PMC4852191_01
Histological appearance of chordoma. (A,B) Sections of primary tumor stained with hematoxylin and eosin (H&E). (C) Parallel section stained for brachyury. Scale bar: 50 mum.
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F1
Histological examination (Figures 1A,B) showed tumor composed predominantly of physaliphorous cells arranged in nests and set in a mucinous matrix. Tumor cells were positive for cytokeratins and S100 (not shown), and also for brachyury (Figure 1C), the molecular hallmark of chordoma.
Objective response of chordoma to linsitinib and erlotinib. (A) Baseline MRI pelvis in June 2009, showing right iliopsoas deposit. Additional marker lesions were measured in the oblique right psoas (29 mm) and left paravertebral region (35 mm) giving a sum for the three lesions of 99 mm. (B) Response MRI of December 2010 showing reduction in size of right iliopsoas deposit. (C) Graph: monitoring of serial MRI scans by RECIST criteria, represented as the sum of measurements of the three marker lesions described in (A). After 44 months (February 2013), these three lesions were no longer clearly measurable, and a new marker lesion (left external iliac, 57 mm) was selected to monitor response to linsitinib dose escalation from 50 mg QD to BID.
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MRI, on the same date, showed recurrent tumor with measurable components within the right iliopsoas muscle (mid-right psoas 35 mm, Figure 2A; right side of L3/4, 29 mm) and to the left side of the intervertebral device at L4 (35 mm). Sequential MRI scans revealed initial disease stabilization, with gradual improvement to partial response by response evaluation criteria in solid tumors (RECIST) in December 2010, after 18 months of study treatment (Figures 2B,C). The patient continued on linsitinib 50 mg BID and erlotinib 100 mg QD with stabilization of the left external iliac deposit (Figure 2C). She came off study in August 2014 after 61 months of trial treatment, and subsequently experienced further symptomatic deterioration accompanied by progression in the left external iliac deposit (Figure 2C), prompting further pelvic surgery in April 2015.
Chordoma of trial patient contains prominent nuclear IGF-1R and membrane EGFR. (A) The specificity of IGF-1R and EGFR IHC was tested using sections of formalin-fixed, paraffin-embedded MCF7 breast cancer cells (high IGF-1R, low EGFR) and SKUT-1 leiomyosarcoma cells (IGF-1R deficient, EGFR positive). (B) Sections of chordoma from the trial patient were stained for IGF-1R and EGFR, including primary tumor from 2003, and recurrent tumor from 2008 (pre-trial) and 2015 (post-trial). Scale bar: 50 mum.
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IGF-1R and EGFR expression, using an IGF-1R IHC protocol recently optimized for sensitivity and specificity. Initial testing in FFPE control cell line pellets confirmed the specificity of the IGF-1R and EGFR antibodies in IHC (Figure 3A). IGF-1R signal that was weaker in the primary biopsy and one of the blocks (C, psoas deposit) of recurrent tumor and stronger and more uniformly positive in two further blocks (A, B; deposits in connective tissue) from the 2008 (pre-trial) recurrence (Figure 3B). In all tumor samples, there was detectable membrane and cytoplasmic IGF-1R, with clearly apparent nuclear positivity that showed heterogeneity between different regions of tumor, and was most evident in the 2008 (pre-trial) recurrence. The tumor also contained detectable EGFR, with weak/moderate membrane and cytoplasmic signal but no nuclear positivity (Figure 3B). Analysis of tumor from the 2015 (post-trial) recurrence showed that EGFR and IGF-1R were still detectable, but with reduced intensity compared with the 2008 recurrence, and reduced nuclear IGF-1R (Figure 3B).
Brain and spinal chordomas contain IGF-1R with variable membrane, cytoplasmic, and nuclear positivity. IGF-1R IHC was performed on the primary tumor, 3 tumor blocks from the 2008 recurrence and one block from the 2015 (post-trial) recurrence of the trial patient, and 15 additional chordoma cases. Each tumor was scored for IGF-1R signal in the membrane, cytoplasm, and nucleus for the percentage showing zero (0), weak (1), moderate (2), or heavy (3) IGF-1R intensity, giving a maximum histoscore in each subcellular location of 100% x 3 = 300.
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To determine the extent to which these appearances are representative, we performed IGF-1R IHC on 15 additional chordoma cases (Figure 4), including 3 cases originating in the sacrococcygeal region and 12 cases arising in the brain.