Poor response to neoadjuvant chemotherapy in metaplastic breast carcinoma

Metaplastic breast carcinoma (MpBC) is a rare special histologic subtype of breast carcinoma characterized by the presence of squamous and/or mesenchymal differentiation. Most MpBCs are of triple-negative phenotype and neoadjuvant chemotherapy (NAC) is frequently utilized in patients with MpBC. The aim of this study was to evaluate response to NAC in a retrospective cohort of MpBCs. We identified 44 patients with MpBC treated with NAC at our center between 2002 and 2018. Median age was 48 years, 86% were clinical stage II–III, and 36% were clinically node-positive. Most (80%) MpBCs were triple-negative or low (1–10%) hormonal receptor positive and HER2 negative on pre-NAC biopsy. While on NAC, 49% showed no clinical response or clinico-radiological progression. Matrix-producing subtype was associated with clinico-radiological response (p = 0.0036). Post NAC, two patients initially ineligible for breast-conserving surgery (BCS) were downstaged to be eligible for BCS, whereas three patients potentially eligible for BCS before treatment became ineligible due to disease progression. Only one (2%) patient had a pathologic complete response (pCR). Among the 16 patients presenting with biopsy-proven clinical node-positive disease, 3 (19%) had nodal pCR. Axillary lymph node dissection was avoided in 3 (19%) patients who had successful axillary downstaging. Residual cancer burden (RCB) was assessed in 22 patients and was significantly associated with disease-free survival and overall survival. We observed a poor response or even disease progression on NAC among patients with MpBC, suggesting that NAC should be reserved for patients with inoperable MpBC.


INTRODUCTION
Metaplastic breast carcinoma (MpBC) is a rare special histologic subtype of breast carcinoma, which includes a heterogeneous group of invasive carcinomas characterized by the presence of squamous and/or mesenchymal differentiation 1 . Histologic variants that are recognized by the World Health Organization (WHO) include spindle cell carcinoma, squamous cell carcinoma, and metaplastic carcinoma with mesenchymal differentiation, which includes matrix-producing carcinoma. Low-grade adenosquamous carcinoma and fibromatosis-like metaplastic carcinoma are the rare low-grade variants with a relatively favorable prognosis which differs from the more common high-grade MpBCs [2][3][4][5] .
MpBC typically presents as a rapidly growing mass that is often palpable and larger in size compared to invasive carcinoma no special type (NST) 6,7 . MpBCs are usually high grade and negative for estrogen receptor, progesterone receptor, and HER2 (i.e. of triple-negative phenotype) 6,[8][9][10] . Despite the large tumor size at presentation, axillary lymph node involvement is infrequent, and local recurrences and distant metastases are thought to occur by hematogenous dissemination 10,11 . Patients with MpBC tend to have worse disease-free survival and overall survival when compared to common forms of triple-negative breast cancer (TNBC) 10,[12][13][14][15][16][17][18][19] .
Given the advanced stage at initial presentation and triplenegative receptor status, neoadjuvant chemotherapy (NAC) is often considered a treatment option for patients with MpBC. Although rates of pathologic complete response (pCR) are relatively high in TNBC NST, at~30-40% with anthracycline and taxane 20,21 , and over 50% with platinum 22,23 , the response to NAC in MpBC has not been well studied and reported rates of pCR are variable, ranging from 10% to 17%, with sample size ranging from 6 to 29 patients [24][25][26][27][28] . Here we sought to study the response to NAC in patients with MpBC and clinical, radiologic, and pathologic features that were associated with response.

Post-NAC pathologic evaluation
Only one patient had pCR, an overall pCR rate of 2% (1/44) in this cohort ( Table 3). The patient was a 32 years old woman with germline BRCA1 del exons 23-24, presented with a 3 cm mass in the right breast at 32 weeks gestation. Core biopsy revealed a matrix-producing metaplastic carcinoma (Fig. 2), poorly differentiated and of triple-negative phenotype, clinical stage II (cT2N0). The patient started NAC with ACT after a c-section. Post-NAC bilateral mastectomy and sentinel lymph node biopsy revealed no residual invasive carcinoma. She remained without evidence of disease at the last follow-up (53 months). One patient had a pCR in the breast but had residual metastatic disease in one lymph node (Table 3). Among the 16 patients with biopsy-proven nodal involvement pre-NAC, 3 (19%) patients had nodal pCR post-NAC (Table 3).
Outcome analysis Follow-up information was available for 39 patients, with a median follow-up of 34.9 months (range: 1.6-140.4). Thirteen patients developed distant metastases. The 3-year disease-free survival and overall survivals were 58% and 65%, respectively. Residual cancer burden (RCB) was the only factor associated with disease-free survival by log-rank test (p = 0.0001) ( Table 4). On univariate analysis, post-NAC tumor size ≤ 2 cm (p = 0.046), post-NAC nodal status (p = 0.037), and RCB (p = 0.0007) were associated with overall survival (Table 4). Multivariate analysis was not performed due to the small sample size and missing data for RCB in a subset of patients.

DISCUSSION
MpBC is usually of triple-negative phenotype and appears more aggressive than conventional TNBC. The National Comprehensive Cancer Network (NCCN) clinical practice guidelines do not have specific treatment recommendations for MpBC distinct from that for invasive breast carcinoma in general 31 . NAC has become  standard practice for patients with clinical stage II-III TNBCs. However, in this study, we observed a poor response to NAC in patients with MpBC, with only one patient achieving pCR, 22% of the patients exhibited no clinical and radiological response, and 27% with progression while on NAC, in contrast to the 30-50% pCR rate in TNBC reported in the literature [20][21][22][23] . Benefits of NAC include downstaging the primary tumor to allow BCS and downstaging the axilla to avoid axillary dissection in patients presenting with node-positive disease. In our study, only two patients were sufficiently downstaged from ineligible to eligible for BCS post-NAC, and three patients progressed from eligible to ineligible for BCS during treatment.
Prior studies from our institution demonstrated nodal pCR rates of 41-48% in patients with cT1-3 biopsy-proven N1 breast cancer treated with NAC 32,33 . Similarly, in the ACOSOG Z1071 trial, of the 694 patients with biopsy-proven node-positive breast cancer (cT0-4 N1-2 M0), the nodal pCR rate was 41% 34 . The nodal pCR rate in patients with TNBC in the ACOSOG Z1071 trial was 49% (84/170) 34 24 . Whereas in our study, we included only cases with predominant metaplastic component, following the WHO classification guidelines 1 , potentially accounting for the lower response rates we observed.
Although the overall response rates in our study were low, differences in response based on histology were observed. The matrix-producing subtype was significantly associated with clinical-radiologic response. Of the 18 patients with matrixproducing subtype and with clinic-radiologic response assessment, 14 (78%) patients had a response to NAC, compared to 27% in non-matrix-producing MpBCs (p = 0.0036). The only patient with pCR had a matrix-producing MpBC. Han et al. also reported that matrix-producing subtype was significantly associated with pCR in their study cohort 25 .
The presence of tumor-infiltrating lymphocytes (TILs) is an independent predictor of response to NAC 35,36 . Denkert et al. reported a pCR rate of over 40% in lymphocyte-predominant breast cancer (LPBC), defined as breast cancer with more than 60% of TILs 35 . TNBC has the highest incidence of LPBC 36,37 . In a systematic review, a median of 20% (range 4-37%, n = 1620 patients) of patients with TNBC demonstrated LPBC, defined as at least 50% or 60% TILs 37 . MpBCs, however, are less frequently associated with high levels of TILs. In one study, a high level of TILs, defined as ≥60% of TILs, was observed in 33% of squamous cell carcinoma but none of the matrix-producing MpBC or spindle cell carcinomas 38 . In our study, high levels of TILs (≥50%) were only seen in 10% (3/31) of MpBCs, including 2 of 10 squamous cell carcinomas and 1 of 13 matrix-producing MpBC with TILs assessment in pre-NAC core biopsy.
Our study has several limitations. It is a retrospective study with the possibility of selection bias. Due to the rarity of MpBC, our sample size is limited. Data analysis was further restricted to patients with complete data in the report or with slides retrievable for review. Despite these limitations, our study is the largest series of MpBC treated with NAC with detailed clinicopathologic annotation.
In conclusion, MpBC had a poor response to conventional NAC. Only one (2%) patient achieved a pathological complete response in our cohort, and 27% had disease progression during NAC. The poor response to NAC in patients with MpBC raises the question of the utility of conventional NAC for these patients. While overall a low rate of clinicoradiologic and pathologic response, variation in response to NAC exists based on the histologic subtype, with some clinical benefit from NAC observed in matrix-producing MpBC, illustrating the heterogeneity within MpBC.

Patient cohort
Patients with MpBC treated with NAC at our center from 2002 to 2018 were retrospectively reviewed. Clinical and radiological features were obtained from the electronic medical record. Clinico-radiologic Table 3. Rates of pathologic complete response in breast and lymph node stratified by histologic subtype of metaplastic carcinoma.

Metaplastic histologic subtype
Overall pCR (ypT0N0) n (%) Breast pCR (ypT0) n (%) Total ypN0 n (%) Nodal pCR in cN +n (%) response was determined by a change in size between pre-NAC and post-NAC on physical examination and/or radiological imaging by the same imaging modality. Clinical lymph node status was determined by physical examination and imaging study, confirmed with fine-needle aspiration or core needle biopsy prior to NAC. Clinical follow-up postsurgical treatment and recurrence events were recorded, and diseasefree survival and overall survival were evaluated. This study was approved by the institutional review board of Memorial Sloan Kettering Cancer Center (protocol #17-287). A waiver of consent was granted by the institutional ethics committee because this work involves no more than minimal risk to the participants or their privacy.

Pathologic evaluation
Pathologic characteristics were retrieved from the pathology report and central slide review. Available diagnostic slides were reviewed by two breast pathologists (W.W. and H.W.) to confirm diagnoses and histologic features. A diagnosis of MpBC was rendered if metaplastic features were predominant in the tumor. The extent of TILs was assessed on pre-NAC biopsy according to the recommendations by the international TILs working group 41 . Pathologic evaluation post-NAC followed the recommendations of the international working group 42 . Pathological complete response (pCR) was defined as no evidence of invasive disease in the breast and lymph nodes, with or without residual ductal carcinoma in situ (ypT0/pTis ypN0). RCB was determined from the primary tumor bed area, overall cancer cellularity, percentage of cancer that is in situ disease, number of positive lymph nodes, and diameter of largest metastasis in lymph nodes, using the MD Anderson RCB calculator 43 . Estrogen receptor, progesterone receptor, and HER2 assessment followed the ASCO/CAP guidelines 44,45 and were recorded in both the pre-NAC biopsy and the post-NAC surgical specimens. Next-generation sequencing data using MSK-IMPACT were recorded when available 29,30 .

Statistics
Fisher's Exact test was used to evaluate associations between categorical variables and response to therapy. Survival outcomes were analyzed with the Kaplan-Meier method and statistical significance were determined by log-rank test. A p value < 0.05 was considered statistically significant.

Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.

DATA AVAILABILITY
The data generated and analyzed during this study are described in the following data record: https://doi.org/10.6084/m9.figshare.14823633 46 . All data are contained in the three Excel files: 'Clinicopathological characteristics.xlsx', 'Rates of pathologic complete response.xlsx' and 'Clinicopathologic factors associated outcomes.xlsx'. These files are housed on institutional storage and are not publicly available for the following reason: data contain information that could compromise research participant privacy and informed consent to share participant-level data was not obtained prior to or during data collection. Data will be made available to authorized researchers who have received approval from the Memorial Sloan Kettering Cancer Center Institutional Review Board. Any enquiries relating to the data should be directed to the corresponding author. Fig. 2 Representative micrograph of the pre-neoadjuvant treatment core biopsy of the sole patient who had pathologic complete response. Note that the core biopsy revealed a matrix-producing metaplastic carcinoma, poorly differentiated. Magnification ×200.