Reproductive and hormonal risk factors for postmenopausal luminal, HER-2-overexpressing, and triple-negative breast cancer

Abstract

BACKGROUND.

Molecular profiling studies have identified subtypes of breast cancer that can be approximately classified by estrogen receptor (ER), progesterone receptor (PR), and HER-2/neu (HER-2) expression. These molecular subtypes are prognostically significant, but to the authors' knowledge, differences in their etiologic profiles have not been established. Reproductive factors may plausibly be differentially correlated with the risk of different breast cancer subtypes because these factors are presumed to impact exposure to endogenous sex hormones.

METHODS.

The authors pooled 2 population-based, case–control studies of breast cancer in women ages 55 to 79 years for an analysis including 1476 controls and 1023 cases of luminal breast cancer, 39 cases of HER-2-overexpressing breast cancer, and 78 cases of triple-negative breast cancer. Polytomous logistic regression was used to compare each case group with controls.

RESULTS.

Associations varied by molecular subtype. Early age at menarche was only found to be associated with risk of HER-2-overexpressing disease (odds ratio [OR] of 2.7; 95% confidence interval [95% CI], 1.4-5.5), whereas breastfeeding for ≥6 months was only found to be protective for luminal and triple-negative disease (OR of 0.8 [95% CI, 0.6-1.0] and OR of 0.5 [95% CI, 0.3-0.9], respectively). Both late age at menopause and the use of estrogen plus progestin hormone therapy were only found to be associated with risk of luminal disease (OR of 1.6 [95% CI, 1.1-2.2] and OR of 1.7 [95% CI, 1.3-2.1], respectively). No differences in risks associated with parity or age at first live birth were observed by subtype.

CONCLUSIONS.

Certain reproductive factors may have a greater impact on the risk of certain molecular subtypes of disease compared with others. Future studies that further define the etiology of breast cancer subtypes will add to the biologic understanding of this disease. Cancer 2008. © 2008 American Cancer Society.

Gene expression studies have identified and validated the existence of 5 “intrinsic” breast cancer subtypes: luminal A, luminal B, HER–2-overexpressing, basal-like, and unclassified.1–3 Luminal A and B tumors are characterized by expression of estrogen receptor–α (ER), typically accompanied by expression of the progesterone receptor (PR). Nonluminal subtypes are characterized by a lack of hormone receptor expression and can be distinguished by the expression of protein tyrosine kinase HER-2/neu (HER-2). HER-2 is up-regulated in HER-2-overexpressing tumors whereas basal-like and unclassified tumors both have the so-called ‘triple-negative’ phenotype (ie, ER-negative [-]/PR-/HER-2-).4, 5 Clinical differences between these breast cancer subtypes have been well-described in the literature.2, 4, 6 Compared with luminal tumors, both HER-2-overexpressing and triple-negative tumors have a poorer prognosis. However, to our knowledge, differences in the epi-demiology of these subtypes are not well-defined. The existing literature suggests that basal-like tumors are more common among African-American women,4, 7 premenopausal women,4, 8 and women with BRCA1 mutations.9, 10 Further assessment of risk factors stratified by tumor subtype is important to identify disease-specific associations that might be masked by grouping together luminal, HER-2-overexpressing, and triple-negative breast cancers.

The focus of the current study was the correlation between reproductive characteristics and risks of luminal, HER-2-overexpressing, and triple-negative breast cancer. Age at menarche and menopause, parity, age at first live birth, and breastfeeding are all established breast cancer risk factors that influence risk primarily through hormonal mechanisms.11 It has been suggested that factors such as these, which are related to endogenous hormone exposure, are more strongly associated with the risk of ER–positive (ER+) (luminal) disease, whereas risk factors for ER- (nonluminal) disease are more likely to involve nonhormonal mechanisms.12–14 A meta-analysis of epidemiologic studies of breast cancer with ER and PR tumor marker data found that parity and age at first birth were associated with risk of hormone receptor-positive but not hormone receptor-negative disease.15 However, to our knowledge, data regarding differences in the associations between these factors and the risk of different molecularly defined breast cancer subtypes are quite limited. Herein, we examined differences in the associations between reproductive and hormonal characteristics and the risk of breast cancer by disease subtype, as defined by joint ER/PR/HER-2 status.

MATERIALS AND METHODS

Given the rarity of HER-2-overexpressing and triple-negative tumors, cases and controls were pooled from 2 population-based, case–control studies of breast cancer. Both studies were approved by the Fred Hutchinson Cancer Research Center (FHCRC) institutional review board, and written informed consent was obtained from all study participants. Both these studies have been described in detail elsewhere.16, 17 Briefly, cases for both studies were identified through the Cancer Surveillance System (CSS) of western Washington State as women with incident invasive breast cancer and no prior history of in situ or invasive breast cancer who resided in King, Pierce, or Snohomish counties at the time of diagnosis. For the earlier of these 2 studies,16 eligible cases were diagnosed between April 1, 1997 and May 31, 1999, were ages 65 through 79 years at the time of diagnosis, and had a Health Care Financing Administration (HCFA) record. Potential controls without a prior breast cancer were identified through HCFA records and frequency matched to cases based on age. The study base for the more recently completed study was identical to that of the earlier study with respect to geography and case ascertainment protocols, although cases eligible for the more recently completed study were diagnosed with breast cancer between January 1, 2000 and March 31, 2004 at ages 55 through 74 years. In addition, given that the original purpose of the latter study was to identify differences in risk factor associations by tumor histology, cases were sampled on the basis of histology; specifically, all lobular cases and a random 25% sample of ductal cases were eligible.17 Controls for this study were identified through random-digit dialing18 and frequency matched to cases based on age. Response rates were similar for both studies (80.6% and 73.6% for cases and controls, respectively, in the earlier study and 83.5% and 71.1% for cases and controls, respectively, in the more recent study), and a total of 2019 eligible cases and 1476 eligible controls were enrolled and interviewed. The current analysis was restricted to controls and ductal cases (n = 1233) given the rarity of ER negativity in lobular disease (<10%) and the oversampling of lobular cases in the more recent study.

The pooled studies used similar protocols and materials. Study questionnaires were administered in-person and obtained data regarding several risk factors including lifestyle factors, reproductive history, use of hormone therapy, and family history of cancer. Women were asked about exposures occurring before their reference date, defined as the date of diagnosis for cases and assigned to controls based on the expected distribution of case reference dates.

Data concerning ER, PR, and HER-2 expression status were identified from CSS records. In addition, centralized pathology testing at the FHCRC had been completed for a sample of cases as part of the protocol from the more recent study; therefore, for a subset of cases for whom data regarding HER-2 expression were not available from CSS records, this information was derived from the results of centralized testing (97 cases). All ER and PR results, and a majority of HER-2 results, were based on immunohistochemistry (IHC) testing. When available, HER-2 data from fluorescent in situ hybridization (FISH) assays recorded in pathology reports (32 cases) were given precedence over IHC results. Overall, ER and PR data were available for 1179 cases (96%), 883 (75%) of whom were ER+ and PR+, 148 (13%) of whom were ER- and PR-, 139 (12%) of whom were ER+ and PR-, and 9 (<1%) of whom were ER- but PR+. Data for HER-2 were available for 847 cases (69%) in total, of whom 730 (86%) were HER-2- and 117 (14%) were HER-2+. Classification of case subtypes was made on the basis of ER, PR, and HER-2 data as follows: luminal (ER+), HER-2-overexpressing (ER-/PR-/HER-2+), and triple-negative (ER-/PR-/HER-2-). Cases with the rare ER-/PR+ phenotype were excluded (9 cases), as were cases with insufficient tumor marker data (52 cases missing ER data, and 32 ER-/PR- cases missing HER-2 data). Final analyses were based on 1476 controls and 1140 cases (1023 [90%] luminal cases, 39 [3%] HER-2-overexpressing cases, and 78 [7%] triple-negative cases).

We used polytomous logistic regression19 to compare luminal, HER-2-overexpressing, and triple-negative case groups with a common control group; this approach is directly comparable to performing a series of simple binary logistic regression models for different case–control comparisons. All analyses were adjusted for age (continuous) and reference year (continuous). Because there was no overlap in reference years between the 2 pooled studies, adjustment for reference year resulted in implicit adjustment for study population. The following nonreproductive breast cancer risk factors were evaluated as potential confounders: education level (≤high school degree/some college/college graduate), smoking status (yes/no), alcohol consumption (nondrinker/≤7 drinks per week/>7 drinks per week), and family history of breast cancer in first-degree relatives (yes/no). We similarly evaluated confounding between each of the reproductive factors under study. Adjustment for potential nonreproductive confounders did not alter risk estimates by >10% for any tumor subtype. However, adjustment for age at first live birth and breastfeeding history was found to have an effect on estimates for the number of live births. Similarly, adjustment for type of menopause impacted effect estimates for hormone therapy use and age at menopause. Therefore, final multivariate analyses for number of live births, age at first live birth, and breastfeeding were coadjusted for one another and restricted to parous women with available data for each of these variables (1226 controls, 802 luminal cases, 32 HER-2-overexpressing cases, and 62 triple-negative cases). Final multivariate analyses for hormone therapy use and age at menopause were adjusted for type of menopause. Analysis of age at menopause was restricted to women who experienced menopause (natural or due to bilateral oophorectomy) at a known age (1087 controls, 763 luminal cases, 27 HER-2-overexpressing cases, and 54 triple-negative cases). Tests for trend were also conducted, modeling variable categories with a single linear term. Heterogeneity of effect estimates between case groups was examined through a series of polytomous regression models comparing HER-2-overexpressing and triple-negative case groups with the luminal case group. All statistical analyses were performed using STATA/SE statistical software (version 9; StataCorp, College Station, Tex).

RESULTS

Compared with controls and the other case groups, luminal cases were somewhat more likely to have a first-degree family history of breast cancer and to be diagnosed at an early stage, HER-2-overexpressing cases were less likely to be non-Hispanic whites and to be college graduates, and triple-negative cases were more likely to be younger (Table 1). With respect to reproductive characteristics, correlations with menarche and parity-related events varied somewhat by tumor subtype (Table 2). We found that early age at menarche was only associated with the risk of HER-2-overexpressing disease (OR for age <13 years vs ≥13 years of 2.7; 95% confidence interval [95% CI], 1.4-5.5). Compared with nulliparous women, women with ≥1 live births experienced a modestly lower risk of luminal disease that was within the limits of chance (OR of 0.8; 95% CI, 0.6-1.0). Among parous women, neither the number of live births nor the age at first live birth were found to be strongly related with the risk of any tumor subtype. Women who breastfed for ≥6 months had reduced risks of both luminal (OR of 0.8; 95% CI, 0.6-1.0) and triple-negative disease (OR of 0.5; 95% CI, 0.3-0.9) compared with parous women who had never breastfed. The risk of luminal breast cancer was lower among women who had a surgical menopause compared with women who experienced a natural menopause (OR of 0.7; 95% CI, 0.6-0.9), and was higher among women who had a late age at menopause (OR for age ≥55 years vs age <45 years of 1.6; 95% CI, 1.1-2.2) or were current users of estrogen and progestin hormone therapy (OR of 1.7; 95% CI, 1.3-2.1). No statistically significant associations between menopausal status, age at menopause, or hormone therapy use and risks of either HER-2-overexpressing or triple-negative tumors were observed. However, there was a suggestion that age at menopause was positively associated with the risk of HER-2-overexpressing disease (P for linear trend = .07).

Table 1. Distribution of Demographic and Risk Factors by Case Status and Tumor Subtype

	Controls	Luminal Cases	HER-2-overexpressing Cases	Triple-negative Cases
	(N = 1476)	(N = 1023)	(N = 39)	(N = 78)
	No. (%)	No. (%)	No. (%)	No. (%)
NA indicates not applicable.
Age, y
55-64	258 (17)	226 (22)	12 (31)	28 (36)
65-74	922 (62)	621 (61)	19 (49)	40 (51)
≥75	296 (20)	176 (17)	8 (21)	10 (13)
Mean	69.3	68.6	67.6	66.0
Race/ethnicity
Non-Hispanic White	1348 (91)	956 (93)	32 (82)	72 (92)
African American	45 (3)	17 (2)	2 (5)	3 (4)
Asian/Pacific Islander	38 (3)	29 (3)	3 (8)	1 (1)
Other	45 (3)	21 (2)	2 (5)	2 (3)
Education, y
≤12	698 (47)	450 (44)	18 (46)	34 (44)
13-15	468 (32)	336 (33)	17 (44)	28 (36)
≥16	309 (21)	237 (23)	4 (10)	16 (21)
First-degree family
History of breast cancer
No	1151 (83)	734 (76)	33 (85)	62 (87)
Yes	230 (17)	226 (24)	6 (15)	9 (13)
Missing data	95	63	0	7
Stage at diagnosis
I	NA	715 (70)	22 (56)	45 (59)
II		271 (27)	12 (31)	25 (33)
III		29 (3)	5 (13)	6 (8)

Table 2. Multivariate-adjusted Odds Ratios for Reproductive and Hormonal Factors by Tumor Subtype*

Reproductive/Hormonal Characteristics	Controls	Luminal Cases		HER-2-overexpressing Cases		Triple-negative Cases
	No. (%)	No. (%)	OR (95% CI)	No. (%)	OR (95% CI)	No. (%)	OR (95% CI)
OR indicates odds ratios; 95% CI, 95% confidence interval; ref, referent. * OR were adjusted for age and diagnosis/reference year. † OR vs controls, P< .05. ‡ OR vs luminal cases, P< .05. § Excludes nulliparous women. Number of live births, age at first live birth, and breastfeeding were coadjusted for each other. ‖ Additionally adjusted for type of menopause experienced. ¶ Excludes women with cessation of menses due to hysterectomy without bilateral oophorectomy.
Age at menarche, y
≥13	817 (55)	552 (54)	1.0 (ref)	12 (31)	1.0 (ref)	41 (53)	1.0 (ref)
<13	658 (45)	467 (46)	1.0 (0.9-1.2)	27 (69)	2.7 (1.4-5.5)†,‡	37 (47)	1.1 (0.7-1.7)
Parity
Nulliparous	130 (9)	114 (11)	1.0 (ref)	3 (8)	1.0 (ref)	7 (9)	1.0 (ref)
Parous	1346 (91)	909 (89)	0.8 (0.6-1.0)	36 (92)	1.2 (0.4-3.9)	71 (91)	1.1 (0.5-2.3)
No. of live births§
1	117 (9)	96 (11)	1.0 (ref)	4 (11)	1.0 (ref)	9 (13)	1.0 (ref)
2	352 (26)	270 (30)	1.0 (0.7-1.3)	16 (44)	1.1 (0.4-3.5)	21 (30)	0.8 (0.3-1.7)
≥3	877 (65)	543 (60)	0.8 (0.6-1.1)	16 (44)	0.4 (0.1-1.3)	41 (58)	0.8 (0.3-1.7)
Age at 1st live birth, y§
<20	274 (20)	178 (20)	1.0 (ref)	4 (11)	1.0 (ref)	14 (20)	1.0 (ref)
20-24	651 (48)	435 (48)	1.0 (0.8-1.3)	24 (67)	2.3 (0.8-6.8)	34 (48)	1.1 (0.6-2.1)
25-29	305 (23)	200 (22)	1.0 (0.7-1.3)	6 (17)	1.0 (0.3-3.8)	19 (27)	1.3 (0.6-2.8)
≥30	113 (8)	93 (10)	1.2 (0.8-1.7)	2 (6)	0.7 (0.1-4.4)	4 (6)	0.7 (0.2-2.3)
Breastfeeding history§
Never	471 (35)	374 (41)	1.0 (ref)	12 (33)	1.0 (ref)	31 (44)	1.0 (ref)
Ever for <6 mo	408 (30)	249 (27)	0.8 (0.6-0.9)†	10 (28)	1.0(0.4-2.3)	25 (35)	0.9 (0.5-1.6)
Ever for ≥6 mo	467 (35)	286 (31)	0.8 (0.6-1.0)†	14 (39)	1.5 (0.7-3.3)	15 (21)	0.5 (0.3-0.9)†
Type of menopause
Natural	873 (60)	663 (66)	1.0 (ref)	22 (56)	1.0 (ref)	45 (59)	1.0 (ref)
Surgical (bilateral oophorectomy)	243 (17)	129 (13)	0.7 (0.6-0.9)†	6 (15)	1.0 (0.4-2.5)	10 (13)	0.8 (0.4-1.6)
Simple hysterectomy	329 (23)	208 (21)	0.8 (0.7-1.0)	11 (28)	1.3 (0.6-2.8)	21 (28)	1.3 (0.7-2.1)
Age at menopause, y‖,¶
<45	250 (23)	117 (15)	1.0 (ref)	4 (15)	1.0 (ref)	11 (20)	1.0 (ref)
45-54	668 (61)	502 (66)	1.4 (1.1-1.9)†	15 (56)	1.4 (0.4-4.6)	32 (59)	0.9 (0.4-2.0)
≥55	169 (16)	144 (19)	1.6 (1.1-2.2)†	8 (30)	2.9 (0.8-10.7)	11 (20)	1.2 (0.5-3.0)
P for linear trend			<.01		.07		.65
Hormone therapy use‖
Never use	537 (37)	343 (35)	1.0 (ref)	10 (27)	1.0 (ref)	31 (41)	1.0 (ref)
Former use only	248 (17)	162 (16)	1.0 (0.8-1.3)	11 (30)	2.2 (0.9-5.4)	13 (17)	0.8 (0.4-1.6)
Current unopposed estrogen use	438 (30)	242 (24)	0.9 (0.7-1.1)	10 (27)	1.0 (0.4-2.6)	21 (28)	0.7 (0.4-1.4)
Current estrogen+progestin use	215 (15)	242 (24)	1.7 (1.3-2.1)†	6 (16)	1.4 (0.5-4.2)	10 (13)	0.6 (0.3-1.3)‡

DISCUSSION

It is important to acknowledge the limitations of the current study before interpreting its results. Despite combining 2 studies, the statistical power of our analyses is limited by the inclusion of only 78 triple-negative and 39 HER-2-overexpressing cases. We also had to exclude 84 (7%) potentially eligible cases who lacked sufficient tumor marker data to be classified into a subtype. In addition, we were unable to distinguish between basal-like and unclassified cases, both of which have the triple-negative phenotype, because we lacked data on epidermal growth factor receptor and cytokeratin 5/6 expression, which are necessary to distinguish between these 2 subtypes. It is also possible that some misclassification of case subtypes resulted from the use of ER, PR, and HER-2 data taken from multiple laboratories within the Seattle-Puget Sound region because the assays and practices for reading and interpreting results vary across institutions and pathologists. However, it is reasonable to assume any such misclassification is nondifferential with respect to reproductive factors.

Consistent with several prior studies that have investigated the association between reproductive factors and the risk of hormone receptor-positive breast cancer,13–15, 20–22 we observed that current users of combined estrogen plus progestin preparations of hormone therapy and women who had a later age at menopause had an increased risk of luminal breast cancer, and women who breastfed had a decreased risk of luminal breast cancer. We also found that certain reproductive and hormonal characteristics were correlated with the risk of HER-2-overexpressing and triple-negative tumors. Age at menarche was only found to be associated with the risk of HER-2-overexpressing disease, and women who breastfed had a reduced risk of triple-negative disease. To our knowledge, only 2 prior studies have assessed differences in the associations between reproductive factors and breast cancer risk according to molecularly defined breast cancer subtypes: the Polish Breast Cancer Study (PBCS)8, 23 and the Carolina Breast Cancer Study (CBCS).23 There are important differences between these studies and ours because both the PBCS and the CBCS included premenopausal and postmenopausal women, and the CBCS also included a substantial proportion of African Americans (41%). Therefore, it is difficult to compare our findings with those of these studies because of differences in age and race/ethnicity distributions, and because both prior studies, as well as ours, are relatively underpowered by the inclusion of limited numbers of basal-like (95 cases and 225 cases, respectively) and HER-2-overexpressing cases (61 cases and 116 cases, respectively). The results of the current study are consistent with the CBCS in their observation that breastfeeding for ≥4 months was associated with a reduced risk of basal-like breast cancer (OR of 0.7; 95% CI, 0.4-0.9). However, contrary to our findings, both the CBCS and the PBCS observed an elevated risk of basal-like breast cancer among nulliparous compared with parous women.

Although definitive conclusions regarding which risk factors are more strongly related to certain molecularly defined subtypes of breast cancer cannot be made based on the current study, our data do support the premise that risk factor profiles vary by breast cancer subtype and that hormonal risk factors have a greater impact on luminal–type breast cancers than HER-2-overexpressing or triple-negative tumors. The observation that breastfeeding has now been shown to reduce the risk of triple-negative disease in 2 studies is intriguing, but the biologic basis for this association is unclear and this finding requires further replication. Given the well-defined clinical differences between breast tumor subtypes, the relatively poor prognoses of HER-2-overexpressing and triple-negative disease,2, 4, 6 and the paucity of existing epidemiologic data, further research is needed to clarify the epidemiology of these tumors because at the current time no consistent risk factors for either of these 2 subtypes have been identified.