Phenotypic analysis of tumor-infiltrating lymphocytes from human breast cancer.

Suspensions of fresh tumor-infiltrating lymphocytes (TIL) were prepared from 30 human breast ductal adenocarcinomas. To evaluate the phenotypic pattern of the isolated TIL, lymphocyte surface markers including CD19, CD3, CD4, CD8, CD16 and HLA-DR were examined by flow cytometry. Lymphocyte recovery ranged from 1.1% to 44%, independent of tumor size. TIL most often scored high for CD3+ with a varying number of CD4+ and CD8+ cells. Three samples out of 30 expressed up to 44% of CD19+ B cells, while CD3-CD16+ NK cells were rare. CD4 and CD8 expression was significantly different between the lymph node metastases group and the lymph node negative group (p < 0.01). 67% of the TIL with a CD4/CD8 ratio greater than 1 showed lymph node metastases. Furthermore, the CD4 expression of TIL and CD4/CD8 ratio correlated with tumor size (p < 0.01), but not with tumor differentiation and hormone receptor expression. Although there was considerable diversity of TIL among breast tumors, our data suggest that a high expression of CD4+ T cells may imply progression of the tumor, and an increased CD4/CD8 ratio of the TIL isolated from human breast adenocarcinoma may indicate development of metastases.     

HER2 testing on core needle biopsy specimens from primary breast cancers: interobserver reproducibility and concordance with surgically resected specimens

Background  

Accurate evaluation of human epidermal growth factor receptor type-2 (HER2) status based on core needle biopsy (CNB) specimens is mandatory for identification of patients with primary breast cancer who will benefit from primary systemic therapy with trastuzumab. The aim of the present study was to validate the application of HER2 testing with CNB specimens from primary breast cancers in terms of interobserver reproducibility and comparison with surgically resected specimens.     

Clinical validity of tumor-infiltrating lymphocytes analysis in patients with triple-negative breast cancer

BackgroundAlthough tumor-infiltrating lymphocytes (TILs) have been associated with a favorable prognosis in triple-negative breast cancer (TNBC) patients, this marker is not currently considered robust enough for entering the clinical practice. In the present study, we assessed the clinical validity of the guidelines recently issued by the International TIL Working Group in a large retrospective series of well-annotated TNBC patients.Patients and methodsTILs were evaluated in all the full-face H&E sections from 897 consecutive TNBC (i.e. tumors with <1% of ER and PgR immunoreactivity and absence of HER2 overexpression or amplification) patients diagnosed and treated at the European Institute of Oncology between 1995 and 2010 (median follow-up 8.2 years, range 6 months to 18 years). All mononuclear cells were evaluated in the stromal area within the borders of the invasive tumor, reported as a percentage value and treated as a continuous variable in survival analysis.ResultsThe median percentage of TILs was 20%, and 21.9% of the cases had ≥50% (lymphocyte predominant breast cancer, LPBC) TILs. At univariable survival analysis, TILs were a significant predictor of better disease-free survival (DFS), distant disease-free survival (DDFS) and overall survival (OS) (P < 0.0001). Multivariable analysis confirmed that each 10% increase in TILs strongly predicted better survival, independent of patients' age, lymph node status, tumor size, histological grade, peritumoral vascular invasion and Ki-67 labeling index. Patients with LPBC had a 10-year survival rate of 71%, 84% and 96% for DFS, DDFS and OS, respectively. Stratified analysis revealed a positive correlation between TILs and OS across all the subgroups analyzed.ConclusionOur data support the analytical validity of the recently issued TILs evaluation guidelines in the clinical practice.     

肿瘤浸润淋巴细胞在三阴性乳腺癌中作用的研究进展

三阴性乳腺癌（TNBC）是一种高度异质性的疾病。因缺乏有效的治疗靶标,故临床预后较差。尽管大量文献报道肿瘤浸润淋巴细胞与TNBC预后之间的关系,但区分不同类型的T淋巴细胞极其重要,因为他们在肿瘤微环境中发挥着不同的作用。此外,有研究发现肿瘤相关巨噬细胞与TNBC预后不良相关。本文将对不同淋巴细胞浸润在TNBC临床预后中的作用作一综述。     

Comparing core needle to surgical biopsies in breast cancer for cell kinetic and ploidy studies

The evaluability and reliability of proliferative activity (expressed as³H-thymidine labeling index,³H-TdR LI) and ploidy determinations on core needle biopsies were compared with those obtained on surgical material from the same breast cancers. The evaluability of³H-TdR LI on core needle biopsies was markedly lower than that on surgical material (53% vs 100%), and the association between³H-TdR LI values in the 16 cases with both evaluable determinations was poor (r_s=0.45). Conversely, determinations of ploidy on core needle biopsy and surgical material provided superimposable results, in terms of evaluability (91% vs 100%) and reliability (r_s=0.99). Further efforts are needed to improve sampling procedures for a reliable assessment of biological markers.     

Prognostic value of tumor-infiltrating lymphocytes on residual disease after primary chemotherapy for triple-negative breast cancer: a retrospective multicenter study

BackgroundThere is a need to develop surrogates for treatment efficacy in the neoadjuvant setting to speed-up drug development and stratify patients according to outcome. Preclinical studies showed that chemotherapy induces an antitumor immune response. In order to develop new surrogates for drug efficacy, we assessed the prognostic value of tumor-infiltrating lymphocytes (TIL) on residual disease after neoadjuvant chemotherapy (NACT) in patients with triple-negative breast cancer (TNBC).Patients and methodsThree hundred four TNBC patients with residual disease after NACT were retrospectively identified in three different hospitals. Hematoxylin and eosin-stained slides from surgical postchemotherapy specimens were evaluated for intratumoral (It-TIL) and stromal (Str-TIL) TIL. Cases were classified as High-TIL if It-TIL and/or Str-TIL >60%.ResultsTIL were assessable for 278 cases. Continuous It-TIL and Str-TIL variables were strong prognostic factors in the multivariate model, both for metastasis-free [hazard ratio (HR) 0.86, 95% confidence interval (CI) 0.77–0.96, P = 0.01 and HR 0.85, 95% CI 0.75–0.98, P = 0.02 for Str-TIL and It-TIL, respectively] and overall survival (HR 0.86, 95% CI 0.77–0.97, P = 0.01 and HR 0.86, 95% CI 0.75–0.99, P = 0.03 for Str-TIL and It-TIL, respectively). The 5-year overall survival rate was 91% (95% CI 68% to 97%) for High-TIL patients (n = 27) and 55% (95% CI 48% to 61%) for Low-TIL patients (HR 0.19, 95% CI 0.06–0.61, log-rank P = 0.0017). The major prognostic impact of TIL was seen for patients with large tumor burden following NACT (residual tumor >2 cm and/or node metastasis). In all but one High-TIL case, It-TIL and Str-TIL values were lower on the prechemotherapy sample.ConclusionsThe presence of TIL in residual disease after NACT is associated with better prognosis in TNBC patients. This parameter may represent a new surrogate of drug efficacy to test investigational agents in the neoadjuvant setting and a new prognostic marker to select patients at high risk of relapse.     

Comparison of gene expression profiles in core biopsies and corresponding surgical breast cancer samples

Introduction  

Gene expression profiling has been successfully used to classify breast cancer into clinically distinct subtypes, and to predict the risk of recurrence and treatment response. The aim of this study was to investigate whether the gene expression profile (GEP) detected in a core biopsy (CB) is representative for the entire tumor, since CB is an important tool in breast cancer diagnosis. Moreover, we investigated whether performing CBs prior to the surgical excision could influence the GEP of the respective tumor.     

Prognostic and predictive value of tumor-infiltrating lymphocytes in breast cancer: a systematic review and meta-analysis

Background

Breast cancer is the most common invasive cancer to affect women in the world. Studies showed tumor-infiltrating lymphocytes can exhibit both beneficial and harmful effects on the biology and clinical outcome of breast cancer, the conclusion still remains incomplete. Here, we conducted a meta-analysis to evaluate the relationship between tumor-infiltrating lymphocytes and breast cancer.

Methods

A comprehensive search strategy was used to search relevant literatures in PubMed and the ISI Web of Science. The correlation among TILs and breast cancer clinicopathological features and prognosis was analyzed by using Review Manager 5.3 and Stata 12.0.

Result

Seventeen eligible studies consisting of 12,968 participants were included. We found that higher value of tumor-infiltrating lymphocytes had no relationship with breast cancer clinicopathological variables. Interestingly, it was correlated with response to neoadjuvant chemotherapy in majority (pooled RR 2.43, 95 % CI 1.99–2.97). Moreover, higher value of total tumor-infiltrating lymphocytes (both intraepithelial and stromal) was associated with better prognosis (pooled HR 0.88, 95 % CI 0.83–0.94), whereas some subtypes predicted a worse prognosis.

Conclusion

This meta-analysis indicated that high value of total TILs is not associated with breast cancer clinicopathological features, but can predict a favorable outcome for neoadjuvant chemotherapy in majority except for hormone receptor (?) subtype. And higher total TILs (both intraepithelial TILs and stromal TILs) may be the potential better prognostic indicators, while some subtypes like PD-1⁺ TILs and Foxp3⁺ TILs show a worse prognosis.

     

粗针穿刺及术后乳腺癌标本的免疫组织化学法检测指标与分子分型的比较研究

  目的  评估粗针穿刺及术后乳腺癌标本的免疫组织化学法检测指标与分子分型一致性,分析导致粗针穿刺标本的免疫组织化学法检测指标差异的因素。  方法  回顾性分析2015年8月至2016年11月324例于天津医科大学肿瘤医院未经新辅助化疗行乳腺癌改良根治术患者的临床病理资料。比较粗针穿刺及术后标本经免疫组织化学法检测的指标ER、PR、HER-2、Ki-67与分子分型一致性。  结果  粗针穿刺及术后乳腺癌标本的免疫组织化学法检测指标ER、PR、HER-2、Ki-67一致率分别为94.1%（305/324）、90.7%（294/324）、61.1%（198/324）、86.7%（281/324）,Kappa值分别为0.84、0.76、0.38、0.34;分子分型一致率为73.4%（91/124）,Kappa值为0.64。  结论  乳腺癌粗针穿刺活检在免疫组织化学法检测指标ER、PR与分子分型评估中准确性较高,在HER-2、Ki-67检测中一致性较低,粗针穿刺结合术后标本的免疫组织化学法结果,可为提高分子分型的准确性及选择最佳治疗提供依据。      

False-negative core needle biopsies of the breast: an analysis of clinical,radiologic, and pathologic findings in 27 concecutive cases of missed breast cancer

BACKGROUND: A benign diagnosis in a core needle biopsy (CNBx) of the breast performed for a clinically and/or radiologically suspicious abnormality is often due to a nonrepresentative sample. However, the discordance may not be recognized, resulting in a logistic delay in the diagnosis. METHODS: Twenty-seven false-negative CNBxs were identified in 952 consecutive CNBxs of the breast (653 benign, 266 malignant, and 33 atypical) performed during a 1-year period. Biopsies were analyzed with respect to clinical and radiologic findings, biopsy type, type of malignancy, and interval between the original CNBx and final diagnosis. Four hundred thirty-eight (67%) of the patients with a benign CNBx diagnosis either underwent excision or had a minimum of 1-year follow-up (mean, 35.6 months; median, 36 months). RESULTS: The cancers missed on CNBx included 6 ductal carcinomas in situ, 17 invasive ductal carcinomas, 3 invasive lobular carcinomas, and 1 non-Hodgkin lymphoma. The overall false-negative rate was 9.1%. For palpable lesions, ultrasound-guided CNBx had a lower rate of missed cancer (3.6%) compared with CNBx without image guidance (13.3%). The false-negative rate for vacuum assisted CNBx biopsy was 7.6% (3.3% for the 11-gauge needle, 22.2% for the 14-gauge needle; 5.6% for nonpalpable mass lesions, 8.2% for microcalcifications). In all seven false-negative CNBxs performed by radiologists, the discordance between the radiologic and pathologic findings was promptly recognized due to their standard follow-up protocol. The discordance between the degree of clinical suspicion, radiologic impression, and the pathologic findings was not immediately recognized in 5 of 20 false-negative CNBxs performed by surgeons (4 without radiologic guidance and 1 with ultrasound guidance), resulting in a delay in the diagnosis ranging from 112-336 days. CONCLUSIONS: A false-negative diagnosis of breast carcinoma was found to be more common in CNBx performed without image guidance but occurred to a lesser degree in image-guided biopsies. A delay in diagnosis can be avoided by establishing a standard post-CNBx follow-up protocol.     

Functional characterization of tumor-infiltrating lymphocytes, lymph-node lymphocytes and peripheral-blood lymphocytes from patients with breast cancer

Mononuclear cell infiltration is frequently seen within human solid tumors. Effector cells within the tumor site usually fail to exhibit cytotoxic or natural killer activity when freshly isolated; however, they develop potent and sometimes specific cytotoxicity after expansion in IL2. Thus, local tumor environment may influence lymphocyte function. In our study, we disaggregated human breast-cancer and lymph-node tissue to obtain lymphocyte-enriched cell fractions. Besides phenotypic analysis, functional characterization with regard to proliferation and cytokine production of tumor-infiltrating lymphocytes (TIL), peripheral-blood lymphocytes (PBL) and lymph-node lymphocytes (LNL) was the aim of our study. TIL showed an enrichment of CD8+ cells with a corresponding decrease in CD4+ cells in comparison with PBL and LNL. In response to PHA, TIL showed decreased 3H-thymidine uptake, but TIL were significantly stimulated by rhIL2. TIL produced low levels of IL2, TNF and IFN gamma upon mitogen/phorbol ester stimulation, while PBL produce high levels of TNF and IFN gamma but low levels of IL2. Under the same experimental conditions, LNL produce high levels of TNF and IL2 but low levels of IFN gamma. Mitogen-mediated TNF secretion was increased after addition of autologous tumor cells in TIL and LNL, whereas IFN gamma secretion tended to be suppressed. Our results indicate different patterns of activities of TIL, LNL and PBL from breast-cancer patients.     

Development of tumor-infiltrating lymphocytes in breast cancer after neoadjuvant paclitaxel chemotherapy.

PURPOSE: Neoadjuvant chemotherapy for breast cancer creates new possibilities for the analysis of biological factors in the tumor and/or host, which may play a role in the response to treatment. In this study we analyzed whether changes in local antitumor immunity take place after neoadjuvant paclitaxel therapy and if they correlate with response to treatment. EXPERIMENTAL DESIGN: Neoadjuvant chemotherapy (paclitaxel, 200 mg/m2 q2w, 4 treatments) was followed by definitive surgical management. Histological sections from the pre- and post-treatment surgical specimens of 25 patients were analyzed for the extent of lymphocytic infiltration and presence of tumor infiltrating lymphocytes (TILs). The cumulative apoptotic response in the tumor after the first dose of paclitaxel was also studied in 10 of 25 patients. RESULTS: Pretreatment lymphocytic infiltrate in the tumor was minimal in the majority of patients and showed no relationship with clinical response. In the patients without TILs before treatment, development of TILs after treatment was noted in 0/3 (0%) patients with stable disease, 3/12 (25%) patients with clinical partial response, and 4/6 (67%) patients with clinical complete response and pathological residual disease. These correlated with the tumor cell apoptotic response to the first dose of paclitaxel. CONCLUSIONS: These results suggest that development of TILs after treatment correlates with clinical response to neoadjuvant paclitaxel therapy. The possible mechanism(s) whereby neoadjuvant chemotherapy may lead to induction of antitumor T cells is discussed. Immunological processes may influence the response of breast cancer patients to neoadjuvant treatment.     

Progression risk of columnar cell lesions of the breast diagnosed in core needle biopsies

Columnar cell lesions (CCLs) of the breast are recognized as putative precursor lesions of invasive carcinoma, but their management remains controversial. We therefore conducted a retrospective study on 311 CCLs, diagnosed in 4,164 14-gauge core needle biopsies (CNB): 221 CCLs without atypia (CCL), 69 with atypia (CCL-A), and 21 atypical ductal hyperplasias originating in CCL (ADH-CCL). Two groups were identified: "immediate treatment" group undergoing excision within four months after the CNB diagnosis of CCL (N = 52) and the "wait-and-see" group followed up to 8 years (median 3.5 years, N = 259). In 7 of 31 women (22.5%, 1 CCL, 4 CCL-A, 2 ADH-CCL) who underwent immediate surgical excision and were initially biopsied for microcalcifications, ductal carcinoma in situ (DCIS) was present and in 2/31 women (6.5%, 1 CCL, 1 CCL-A) invasive carcinoma. In 2/21 excisions (9.5%, 1 CCL, 1 CCL-A) initially biopsied for a density, DCIS was present and invasive carcinoma in 5/21 excisions (23.8%, 2 CCL, 3 CCL-A). In the wait-and-see group, 9/259 women (3.5%) developed invasive carcinoma, 6 ipsi, and 3 contralaterally. Progression risks of CCL-A and ADH-CCL were 18% and 22%,versus 2% for CCL without atypia (p < 0.001). In conclusion, CCL-A or ADH-CCL in a CNB were associated with a high risk of DCIS/invasive carcinoma in immediate surgical excision biopsies. The 8-years progression risks for CCL-A and ADH-CCL were around 20%. This illustrates that an atypical CCL in a CNB may signal the presence of concurrent lesions or development of advanced lesions in future and may justify ("mini") surgical excision.     

Modified core wash cytology procedure for the immediate diagnosis of core needle biopsies of breast lesions

BACKGROUND:

Core wash or touch imprint cytology is often used to obtain a quick, preliminary diagnosis on a core needle biopsy (CNB) of breast lesions, essential for the management of the 1‐day breast clinic. Contradictory results of both techniques in the literature led to this preclinical study investigating an alternative method of touch imprint and core wash cytology.

METHODS:

Thirty breast lesions were biopsied by a core needle in a laboratory setting. The CNBs were collected in RPMI fluid (Roswell Park Memorial Institute fluid). The touch imprint cytology was performed taking the biopsy out of the fluid and smearing it on a microscopic slide and May‐Grunwald Giemsa stained. The core wash cytology was made by fixating the remaining cells in Fixcyt and prepared with a liquid‐based preparation method and Papanicolaou stained. The cytologic findings were categorized into benign, atypical favoring benign, atypical, suspicious, and malignant and compared with the histologic CNB results.

RESULTS:

The CNBs showed 20 of 30 samples to be malignant, 2 to be phylloides tumors, 7 to be benign, and 1 to be unsatisfactory. Both techniques showed a sensitivity of 95% and specificity of 100%. Touch imprint yielded insufficient diagnoses (13.3%), compared with core wash (6.6%). Of the core wash cases, 86% showed a good quality versus 30% in touch imprint cytology.

CONCLUSIONS:

This preclinical study on modified touch imprint and core wash techniques led to results that were comparable to or better than those in the literature. The core wash cytology is preferred to touch imprint because of the better morphology. Cancer (Cancer Cytopathol) 2009. © 2009 American Cancer Society.     

Ultrasound-guided core needle biopsy for breast cancer: preliminary report

BACKGROUND: Ultrasound-guided automated percutaneous core needle biopsy (US-CNB) for breast tumors has been introduced into clinical practice, but it has not yet been used routinely. We evaluated its usefulness, especially in terms of histological accuracy. METHODS: Thirty-one consecutive patients underwent mammography followed by breast biopsy with the automated core needle biopsy device. RESULTS: Mammography was highly suggestive of malignancy or suspicious abnormalities in 17 cases whose histological findings from US-CNB specimens were invasive ductal carcinoma without exception. The other 14 cases with benign or probably benign mammography findings showed no malignancy histologically in the US-CNB specimens. In cases of malignancy, the accuracy rates of histological findings for the specimens obtained by US-CNB were 94.1% in histological type, 100% in direct infiltration, 82.4% in lymphatic infiltration, 82.4% in venous infiltration, 94.1% in histological grading and 82.4% in intraductal spread. CONCLUSION: US-CNB was useful for making reliable preoperative histopathological diagnosis and may substitute fine needle aspiration biopsy and surgical biopsy.     

乳腺癌核芯针穿刺印片与组织学诊断的比较研究

目的 探讨印片细胞学(TIC)与组织病理诊断以及印片免疫细胞化学与免疫组织化学检测结果的一致性,评价在乳腺癌新辅助化疗前TIC诊断的临床应用价值.方法 收集行核芯针穿刺组织TIC诊断的乳腺肿物患者289例,其中287例有核芯针活检(CNB)病理结果对照,190例有术后病理结果对照.289例中,64例行印片的雌激素受体(ER)、孕激素受体(PR)和人表皮生长因子受体2(HER-2)免疫细胞化学检测,其中52例有CNB免疫组织化学检测结果,43例有术后病理免疫组织化学检测结果.结果 TIC诊断良性22例,恶性263例,不满意标本4例.假阴性率和不满意率均为1.4%,假阳性率为0.35%.与术后病理结果比较,TIC和CNB诊断乳腺癌的敏感性分别为96.2%和95.0%(P=0.601),特异性分别为87.5%和100%(P=0.471),准确率分别为95.8%和95.3%(P=0.804),差异均无统计学意义.ER、PR和HER-2印片免疫细胞化学检测结果与CNB的免疫组织化学检测结果的符合率分别为86.5%、75.0%和78.8%,与术后病理免疫组织化学检测结果的符合率分别为88.4%、74.4%和75.6%,CNB与术后病理免疫组织化学检测结果的符合率分别为83.7%、74.4%和76.5%,三者差异无统计学意义(P＞0.05).结论 TIC诊断乳腺癌的敏感性、特异性和准确率较高,与CNB组织病理诊断无明显差异,可以辅助CNB为乳腺肿物患者提供快速的细胞学诊断.印片免疫细胞化学方法可以辅助CNB作为新辅助化疗前检测乳腺癌患者受体水平的手段之一.