Morphology to molecular subtyping: Retrospective study to correlate the molecular subtypes of breast cancer with mammography and ultrasound imaging features

INTRODUCTION

Breast cancer is one of the leading causes of morbidity and mortality among females. With increasing awareness about breast cancer, the number of patients presenting with breast symptoms such as mastalgia, palpable lump, nipple discharge, etc. has also increased. Current-day breast imaging is not only limited to benign-malignant differentiation based on American College of Radiology (ACR) Breast Imaging Reporting and Data Systems (BI-RADS) guidelines but also includes molecular profiling, which is essential for personalized treatment strategies and guided diagnostic and therapeutic procedures.^[1]

Histopathological examination (HPE) and immunohisto-chemical analysis have been indispensable prerequisites for breast cancer treatment. However, tumor heterogeneity, such as mosaicism and variability in receptor status across multifocal or multicentric lesions, poses a significant challenge. Thus, image-based prediction may help in deciding if more than one lesion should be biopsied. Additionally, especially for HER2neu status, imaging-based subtyping may help us in deciding if fluorescence in situ hybridization is necessary for confirmation for receptor status. Moreover, in the era of machine learning and artificial intelligence, this information may play a significant role in designing appropriate models. Therefore, this study was designed to correlate the morphological features of malignant masses on mammogram (MG) and ultrasound (USG) with immunohistochemical markers for different molecular subtypes.

MATERIALS AND METHODS

The study was approved by the hospital’s Publication Oversee Committee (IEC Number CSP-MED/20/SEP/61/90), and the need for informed consent was waived off in view of the retrospective study design. The data of known breast cancer patients from April 2018 to March 2022 was retrieved from Hospital information system/Picture archiving and communication System (HIS/PACS), and only those patients with complete data sets, including MG, USG and HPE as well as receptor status, were included. The demographic data, MG findings, breast density, and HPE details, including receptor status, were tabulated.

RESULTS

In this study, a total of 192 patients were included with an age range of 51–60 years and a mean age of 56 years. Amongst these, 29 patients had Luminal A-type cancer, 82 were Luminal B, 36 had HER2-enriched, and 45 were triple-negative breast cancer. The most frequently observed breast density pattern was ACR category B, characterized by scattered areas of fibroglandular parenchyma.

Tables 1 and 2 represent MG and US features for each molecular subtype, respectively, based on the ACR lexicon criteria.

DISCUSSION

Over the last decade, the focus of diagnosing breast cancer has shifted from histopathological examination (HPE) alone to combined HPE and IHC analysis. Hormonal and oncogene-targeted therapies are important steps in the management of patients as part of preoperative planning or even for non-resectable metastatic tumors as palliative treatment. Hence, the contribution of radiologists is to aid in the diagnosis and management of such patients by identifying the unique features of imaging in each type.

In our study, Luminal A cases most commonly presented as irregular, spiculated masses on MGs [Figure 1a and 1b] and appeared hypoechoic and antiparallel on USG with posterior acoustic shadowing [Figure 1c]. AD was observed more frequently in this subtype, although the difference was not statistically significant. Our findings were in concordance with the other studies, which suggested that most Luminal type A masses had spiculated margins and more frequent association with AD as compared to other subtypes.^[2–10] We observed that microcalcifications were less common in Luminal A (24%) compared to other subtypes, which is in discordance with a study done by Cen et al., who reported grouped or clustered calcifications in 70% of hormone receptor-positive cancers.^[11] Similarly, studies by Rashmi et al. and Mi Young Kim et al. described the presence of coarse heterogeneous calcifications in Luminal A subtypes.^[12,13]

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Figure 1:

Luminal A breast cancer: Mammogram (a) CC and (b) MLO views of a 50-year-old female with Type B breast parenchyma show an irregular high-density mass with spiculated margins in upper central quadrant of the right breast with nipple retraction. No significant right axillary lymph adenopathy. (c) US shows an irregular anti-parallel oriented hypo-echoic mass with spiculated margins and posterior acoustic shadowing in the sub-areolar region of the right breast. (d) Increased internal vascularity is seen within. (e) No significant lymph nodes are seen in the right axilla. (f) Histopathology shows invasive mammary carcinoma- Nottingham histological grade II (H&E x 100). Tumor cells showing (g) ER positivity, (h) PgR positivity, (i) HER2 negativity (j) and Ki67 index <14 % (IHC x100). CC: Cranio-caudal, MLO: Mediolateral oblique, US: Ultrasound, H&E: Hematoxylin and eosin, ER: Estrogen receptor, PgR: Progesterone receptor, HER2: Human epidermal growth factor receptor 2, IHC: Immunohistochemistry.

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6% cases showed nipple retraction compared to Luminal B and HER2 enriched breast cancer [Figure 1d]. Luminal A cases in our study had fewer number of patients with axillary lymph adenopathy compared to other subtypes [Figure 1e]. Representative Figure 1f–j shows pathological findings for Luminal A mass which was invasive mammary cacinoma on histology with ER, PgR positivity, HER2 negativity and Ki67% index <14% on IHC.

In Luminal B, the masses were predominantly irregular in shape, spiculated, and exhibited high density on mammography. This aligns with the findings of Algazzar et al., who reported that 100% of Luminal B lesions in their study were irregular in shape.^[14] On USG also, 96.3% (n = 79) of masses were irregular in shape, similar to 76.7% of irregular Luminal B masses in a study by Boisserie-Lacroix et al.^[2] In this study, 53% of cases (n = 44) of Luminal B demonstrated spiculated margins [Figure 2a and b], and 28% of cases exhibited indistinct margins, a finding consistent with the results reported by Tong Wu et al.^[8] Nearly 96% (n = 79) of the masses identified on USG were anti-parallel in our study [Figure 2c], similar to the study by Shaikh et al. and Khalaf et al.^[3,6] 77% (n = 63) of masses were hypoechoic in our study, similar to the findings documented by Boisserie-Lacroix et al.^[2] A significant number of patients in this group showed AD, which is in line with observations made by Wojcinski et al.^[15] Calcifications, although less frequent, were primarily intralesional. Among these, fine linear and amorphous patterns were the most commonly observed, suggestive of malignancy, (Figure 2d US image showing no calcification within mass), which is similar to the findings described by Cen et al.^[11] Focal asymmetry was an uncommon feature, in agreement with the observations made by Ian et al.^[5] Tru-cut biopsy is required for pathological analysis due to suspcious imaging features [Figure 2e]. Representative Figure 2f–j shows pathological findings for Luminal B mass which was invasive mammary cacinoma on histology with ER, PgR positivity, HER2 negativity and Ki67% index >14% on IHC.

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Figure 2:

Luminal B breast cancer: Mammogram (a) CC and (b) MLO views of a 53-year-old female with Type B breast parenchyma showing an irregular high-density mass with spiculated margins in central inner quadrant of the right breast. (c) US shows an irregular anti-parallel oriented heterogeneously hypoechoic mass with angular margins and posterior shadowing in the right breast with no significant internal vascularity. (d) No calcifications seen within. (e) US guided Tru-cut biopsy image of the right breast mass. (f) Histopathology shows invasive mammary carcinoma, no special type, with islands of tumor cells. (H&E x 100). Tumor cells showing (g) ER positivity, (h) PgR positivity, (i) HER2 negativity and (j) Ki67 index >14 % (IHC x100). CC: Cranio-caudal, MLO: Mediolateral oblique, US: Ultrasound, H&E: Hematoxylin and eosin, ER: Estrogen receptor, PgR: Progesterone receptor, HER2: Human epidermal growth factor receptor 2, IHC: Immunohistochemistry.

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The HER2-enriched subtype typically presented as irregular, high-density masses on mammography [Figure 3a and b], a finding supported by previous studies.^[14,16] Margins were frequently micro-lobulated and non-circumscribed, consistent with observations reported by Wojcinski et al.^[15] This group had the highest incidence of calcifications among all subtypes, with [Figure 3a and b]. Among 20 cases, 16 (80%) exhibited intralesional calcifications. Amorphous calcifications were the most common, observed in 45% (n = 9) of patients, followed by fine pleomorphic calcifications in 40% (n = 8). These findings are in alignment with a study done by Cen et al., who reported amorphous or coarse heterogeneous calcifications in 56.5% of HER2-positive cases.^[11] However, other studies have also indicated an association of linear or linear branching calcification patterns with HER2-enriched tumors.^[4,17] On USG, 94% of cases appeared as irregular masses [Figure 3c], a finding that is in agreement with previously published studies.^[2,4,5] In contrast, Ḉelebi et al. reported that 55% of HER2-positive masses were circumscribed, highlighting variability in USG characteristics.^[10] We also found that nearly 97% (n = 28) of HER2-enriched masses were arranged in an anti-parallel orientation on USG [Figure 3c], consistent with earlier reports.^[3,6–8,18] In our study, nearly 72% HER2 cases had associated axillary lymphadenopathy [Figure 3d], which was concordant with the results of study by Khalaf et al.^[6] Tru-cut biopsy is performed for H&E and IHC analysis for molecular subtyping [Figure 3e]. Representative Figure 3f–j shows pathological findings for HER2 Enriched mass which was invasive mammary cacinoma on histology with ER and PgR negativity, HER2 positivity, and Ki67% index 20% on IHC.

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Figure 3:

HER2 Enriched breast cancer: Mammogram (a) CC and (b) MLO views of a 47-year-old female with Type B breast parenchyma showing an irregular high-density mass with micro-lobulated margins in upper outer quadrant of the right breast. Amorphous microcalcifications are seen within the mass. Mild diffuse skin thickening noted. (c) US shows an irregular anti-parallel oriented heterogeneously hypoechoic mass with micro-lobulated margins and few calcifications in the right breast. (d) Few enlarged lymph nodes with increased cortical thickness and loss of fatty hilum are seen in the right axilla. (e) US guided Tru-cut biopsy image of the right breast mass. (f) Histopathology shows invasive mammary carcinoma with a focal micropapillary pattern. (H&E x 100). Tumor cells with (g) ER negativity, (h) PgR negativity, (i) HER2 positivity and (j) Ki67 index 20 % (IHC x100). CC: Cranio-caudal, MLO: Mediolateral oblique, US: Ultrasound, H&E: Hematoxylin and eosin, ER: Estrogen receptor, PgR: Progesterone receptor, HER2: Human epidermal growth factor receptor 2, IHC: Immunohistochemistry.

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TNBC in our study commonly presented as irregular or round-to-oval masses, consistent with findings reported by previous studies.^{[7,14,19–25]} Circumscribed margins were frequently observed [Figure 4a and b], a feature also studied by Cho et al., who found that TNBC masses often demonstrate more circumscribed borders.^[4] Approximately 38% (n = 17) of the masses in our study exhibited parallel orientation [Figure 4c], aligning with the literature.^[7,20] AD was observed in 33% of cases, in contrast to the study by Ian et al., which reported no significant presence of this feature.^[5]Masses were heterogeneously hypoechoic in our study on USG, whereas a hypoechoic echo pattern is the most commonly described feature of TNBC.^{[7,8,16,19,24,25]} Although 24% of patients showed intralesional calcifications on USG, Ko et al.^[20] and Dogan et al.^[23] did not find any significant calcifications in TNBC masses on USG.^[19,20] Nearly 31% of cases had a cystic component within masses on USG imaging [Figure 4d] which may be attributed to the higher grade of these tumors. Schopp et al.^[21] has also suggested the appearance of TNBC masses as complex solid cystic masses and mimicking benign cystic entities.^[21] Skin thickening was seen in 26% of TNBC; however, Evans et al.^[19] reported that skin thickening has limited prognostic value in triple-negative breast cancer relative to other subtypes.^[22] TNBC cases also showed a higher incidence of lymph node involvement, corroborating with the findings of Evans et al.^[19], Tandon et al.^[7] found a notable portion of TNBC cases with lymph node involvement, while Cen et al.^[11] observed a lower incidence, and San-Gang Wu et al.^[26] reported an even smaller proportion.^[7,11,23] This variation suggests that the hematogenous route of metastasis might be more common in high-grade TNBC tumors compared to lymph nodal spread. Representative Figure 4e–i shows pathological findings for Triple-negative breast mass which was invasive mammary cacinoma on histology with ER, PgR and HER2 negativity with Ki67% index 45% on IHC.

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Figure 4:

Triple-negative breast cancer: Mammogram (a) CC and (b) MLO views of a 60-year-old female with Type A breast parenchyma showing an oval high-density mass with circumscribed margins in the subareolar region of the right breast. Two focal microcalcifications are seen within the lesion. Popcorn calcification is seen in the upper central quadrant of the right breast posterior to the aforementioned mass and within two high-density masses seen in the upper outer and lower inner quadrant respectively in the left breast– suggestive of involuting fibroadenoma. (c) US shows an oval circumscribed heterogeneously hypoechoic mass with posterior acoustic enhancement in the subareolar region of the right breast. (d) Mild internal vascularity and cystic components are seen in the mass. (e) Histopathology shows invasive mammary carcinoma, no special type. (H&E x 100). Tumor cells with (f) ER negativity, (g) PgR negativity, (h) HER2 negativity and (i) Ki67 index 45 % (IHC x100). CC: Cranio-caudal, MLO: Mediolateral oblique, US: Ultrasound, H&E: Hematoxylin and eosin, ER: Estrogen receptor, PgR: Progesterone receptor, HER2: Human epidermal growth factor receptor 2, IHC: Immunohistochemistry.

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Although this study is limited by small sample size and retrospective design from a single institute, it has highlighted the role of IHC correlation with imaging markers, which may assist in further prospective validation studies. This study demonstrated Luminal A to have predominantly irregular masses with spiculated margins and posterior acoustic shadowing; Luminal B as irregular masses with spiculated or micro-lobulated margins and no specific posterior features; and HER2-enriched tumors as predominantly irregular masses with micro-lobulated margins, mixed posterior features, and intra-lesional calcifications. TNBC cases were relatively round to oval, well-circumscribed masses with parallel orientation and posterior acoustic enhancement.

CONCLUSION

To conclude, a diligent approach in interpreting suspicious breast masses on MGs and USGs according to the ACR-BI-RADS 5^th edition Lexicon shall lead to the correct interpretation and correlation of imaging with molecular subtypes of breast cancer in concordance with gold standard IHC biomarkers.