Association between real-time shear wave elastography findings and HER-2 expression in breast cancer

Introduction

Breast cancer is the most common diagnosed cancer and the leading cause of cancer death among women (1). In China, the age-standardized incidence rate and age-standardized mortality rates of female breast cancer in individuals over 20 years old increased from 46.34 per 100,000 in 2003 to 68.78 per 100,000 in 2017, and from 11.12 per 100,000 in 2003 to 11.67 per 100,000 in 2017, respectively (2). Higher incidence and mortality rates were seen in coastal cities when compared to inland areas, with urban centers also surpassing rural regions in breast cancer prevalence.

With the advancement of molecular profiling, breast cancer is classified into distinct molecular subtypes according to the expression of immunohistochemical markers, including estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor-2 (HER-2), and the proliferation marker Ki-67 (3). These subtypes include luminal A, luminal B, HER-2-overexpressing, and basal-like breast cancers (3). Emerging evidence suggests that the pathogenesis and mechanism underlying the progression of disease among these molecular subtypes are different, thus leading to various treatment approaches and prognostic outcomes (4). Notably, HER-2 gene amplification occurs in approximately 15–20% of breast cancers. HER-2 overexpressing breast cancer is associated with more aggressive tumor characteristics and poorer clinical outcomes as HER-2 promotes tumor cell proliferation, differentiation, and survival (5). The primary treatment approach for HER-2-positive breast cancer is to inhibit the activity of HER-2 receptor (6). Monoclonal antibodies such as trastuzumab and small-molecule tyrosine kinase inhibitors are widely used targeted cancer therapies (6). Their ability to bind to HER-2 receptors overexpressed on the surface of cancer cells inhibits subsequent cellular activation and thereby suppresses tumor growth and proliferation (6). Given the pivotal role of HER-2 expression in disease prognosis and therapeutic responsiveness, it is important to determine HER-2 status of breast cancer for better treatment planning.

Ultrasonography is a commonly used non-invasive imaging technique for screening and diagnosis of breast cancer especially in low-resource countries (7). Tumor boundary, tumor margin, presence of calcification, spatial vascularity pattern, and surrounding tissue change could be observed on conventional ultrasound and correlated with the specific molecular profiles of the breast cancer (8). There is growing evidence that information such as tissue stiffness, blood flow and perfusion obtained from multimodal ultrasound are associated with biological markers and molecular subtypes of breast cancer (9). Shear wave elastography (SWE) is another recently developed ultrasound-based technique that enables the quantitative assessment of tissue elasticity (10). As malignant tissues typically exhibit high Young’s modulus and showed “stiff ring sign” at the edges of the tumor (10), SWE will be a useful imaging modality for diagnosis of breast cancer. However, the association between SWE features and the expression of key molecular biomarkers, such as HER-2, remains limited. The present study therefore aims to further elucidate the relationship between the SWE characteristics of breast tumors and HER-2 status, and investigate the association between SWE parameters and HER-2 status, contributing to the understanding of imaging biomarkers in breast cancer. We present this article in accordance with the STARD reporting checklist (available at https://tcr.amegroups.com/article/view/10.21037/tcr-2024-2359/rc).

Methods

Study participants

A total of 67 female patients (70 lesions) at mean age of 55.29±10.87 years (range, 37–82 years) who underwent surgery for breast cancer at The Affiliated People’s Hospital of Ningbo University between January 2017 and January 2021 were retrospectively studied. The diagnosis of breast cancer was confirmed by pathological examination of samples obtained from either biopsy or surgery. SWE was conducted prior to surgical treatment for all patients and the Young’s modulus values of the lesions were assessed. All cases must meet the following inclusion criteria: (I) availability of SWE imaging, pathology and immunohistochemistry data; (II) availability of SWE data prior to any surgical intervention, radiotherapy, chemotherapy, or immunotherapy; (III) good SWE quality and adherence to diagnostic requirements; and (IV) no history of breast augmentation surgery, mastitis, radiotherapy, chemotherapy, or breast-related surgery. Exclusion criteria were as follows: (I) patients were in the lactation or pregnancy period; (II) presence of any breast nodule at a diameter more than 50 mm; (III) the interval between ultrasound examination and surgery was longer than 1 month; or (IV) presence of large calcification foci or breast cystic lesions that could interfere with the analysis. The study was conducted in accordance with the Declaration of Helsinki and its subsequent amendments. The study was approved by the institutional ethics committee of The Affiliated People’s Hospital of Ningbo University (No. 2021-064). Informed consent was obtained from all participants.

SWE imaging

SWE images were obtained using the AixPlorer ultrasound system (SuperSonic Imagine, Aix-en-Provence, France) equipped with an L14-5 linear array probe operating at a frequency range of 5–14 MHz. The “Elasto” mode was selected and the display scale was set at 140 kPa. During the imaging examinations, patients were positioned in the supine or semi-recumbent position. In addition to the SWE measurements, a routine breast ultrasound examination was conducted to record the location, size, morphology, border characteristics, and vascular flow of the lesions.

During the imaging assessment, the region of interest (ROI) for feature extraction was delineated on the largest cross section of B-model image. The size of the sampling frame was adjusted to include the lesion and its surrounding normal breast glandular and adipose tissues. At the time of image acquisition, the ultrasound probe set between 5–14 MHz was applied with minimal pressure for at least 3 seconds and patients were instructed to remain still and hold their breath as much as possible to minimize motion-related artifacts. The examination area was adjusted to approximately twice the size of the lesion, and the quantitative elastic modulus value of the lesion was measured. Each lesion underwent three repeated measurements, and the average value was calculated for subsequent analysis.

Image analysis

The ROI automatically displayed on the color-coded elasticity map was labeled as ROI-M. Additionally, concentric shells were applied to select the edge areas at 1, 2, and 3 mm from the lesion border, designated as ROI-1 mm, ROI-2 mm, and ROI-3 mm, respectively. Upon delineation of lesions and surrounding edge regions, the ultrasound system automatically generated the elastic parameters, including the mean elastic modulus (E_mean), maximum elastic modulus (E_max), and minimum elastic modulus (E_min), for each of the defined ROIs.

Biomarker assessment

According to the established guidelines for HER-2 determination in breast cancer (11), immunohistochemical expression of HER-2 can be classified into four distinct levels: 0, 1+, 2+, and 3+. Cases with HER-2 expression levels of 0 or 1+ are considered HER-2-negative, while those graded as 3+ are considered HER-2-positive. Tumors with immunohistochemical HER-2 score 2+ require further evaluation by fluorescence in situ hybridization (FISH) to ascertain the presence or absence of HER-2 gene amplification. Additionally, ER positivity is defined by the percentage of tumor cells with nuclear staining by immunohistochemistry, i.e., <10% (−), 10–25% (+), 26–50% (++), and >50% (+++). For PR and Ki-67, the established cutoff for positivity is 15%.

Statistical analysis

SPSS 26.0 software was used for statistical analysis. Since this was a retrospective study using available clinical data, no formal sample size calculation was performed. The relatively high proportion of HER-2-positive cases may reflect selection bias inherent in patients referred for preoperative elastography. Chi-squared test was used for analysis of categorical data. Continuous data were expressed as mean ± standard deviation and analyzed using independent sample t-test. MedCalc for Windows, version 16.8 (MedCalc Software, Mariakerke, Belgium) was used to generate and analyze the receiver operating characteristic (ROC) curve of each diagnostic method for determination of HER-2 expression. The Z test was used to compare the area under the curve (AUC) and diagnostic efficacy between those methods. P value of less than 0.05 was considered statistically significant.

Results

A total of 100 pathologically confirmed breast cancer patients who underwent ultrasound SWE prior to any anti-cancer treatment in The Affiliated People’s Hospital of Ningbo University were selected. Among them, 33 patients were excluded from the retrospective analysis because either molecular subtyping of the breast cancer could not be performed or pathological findings were incomplete. Ultimately, the study cohort consisted of 67 patients with 70 breast cancer lesions were included in the final analysis. Of the 70 lesions evaluated, 31 were classified as HER-2 negative (negative group) and the remaining 39 lesions were classified as HER-2 positive (positive group) according to the biomarker assessment criteria as aforementioned. No significant difference was observed for basic characteristics and pathological diagnosis of breast cancer between HER-2 negative and positive groups (Table 1).

Compared with HER-2 negative group, HER-2 positive group had a higher proportion of lesions with a diameter of ≥20 mm, irregular shapes, spiculated edges, and calcification signs (P<0.05). However, there was no significant difference in the “stiff ring sign” and aspect ratio of breast cancer between the HER-2 negative and positive groups (Table 2). The E_max of HER-2 positive group was significantly lower than that of the negative group (P<0.05), but there was no difference in Young’s modulus values E_mean and E_min between HER-2 negative and positive groups (Table 3). Details of ultrasound images are depicted in Figure 1. Relationship between elasticity modulus values and expression of biomarkers were further explored. Over-expression of HER-2 was found statistically associated with the E_max values obtained from SWE measurements (P=0.02) whereas the expression level of other biomarkers including ER, PR, Ki-67 were not associated with the E_max values (Table 4).

Figure 1 Ultrasound images of HER-2 negative and positive breast cancer. HER-2 negative group: (A) grayscale ultrasound image at size of 1.0 cm × 7.0 cm; (B) SWE image plus Q-box: E_max =78.8 kPa, E_mean =42.1 kPa, E_min =17.7 kPa. HER-2 positive group: (C) grayscale ultrasound image at size of 1.7 cm × 1.5 cm; (D) SWE image plus Q-box: E_max =192.0 kPa, E_mean =38.3 kPa, E_min =3.7 kPa. E_max, maximum elastic modulus; E_mean, mean elastic modulus; E_min, minimum elastic modulus; HER-2, human epidermal growth factor receptor-2; SWE, shear wave elastography.

ROC curves of different methods for determination of HER-2 status of breast cancer were compared (Figure 2). The AUC value of E_max combined with tumor size and calcification status was significantly higher than individual parameters (Z =1.97, P=0.049; Z =2.26, P=0.02; Z =2.22, P=0.03). The AUC value of E_max combined with tumor size was not significantly higher than that of E_max and tumor size alone (P=0.33, Z =0.983; P=0.25, Z =1.14). The AUC value of E_max combined with calcification was significantly higher than that of calcification alone (P=0.04, Z =2.092) while the combined AUC value was also higher than E_max alone (P=0.66, Z =1.84). The AUC value of tumor size combined with calcification was significantly higher than that of calcification alone (P=0.002, Z =3.040) while the combined AUC value was not significantly higher than that of tumor (P=0.10, Z =1.64) (Table 5).

Figure 2 ROC curves of tumor E_max, size, calcification and combined methods in determination HER-2 status in breast cancer. E_max, maximum elastic modulus values; HER-2, human epidermal growth factor receptor-2; ROC, receiver operating characteristic.

Discussion

The distinct pathological and morphological changes driven by the intrinsic biological behavior of breast cancer contribute to diverse imaging manifestations (12,13). Conventional B-mode ultrasound allows continuous real-time scanning to detect breast lesions and assess their morphological features such as shape, echogenicity, and size, but it cannot assess the stiffness or hardness of the lesions alone. It is noteworthy that fibrosis surrounding the breast tumor typically results in increased tissue hardness compared to benign lesions. Therefore, there exists a correlation between the expression of specific protein molecules in breast cancer and the corresponding ultrasound imaging characteristics.

SWE has demonstrated diagnostic performance in the evaluation of both thyroid and breast tumors (12,14). Changes in the elasticity of breast cancer lesions are closely linked to the proliferation and infiltration of cancer cells within the tumor. Specifically, proliferation of cancer cells leads to increased tissue hardness, while tumor infiltration to surrounding tissues can further increase stiffness of the breast (15). Compared to other elastographic techniques, SWE boasts distinct advantages, including its quantitative, independent, and highly reproducible nature (16). Following the unique “Mach cone” principle, this advanced modality harnesses the differential propagation velocities of shear waves across diverse tissue types to derive the absolute value of tissue elasticity, which is expressed as the Young’s modulus values in units of Pa or kPa. This Young’s modulus value is positively associated with the hardness of the examined tissue (17). Within the context of breast cancer, there is growing evidence that the molecular characteristics of tumor can influence its overall biological behavior, tissue properties, and associated pathological changes, resulting in various degree of tissue softness or hardness (11). By employing SWE to objectively and accurately measure the stiffness of breast cancer masses, researchers can further elucidate the relationships between distinct genotypic profiles and the corresponding elasticity characteristics of the tumors.

Regarding the study of HER-2 expression levels and SWE parameters, the present findings indicate that HER-2-positive breast cancers exhibit higher elasticity measurements, which is consistent with the existing literature (18). Specifically, this study identified that E_max within the lesion itself are important risk factors. This observation may be attributed to the highly proliferative and aggressive nature of HER-2-positive breast cancer which usually has a higher histological grade. Lesions of HER-2-positive breast cancer therefore demonstrate reduced tissue elasticity compared to their HER-2-negative counterparts. These results underscore the potential utility of quantitative SWE examination in assessing the underlying biological characteristics and behavior of breast tumors based on their elasticity profiles.

The present study revealed that HER-2-positive breast cancer exhibits distinct radiological characteristics with large tumor size, irregular shape, blurred margins, and internal calcifications. HER-2-positive tumors are characterized by irregular contours and spiculated margin which may be attributed to aggressive invasion to peripheral tissues, which are consistent with previous reported study (19). In addition, the presence of calcifications is significantly more prevalent among HER-2-positive patients compared to their HER-2-negative counterparts. This may be explained by the rapid growth kinetics of HER-2-positive tumors which can result in localized tissue ischemia and subsequent liquefactive necrosis, ultimately leading to the accumulation of calcium salts in the tumor. Collectively, these findings suggest that tumor size and presence of calcifications within breast tumors may be associated with HER-2 positivity.

Further analysis on the influence of other clinically relevant biomarkers on E_max was also conducted. In contrast to HER-2, the expression of ER/PR and the proliferation marker Ki-67 did not demonstrate any significant correlation with E_max. These findings may be attributed to the unique pathological characteristics of HER-2-positive tumors, which often exhibit stromal edema, heightened cellularity, increased microvessel density, elevated tissue stiffness, and the presence of intratumoral (4,20-22). Therefore, the present data suggest that the E_max parameter derived from SWE may potentially serve as a non-invasive surrogate marker for evaluating the HER-2 expression status of breast tumor.

To explore the accuracy of E_max in the determination of HER-2 status, comparative analyses on ROC curves of individual and combined utilities of Young’s modulus E_max, tumor size, and the presence of calcifications for diagnosing HER-2-positive breast cancer were performed. The results demonstrated that the combined application of these parameters significantly enhanced the diagnostic efficiency compared to using any individual parameter alone. Specifically, this multiparametric approach exhibited high specificity, albeit with slightly lower sensitivity. These findings suggest that the combined assessment of E_max, tumor size, and calcifications can provide a more objective and efficient non-invasive evaluation of HER-2 positivity in clinical practice. However, there are several limitations in this study. Small sample size and data obtained from a single institution may introduce potential bias. More so, there is absence of information on lymph node metastasis and tumor vascularity. Further detailed investigation is therefore required. The accuracy of SWE parameters may be influenced by patient selection criteria and prior screening history, which could affect the generalizability of these findings. Moreover, future studies with larger cohorts and controlled stratification of molecular subtypes will be necessary to further investigate if this relationship between SWE-E_max and Ki-67 is independent of HER-2 status.

Conclusions

In summary, the combined assessment of E_max derived from SWE, along with tumor size and the presence of calcifications, may assist in identifying imaging characteristics associated with HER-2-positive breast cancer, but its predictive accuracy remains limited. Given the potential for misclassification, further validation in larger cohorts is necessary before clinical application. While this approach may contribute to a more comprehensive understanding of tumor characteristics, it should not be considered a substitute for biopsy in treatment decision-making. The findings suggest that such imaging-based evaluations could support, but not replace, existing diagnostic strategies in assessing tumor invasiveness and guiding treatment planning for patients with HER-2-positive breast cancer.

Acknowledgments

None.

Footnote

Reporting Checklist: The authors have completed the STARD reporting checklist. Available at https://tcr.amegroups.com/article/view/10.21037/tcr-2024-2359/rc

Data Sharing Statement: Available at https://tcr.amegroups.com/article/view/10.21037/tcr-2024-2359/dss

Peer Review File: Available at https://tcr.amegroups.com/article/view/10.21037/tcr-2024-2359/prf

Funding: None.

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tcr.amegroups.com/article/view/10.21037/tcr-2024-2359/coif). The authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. The study was conducted in accordance with the Declaration of Helsinki and its subsequent amendments. The study was approved by the institutional ethics committee of The Affiliated People’s Hospital of Ningbo University (No. 2021-064). Informed consent was obtained from all participants.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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