Multiparametric magnetic resonance imaging for radiation therapy response monitoring in soft tissue sarcomas: a histology and MRI co-registration algorithm

Abstract

Rationale: To establish a spatially exact co-registration procedure between in vivo multiparametric magnetic resonance imaging (mpMRI) and (immuno)histopathology of soft tissue sarcomas (STS) to identify imaging parameters that reflect radiation therapy response of STS. Methods: The mpMRI-Protocol included diffusion-weighted (DWI), intravoxel-incoherent motion (IVIM), and dynamic contrast-enhancing (DCE) imaging. The resection specimen was embedded in 6.5% agarose after initial fixation in formalin. To ensure identical alignment of histopathological sectioning and in vivo imaging, an ex vivo MRI scan of the specimen was rigidly co-registered with the in vivo mpMRI. The deviating angulation of the specimen to the in vivo location of the tumor was determined. The agarose block was trimmed accordingly. A second ex vivo MRI in a dedicated localizer with a 4 mm grid was performed, which was matched to a custom-built sectioning machine. Microtomy sections were stained with hematoxylin and eosin. Immunohistochemical staining was performed with anti-ALDH1A1 antibodies as a radioresistance and anti-MIB1 antibodies as a proliferation marker. Fusion of the digitized microtomy sections with the in vivo mpMRI was accomplished through nonrigid co-registration to the in vivo mpMRI. Co-registration accuracy was qualitatively assessed by visual assessment and quantitatively evaluated by computing target registration errors (TRE). Results: The study sample comprised nine tumor sections from three STS patients. Visual assessment after nonrigid co-registration showed a strong morphological correlation of the histopathological specimens with ex vivo MRI and in vivo mpMRI after neoadjuvant radiation therapy. Quantitative assessment of the co-registration procedure using TRE analysis of different pairs of pathology and MRI sections revealed highly accurate structural alignment, with a total median TRE of 2.25 mm (histology - ex vivo MRI), 2.22 mm (histology - in vivo mpMRI), and 2.02 mm (ex vivo MRI - in vivo mpMRI). There was no significant difference between TREs of the different pairs of sections or caudal, middle, and cranial tumor parts, respectively. Conclusion: Our initial results show a promising approach to obtaining accurate co-registration between histopathology and in vivo MRI for STS. In a larger cohort of patients, the method established here will enable the prospective identification and validation of in vivo imaging biomarkers for radiation therapy response prediction and monitoring in STS patients via precise molecular and cellular correlation.

Figure 1

Flowchart of the co-registration procedure.

Figure 2

Selection of 3 representative STS sections (cranial, middle, and caudal part) of patient 1. (A) In vivo MRI; (B) Ex vivo MRI; (C) Macropathological specimen; (D) Histopathological H&E staining. Cau: caudal; cra: cranial; mid: middle.

Figure 3

Representative caudal section of a pleomorphic sarcoma (patient 2). Color-coded landmarks were defined by a pathologist and radiologist on (A) in vivo MRI, (B) ex vivo MRI, the specimen (not shown), and (C) the H&E microtomy section. (D) Exemplary illustration of the TRE computation: landmarks were randomly split into two sets, one for nonrigid co-registration (green) and the other for evaluating the co-registration accuracy as the Euclidian distance in millimeters (white connections of landmarks). TRE: target registration error

Figure 4

Morphological MRI sequences of the three STS patients. Exemplary axial T2w images (A, B, C) and corresponding coronal T2w-STIR images (A', B', C') of the three STS patients included. The dashed line in the lower row indicates the sectioning plane of axial T2w images in the upper row. STIR, short tau inversion recovery.

Figure 5

Representative cranial section of an MFS (patient 1). Visual assessment of (A) axial in vivo T2w, (B) axial ex vivo T2w, (C) macroscopic section, and (D) H&E stained microtomy section shows a high spatial and morphological correlation. Note the light deformations of the microtomy section (D) at the cutting edges (black arrows) as well as split and microtomy artifacts (white arrows).

Figure 6

Results of the target registration error analysis illustrated as boxplots combined with jitter plots. (A) Pairs of MRI and histology sections: left, histology - ex vivo MRI; middle, histology - in vivo MRI; right, ex vivo - in vivo MRI. (B) Sections of the caudal (left), middle (middle), and cranial (right) STS parts. Boxplots: middle line represents the median; the upper and lower ends of the box represent the 75th and 25th percentiles, respectively. Jitter plot: Dots indicate landmarks color-coded for each patient. Y-axis: target registration error (TRE) in mm.

Figure 7

Initial comparison between (immuno)histology and quantitative mpMRI parameters. (A-D) H&E and (A'-D') Mib1 stainings in 100x magnification show the four predominant histology patterns on the section: Cell-rich - vital tumor cells (A and A'); sclerotic background - vital tumor cells (B and B'); myxoid - vital tumor cells (C and C'); predominant myxoid - single vital tumor cells (D and D'). (E-H) Exemplary parametric maps for each quantitative MRI sequence: DCE - K^trans (E); DWI - ADC (F); IVIM - fD* (G). (H) Result of the K-means clustering based on all 8 available parametric Maps; White squares indicate the area of the histological images.

Figure 8

Boxplots of all parametric MRI parameters for each distinct Region A-D as shown in Figure 7. Boxplots: middle line represents the median; the upper and lower ends of the box represent the 75th and 25th percentiles, respectively. Black dot in Box represents the mean. Color-coding identical to Figure 7: region A, yellow; region B blue; region C, red, region D, cyan. P values were obtained using one-way ANOVA, followed by the t-tests (Bonferroni-Holm adjusted) for comparison of mean values between regions. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001; ns, not significant.