1. Introduction
Neoadjuvant or pre-operative chemotherapy (NAC) is classically administered to patients with inflammatory or locally advanced breast cancer (BC). Beyond increasing breast-conserving surgery rates [
1], it also serves as an in vivo chemosensitivity test and the analysis of residual tumor burden may help understanding treatment resistance mechanisms [
2]. In addition, it helps refining the prognosis of patients after NAC, as pathological complete response (pCR) after NAC is associated with a better long-term survival [
1,
3].
Nearly 5% of breast cancers occur in a context of genetic predisposition, mostly represented by monoallelic pathogenic variants of
BRCA1,
BRCA2 or
PALB2 genes [
4]. Patients with loss-of-function of the
BRCA1 or
2 proteins have a higher cumulated breast cancer risk, with a cumulated lifetime risk at eighty years old of 72% (
BRCA1) and 69% (
BRCA2) [
5]. The peak incidence for
BRCA1 mutation carriers occurs between 41 and 50 years old (28.3 per 1000 person-years), whereas it occurs ten years later for
BRCA2 mutation carriers (30.6 per 1000 person-years between 51 and 60) [
5].
BRCA1 and
BRCA2 are tumor-suppressor genes that code for proteins involved in homologous recombination (HR) repair. HR deficiency (HRD) occurs when the second allele is inactivated by allelic deletion (often detected by LOH), genic alteration or promoter methylation (for
BRCA1 only). Biallelic
BRCA1/2 inactivation results in genomic instability and theoretically increases the somatic mutational load [
6].
Tumors associated with germline or somatic
BRCA1/2 pathogenic mutations display different patterns when compared with sporadic BCs. Cancers occurring among
BRCA1 carriers are more frequently classified as medullary [
7], whereas histological subtypes among
BRCA2 carriers tend to be more heterogeneous [
8]. In addition,
BRCA1 carriers are more frequently ER-negative, PR-negative and lack
HER2 amplification (i.e., display a triple negative (TNBCs) phenotype [
9]) whereas in
BRCA2 carriers, a similar prevalence of ER-positive tumors has been described when compared with sporadic controls [
10,
11,
12,
13].
Most of patients with TNBCs receive chemotherapy [
14,
15]. Due to the alteration of
BRCA1 and
BRCA2 proteins in tumor cells,
BRCA-mutated cells are unable to properly repair double-strand breaks, classically induced by DNA-alkylating agents [
16]. Hence,
BRCA deficiency has sometimes been associated with a higher sensitivity to platinum agents when compared to other types of neoadjuvant chemotherapy regimens [
17,
18,
19]. In a recent meta-analysis of platinum-based neoadjuvant chemotherapy in TNBC, the addition of carboplatin was not associated with significantly increased pCR rate in
BRCA-mutated patients (OR = 1.17, CI95% [0.51–2.67],
p = 0.711) [
20]. So far, the benefit of adding a platinum agent in
BRCA-mutated patients receiving standard neoadjuvant chemotherapy remains a matter of debate. Nevertheless, beyond the controversy upon platinum-based agents in
BRCA-deficient tumors, the effectiveness of standard NAC in all BC subtypes associated with
BRCA pathogenic variants compared to controls has been poorly explored so far.
The role of tumor infiltrating lymphocytes (TILs) in BC has been extensively studied over the last decade. High levels of TILs before NAC are associated with higher pCR rates and better survival, especially for TNBC and
HER2-positive BCs [
21,
22]. However, despite a growing interest in the field of immunity and oncology, characterization and quantification of TILs across all BC subtypes according to
BRCA status has not been extensively described. Similarly, no study has evaluated so far, the evolution of immune infiltration after NAC according to
BRCA status.
The objective of the current study is to determine if pre and post-NAC TILs, chemosensitivity and prognosis differ according to BRCA status in a cohort of BC patients treated with NAC.
3. Discussion
In the current study, we did not identify any association between BRCA status and immune infiltration, whatever the type of TILs (IT, str). We found a better response to neoadjuvant chemotherapy in tumors associated with a germline BRCA pathogenic variant when compared to BRCA-WT, however the latter was limited to the restricted group of luminal BCs (BRCA-proficient n = 75; BRCA-deficient, n = 15) and was not statistically significant after multivariate analysis, possibly due to the small sample size of the population. Probably in relation, we recovered higher post-NAC lymphocyte infiltration in BRCA-deficient tumors in the luminal BC subgroup.
Regarding pre-treatment immune infiltration, Sønderstrup and colleagues [
23] analyzed str TIL levels in a nationwide cohort of
BRCA1 and
BRCA2 carriers with primary BCs. They found a greater prevalence of high stromal TILs (defined as TILs-positive tumors with ≥ 60% str TILs) in
BRCA1-deficient tumors (
n = 243) when compared with
BRCA2-deficient tumors (
n = 168) (36% versus 15% respectively,
p < 0.0001). However, no control group with
BRCA-WT tumors was available in this study. In a small study of 85 TNBC patients, Solinas and colleagues [
24] investigated the distribution of TILs subpopulations. The tumors of patients in the
BRCA1 or
BRCA2-mutated group displayed a higher prevalence of TILs-positive tumors (defined as tumors with ≥ 10% str or IT TILs) when compared with the
BRCA-WT (93.2% versus 75.6% respectively,
p = 0.037). No other statistically significant differences were identified between
BRCA-carriers and non-carriers, neither in TILs subpopulations nor their location. More recently, Telli and colleagues [
25] investigated the association between TILs, homologous recombination deficiency (HDR) and
BRCA1/2 status in a cohort of 161 TNBC patients pooled from five phase II neoadjuvant clinical trials of platinum-based therapy. They found that IT TILs and str TILs density were not associated with
BRCA1/2 status (
p = 0.312 and
p = 0.391, respectively). Consistently with Telli et al., we did not observe any difference in baseline immune infiltration according to
BRCA status.
Some retrospective studies suggested that tumors displayed higher chemosensitivity according to
BRCA-mutation status [
17,
18,
19,
26,
27,
28,
29,
30,
31,
32]. Arun et al. [
30] compared pCR rates after NAC between
BRCA1 or
BRCA2-carriers (
n = 57 and
n = 23, respectively) and WT controls (
n = 237). The majority of patients (82%) received an anthracycline-taxane containing regimen as NAC. The authors found that
BRCA1 mutation was an independent positive predictor of pCR (OR = 3.16, 95%CI 1.55–6.42,
p = 0.002). In the largest study so far, Wunderle et al. [
18] investigated efficacy of chemotherapy among a cohort of 355 patients composed with 16.6% (59/355) of
BRCA-carriers. Across all BC subtypes, 64.4% of patients with a
BRCA1/2 pathogenic variant received anthracycline-based treatments, while the rest received carboplatin. pCR was observed in 54.3% (32/59) of all
BRCA1/2 mutation carriers, and in 39.5% (15/34) of the
BRCA-carriers versus 13% of the WT BCs in the anthracycline-regimen (
Table 2). In our cohort, we found similar results after univariate analysis, and we additionally evidenced a nearly significant interaction with BC subtype. In addition, ongoing trials should determine whether PARP inhibitors might improve outcome when administered in the adjuvant or neoadjuvant setting in early luminal breast cancer patients with
BRCA1/2 mutation [
33]. The fact that our results were no longer significant after multivariate analysis is possibly due to a lack of statistical power.
Furthermore, we found that both str and IT TIL levels were higher after NAC completion in the luminal BCs. Whether this difference in post treatment TILs is a cause, a consequence, or unrelated to response to chemotherapy remains unknown. Indeed, post-NAC TIL levels have been shown to be strongly related to response to chemotherapy in BC cohorts including all BC subtypes [
34,
35,
36,
37] and response to checkpoint inhibitors (IC) in early TNBC [
37]. Moreover, Anurag et al. [
38] identified upregulation of the targetable immune-checkpoint components (IDO1, LAG3 and PD1) in AI-resistant luminal B tumors suggesting that luminal BC could also be immunologically “hot”. Besides, only a few studies have investigated the dynamic of TIL levels in response to NAC. Hamy et al. [
36] noticed that mean TIL levels decreased after chemotherapy completion across all the BC subtype (pre-NAC TILs: 24.1% vs. post-NAC TILs: 13.0%,
p < 0.001).
Table 2.
Literature Review.
Table 2.
Literature Review.
Study | Setting/Design | Control Group | Number of Patients (n) | TNBC (n) | HER2-Positive (n) | Luminal (n) | BRCA1 | BRCA2 | BRCA 1 and 2 | Chemotherapy Regimen | sTILS Evaluation | pCR in BRCA-Carriers vs. Non-Carriers | Survival Analyses | Comments |
---|
Byrski (2014) [26] {BCRT | Neoadjuvant epidemiologic prospective cohort | No | 10 | 10 | 0 | 10 | 0 | 0 | 0 | Cis | No | 90% | No | 90% (9/10) in BRCA1-mutated BC patients achieved a pCR after NAC with cisplatin chemotherapy |
Byrski (2015) [27] HCCP | Neoadjuvant epidemiologic prospective cohort | No | 107 | 82 | 2 | NA | 107 | 0 | 0 | Cis | No | 61% | No | 61% (65/107) in BRCA1-mutated BC patients achieved pCR after NAC with cisplatin chemotherapy. In this study of BRCA1-mutation carriers, a pCR was also achieved in 56% of 16 patients with ER-positive BC. No survival analysis were provided in the current study. |
Hanhnen (2017) [28] JAMA Oncology | Neoadjuvant secondary analysis of the GeparSixto randomized clinical trial | Yes | 291 | 291 | 0 | 0 | 50 | 0 | P + Dox + Bev ± Cb | No | 66.7% vs. 36.4% | Yes | Patients with BRCA-mutation did not derive a pCR benefit from the addition of carboplatine (65.4% vs. 66.7%) compared to non-BRCA carriers (55% vs. 36.4%). No significant difference in overall prognosis observed in the BRCA-mutated subgroup. |
Sharma (2017) [39] CCR | Neoadjuvant prospective, multicenter, non-randomized trial | Yes | 190 | 190 | 0 | 0 | 30 | 0 | Cb + D | No | 59% vs. 56% | No | No significative difference in pCR between BRCA-carriers and WT TNBC (59% and 56%, respectively (p = 0.83)). The Cb-D regimen was well tolerated and yielded high pCR rates in both BRCA associated and WT TNBC. These results are comparable to pCR of previous studies (who investigated pCR after NAC with addition of Cb to AT regimen in TNBC cohort). |
Poggio (2018) [20] Annals of Oncology | Neoadjuvant meta-analysis of nine randomized controlled trials | No | 96 | 96 | 0 | 0 | 96 | 0 | P + Dox + Bev ± Cb P + AC ± Cb | No | 54.3% | No | Among 96 BRCA-mutated patients included in 2 controlled trials, the addition of carboplatin was not associated with increased pCR rate (OR 1.17, 95% CI 0.51–2.67, p = 0.711). No survival analyses were available according to BRCA status. |
Telli (2019) [25] CCR | Five randomized controlled trials | Yes | 161 | 161 | 0 | 0 | 34 | 0 | Cb + Gem + Iniparib; Cis; Cis + Bev; Cb + Eribulin; Cb + nab-P ± Vorinostat | Yes | No | No | pCR was achieved in 51 (31.7%) patients. In patients with TNBC treated with neoadjuvant platinum-based therapy, iTIL and sTIL densities were not significantly associated with BRCA1/2-mutated tumor status (p = 0.312 and p = 0.391). In multivariate analyses, sTIL density (OR 1.23, 95% CI 0.94–1.61, p = 0.139) was not associated with pCR, but was associated with RCB 0/I status (OR 1.62, 95% CI 1.20–2.28, p = 0.001). |
Sønderstrup (2019) [23] Acta Oncologica | Epidemiologic prospective mulitcentric cohort (nationwide) | No | 411 | NA | 24 | NA | 243 | 168 | 0 | NA | Yes | No | Yes | High sTILs (defined as TILs > 60%) were observed in 36% in BRCA1- and 15% in BRCA2-mutated tumors (p < 0.0001). Significant association with survival (OS and DFS) was observed in BRCA1 subgroup. sTILs are an important prognostic factor in BRCA BC and increasing sTILs is associated with a better prognosis. |
Byrski (2009) [17] JCO | Neoadjuvant Epidemiologic epidemiologic retrospective cohort | No | 102 | NA | 6 | NA | 102 | 0 | 0 | CMF; AT; AC FAC or Cis | No | 23.5% | No | pCR was achieved in 23.5% of 102 patients with a BRCA1 mutation who received NAC. Especially, a complete pCR was observed in 8% (2/25) with AT- regimen (standard of care) compared to 83% (10/12) with cisplatin. |
Chappuis (2002) [29] JMG | Neoadjuvant Retrospective retrospective multicentric clinical trial | Yes | 38 | NA | NA | NA | 7 | 4 | 0 | FAC; AC; CEF AC + CMF AC + D | No | 44% vs. 4% | No | pCR was achieved in 44% (4/11) of the BRCA-carriers and 4%(1/27) of the non-carriers (p = 0.009). No survival analysis were experienced in this study. |
Arun (2011) [30] JCO | Neoadjuvant Epidemiologic epidemiologic retrospective cohort | Yes | 317 | 77 | 60 | NA | 57 | 23 | 0 | A-single agent; AT or T-single-agent | No | 46% vs. 22% | Yes | pCR was achieved in 46% of BRCA1-carriers and 13% of BRCA2-carriers and 22% of BRCA non-carriers (<0.001). In the multivariate logistic model, BRCA1 status (OR = 1.96, p = 0.03) remained as independant significant predictors of a pCR. No significant difference in overall prognosis. |
Wang (2014) [40] Annals of Oncology | Neoadjuvant Epidemiologic retrospective cohort | Yes | 652 | 652 | 0 | 0 | 52 | NA | 0 | A-single agent; AT or T-single-agent | No | 53.8% vs. 29.7% | Yes | The pCR rate was 31.6% in the 652 patients who received NAC. BRCA1 carriers had a significantly higher pCR rate than non-carriers (BRCA1 carriers versus non-carriers, 53.8% versus 29.7%, p < 0.001). Among women treated with anthracycline with or without taxane regimens, the pCR rate was 57.1% for BRCA1 carriers, 29.0% for non-carriers (p < 0.001). The RFS was similar according to BRCA status. |
Paluch-Shimon(2016) [31] BCRT | Neoadjuvant epidemiologic retrospective cohort | Yes | 80 | 80 | 0 | 0 | 34 | 0 | 0 | AT | No | 68% vs. 37% | Yes | The BRCA1-carriers had pCR rate of 68% compared with 37% among non-carriers, p = 0.01. Yet this did not translate into superior survival for BRCA1 carriers compared with non-carriers. |
Bignon (2017) [41] Breast | Neoadjuvant epidemiologic retrospective cohort | No | 53 | 53 | 0 | 0 | 46 | 6 | 1 | A-single agent or AT | No | 66% | Yes | The pCR rate was 38.3% [95% CI, 26%–55%] among BRCA1 mutation carriers, and 66% among the 6 BRCA2 mutation carriers. 15 relapses and 6 s cancers were recorded during the follow-up period. 11 deaths occurred, all of which were in the non-pCR group. DFS (p < 0.01) and OS (p < 0.01) were significantly better in the pCR group than the non-pCR group. |
Wunderle (2018) [18] BCRT | Neoadjuvant Epidemiologic retrospective cohort | Yes | 355 | 138 | 58 | 159 | 43 | 16 | 0 | AT; Cb | No | 54.3% vs. 12.6% | Yes | pCR was observed in 54.3% of BRCA1/2 mutation carriers, but only in 12.6% of non-carriers. The adjusted odds ratio was 2.48 (95% CI 1.26–4.91) for BRCA1/2 carriers versus non-carriers. No difference in overall survival was observed. |
Saether (2018) [32] HCCP | Neoadjuvant Epidemiologic retrospective cohort | No | 12 | NA | NA | NA | 12 | 0 | 0 | Cis + Dox or Cb + D | No | 83% | No | 11 patients received a combination of cisplatin and doxorubicin, and 1 patient received carboplatin and docetaxel. 83% (10/12) of the BRCA1-carriers achieved pCR. This results were comparable to existing results found in similar studies. No information about BC subtype among the study population and the toxicity of the chemotherapy was not evaluated. |
Sella (2018) [19] Breast | Neoadjuvant Epidemiologic retrospective cohort | Yes | 43 | 43 | 0 | 0 | 14 | 0 | 0 | AT ± Cb | No | 67% vs. 38% | No | pCR was achieved in 38% in BRCA WT compared to 67% in BRCA-associated TNBC (p = 0.232). No benefit from the addition of carboplatine in BRCA-carriers (64.3% vs. 67%) compared to non-BRCA carriers (44.8% vs. 38%) when compared to historic institutional rates with AT. |
Solinas (2019) [24] Cancer Letters | Epidemiologic retrospective cohort | Yes | 85 | 85 | 0 | 0 | 38 | 6 | 0 | NA | Yes | No | Yes | The BRCA-mutated tumors had a significantly higher incidence of TIL-positive levels compared to WT (44% and 41%, respectively p = 0.037). No significant difference between BRCA-mutated and WT groups neither in TIL subpopulation nor their location. No difference in I-DFS and OS after stratification on TIL infiltration levels. |
Our study (2020) | Epidemiologic retrospective cohort | Yes | 267 | 110 | 67 | 90 | 31 | 14 | 1 | A-single agent; AT or T-single-agent | Yes | 45.7% vs 28% | Yes | Among the whole population, 84 tumors achieved a pCR (31.5%). After stratification by BC subtype, pCR rates were significantly higher in luminal BRCA-mutated BCs when compared with WT tumors (33.3% vs. 5.4%, p = 0.006).Pre and post-NAC str or IT TILs were not significantly different between BRCA-carriers and non-carriers in whole population. In the luminal BC, both str and IT post-NAC TIL levels were significantly higher in BRCA-mutated tumors when compared with WT tumors but was no longer significant after multivariate analysis. No difference in RFS or OS between BRCA-mutated and BRCA-WT patients. |
This decrease was strongly associated with high pCR rates, and the variation of TIL levels was strongly inversely correlated with pre-NAC TIL levels (and the variation of TIL levels was strongly inversely correlated with pre-NAC TIL levels (r = −0.80, p < 0.001).
Finally, in line with several recently published clinical studies [
42,
43,
44], we found that survival outcomes were not different between
BRCA-carriers and non-carriers. A multivariate study, including 223 BC patients carrying
BRCA pathogenic variants and 446 controls with sporadic BC matched for age and year of diagnosis, showed no difference in terms of specific BC survival between
BRCA1 or
BRCA2 mutation carriers and controls [
45]. Templeton et al. evaluated a total of 16 studies comprising data from 10,180 patients and concluded that
BRCA pathogenic mutations were not associated with a worse overall survival [
46]. The difference between the pCR rate and survival analysis could be due to several factors. First,
BRCA mutation carriers are commonly offered additional treatment, including a bilateral mastectomy. Second, the increase of TILs in the surgical piece might reveals a higher immunogenic tumor, which may involve a more sustained response to treatment over time. Besides, carriers of a
BRCA pathogenic variant were more likely to be diagnosed with TNBC. Copson et al. [
44] have shown that
BRCA mutation carriers with triple-negative breast cancer might have a survival advantage during the first few years after diagnosis compared with non-carriers. This benefit might reflect greater sensitivity of
BRCA-mutant breast cancers to chemotherapy or the greater visibility to host immune attack.
Limits of our study include its retrospective observational design as well as small effectives potentially leading to a lack of statistical power. Therefore, our results might be submitted to evaluation biases, especially for the time-to-event analysis. Indeed, we present a study of patients with two rare conditions. First, according to French national guidelines, neoadjuvant chemotherapy is currently prescribed only in 15% of the patients with locally advanced breast cancers. Second, screening of inherited
BRCA mutation is performed in a highly selected population representing nearly one quarter of breast cancer [
47]. Our study design does not allow us to draw firm conclusions and future studies are warranted to confirm the hypotheses generated. Moreover, the incidence of bi-allelic pathogenic alterations in HR-related genes according to somatic origin is well-known and ranches from 1 to 2% [
48] but we did not explore somatic mutational status in the tumor tissues in the current study. The study also has several strengths, for instance from being the largest cohort with a
BRCA-WT control group, and analyses performed after stratification by BC subtype. Finally, to our knowledge, we provide data on post-NAC immune infiltration according to
BRCA status for the first time.