Molecular profiles of asynchronous breast cancer metastases are of clinical relevance to individual patients' treatment, whereas the role of profiles in synchronous lymph node metastases is not defined. The present study aimed to assess individual biomarkers and molecular subtypes according to the St Gallen classification in primary breast tumours, synchronous lymph node metastases and asynchronous relapses and relate the results to 10-year breast cancer mortality (BCM). Tissue microarrays were constructed from archived tissue blocks of primary tumours (N = 524), synchronous lymph node metastases (N = 147) and asynchronous relapses (N = 36). The samples were evaluated by two independent pathologists according to oestrogen receptor (ER), progesterone receptor (PR), Ki67 and human epidermal growth factor receptor 2 (HER2) by immunohistochemistry and in situ hybridisation. The expression of biomarkers and molecular subtypes in the primary tumour was compared with that in the synchronous lymph node metastases and relapses, and related to 10-year BCM. Discordances were found between primary tumours and relapses (ER: p = 0.006, PR: p = 0.04, Ki67: p = 0.02, HER2: p = 0.02, St Gallen subtypes: p = 0.07) but not between primary tumours and metastatic lymph node. Prognostic information was gained by the molecular subtype classification in primary tumours and nodal metastases; triple negative subtype had the highest BCM compared with the luminal A subtype (primary tumours: HR 4.0; 95 % CI 2.0-8.2, p < 0.001, lymph node metastases: HR 3.5; 95 % CI 1.3-9.7, p = 0.02). When a shift in subtype inherence between primary tumour and metastatic lymph node was identified, the prognosis seemed to follow the subtype of the lymph node. Molecular profiles are not stable throughout tumour progression in breast cancer. Prognostic information for individual patients appears to be available from the analysis of biomarker expression in synchronous metastatic lymph nodes. The study supports biomarker analysis also in asynchronous relapses.

BackgroundThe St Gallen surrogate molecular subtype definitions classify the oestrogen (ER) positive breast cancer into the luminal A and luminal B subtypes according to proliferation rate and/or expression of human epidermal growth factor receptor 2 (HER2) with differences in prognosis and chemo-responsiveness. Primary tumours and lymph node metastases might represent different malignant clones, but in the clinical setting only the biomarker profile of the primary tumour is used for selection of adjuvant systemic treatment. The present study aimed to classify primary breast tumours and matched lymph node metastases into luminal A, luminal B, HER2-positive and triple-negative subtypes and compare the distributions.MethodsEighty-five patients with available tumour tissue from both locations were classified. The distribution of molecular subtypes in primary tumours and corresponding lymph node metastases were compared, and related to 5-year distant disease-free survival (DDFS).ResultsThe St Gallen molecular subtypes were discordant between primary tumours and matched lymph node metastases in 11% of the patients (p = 0.06). The luminal A subtype in the primary tumour shifted to a subtype with a worse prognostic profile in the lymph node metastases in 7 of 45 cases (16%) whereas no shift in the opposite direction was observed (0/38) (p = 0.02). All subtypes had an increased hazard for developing distant metastasis during the first 5 years after diagnosis in both primary breast tumours and matched lymph node metastases, compared with the luminal A subtype.ConclusionThe classification according to the St Gallen molecular subtypes in primary tumours and matched lymph node metastases, implicates a shift to a more aggressive subtype in synchronous lymph node metastases compared to the primary breast tumour. The selection of systemic adjuvant therapy might benefit from taking the molecular subtypes in the metastatic node into account.

BackgroundDiagnosis by screening mammography is considered an independent positive prognostic factor, although the data are not fully in agreement. The aim of the study was to explore whether the mode of detection (screening‐detected versus symptomatic) adds prognostic information to the St Gallen molecular subtypes of primary breast cancer, in terms of 10‐year cumulative breast cancer mortality (BCM).MethodsA prospective cohort of patients with primary breast cancer, who had regularly been invited to screening mammography, were included. Tissue microarrays were constructed from primary tumours and lymph node metastases, and evaluated by two independent pathologists. Primary tumours and lymph node metastases were classified into St Gallen molecular subtypes. Cause of death was retrieved from the Central Statistics Office.ResultsA total of 434 patients with primary breast cancer were included in the study. Some 370 primary tumours and 111 lymph node metastases were classified into St Gallen molecular subtypes. The luminal A‐like subtype was more common among the screening‐detected primary tumours (P = 0·035) and corresponding lymph node metastases (P = 0·114) than among symptomatic cancers. Patients with screening‐detected tumours had a lower BCM (P = 0·017), and for those diagnosed with luminal A‐like tumours the 10‐year cumulative BCM was 3 per cent. For patients with luminal A‐like lymph node metastases, there was no BCM. In a stepwise multivariable analysis, the prognostic information yielded by screening detection was hampered by stage and tumour biology.ConclusionThe prognosis was excellent for patients within the screening programme who were diagnosed with a luminal A‐like primary tumour and/or lymph node metastases. Stage, molecular pathology and mode of detection help to define patients at low risk of death from breast cancer.

The concordance for the biomarkers analyzed in matched pairs of primary tumors and lymph node metastases was high. Moreover, survival analyses showed that the expression of biomarkers in lymph node metastases can provide prognostic information when no analyses of the primary tumor can be done. Treatment selection based on biomarkers in the lymph node is a topic for further studies.

BackgroundTriple-negative breast cancer (TNBC) is a heterogeneous subgroup of breast cancer with poor prognosis and no targeted therapy available. Receptor tyrosine kinases (RTKs) are emerging targets in anticancer therapy and many RTK-inhibiting drugs are currently being developed. The aim of this study was to elucidate if there is a correlation between the protein expression of three RTKs c-KIT, VEGFR2 and PDGFRα, their gene copy number, and prognosis in TNBC compared to non-TNBC.MethodsTumor tissue samples from patients diagnosed with primary breast cancer were stained with immunohistochemistry (IHC) for protein assessment, and with fluorescence in situ hybridization (FISH) for gene copy number determination. Breast cancer mortality (BCM), measured from the date of surgery to death, was used as endpoint.ResultsThe cohort included 464 patients, out of which 34 (7.3%) had a TNBC. High expression of the three RTKs was more common in TNBC compared to non-TNBC: c-KIT 49% vs. 10% (P<0.001), PDGFRα 32% vs. 19% (P = 0.07) and VEGFR2 32% vs. 6% (P<0.001). The odds ratio (OR) of c-KIT, VEGFR2 and PDGFRα positivity, adjusted for tumor characteristics, was 6.8, 3.6 and 1.3 times higher for TNBC than for non-TNBC. 73.5% of the TNBC had high expression of at least one of the three investigated receptors, compared to 30.0% of the non-TNBC (P<0.001). Survival analysis showed no significant difference in BCM for TNBC patients with high vs. low c-KIT, PDGFRα or VEGFR2 protein expression. 193 (42%) tumors were evaluated with FISH. No correlation was seen between increased gene copy number and TNBC, or between increased gene copy number and high protein expression of the RTK.Conclusionc-KIT, VEGFR2 and PDGFRα show higher protein expression in TNBC compared to non-TNBC. Further investigation clarifying the importance of these RTKs in TNBC is encouraged, as they are possible targets for anticancer therapy.

BackgroundDisseminated tumour cells (DTCs) in the bone marrow of patients with breast cancer have been identified as an independent predictor of poor prognosis in patients with non-metastatic disease. This prospective study aimed to evaluate the presence and prognostic value of DTCs in the bone marrow of female patients with primary breast cancer.MethodsBetween 1999 and 2003, bone marrow aspirates were obtained from patients at the time of surgery for primary invasive breast cancer. DTCs in bone marrow were identified using monoclonal antibodies against cytokeratins for detection of epithelial cells. The detection of DTCs was related to clinical follow-up with distant disease-free survival (DDFS) and breast cancer-specific survival as endpoints. Bone marrow aspirates from adult healthy bone marrow donors were analysed separately.ResultsDTCs were analysed in 401 patients, and cytokeratin-positive cells were found in 152 of these (38%). An immunofluorescence (IF) staining procedure was used in 327 patients, and immunocytochemistry (IC) was performed in 74 patients. The IF-based method resulted in 40% DTC-positive cases, whereas 30% were positive using IC (p = 0.11). The presence of DTCs in bone marrow was not significantly related to patient or tumour characteristics. The presence of DTCs was not a prognostic factor for DDFS (IF: hazards ratio [HR], 2.2; 95% confidence interval [CI], 0.63–2.2; p = 0.60; IC: HR, 0.84; 95% CI, 0.09–8.1; p = 0.88). Significant prognostic factors were lymph node metastases, oestrogen receptor positivity, Nottingham histological grade, and tumour size using Cox univariate analysis. The analyses were positive for epithelial cells in bone marrow from adult healthy donors in 19 (25%) samples.ConclusionsThe detection of DTCs in bone marrow in primary breast cancer was previously shown to be a predictor of poor prognosis. We were not able to confirm these results in a prospective cohort including unselected patients before the standard procedure was established. Future studies with a standardised patient protocol and improved technique for isolating and detecting DTCs may reveal the clinical applications of DTC detection in patients with micrometastases in the bone marrow.

BackgroundThe aim was to confirm a previously defined prognostic index, combining a proliferation marker, histological grade, and estrogen receptor (ER) in different subsets of primary N0/N1 chemo-naïve breast cancer patients.Methods/designIn the present study, including 1,854 patients, Ki67 was used in the index (KiGE), since it is the generally accepted proliferation marker in clinical routine. The low KiGE-group was defined as histological grade 1 patients and grade 2 patients which were ER-positive and had low Ki67 expression. All other patients made up the high KiGE-group. The KiGE-index separated patients into two groups with different prognosis. In multivariate analysis, KiGE was significantly associated with disease-free survival, when adjusted for age at diagnosis, tumor size and adjuvant endocrine treatment (hazard ratio: 3.5, 95% confidence interval: 2.6–4.7, P<0.0001).DiscussionWe have confirmed a prognostic index based on a proliferation marker (Ki67), histological grade, and ER for identification of a low-risk group of patients with N0/N1 primary breast cancer. For this low-risk group constituting 57% of the patients, with a five-year distant disease-free survival of 92%, adjuvant chemotherapy will have limited effect and may be avoided.

Objective: According to St Gallen recommendations from 2013, estrogen receptor (ER), progesterone receptor (PR), HER-2, and Ki-67 defines two subtypes of ER-positive and HER-2 normal breast cancer (BC): Luminal A-like and Luminal B-like. Patients with Luminal B-like BC are often recommended chemotherapy in addition to endocrine therapy, whereas endocrine therapy may be sufficient for patients with Luminal A-like BC. Histological grade (G) 1, 2 and 3 are not included in the St Gallen recommendations. Our unpublished data from a series of 161 premenopausal N0 BC patients with long-term follow-up show that the classification of Luminal A-like vs Luminal B-like HER-2 normal BC is strongly associated to G. Luminal A-like BC is often G1 or G2 and Luminal B-like BC is usually G2 or G3. We also found that the few G3 (n=6) Luminal A-like cases had a prognosis more similar to Luminal B-like and that the few G1 (n=2) Luminal B-like HER-2 normal cases had a prognosis more similar to Luminal A-like. The aim of this study is to evaluate in other cohorts if these findings can be confirmed. Methods: G and St Gallen subtypes were evaluated in three BC cohorts from altogether 547 pre- and postmenopausal chemotherapy naïve T1-2N0-N1M0 patients. The endpoint was distant disease-free survival with 10 years of follow-up. We compared the Luminal A-like and the Luminal B-like HER-2 normal subtype definition according to the original St Gallen recommendation from 2013 based on ER, PR, HER-2 and Ki-67 (St Gallen 2013) with our proposal, where ER-positive, HER-2 normal, G1 BC is defined as Luminal A-like, independent of Ki-67 and PR, and ER-positive, G3 BC is defined as Luminal B-like, independent of Ki-67 and PR (St Gallen 2013+G). The importance of Ki-67 and PR for subtyping was thus restricted to G2 BC. Results: The hazard ratio (HR) between Luminal B-like HER-2 normal (n=185) and Luminal A-like (n=362) defined according to St Gallen 2013+G, was 2.3 (95% confidence interval (CI) 1.6-3.4; p&lt;0.0001) compared to 1.6 (95% CI: 1.1-2.4; p=0.025) according to St Gallen 2013. Twenty-five patients, classified as Luminal B-like HER-2 normal with St Gallen 2013 were G1 and consequently reclassified as Luminal A-like with St Gallen 2013+G. None of these twenty-five patients developed metastases during the follow-up period. Thirty-eight patients showed the opposite pattern (G3 Luminal A-like with St Gallen but Luminal B-like according to St Gallen 2013+G). Seventeen of these patients developed distant metastases already during the first five years. Conclusion: Our findings strongly suggest that ER-positive, HER-2 normal, and G1 BC is a good prognosis group, independent of Ki-67 and PR, and should be treated as Luminal A-like BC, whereas ER-positive, HER-2 normal, and G3 BC should be considered as a worse prognosis group, independent of Ki-67 and PR, and should be treated as Luminal B-like BC. Based on our findings the importance of Ki-67 and PR is restricted to G2 BC for the discrimination between Luminal A-like and Luminal B-like HER-2 normal subtypes of BC. Citation Format: Anna Ehinger, Per Malmström, Pär-Ola Bendahl, Christopher W Elston, Anna-Karin Falck, Carina Forsare, Dorthe Grabau, Lisa Rydén, Olle Stål, Mårten Fernö. Histological grade provides significant prognostic information in the discrimination between luminal A-like and luminal B-like HER-2 normal subtypes of breast cancer according to St Gallen 2013 [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-08-43.