PMC6719522_01
Clinicoradiological and pathological findings in case 1. The clinical progression of case 1 is summarized in uppermost. In 2002, the lesion was composed of round monotonous tumor cells with perinuclear halos (A) and showed increased cellularity with microvascular proliferation in 2009 (B). The most recent tumor, in 2017, displayed relatively low cellularity but increased cytological atypia (C) (A-C, hematoxylin and eosin staining; original magnification, x 200). The tumor cells showed well-preserved IDH1-R132H positivity at each recurrence (D-F), and P53 staining intensity became stronger with further recurrences (G-I). In comparison, ATRX expression was well-maintained according to two earlier biopsies (J,K) but was lost in the recent recurrence episode (L). Interestingly, the specimen in 2009 demonstrated strong ATRX expression but also contained tumor cells showing loss of ATRX expression in small areas (O, arrowheads) (D-L,O, IHC results; original magnification, x 400). Chromosome 1 FISH performed on the 2002 specimen demonstrated loss of 1p (red) in the tumor cells (M). Chromosome 19 FISH performed on the 2017 specimen demonstrated two copies of 19q (red) and 19p (green) in the tumor cells (N) (M,N, FISH; original magnification, x 1,000). *ATRX, alpha-thalassemia/mental retardation X-linked; FISH, fluorescence in situ hybridization; IDH, isocitrate dehydrogenase; IHC, immunohistochemistry.
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F1
Clinical course and radiological features are summarized in Figure 1 (uppermost). The original lesion at the age of 29 years was a diffusely infiltrating tumor composed of round, uniform cells with perinuclear halos (Figure 1A). Intratumoral microvessels were proliferative, with increased endothelial layers (Figure 1B). The second recurrent tumor, obtained from the third surgery, was composed of oval-to-round nuclei with mild nuclear atypia (Figure 1C). The tumor cells of initial and recurrent were positive for IDH1-R132H (Figures 1D-F). P53 staining intensity was negative in most of the initial tumor cells and became stronger with further recurrences (Figures 1G-I). ATRX preservation was observed in the initial and first recurrent tumor (Figures 1J,K). The second recurrent tumor showed the loss of ATRX (Figure 1L). Interestingly, a small number of tumor cells of the first recurrent mass demonstrated ATRX loss and were intermingled with ARTX-preserving cells (Figure 1O). 1p/19q FISH examination performed on the first two tumors revealed 1p/19q codeletion (Figure 1M), while the second recurrent tumor did not (Figure 1N).
Illustration of mutations detected by next-generation sequencing analysis in case 1, visualized in Integrative Genome Viewer. IDH1 mutation R132H was preserved at the same loci as that of the original tumor in the most recent recurrence (A). TERT promoter mutation C228T and KRAS mutation G12D were maintained from the original to the recurrent anaplastic oligodendroglioma, but were lost in the recurrent astrocytoma (B,C). By contrast, mutations of ATRX, TP53, and CDKN2A were newly acquired at the most recent astrocytoma recurrence. Arrows indicate the retention of the same mutations at each recurrence (D-F). *ATRX, alpha-thalassemia/mental retardation X-linked; IDH, isocitrate dehydrogenase.
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F2
The mutations detected by NGS are summarized in Figure 2 and Supplementary Data Sheet 2. To exclude accidental mix-up of samples, Y-chromosome specific short tandem repeat (Y-STR) analysis was performed on three sequential samples of the first patient. The analysis result showed that all FFPE samples belonged to the same patient (Supplementary Data Sheet 3).
PMC6719522_02
Clinicoradiological and pathological findings in case 2. The clinical progression of case 2 is summarized in uppermost. The original lesion displayed microvascular proliferation among monotonous tumor cells with perinuclear halos in 2008 (A) and then converted to a tumor with marked cytological atypia in 2016 (B) (A,B, hematoxylin and eosin stain; original magnification, x 200). The tumor cells showed IDH1-R132H positivity by IHC at both occasions (C,D). P53 immunoreactivity converted from weak heterogeneous positivity of the wild type in 2008 to strong positivity of the mutant type at recurrence (E,F). ATRX expression was strongly positive in the original tumor (G) but was lost at recurrence (H). Similar to case 1, the specimen in 2009 demonstrated ATRX-negative cells that intermingled with the cells showing strong ATRX expression (I, arrowheads) (C-H,I, IHC results; original magnification, x 400). Chromosome 1 FISH performed on the 2002 specimen demonstrated loss of 1p (red) in the tumor cells (J). Chromosome 19 FISH performed on the 2016 specimen demonstrated two copies of 19q (red) and 19p (green) in the tumor cells (K) (J,K, FISH; original magnification, x 1,000). *ATRX, alpha-thalassemia/mental retardation X-linked; FISH, fluorescence in situ hybridization; IDH, isocitrate dehydrogenase; IHC, immunohistochemistry.
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F3
The tumor showed geographic necrosis and microvascular proliferation in compact tumoral areas (Figure 3A). The lesion was composed of highly pleomorphic tumor cells and microvascular proliferation was present (Figure 3B). IHC of origin & recurrent tumors showed positive for IDH1-R132H (Figures 3C,D). P53 staining of origin tumor was that of the wild type but the pleomorphic cells of recurrent tumor were strongly positive for P53 (Figures 3E,F). While the origin tumor cells showed preservation of ATRX expression, the recurrent tumor displayed ATRX loss (Figures 3G,H). Similar to the first recurrent tumor of case 1, a small number of tumor cells in the recurrent tumor exhibited ATRX loss among those with strong ATRX expression (Figure 3I). 1p/19q FISH examination performed on the initial tumors revealed 1p/19q codeletion (Figure 3J), while the recurrent tumor did not (Figure 3K).
PMC6719522_03
Clinicoradiological and pathological findings in case 2. The clinical progression of case 2 is summarized in uppermost. The original lesion displayed microvascular proliferation among monotonous tumor cells with perinuclear halos in 2008 (A) and then converted to a tumor with marked cytological atypia in 2016 (B) (A,B, hematoxylin and eosin stain; original magnification, x 200). The tumor cells showed IDH1-R132H positivity by IHC at both occasions (C,D). P53 immunoreactivity converted from weak heterogeneous positivity of the wild type in 2008 to strong positivity of the mutant type at recurrence (E,F). ATRX expression was strongly positive in the original tumor (G) but was lost at recurrence (H). Similar to case 1, the specimen in 2009 demonstrated ATRX-negative cells that intermingled with the cells showing strong ATRX expression (I, arrowheads) (C-H,I, IHC results; original magnification, x 400). Chromosome 1 FISH performed on the 2002 specimen demonstrated loss of 1p (red) in the tumor cells (J). Chromosome 19 FISH performed on the 2016 specimen demonstrated two copies of 19q (red) and 19p (green) in the tumor cells (K) (J,K, FISH; original magnification, x 1,000). *ATRX, alpha-thalassemia/mental retardation X-linked; FISH, fluorescence in situ hybridization; IDH, isocitrate dehydrogenase; IHC, immunohistochemistry.
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F3
The tumor showed geographic necrosis and microvascular proliferation in compact tumoral areas (Figure 3A). The lesion was composed of highly pleomorphic tumor cells and microvascular proliferation was present (Figure 3B). IHC of origin & recurrent tumors showed positive for IDH1-R132H (Figures 3C,D). P53 staining of origin tumor was that of the wild type but the pleomorphic cells of recurrent tumor were strongly positive for P53 (Figures 3E,F). While the origin tumor cells showed preservation of ATRX expression, the recurrent tumor displayed ATRX loss (Figures 3G,H). Similar to the first recurrent tumor of case 1, a small number of tumor cells in the recurrent tumor exhibited ATRX loss among those with strong ATRX expression (Figure 3I). 1p/19q FISH examination performed on the initial tumors revealed 1p/19q codeletion (Figure 3J), while the recurrent tumor did not (Figure 3K).