Clinicopathological features of BC patients
The clinicopathological features of patients (n = 356) treated with POC have been summarized in Table 1. The patients were operated at median age of 55 years (range, 24–78 years) and the median tumor diameter 28.7 mm (range, 9.2–119.8 mm). Skin infiltration was observed in 58 patients (16.3%). Further, imaging methods of diagnosis did not indicate lymph node metastasis in 121 patients (34.0 %). The number of ER-negative, PgR-negative, and HER2-positive patients was 187 (52.5 %), 242 (68.0 %), and 125 (35.1 %), respectively. Moreover, Ki67-high (above 14%) was observed in 239 patients (67.1 %). Based on these results, the BC subtypes were classified as follows– HR + HER2-: 126 patients (35.4 %), HR + HER2+: 47 patients (13.2 %), HER2-enriched: 78 patients (21.9 %), and TNBC: 105 patients (29.5 %). Furthermore, the Responders for ORR reached 88.8%, the rate of pCR post-operative pathology was 33.1%, and 161 patients (45.2%) showed high TILs density.
Table 1
Clinicopathological features of 356 patients who were treated with preoperative chemotherapy
Parameters
|
All patients (n = 356) (%)
|
Younger (n = 75) (%)
|
Elderly (n = 116) (%)
|
Age (years old)
|
55 (24–78)
|
41 (24–45)
|
67 (61–78)
|
Tumor size (mm)
|
28.7 (9.2–119.8)
|
29.5 (9.9–82.6)
|
27.3 (9.2–89.8)
|
Skin infiltration
Negative / Positive
|
298 (83.7%) / 58 (16.3%)
|
68 (90.7%) / 7 (9.3%)
|
90 (77.6%) / 26 (22.4%)
|
Lymph node metastasis
N0 / N1 / N2 / N3
|
121 (33.9%) / 133 (37.4%)
/ 68 (19.1%) / 34(9.6%)
|
28 (37.3%) / 28 (37.3%)
/ 14 (18.7%) / 5 (6.7%)
|
44 (37.9%) / 36 (31.0%)
/ 22 (19.0%) / 14(12.1%)
|
Estrogen receptor
Negative / Positive
|
187 (52.5%) / 169 (47.5%)
|
37 (49.3%) / 38 (50.7%)
|
67 (57.8%) / 49 (42.2%)
|
Progesterone receptor
Negative / Positive
|
242 (68.0%) / 114 (32.0%)
|
42 (56.0%) / 33 (44.0%)
|
89 (76.7%) / 27 (23.3%)
|
HER2
Negative / Positive
|
231 (64.9%) / 125 (35.1%)
|
47 (62.7%) / 28 (37.3%)
|
69 (59.5%) / 47 (40.5%)
|
Ki67
≤ 14 % / >14 %
|
117 (32.9%) / 239 (67.1%)
|
22 (29.3%) / 53 (70.7%)
|
40 (34.5%) / 76 (65.5%)
|
Intrinsic subtype
HR + HER2-BC / HR + HER2 + BC /
HER2BC / TNBC
|
126 (35.4%) / 47 (13.2%)
/ 78 (21.9%) / 105 (29.5%)
|
24 (32.0%) / 16 (21.3%)
/ 12 (16.0%) / 23 (30.7%)
|
39 (33.6%) / 11 (9.5%)
/ 36 (31.0%) / 30 (25.9%)
|
Objective response rate
Non-Responders / Responders
|
40 (11.2%) / 316 (88.8%)
|
5 (6.7%) / 70 (93.3%)
|
17 (14.7%) / 99 (85.3%)
|
Pathological response
Non-pCR / pCR
|
238 (66.9%) / 118 (33.1%)
|
46 (61.3%) / 29 (38.7%)
|
78 (67.2%) / 38 (32.8%)
|
TILs
Low / High
|
195 (54.5%) / 161 (45.2%)
|
31 (41.3%) / 44 (58.7%)
|
65 (56.0%) / 51 (44.0%)
|
HER: human epidermal growth factor receptor. CR: complete response. TILs: tumor- infiltrating lymphocytes. |
Further, while most of the clinicopathological factors were not significantly different, the rate of skin infiltration and PgR-negative status were significantly higher in the older that in the younger patients (P = 0.002 and P = 0.003, respectively) (Table 2). Moreover, the ORR, although statistically insignificant, was found to be higher in the younger than in the older patients (P = 0.091).
Table 2
Difference in clinicopathological features due to TILs in younger and elderly patients
Parameters
|
tumor- infiltrating lymphocytes (n = 191)
|
Low (n = 96)
|
High (n = 95)
|
p value
|
Age (years old)
≤ 45
> 60
|
31 (32.3%)
65 (67.7%)
|
44 (46.3%)
51 (53.7%)
|
0.047
|
Tumor size (mm)
≤ 20.0
> 20.0
|
20 (20.8%)
76 (79.2%)
|
14 (14.7%)
81 (85.3%)
|
0.271
|
Skin infiltration
Negative
Positive
|
71 (74.0%)
25 (26.0%)
|
87 (91.6%)
8 (8.4%)
|
0.001
|
Lymph node status
Negative
Positive
|
33 (34.4%)
63 (65.6%)
|
39 (41.1%)
56 (58.9%)
|
0.341
|
Estrogen receptor
Negative
Positive
|
37 (38.5%)
59 (61.5%)
|
67 (70.5%)
28 (29.5%)
|
< 0.001
|
Progesterone receptor
Negative
Positive
|
55 (57.3%)
41 (42.7%)
|
76 (80.0%)
19 (20.0%)
|
0.001
|
Hormone receptor
Negative
Positive
|
35 (36.5%)
61 (63.5%)
|
66 (69.5%)
29 (30.5%)
|
< 0.001
|
HER2
Negative
Positive
|
69 (71.9%)
27 (28.1%)
|
47 (49.5%)
48 (50.5%)
|
0.002
|
Ki67
≤14 %
>14 %
|
37 (38.5%)
59 (61.5%)
|
25 (26.3%)
70 (73.7%)
|
0.071
|
ORR
Non-Responders
Responders
|
18 (18.8%)
78 (81.2%)
|
4 (4.2%)
91 (95.8%)
|
0.002
|
Pathological response
Non-pCR
pCR
|
79 (82.3%)
17 (17.7%)
|
45 (47.4%)
50 (52.6%)
|
< 0.001
|
TILs: tumor- infiltrating lymphocytes. HER: human epidermal growth factor receptor. ORR: objective response rate. CR: complete response. |
Correlation of TILs density with clinicopathological features and prognosis of patients
First, the 356 patients were divided into high and low TILs density groups, and their correlation with clinicopathological factors was examined (Supplementary Table S1). Following characteristics were observed in the low TILs than the high TILs group: ≥ 45 years (P = 0.008), skin invasion (P = 0.001), ER-positive (P < 0.001), PgR-positive (P < 0.001), HER2-negative (P = 0.011), Ki67-high (P < 0.001), low ORR (P = 0.001), and low pCR rate (P < 0.001).
Further, the high TILs density group showed significantly better DFS than the low TILs density group in HER2-enriched (P = 0.012, log-rank) and TNBC (P = 0.002, log-rank) categories (Supplementary Fig. S2). Therefore, DFS was better in the high TILs density group despite no significant difference in HR + BC (P = 0.011, log-rank). However, the high TILs density group had better OS, although not statistically significant, than the low TILs density group in TNBC category (P = 0.057, log-rank), but there was no significant difference between the difference of TILs density (Supplementary Fig. S3). Further, in the univariate analysis for DFS, high TILs density group associated with significantly better DFS (P = 0.010, HR = 0.512) (Supplementary Table S2). However, in the multivariate analysis for DFS, TILs density was not an independent factor (P = 0.227, HR = 0.699), since skin invasion (P = 0.012, HR = 2.180), lymph node metastasis (P = 0.001, HR = 2.918), HER2-positive (P = 0.020, HR = 0.498), Responders in ORR (P < 0.001, HR = 0.247), and pCR (P < 0.001, HR = 0.315) influenced the DFS. Additionally, difference in OS due to TILs was insignificant even in the univariate analysis (P = 0.214, HR = 0.660) (Supplementary Table S3).
Further, the patients were classified based on age as < 45 years, 46–60 years, and ≥ 61 years, and the distribution of TILs density was analyzed using t-test (Fig. 1). Our analysis did not indicate significant difference in HR + BC for any of the age groups. However, in HER2-enriched BC, the patients aged < 45 years had significantly higher TILs density than patients in other age groups (vs. 46–60 years: P = 0.002, and vs. ≥ 61 years: P = 0.018). Furthermore, in the TNBC category, the patients aged ≥ 61 years had significantly higher TILs density than patients in other age groups (vs. ≤ 40 years: P = 0.035, and vs. 46–60 years: P = 0.047).
Examination of clinicopathological factors and prognosis in the younger and older BC patients
First, we studied the correlation between TILs density and clinicopathological factors in the younger and older patients (Table 2). Although patients aged 46–60 years were excluded from the analysis, the characteristics of the high TILs density group were similar to those for all patients: > 60 years (P = 0.047), skin infiltration (P = 0.001), ER-positive (P < 0.001), PgR-positive (P = 0.001), HER2-negative (P = 0.002), lower ORR (P = 0.002), and lower pCR rate (P < 0.001).
Further, younger patients showed significantly higher pCR rates than older patients in the HR + HER2- and HER2-enriched BC category (P = 0.021 and P = 0.048, respectively) (Table 3). Moreover, in HR + HER2 + BC, the responder rate for ORR was significantly higher in the younger patients than in older patients (P = 0.009). However, no significant difference was observed in the effect of POC on TNBC.
Table 3
Difference in clinicopathological features due to age
Parameters
|
All intrinsic subtype (n = 191)
|
HR + HER2-BC (n = 61)
|
HR + HER2 + BC (n = 27)
|
HER2BC (n = 48)
|
TNBC (n = 53)
|
Young
(n = 75)
|
Elderly
(n = 116)
|
p value
|
Young
(n = 24)
|
Elderly
(n = 39)
|
p value
|
Young
(n = 16)
|
Elderly
(n = 11)
|
p value
|
Young
(n = 12)
|
Elderly
(n = 36)
|
p value
|
Young
(n = 23)
|
Elderly
(n = 30)
|
p value
|
Tumor size (mm)
≤ 20.0
> 20.0
|
10 (13.3%)
65 (86.7%)
|
24 (20.7%)
92 (79.3%)
|
0.194
|
2 (8.3%)
22 (91.7%)
|
7 (17.9%)
32 (82.1%)
|
0.290
|
3 (18.8%)
13 (81.2%)
|
2 (18.2%)
9 (81.8%)
|
0.970
|
2 (16.7%)
10 (83.3%)
|
7 (19.4%)
29 (80.6%)
|
0.831
|
3 (13.0%)
20 (87.0%)
|
8 (26.7%)
22 (73.3%)
|
0.225
|
Skin infiltration
Negative
Positive
|
68 (90.7%)
7 (9.3%)
|
90 (77.6%)
26 (22.4%)
|
0.020
|
20 (83.3%)
4 (16.7%)
|
29 (74.4%)
10 (25.6%)
|
0.405
|
14 (87.5%)
2 (12.5%)
|
6 (54.5%)
5 (45.5%)
|
0.055
|
12 (100.0%)
0 (0.0%)
|
29 (80.6%)
7 (19.4%)
|
0.098
|
22 (95.7%)
1 (4.3%)
|
26 (86.7%)
4 (13.3%)
|
0.267
|
Lymph node status
Negative
Positive
|
28 (37.3%)
47 (62.7%)
|
44 (37.9%)
72 (62.1%)
|
0.934
|
8 (33.3%)
16 (66.7%)
|
12 (30.8%)
27 (69.2%)
|
0.832
|
9 (56.2%)
7 (43.8%)
|
2 (18.2%)
9 (81.8%)
|
0.048
|
4 (33.3%)
8 (66.7%)
|
17 (47.2%)
19 (52.8/%)
|
0.401
|
7 (30.4%)
16 (69.6%)
|
13 (43.3%)
17 (56.7%)
|
0.337
|
Estrogen receptor
Negative
Positive
|
37 (49.3%)
38 (50.7%)
|
67 (57.8%)
49 (42.2%)
|
0.254
|
2 (8.3%)
22 (91.7%)
|
0 (0.0%)
39 (100.0%)
|
0.067
|
0 (0.0%)
16 (100.0%)
|
1 (9.1%)
10 (90.9%)
|
0.219
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
Progesterone receptor
Negative
Positive
|
42 (56.0%)
33 (44.0%)
|
89 (76.7%)
27 (23.3%)
|
0.003
|
5 (20.8%)
19 (79.2%)
|
16 (41.0%)
23 (59.0%)
|
0.099
|
2 (12.5%)
14 (87.5%)
|
7 (63.6%)
4 (36.4%)
|
0.006
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
Hormone receptor
Negative
Positive
|
35 (46.7%)
40 (53.3%)
|
66 (56.9%)
50 (43.1%)
|
0.167
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
HER2
Negative
Positive
|
47 (62.7%)
28 (37.3%)
|
69 (59.5%)
47 (40.5%)
|
0.660
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
-
-
|
-
-
|
|
Ki67
≤14 %
>14 %
|
22 (29.3%)
53 (70.7%)
|
40 (34.5%)
76 (65.5%)
|
0.458
|
12 (50.0%)
12 (50.0%)
|
21 (53.8%)
18 (46.2%)
|
0.767
|
7 (43.8%)
9 (56.2%)
|
2 (18.2%)
9 (81.8%)
|
0.166
|
1 (8.3%)
11 (91.7%)
|
12 (33.3%)
24 (66.7%)
|
0.091
|
2 (8.7%)
21 (91.3%)
|
5 (16.7%)
25 (83.3%)
|
0.396
|
ORR
Non-Responders
Responders
|
5 (6.7%)
70 (93.3%)
|
17 (14.8%)
99 (85.2%)
|
0.091
|
2 (8.3%)
22 (91.7%)
|
8 (20.5%)
31 (79.5%)
|
0.199
|
0 (0.0%)
16 (100.0%)
|
4 (36.4%)
7 (63.6%)
|
0.009
|
0 (0.0%)
12 (100.0%)
|
1 (2.8%)
35 (97.2%)
|
0.560
|
3 (13.0%)
20 (87.0%)
|
4 (13.3%)
26 (86.7%)
|
0.975
|
Pathological response
Non-pCR
pCR
|
46 (61.3%)
29 (38.7%)
|
78 (67.2%)
38 (32.8%)
|
0.403
|
18 (75.0%)
6 (25.0%)
|
37 (94.9%)
2 (5.1%)
|
0.021
|
13 (81.2%)
3 (18.8%)
|
10 (90.9%)
1 (9.1%)
|
0.488
|
1 (8.3%)
11 (91.7%)
|
14 (38.9%)
22 (61.1%)
|
0.048
|
14 (60.9%)
9 (39.1%)
|
17 (56.7%)
13 (43.3%)
|
0.758
|
TILs
Low
High
|
31 (41.3%)
44 (58.7%)
|
65 (56.0%)
51 (44.0%)
|
0.047
|
14 (58.3%)
10 (41.7%)
|
31 (79.5%)
8 (20.5%)
|
0.071
|
7 (43.8%)
9 (56.2%)
|
9 (81.8%)
2 (18.2%)
|
0.048
|
1 (8.3%)
11 (91.7%)
|
10 (27.8%)
26 (72.2%)
|
0.165
|
9 (39.1%)
14 (60.9%)
|
15 (50.0%)
15 (50.0%)
|
0.431
|
HER: human epidermal growth factor receptor. ORR: objective response rate. CR: complete response. TILs: tumor- infiltrating lymphocytes. |
Next, when DFS was compared between the younger and older patients, no significant difference was found overall or in any subtype (Fig. 2). Moreover, our analysis indicated that age or TILs was not a predictor of DFS in the univariate analysis (P = 0.619 and P = 0.066, respectively) (Table 4). Although upon comparison of OS, a significant difference was observed between younger and older patients with TNBC (P = 0.039, log-rank) (Fig. 3), the results were contrasting and suggested better OS in older patients than in younger patients. Additionally, in univariate analysis with OS, no significant difference in age and TILs density was observed (P = 0.346 and P = 0.216, respectively) (Table 5).
Table 4
Univariate and multivariate analysis with respect to DFS in younger and elderly patients
|
Univarite analysis
|
Multivariate analysis
|
Parameters
|
Hazard ratio
|
95% CI
|
p value
|
Hazard ratio
|
95% CI
|
p value
|
Age at opetation (yr)
≤ 45 vs > 60
|
0.916
|
0.651-1.300
|
0.619
|
|
|
|
Tumor size (mm)
≤ 20 vs > 20
|
0.674
|
0.309–1.684
|
0.373
|
|
|
|
Skin infiltration
Negative vs Positive
|
2.629
|
1.140–5.582
|
0.025
|
2.597
|
1.075–5.858
|
0.035
|
Lymph node status
Negative vs Positive
|
4.935
|
1.756–20.600
|
0.001
|
3.981
|
1.385–16.828
|
0.008
|
Estrogen receptor
Negative vs Positive
|
0.738
|
0.358–1.469
|
0.390
|
|
|
|
Progesterone receptor
Negative vs Positive
|
0.733
|
0.322–1.524
|
0.418
|
|
|
|
Hormone receptor
Negative vs Positive
|
0.675
|
0.327–1.344
|
0.265
|
|
|
|
HER2
Negative vs Positive
|
0.237
|
0.070–0.602
|
0.001
|
0.479
|
0.130–1.423
|
0.193
|
Intrinsic subtype
Not TNBC vs TNBC
|
2.710
|
1.356–5.392
|
0.005
|
2.418
|
1.080–5.456
|
0.032
|
Ki67
≤14 % vs > 14 %
|
2.339
|
1.066–5.872
|
0.033
|
2.489
|
1.089–6.417
|
0.030
|
Objective response rate
Non-Responders vs Responders
|
0.309
|
0.145–0.734
|
0.010
|
0.381
|
0.159–0.984
|
0.047
|
Pathological response
Non-pCR vs pCR
|
0.195
|
0.058–0.499
|
< 0.001
|
0.238
|
0.065–0.685
|
0.006
|
TILs
Low vs High
|
0.523
|
0.253–1.045
|
0.066
|
0.991
|
0.431–2.231
|
0.982
|
DFS: Disease-free survival. CI: confidence intervals. HER: human epidermal growth factor receptor. pCR: pathological complete response. TILs: tumor- infiltrating lymphocytes. |
Table 5
Univariate and multivariate analysis with respect to OS in younger and elderly patients
|
Univarite analysis
|
Multivariate analysis
|
Parameters
|
Hazard ratio
|
95% CI
|
p value
|
Hazard ratio
|
95% CI
|
p value
|
Age at opetation (yr)
≤ 45 vs > 60
|
0.813
|
0.524–1.255
|
0.346
|
|
|
|
Tumor size (mm)
≤ 20 vs > 20
|
1.188
|
0.402–5.074
|
0.778
|
|
|
|
Skin infiltration
Negative vs Positive
|
5.034
|
1.940-12.433
|
0.002
|
6.899
|
2.467–18.908
|
< 0.001
|
Lymph node status
Negative vs Positive
|
4.239
|
1.227–26.631
|
0.019
|
2.999
|
0.815–19.389
|
0.106
|
Estrogen receptor
Negative vs Positive
|
0.474
|
0.169–1.167
|
0.107
|
|
|
|
Progesterone receptor
Negative vs Positive
|
0.475
|
0.137–1.285
|
0.151
|
|
|
|
Hormone receptor
Negative vs Positive
|
0.441
|
0.157–1.085
|
0.076
|
|
|
|
HER2
Negative vs Positive
|
0.283
|
0.066–0.844
|
0.021
|
0.721
|
0.149–2.809
|
0.645
|
Intrinsic subtype
Not TNBC vs TNBC
|
3.966
|
1.640–10.130
|
0.002
|
3.703
|
1.323–11.575
|
0.012
|
Ki67
≤14 % vs > 14 %
|
2.730
|
1.004–9.518
|
0.049
|
2.271
|
0.768–8.314
|
0.144
|
Objective response rate
Non-Responders vs Responders
|
0.244
|
0.097–0.692
|
0.010
|
0.259
|
0.090–0.797
|
0.020
|
Pathological response
Non-pCR vs pCR
|
0.241
|
0.056–0.718
|
0.009
|
0.384
|
0.082–1.332
|
0.137
|
TILs
Low vs High
|
0.578
|
0.232–1.380
|
0.216
|
|
|
|
OS: Overall survival. CI: confidence intervals. HER: human epidermal growth factor receptor. pCR: pathological complete response. TILs: tumor- infiltrating lymphocytes. |