CSF, respectively. All PLAP levels measured in the serum of patients with various brain tumors were 0.18 IU/liter or less. This study strongly suggests that PLAP is a clinically useful tumor marker for primary intracranial germinoma.

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Jun Shinoda, Hirohito Yano, Shin-Ichi Yoshimura, Ayumi Okumura, Yasuhiko Kaku, Toru Iwama and Noboru Sakai

✓ The authors have recently performed a fluorescence-guided tumor resection procedure by using high-dose fluorescein sodium without any special surgical microscopes for the intraoperative visualization of glioblastoma multiforme (GBM), and they report on the actual procedure and clinicopathological findings.

Thirty-two patients with GBMs underwent tumor resection during which this fluorescence-guided procedure was used. Fluorescein sodium (20 mg/kg) was intravenously injected after dural opening at the craniotomy site. The tumor was stained almost homogeneously yellow and the color was intense enough to be readily perceived for resection. The center of the solid lesion was stained a deep yellow and surrounded by a transition zone that was faintly stained. The colored lesion was clearly distinguishable from the unstained zone outside the GBM, particularly in the white matter. Both the deeply and faintly stained regions included endothelial proliferation and dense tumor cells. In the unstained region, less dense tumor cells were consistently revealed; however, no endothelial proliferation could be seen. Grosstotal resection (GTR) was successful in 84.4% of the patients who received an injection of fluorescein sodium, which accounted for 100% of those in whom all the visible yellow color (both the deeply and faintly stained regions) was judged to have been resected during operation. Gross-total resection was performed in 100% of the patients who underwent the fluorescence-guided procedure and assigned to Stage I, a GBM stage in which, as a therapeutic policy, the tumor should be resected as radically as possible. The GTR rates in patients who received fluorescein sodium were significantly higher than those in patients who did not (73 patients with GBMs who underwent tumor resection without the fluorescence-guided procedure). Although the extent of surgery was revealed to be one of the significant and independent prognostic factors for GBM, the fluorescein sodium—guided resection procedure was not a significant or independent prognostic factor in this series.

This surgical procedure does not require any special surgical microscopic equipment and is simple, safe, useful, readily accomplished, and universally available for resection of GBMs. Its efficacy simplifies the surgical procedure of navigating the stained lesion from the unstained area to achieve GTR of GBMs, which can be demonstrated on magnetic resonance images.

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Jun Shinoda, Yoshiaki Miwa, Noboru Sakai, Hiromu Yamada, Hiroto Shima, Kazuo Kato, Masayoshi Takahashi and Kuniyasu Shimokawa

✓ Indirect immunoperoxidase staining by the peroxidase-antiperoxidase (PAP) technique was carried out on 23 human primary intracranial germ-cell tumors (17 germinomas, one embryonal carcinoma, one yolk-sac tumor, three teratomas, and one teratoma with embryonal carcinoma) and on six human primary pineal nongerm-cell tumors (one pineocytoma, two pineoblastomas, two astrocytomas, and one glioblastoma multiforme). The technique used specific rabbit antisera against placental alkaline phosphatase (PLAP), alpha-fetoprotein (AFP), and human chorionic gonadotropin (HCG). Thirteen of 17 intracranial germinomas (76.5%) showed positive staining for PLAP mainly on the tumor cell membrane. In six primary intracranial non-seminomatous germ-cell tumors, there was weak positive staining indicating the presence of PLAP in only a few cells of one embryonal carcinoma, and in some glandular epithelial cells of one teratoma; this staining was limited to the cytoplasm. None of the other six primary pineal non-germ-cell tumors showed any positive PLAP reaction. From these results, PLAP was shown to be very useful in histopathology as a diagnostic tumor marker of intracranial germinoma.

Positive AFP staining was seen in several yolk-sac tumor cells and a few embryonal carcinoma cells. However, no intracranial germinomas and non-germ-cell tumors of the pineal region showed positive reaction. As for HCG, only one suprasellar germinoma and one pineal embryonal carcinoma among 29 specimens contained a few positive-staining tumor cells.

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Hiromichi Ando, Masanao Saio, Noriyuki Tamakawa, Naoyuki Ohe, Takashi Nakayama, Hai Yu, Yasuhiko Kaku, Toru Iwama, Jun Shinoda, Noboru Sakai and Tsuyoshi Takami

Object. It is well known that the central nervous system (CNS) is an immunologically privileged site. To characterize CD8+ tumor-infiltrating lymphocytes (TILs) recovered from the CNS, the authors compared these cells with TILs recovered from subcutaneous tissue by using a B7.1 gene—modified tumor implantation model.

Methods. The authors established a B7.1 gene—modified EL4 murine lymphoma cell line (EL4-B7.1) and implanted the cells into the CNS to observe the duration of tumor-free survival. Although EL4-B7.1 cells were completely rejected in a subcutaneous implantation model, 40% of animals died after the CNS implantation (all animals in which the parent tumor was implanted died within 16 days). Therefore, the authors isolated TILs from each implantation site and analyzed the expressions of activation antigens CD25 and CD69 by performing the anti-CD8 magnetic beads separation method and flow cytometric analysis. After implantation of the parent tumor, there was no difference in the number of TILs from each site (CD25 1.7–3.2%, CD69 21.9–34.3%). After implantation of the B7.1-modified tumor, the CD25-expressing TIL population from the subcutaneous site was 4.68 times higher than that from the CNS site (17.8% compared with 3.8%). Based on these findings, the authors used a mitomycin C—treated EL4-B7.1 subcutaneous vaccination with various protocols. Vaccination before tumor challenge was sufficient to prevent the development of the tumor. For animals with established tumor, the vaccination protocol was able to prolong host survival (p = 0.0053).

Conclusions. The data clearly demonstrate that the CNS environment fails to activate CD8+ TILs fully. These are the first data indicating in detail a difference between CD8+ TILs from the CNS and those from other sites based on a B7.1-modified tumor model.

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Hiroaki Takei, Jun Shinoda, Soko Ikuta, Takashi Maruyama, Yoshihiro Muragaki, Tomohiro Kawasaki, Yuka Ikegame, Makoto Okada, Takeshi Ito, Yoshitaka Asano, Kazutoshi Yokoyama, Noriyuki Nakayama, Hirohito Yano and Toru Iwama

OBJECTIVE

Positron emission tomography (PET) is important in the noninvasive diagnostic imaging of gliomas. There are many PET studies on glioma diagnosis based on the 2007 WHO classification; however, there are no studies on glioma diagnosis using the new classification (the 2016 WHO classification). Here, the authors investigated the relationship between uptake of 11C-methionine (MET), 11C-choline (CHO), and 18F-fluorodeoxyglucose (FDG) on PET imaging and isocitrate dehydrogenase (IDH) status (wild-type [IDH-wt] or mutant [IDH-mut]) in astrocytic and oligodendroglial tumors according to the 2016 WHO classification.

METHODS

In total, 105 patients with newly diagnosed cerebral gliomas (6 diffuse astrocytomas [DAs] with IDH-wt, 6 DAs with IDH-mut, 7 anaplastic astrocytomas [AAs] with IDH-wt, 24 AAs with IDH-mut, 26 glioblastomas [GBMs] with IDH-wt, 5 GBMs with IDH-mut, 19 oligodendrogliomas [ODs], and 12 anaplastic oligodendrogliomas [AOs]) were included. All OD and AO patients had both IDH-mut and 1p/19q codeletion. The maximum standardized uptake value (SUV) of the tumor/mean SUV of normal cortex (T/N) ratios for MET, CHO, and FDG were calculated, and the mean T/N ratios of DA, AA, and GBM with IDH-wt and IDH-mut were compared. The diagnostic accuracy for distinguishing gliomas with IDH-wt from those with IDH-mut was assessed using receiver operating characteristic (ROC) curve analysis of the mean T/N ratios for the 3 PET tracers.

RESULTS

There were significant differences in the mean T/N ratios for all 3 PET tracers between the IDH-wt and IDH-mut groups of all histological classifications (p < 0.001). Among the 27 gliomas with mean T/N ratios higher than the cutoff values for all 3 PET tracers, 23 (85.2%) were classified into the IDH-wt group using ROC analysis. In DA, there were no significant differences in the T/N ratios for MET, CHO, and FDG between the IDH-wt and IDH-mut groups. In AA, the mean T/N ratios of all 3 PET tracers in the IDH-wt group were significantly higher than those in the IDH-mut group (p < 0.01). In GBM, the mean T/N ratio in the IDH-wt group was significantly higher than that in the IDH-mut group for both MET (p = 0.034) and CHO (p = 0.01). However, there was no significant difference in the ratio for FDG.

CONCLUSIONS

PET imaging using MET, CHO, and FDG was suggested to be informative for preoperatively differentiating gliomas according to the 2016 WHO classification, particularly for differentiating IDH-wt and IDH-mut tumors.

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Oral Presentations

2010 AANS Annual Meeting Philadelphia, Pennsylvania May 1–5, 2010