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PD-L1 expression in invasive solid papillary breast carcinomas Mungan SA, Yılmaz ZS, Saygın I, Cakır E, Ersoz S, Cobanoglu U


Background: Invasive solid papillary carcinomas (ISPC) are rare malignant neoplasms in the classification of WHO 2019 breast tumors. Aims: We aimed to investigate the correlations between programmed cell death ligand-1 (PD-L1) expression status of tumor and immune cells and clinicopathological parameters by molecular classification of this rare morphological subtype. This study will contribute to the literature about the PD-L1 expression state of ISPCs for the first time. Material and Methods: The study included 19 invasive solid papillary carcinoma cases diagnosed between 2009 and 2019 in Pathology Department. Molecular subtyping was performed in 19 cases by immunohistochemical studies (ER/PR, Her-2/neu, Ki-67), and PD-L1 expression was evaluated in neoplastic and immune cells. Results: PD-L1 expression was detected in 4 (21%) cases, 3 (75%) of them were in luminal B and 1 (25%) were in the luminal A group. The correlation between molecular subtypes and PD-L1 expression was statistically significant (P = 0.016). Patients with PD-L1 expression had a higher Ki-67 index than patients without PD-L1 expression (P = 0.037). In addition, there was a statistically significant correlation between PD-L1 expressions of intratumoral lymphocytes and PD-L1 expressions of neoplastic cells (P = 0.004). Conclusions: While predicting the group that will benefit more from immunotherapy in solid papillary carcinoma cases, not only PD-L1 expression of tumor cells but also PD-L1 expression in tumor infiltrating lymphocyte (TIL) can help. In addition, PD-L1 staining rates of tumor cells as well as clinicopathological parameters (molecular subtype, high Ki-67 index, presence of TIL) can be predictive about immunotherapy.

Keywords: Breast carcinoma, immunohistochemistry, programmed cell death ligand-1, solid papillary carcinoma

How to cite this article:
Mungan SA, Yılmaz ZS, Saygın I, Cakır E, Ersoz S, Cobanoglu U. PD-L1 expression in invasive solid papillary breast carcinomas. Indian J Pathol Microbiol 2022;65:311-5

How to cite this URL:
Mungan SA, Yılmaz ZS, Saygın I, Cakır E, Ersoz S, Cobanoglu U. PD-L1 expression in invasive solid papillary breast carcinomas. Indian J Pathol Microbiol [serial online] 2022 [cited 2022 May 5];65:311-5. Available from: https://www.ijpmonline.org/text.asp?2022/65/2/311/343178

   Introduction   Top

Invasive solid papillary carcinoma (ISPC) is included in the 2019 WHO classification as a histomorphological subtype of epithelial breast carcinomas.[1] ISPC is a slow-growing malignant neoplasm with better prognosis than other breast carcinomas. It constitutes 1.3% of all breast carcinomas in women. It is generally detected in the 7th decade and postmenopausal women.[2] Prognostic factors such as lymphovascular invasion, lymph node, or distant metastasis correlate with the grade of the invasive component of the tumor.[3] Neuroendocrine and mucinous differentiation can accompany at variable rates.[1]

It is widely accepted that breast carcinomas are a heterogeneous cancer group.[4] There are cancers that have different biological behaviors in the same histological subtype. Therefore, histological classification may be insufficient to explain some parameters. There has been a need for more explanatory classification for personalized medicine and being more beneficial to current oncological approaches. Recent studies focused on genomic, transcritomic, epigenetic and proteomic data obtained by advanced technologies while making the current molecular classification of breast carcinomas.[5] However, accessing this data is expensive and may not always be possible. With immunohistochemical studies that based on these data, but cheaper and feasible method, breast carcinomas are molecularly divided into 4 main groups: Luminal A, Luminal B, Her-2/neu and Triple Negative Breast Carcinoma (TNBC).[4],[6] The purpose of the molecular classification of breast carcinomas is to give an accurate diagnosis that assist the decision in the oncological approach and also to predict tumor behavior.[4] A study on the molecular classification of ISPCs, a rare histological subtype in the literature, has not been reported so far.

One of the most current treatment modalities among oncological approaches is immunotherapy using checkpoint inhibitor agents that target programmed cell death-1 (PD-1) and programmed cell death ligand-1 (PD-L1) blockade. By using checkpoint inhibitor drugs (PD-1/PD-L1 blockade), improvement in overall survival has been published in many malignancies such as melanoma, non-small cell lung carcinomas, head and neck and renal tumors.[7],[8]

Almost all of the current PD-L1 studies have been performed in the TNBC group, which is known to have poor prognosis in breast carcinomas. In this study, we analyzed the molecular subtyping and PD-L1 expressions of ISPCs which are rare subtypes of breast carcinomas having good prognosis and slow course. We investigated the possible correlations between PD-L1 expressions of ISPCs, their molecular subtypes and clinicopathological parameters such as concomitant ductal carcinoma in situ (DCIS) component, tumor infiltrating lymphocyte (TIL), tumor size, lymphovascular invasion, neuroendocrine and mucinous differentiation, Her-2/neu status, and hormone expressions.

   Materials and Methods   Top

Patient selection

22 cases who were diagnosed as invasive solid papillary breast carcinoma between 2009-2019 years were selected from the archive of the Pathology Department. Three cases were excluded from the study because the invasive area was very small (<4 mm) and could not be represented in serial sections. Histopathological diagnosis was confirmed by two pathologists experienced in breast pathology. 19 cases diagnosed by excision materials were included in the study.

Age and tumor size at the time of diagnosis were obtained from the report samples for each patient. In addition, DCIS, TIL, lymphovascular invasion, neuroendocrine and mucinous differentiation status were evaluated again from hematoxylin eosin (HE) stained sections. Immunohistochemical studies were performed to determine the molecular subtype, hormone expression (ER/PR), Her-2/neu and PD-L1 protein expression states of each case. According to the tumor size the cases were divided into 3 groups as ≤2 cm, >2- ≤5 cm and >5 cm.


For immunohistochemical studies, 4 micron thick sections prepared from formalin fixed paraffin embedded (FFPE) tumoral tissues were used. All immunostaining studies were performed on a Roche/Ventana BenchMark Ultra platform [For details see [Table 1]]. When evaluating PD-L1 expression, reactive tonsil tissue was used as positive control tissue. The results of immunohistochemical studies were evaluated according to the reported breast carcinoma biomarker guideline updated by the American College of Pathologists (CAP) in February 2020.[9] The immunexpressions of neoplastic cells are shown in [Figure 1]. The complete/incomplete membranous staining pattern in neoplastic cells was considered positive in evaluation of PD-L1 expression. Cases containing neoplastic cell expressing more than 1% PD-L1 were noted positive. For TIL, the cut of value was 1% like neoplastic cells.[8]

Figure 1: (a) Infiltrative pattern of solid papillar carcinoma of breast, (b) Neuroendocrine differentiation of invasive solid papillar carcinoma (HEX100)

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ER expression rates were grouped as negative (0%), low positive (1-10%) and positive (>10%). PR expressions were evaluated as negative (0%) and positive (≥1%).

Her-2/neu expression of invasive tumor cells was scored in 4 categories between 0-3. Score 0: No staining or membrane stating that is incomplete and is faint/barely perceptible and within ≤10% of tumor cells, Score 1: Incomplete membrane staining that is faint/barely Score 2: Weak to moderate complete membrane staining in >10% of tumor cells or complete membrane staining that is intense but within ≤10% of tumor cells, Score 3: Complete membrane staining that is intense and >10% of tumor cells. Score 0 and 1 were considered negative and Score 3 was considered positive. Score 2 was directed for the Flourecent In Situ Hybridization (FISH)-Her-2/neu study. In FISH- Her-2/neu evaluation, those with an average Her-2/neu copy number <4.0 signal in the tumor cells were considered negative and those with a ≥ 6 signal were considered positive. Immunohistochemical and FISH- Her-2/neu tests were repeated in those with an average signal count of 4-6.[9]

According to Ki-67 staining rates, we divided 19 cases into two groups, less than 15% and equal/high stained. This rate is the threshold value used in Luminal A and B group separation in molecular subtyping of breast carcinomas.[4],[5],[6]

TIL evaluation

The lymphocytes and plasma cells in the invasive tumor areas were counted. The cases containing 40 or more intratumoral immune cells were considered TIL positive. While evaluating TIL, the most presentative tumor block was used and counted areas were averaged. The stroma of the invasive tumor parenchyma was focused. Lymphocyte and plasma cells infiltrated in the stromal area were counted. It was not decided with hotspot areas with aggregates. Peripheral tumor and necrotic areas containing neutrophils were not included in the TIL examination.[1]

Statistical analysis

Distribution of continuous variables was assessed by Shapiro Wilk’s test, and Student t-test was used for the comparison of normally distributed variable. Fisher exact test was used for the comparison of the categorical variables. R Software (R Foundation for Statistical Computing, Vienna, Austria) was used for statistical analyses.

Ethical considerations

Ethics committee approval was received for this retrospective study by the Faculty of Medicine Scientific Research Ethics Committee with the document number 24237859-219.

   Results   Top

The average age of the patients was 64.0 (±12.8). The age range of 19 cases was 45-90. All patients were women. A total of 19 patients were provided with paraffin-embedded tissues from mastectomy, lumpectomy or excisional specimens for immunohistochemistry. The clinicopathological data of the patients included in the study are shown in [Table 2].

Table 2: Relationship of PD-L1 expression and clinicopathological characteristics

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Hormone expression profile: All of the cases were positive for both estrogen (ER) and progesteron (PR). There was no low estrogen group (<%10) according to the lastTemplate for Reporting Results of Biomarker Testing of Specimens From Patients With Carcinoma of the Breast ” reported by college of American pathologists.[9] Each case had estrogen and progesteron staining in neoplastic cells at least 40% immunohistochemically.

Her-2/neu status: All of the cases were Her-2/neu negative. Her-2/neu expression states of all cases were scored by immunohistochemical technique first. The score was 0 and 1 in 18 cases. In only 1 case, score 2 results were obtained. In this case, Her-2/neu amplification with FISH was evaluated and negative result was obtained.

Ki-67 index: 14 of 19 cases (73%) showed less than 15% staining. One of 14 cases (7.1%) with low proliferation index was PD-L1 positive. PD-L1 expression was observed in 3 (60%) of 5 cases whose Ki-67 index was 15% or more. There was a statistically significant correlation between Ki-67 index and PD-L1 expressions (P = 0.037).

Molecular Subtype: We performed molecular subtyping using the immunohistochemistry technique in 19 cases diagnosed with invasive solid papillary carcinoma. All cases were ER/PR positive, that belong to Luminal breast carcinomas group. None of our cases showed Her-2/neu positivity. Therefore, we did not have any case meeting Her-2/neu type breast carcinoma and TNBC molecular subtypes. Of the 19 cases, all of which expressed ER/PR, 14 were included in the Luminal A subtype due to their low Ki-67 indexes and 5 were included in the Luminal B classes due to their higher proliferation rate. One of 14 (7.1%) Luminal A cases expressed PD-L1 by more than 1% (PD-L1 positive), 3 of 5 Luminal B breast carcinomas (60%) were immunohistochemically PD-L1 positive. PD-L1 positivity of Luminal B type ISPC was statistically significant (P = 0.016).

Tumor size: The tumor sizes of the patients who participated in the study were between 1 and 4.5 cm. According to the TNM staging system, 8 cases pT1; tumor diameter is ≤2 cm, and 11 cases pT2; tumor diameters are between 2-5 cm. While 3 (37%) of 8 cases in the pT1 group were PD-L1 positive (>1% expression), 1 of 11 cases (9%) in the pT2 group showed more than 1% expression with PD-L1. There was no statistically significant correlation between tumor size and PD-L1 expression (P = 0.262).

Lymphovascular invasion: Three (15%) of 19 ISPC cases had lymphovascular invasion. Two (66%) of 3 cases with lymphovascular invasion had more than 1% PD-L1 expression (PD-L1 positive). In 2 (12%) of 16 cases without lymphovascular invasion, PD-L1 expression is more than 1%. In ISPC cases, there is no statistically significant difference in the presence or absence of lymphovascular invasion and PD-L1 expression (P = 0.097).

Neuroendocrine differentiation: In 7 (36%) of 19 cases, focal neuroendocrine (<50%) differentiation is available. In 2 (28%) of 7 cases with neuroendocrine differentiation, PD-L1 expression is above 1% (PD-L1 positive). Of the 12 cases without neuroendocrine differentiation, 2 (16%) expressed PD-L1 expression more than 1% (PD-L1 positive) and 10 were PD-L1 negative. No statistically significant correlation was found between neuroendocrine differentiation and PD-L1 expression of neoplastic cells (P = 0.603).

Mucinous differentiation: In 8 (42%) of 19 cases, focal mucinous (<50%) differentiation is present. In 3 (37%) of 8 cases with mucinous differentiation, PD-L1 expression is above 1% (PD-L1 positive). In 1 (9%) of 11 cases without mucinous differentiation, PD-L1 expressed more than 1% (PD-L1 positive) and 10 were PD-L1 negative. There was no statistically significant correlation between mucinous differentiation and PD-L1 expression of neoplastic cells (P = 0.262).

Ductal carcinoma in situ (DCIS): 13 (68%) of 19 cases had ductal carcinoma in situ (DCIS) foci. In 2 (15%) of 13 cases with DCIS foci, PD-L1 expression is above 1% (PD-L1 positive). In 2 (33%) of 6 cases with no DCIS focus, PD-L1 expressed more than 1% (PD-L1 positive), and 4 were PD-L1 negative. There was no statistically significant correlation between the presence of DCIS and PD-L1 expression of neoplastic cells (P = 0.557).

Tumor infiltrating lymphocyte (TIL): 9 of 19 cases (47.4%) had TIL. In 4 (44.4%) cases with TIL, more than 1% PD-L1 expression was detected in neoplastic cells (PD-L1 positive) [Figure 2]. In 10 (52.6%) cases, there was no TIL. PD-L1 expression was not detected in cases without TIL. There was a significant correlation between the presence of TIL and PD-L1 expression (P = 0.033).

Figure 2: Immunohistochemical staining of PD-L1 of four positive cases. (a-d) Tumor cells expressed PD-L1 (IHC X200), (e) Epithelial staining of PD-L1 in neoplastic cells (long arrow) and PD-L1 expression of tumor infiltrating lymphocytes (short arrows)

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PD-L1 expressions were also evaluated in infiltrative lymphocytes in patients with TIL. We compared PD-L1 expression in lymphocytes infiltrating the tumor with PD-L1 expression in neoplastic cells. In 3 (33.3%) of 9 cases with TIL, more than 1% PD-L1 expression in neoplastic cells was noticed. In 1 (11%) patient without PD-L1 expression in lymphocytes, neoplastic cells were PD-L1 positive (>1% expression). A statistically significant correlation was found between PD-L1 expressions of intratumoral lymphocytes and PD-L1 expressions of neoplastic cells (P = 0.004).

   Discussion   Top

The PD-L1 gene is well known in cancer immunology and has been under investigation for a while. The purpose of the checkpoint inhibitor drugs is to prevent the immune escape mechanism created by binding with the ligand of the PD-L1 protein encoded by this gene.[7],[10] PD-L1 inhibitor drugs, also known as checkpoint inhibitors, give hope for many cancers in targeted oncological treatment.[8] There are no reports in the literature of PD-L1 expressions of invasive solid papillary carcinomas yet. It is known that PD-L1 expression is a positive predictive marker in other solid carcinomas. Information about the relationship between PD-L1 expression in breast carcinomas and prognosis differs from other cancers in the literature. Sabatier et al.[11] reported that PD-L1 expression is a negative prognostic marker related to large tumor size, high grade, ER-negative, PR-negative, high proliferation, ERBB2-positivity in breast cancers. Wang et al. studied PD-L1 in TNBCs and published that PD-L1 positivity in TILs, not in neoplastic cells is a negative prognostic marker.[8] We found a statistically significant correlation between PD-L1 expression and Ki-67 index (proliferation rate) in the slow-growing, well-prognosed ISPC group. In our study, there was no significant correlation between PD-L1 expression in neoplastic cells of ISPCs and age, gender, hormone expression status, Her-2/neu amplification, tumor size, lymphovascular invasion, neuroendocrine or mucinous differentiation, and concomitant DCIS. However, there are publications in the literature showing that there is a correlation between large tumor size, ER/PR negativity, Her-2/neu positivity and PD-L1 upregulation.[10],[11] The statistically significant relationship found by this study between tumor proliferation rate and PD-L1 expression is compatible with the literatüre.[10],[11],[12]

Molecular subtyping which is the current classification in breast carcinomas, and PD-L1 expression status and immunotherapy are among the current research topics. There are PD-L1 studies of some subtypes such as basal type in the literature. It has been stated that PD-L1 expression rates are higher in basal type and TNBC groups, so they can benefit from immunotherapy more than the others.[7],[12],[13]

Our study was carried out in ISPCs. This histomorphological subtype of breast carcinoma is rare and is known to have good prognosis. Therefore, this study did not have all molecular classification subgroups. All of the cases included in the study belong to Luminal A and Luminal B groups according to molecular classification. However, when these two groups were compared, it was seen that most of the cases expressing PD-L1 belonged to the Luminal B subtype.

TIL is present in 9 of the 19 patients who participated in the study. Presence of TIL and PD-L1 expression of lymphocytes participating in TIL are also parameters used to predict the response to immunotherapy. In our study, a statistically significant correlation was found between PD-L1 expression of tumor cells and the presence of TIL and the expression of PD-L1 of TIL (P = 0.033 and P = 0.004). In other words, when PD-L1 is expressed in cases with TIL or in lymphocytes involved in TIL, tumor cells also express PD-L1 and the probability of success for immunotherapy increases. Wang et al.[8] published that in TNBCs, a poor prognostic group of breast carcinomas, that PD-L1 expression in TILs can predict survival. Many recent studies have shown that PD-L1 protein is expressed more in TILs than neoplastic cells.[14],[15] The first data reported about the PD-L1 expression of ISPCs, in tumor cells and TIL belong to our study.

   Conclusions   Top

Invasive SPCs, which are known to have good prognosis may express PD-L1 even at a low rate, which may be useful in detecting cases with a relatively high rate of proliferation and planning the treatment. In addition, not only the PD-L1 expression of neoplastic cells, but also the presence of TIL, the TIL’s PD-L1 expression and the Luminal B subtype in the molecular classification can give an idea to determine the predictivity of immunotherapy. Due to the retrospective nature of the study and the low number of cases, confirmation of the current hypothesis with repeated data is required.

Financial support and sponsorship


Conclicts of interest

There are no conflicts of interest.


   References   Top

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Sabatier R, Finetti P, Mamessier E, Adelaide J, Chaffanet M, Ali HR, et al. Prognostic and predictive value of PDL1 expression in breast cancer. Oncotarget 2015;6:5449-64.  Back to cited text no. 11
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Sun S, Fei X, Mao Y, Wang X, Garfield DH, Huang O, et al. PD-1 (+) immune cell infiltration inversely correlates with survival of operable breast cancer patients. Cancer Immunol Immunother 2014;63:395-406.  Back to cited text no. 14
Schalper KA, Velcheti V, Carvajal D, Wimberly H, Brown J, Pusztai L, et al. In situ tumor PD-L1 mRNA expression is associated with increased TILs and better outcome in breast carcinomas. Clin Cancer Res 2014;20:2773-82.  Back to cited text no. 15


Correspondence Address:
Sevdegul Aydın Mungan
Pathology Department, Karadeniz Technical University Faculty of Medicine, Kalkinma Mah Farabi Hastanesi Tıbbi Patoloji AD 61080 Trabzon
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/IJPM.IJPM_1229_20

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[Table 1], [Table 2]



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