Estrogen receptor (ER), progesterone receptor (PR), and HER2 expression pre- and post- neoadjuvant chemotherapy in primary breast carcinoma: a single institutional experience

Background

The estrogen receptor (ER), progesterone receptor (PR), and HER2 profile of a primary breast carcinoma plays a significant role in patient management and treatment. Because of the increasing utilization of neoadjuvant chemotherapy or hormone therapy, surgically-resected carcinomas often show marked treatment effect. The aim of this study was to compare immunohistochemical (IHC) profiles (ER, PR, HER2, HER2 FISH) of primary breast carcinomas before and after neoadjuvant chemotherapy to assess the subsequent effects on hormone receptor status.

Design

Primary breast carcinomas from 38 female patients treated with neoadjuvant therapy after needle core biopsy or fine needle aspiration diagnosis were included. Histologic data was collected for each case, including site, type, grade, tumor size (cm), pre- and post- neoadjuvant treatment IHC panel (ER, PR, HER2), and fluorescence in-situ hybridization (FISH) for HER2.

Results

Of the 38 carcinomas studied, 45 % were positive for ER by IHC both pre- and post- neoadjuvant treatment (P=1.00). IHC studies for PR in these 38 patients showed 37% positivity for PR pre-neoadjuvant therapy and 21% positivity post-treatment (p=0.03). For 37 patients with HER2 IHC, 32% were positive pre-treatment, and 22% were positive post-treatment (P = 0.20). For 7 patients, HER2 FISH was positive in 71% pre-therapy and in 57% post-treatment (P=0.32).

Conclusions

Profiles for ER, HER2 IHC, and HER2 FISH were not significantly different in primary breast carcinomas before and after neoadjuvant chemotherapy. Further investigation is warranted to assess reproducibility of technique and investigate clinical implications of significant loss of PR status in treated patients.     

In situ immune response after neoadjuvant chemotherapy for breast cancer predicts survival

Accumulating preclinical evidence suggests that anticancer immune responses contribute to the success of chemotherapy. However, the predictive value of tumour-infiltrating lymphocytes after neoadjuvant chemotherapy for breast cancer remains unknown. We hypothesized that the nature of the immune infiltrate following neoadjuvant chemotherapy would predict patient survival. In a series of 111 consecutive HER2- and a series of 51 non-HER2-overexpressing breast cancer patients treated by neoadjuvant chemotherapy, we studied by immunohistochemistry tumour infiltration by FOXP3 and CD8 T lymphocytes before and after chemotherapy. Kaplan-Meier analysis and Cox modelling were used to assess relapse-free survival (RFS) and overall survival (OS). A predictive scoring system using American Joint Committee on Cancer (AJCC) pathological staging and immunological markers was created. Association of high CD8 and low FOXP3 cell infiltrates after chemotherapy was significantly associated with improved RFS (p = 0.02) and OS (p = 0.002), and outperformed classical predictive factors in multivariate analysis. A combined score associating CD8/FOXP3 ratio and pathological AJCC staging isolated a subgroup of patients with a long-term overall survival of 100%. Importantly, this score also identified patients with a favourable prognosis in an independent cohort of HER2-negative breast cancer patients. These results suggest that immunological CD8 and FOXP3 cell infiltrate after treatment is an independent predictive factor of survival in breast cancer patients treated with neoadjuvant chemotherapy and provides new insights into the role of the immune milieu and cancer.     

乳腺癌新辅助化疗前后组织病理学与分子标志物表达意义的探讨及对比分析

目的 探讨乳腺癌新辅助化疗前核芯针穿刺活检(CNB)标本和化疗后手术切除标本的病理类型、病理分级、分子标志物表达及其改变与治疗反应病理评价的关系.方法 收集209例接受新辅助化疗的乳腺癌患者CNB和手术切除标本,评价其病理类型、病理分级、治疗反应病理评价信息(MP分级系统);应用免疫组织化学方法 (MaxVision二步法)检测上述标本分子标志物雌激素受体(ER)、孕激素受体(PR)和HER2的表达信息,应用SPSS 15.0软件进行相关统计学分析.结果 (1)新辅助化疗后治疗反应病理评价分别为MP1级14例,MP2级35例,MP3级106例,MP4级36例,MP5级18例.(2)CNB标本的ER表达与治疗反应病理评价呈负相关(χ2=33.083,P=0.001);手术切除标本各类信息与治疗反应病理评价未见统计学相关性(P>0.05);(3)化疗后手术切除标本信息均可发生改变,病理类型、病理分级发生变化病例所占比例分别为6.8%(9/132)和34.9%(30/86);ER、PR、HER2表达发生改变的比例分别为42.4%(75/177)、55.4%(98/177)和26.6%(46/173),仅HER2表达改变的差异有统计学意义(P=0.049).上述信息改变与治疗反应病理评价无关(P>0.05).结论 CNB标本对预测肿瘤治疗反应的病理评价具有重要价值.化疗后肿瘤的信息均可发生改变,因此有必要在新辅助化疗后重复确认肿瘤组织的病理类型及病理分级,并应用免疫组织化学方法 重复检测化疗后手术切除标本的分子标志物表达.

Abstract：

Objective To investigate the relationship between the pathologic responses and histologic type, grade, the expression of ER, PR and HER2 and their changes in breast carcinoma before and after neoadjuvant chemotherapy (NAC). Methods Two-hundred and nine cases of breast cancer with NAC were analyzed and clinical, pathologic data were evaluated based on the Miller and Payne (MP) grading system. The expression of ER, PR and HER2 in the cancers before and after NAC were detected by immunohistochemistry (MaxVision method). SPSS 15.0 software was used to conduct statistical analysis. Results (1) Pathologic responses to the NAC were graded as MP1 (14 cases), MP2 (35 cases), MP3 (106 cases), MP4 (36 cases) and MP5 (18 cases); (2) The expression of ER in core needle biopsy had related negatively to the pathologic response (χ2=33.083, P=0.001). However, the histologic type, grade, ER and PR status, and HER2 expression in surgically-removed specimens had not related to the pathologic response (P>0.05); (3) After NAC, the pathologic type and grade changed in 6.8% (9/132) and 34.9% (30/86) of the cases, and the rates of changes in the expression of ER, PR and HER2 were 42.4% (75/177), 55.4% (98/177) and 26.6% (46/173), respectively. Only the expression of HER2 had significant difference between before and after neoadjuvant chemotherapy (P=0.049). The changes in other data had no relationship with the pathologic response (P>0.05). Conclusions Analysis of core needle biopsy can provide important information to predict the pathologic responses to the NAC. The pathologic appearance, grade, ER, PR and HER2 in breast carcinoma may change after NAC. It is necessary to examine the histologic type, grade and the expression of ER, PR and HER2 after NAC once more.     

Predictive value of CD44 and CD24 for prognosis and chemotherapy response in invasive breast ductal carcinoma

Objective: Cells with unique phenotypes and stem cell-like properties have been found to exist in breast cancer. The aim of the present study was to study the relationship of CD24, CD44, CD44(+)/CD24(-/low) and CD44(-)/CD24(+) tumor phenotypes’ with clinico-pathological features, chemotherapy response and with prognosis. Methods: The study included paraffin-embedded tissues of 140 primary and secondary invasive ductal carcinoma samples. All the patients received routine chemotherapy. Expression of CD24, CD44, ER, PR, and Her2 were assayed immunohistochemically. We applied double-staining immunohistochemistry for the detection of CD44(+)/CD24(-/low), CD44(+)/CD24 (+), CD44(-)/CD24(-) and CD44(-)/CD24(+) cells. The association between the proportions of CD44(+)/CD24(-/low) and CD44(-)/CD24(+) and clinicopathological features, chemotherapy response and with prognosis of these patients was evaluated. Results: CD24 expression was not significantly associated with tumor characteristics, but was significantly associated with poor prognostic variables including ER-, PR-, HER2(+) and triple negative (TN) phenotype; There was no association of CD44 with nodal status, age or HER2 expression. In the correlation analysis, CD24 expression was positively associated with chemotherapy response (P = 0.018), however, CD44 expression was not associated with pathological response to chemotherapy When both markers are considered, the CD44(+)/CD24(-) phenotype had the poor prognosis. The proportion of CD44+/CD24- tumor cells was significantly associated with lymph node involvement, recurrent or metastatic tumors and ER/PR status. High CD44(+)/CD24(-) phenotype had poor response to chemotherapy. The median disease-free survival (DFS) of patients with and without CD44(+)/CD24(-/low) tumor cells were 19.8 ± 2.6 months and 31.7 ± 4.2 months, and the median overall survival (OS) of patients with and without CD44(+)/CD24(-/low) tumor cells were 33.5 ± 2.8 months and 51.4 ± 3.9 months, respectively, and with both univariate and multivariate analyses showing that the proportion of CD44(+)/CD24(-/low) tumor cells was strongly correlated with DFS and OS. However, the CD44(-)/CD24(+), CD44(+)/CD24(+), CD44(-)/CD24 (-) phenotype had no relation with prognosis. Conclusion: There was significant correlation between CD44(+)/CD24(-/low) tumor cell prevalence and tumor metastasis, prognosis and chemotherapy response. The CD44(+)/CD24(-) phenotype may be an important factor for malignant relapse following surgical resection and chemotherapy in patients with invasive ductal carcinoma.     

Tandem high-dose chemotherapy in high-risk primary breast cancer: a multivariate analysis and a matched-pair comparison with standard-dose chemotherapy.

Stem cell-supported high-dose chemotherapy (HDCT) is currently being evaluated in patients with high-risk primary breast cancer (HRPBC), as defined by extensive axillary lymph node involvement. Conclusive results from randomized studies with sufficient patient numbers and follow-up are pending. We retrospectively analyzed 144 HRPBC patients enrolled in a single-arm trial of tandem HDCT at the University of Heidelberg to evaluate the prognostic value of nodal ratio, HER2/neu status, and cytokeratin-positive bone marrow cells and to compare the outcomes of these patients with those of a conventionally treated control group of 91 patients matched by nodal ratio, tumor size, combined hormone-receptor status, and HER2/neu status. The tandem HDCT regimen consisted of 2 cycles of induction chemotherapy followed by 2 cycles of blood stem cell-supported high-dose ifosfamide, 12 g/m2; carboplatin, 900 mg/M2; and epirubicin, 180 mg/m2. Conventionally treated patients received a regimen containing anthracycline without taxanes (52 patients) or CMF (cyclophosphamide, methotrexate, and 5-flurouracil; 39 patients). With a median follow-up of 3.8 years, disease-free, distant disease-free, and overall survival rates were 62%, 65%, and 84%, respectively. In univariate analysis, besides the hormone receptor status (P = .007), HER2/neu overexpression was the strongest predictor of earlier death (P = .017). In multivariate analysis, a nodal ratio of > or =0.8 was found to be the only independent predictor of relapse (relative risk [RR] = 2.09; 95% confidence interval [CI], 1.21-3.60; P = .008) and only the absence of hormone receptors was associated with earlier death (RR = 3.59; 95% CI, 1.45-8.86; P = .006). Despite a trend toward later distant relapse after HDCT compared with standard-dose chemotherapy with a median follow-up of 3 years (P = .059), thus far, matched-pair analysis has not demonstrated significantly better survival rates after HDCT in all matched patients (P = .786) or in the subgroups of anthracycline-treated patients and patients with and without overexpression of HER2/neu. So far, the follow-up time has been too short to draw definite conclusions; however, patients with a nodal ratio of > or =0.8, receptor-negative tumors, or HER2/neu overexpression are at high risk for relapse and death, irrespective of the kind of adjuvant chemotherapy.     

Metronomic chemotherapy in the neoadjuvant setting: results of two parallel feasibility trials (TraQme and TAME) in patients with HER2+ and HER2? locally advanced breast cancer

Neoadjuvant chemotherapy has practical and theoretical advantages over adjuvant chemotherapy strategy in breast cancer (BC) management. Moreover, metronomic delivery has a more favorable toxicity profile. The present study examined the feasibility of neoadjuvant metronomic chemotherapy in two cohorts [HER2+ (TraQme) and HER2− (TAME)] of locally advanced BC. Twenty patients were prospectively enrolled (TraQme, n=9; TAME, n=11). Both cohorts received weekly paclitaxel at 100 mg/m² during 8 weeks followed by weekly doxorubicin at 24 mg/m² for 9 weeks in combination with oral cyclophosphamide at 100 mg/day (fixed dose). The HER2+ cohort received weekly trastuzumab. The study was interrupted because of safety issues. Thirty-six percent of patients in the TAME cohort and all patients from the TraQme cohort had stage III BC. Of note, 33% from the TraQme cohort and 66% from the TAME cohort displayed hormone receptor positivity in tumor tissue. The pathological complete response rates were 55% and 18% among patients enrolled in the TraQme and TAME cohorts, respectively. Patients in the TraQme cohort had more advanced BC stages at diagnosis, higher-grade pathological classification, and more tumors lacking hormone receptor expression, compared to the TAME cohort. The toxicity profile was also different. Two patients in the TraQme cohort developed pneumonitis, and in the TAME cohort we observed more hematological toxicity and hand-foot syndrome. The neoadjuvant metronomic chemotherapy regimen evaluated in this trial was highly effective in achieving a tumor response, especially in the HER2+ cohort. Pneumonitis was a serious, unexpected adverse event observed in this group. Further larger and randomized trials are warranted to evaluate the association between metronomic chemotherapy and trastuzumab treatment.     

Identification of a basal-like subtype of breast ductal carcinoma in situ

Microarray profiling of invasive breast carcinomas has identified subtypes including luminal A, luminal B, HER2-overexpressing, and basal-like. The poor-prognosis, basal-like tumors have been immunohistochemically characterized as estrogen receptor (ER)-negative, HER2/neu-negative, and cytokeratin 5/6-positive and/or epidermal growth factor receptor (EGFR)-positive. The aim of this study was to determine the prevalence of basal-like ductal carcinoma in situ in a population-based series of cases using immunohistochemical surrogates. A total of 245 pure ductal carcinoma in situ cases from a population-based, case-control study were evaluated for histologic characteristics and immunostained for ER, HER2/neu, EGFR, cytokeratin 5/6, p53, and Ki-67. The subtypes were defined as: luminal A (ER+, HER2-), luminal B (ER+, HER2+), HER2 positive (ER-, HER2+), and basal-like (ER-, HER2-, EGFR+, and/or cytokeratin 5/6+). The prevalence of breast cancer subtypes was basal-like (n = 19 [8%]); luminal A, n = 149 (61%); luminal B, n = 23 (9%); and HER2+/ER-, n = 38 (16%). Sixteen tumors (6%) were unclassified (negative for all 4 defining markers). The basal-like subtype was associated with unfavorable prognostic variables including high-grade nuclei (P < .0001), p53 overexpression (P < .0001), and elevated Ki-67 index (P < .0001). These studies demonstrate the presence of a basal-like in situ carcinoma, a potential precursor lesion to invasive basal-like carcinoma.     

Increased sodium-proton antiporter activity in patients with obstructive sleep apnoea

Cytosolic pH (pH(i)) and the activity of the sodium-proton antiporter (Na(+)/H(+) antiporter) were measured in lymphocytes from 22 patients with obstructive sleep apnoea and from 24 age-matched healthy subjects (Controls). The cellular Na(+)/H(+) antiporter was measured spectrophotometrically using a pH-sensitive fluorescent dye after intracellular acidification using sodium propionate. Resting pHi was similar in lymphocytes from patients with obstructive sleep apnoea and from controls (7.36 +/- 0.20, n=22; vs. 7.35 +/- 0.19, n=24; mean +/- SD). The Na(+)/H(+) antiporter activity was significantly higher in patients with obstructive sleep apnoea than in controls (11.87 +/- 3.26 x 10(-3) pH(i)/s vs. 4.38 +/- 1.40 x 10(-3) pH(i)/s; P < 0. 0001). The apparent affinity of the Na+/H+ antiporter was not significantly different between the groups (6.90 +/- 0.23 vs. 6.87 +/- 0.20). In patients with obstructive sleep apnoea the activity of the Na(+)/H(+) antiporter remained stable during the night. The activity of the Na(+)/H(+) antiporter was 13.49 +/- 4.80 x 10(-3) pH(i)/s at 20.00 and 13.26 +/- 6.13 x 10(-3) pH(i)/s at 02.00. From the present results it is concluded that an increased cellular Na(+)/H(+) antiporter activity may be a genetic marker for patients who are predisposed to obstructive sleep apnoea.     

乳腺癌新辅助化疗前后各预测因子的变化

目的：探讨浸润性乳腺癌新辅助化疗前后预测因子的变化规律。方法：应用Pubmed及CNKI数据库系统以“浸润性乳腺癌及新辅助化疗,预测因子”为关键词检索文献,筛选其中近年来包含雌激素受体（ER）、孕激素受体（PR）、人类表皮生长因子2（HER2）、抑癌基因（p53）和增殖细胞核抗原（Ki-67）变化的文献进行分析。结果：关于乳腺癌新辅助化疗前后各预测因子的变化已展开了大量研究,这些研究结果肯定了新辅助前后对预测因子ER、PR、HER2、p53、Ki-67检测对治疗方法的选择及患者预后的重要意义。结论：由于种种原因,对于各预测因子的变化还未能得出一个肯定的变化规律,需要进一步大样本的研究及实验分析。     

Predictive and prognostic factors in locally advanced breast cancer: effect of intratumoral FOXP3+ Tregs

This study aimed to investigate the prognostic and predictive effect of FOXP3+ Tregs together with clinicopathologic factors in locally advanced breast cancer (LABC) patients. The medical records of 101 LABC patients who received neoadjuvant chemotherapy (NAC) between 2005 and 2012 were evaluated retrospectively. The density of intratumoral FOXP3+ lymphocytes in paraffin-embedded tissues was assessed by immunohistochemical analyses in appropriate cases. The relationship with clinicopathologic features, prognosis and chemotherapy response was investigated. HR(–) and HER2(+) tumors tended to have higher pre-chemotherapy Tregs than HR(+) tumors, and significantly higher pathologic complete response (PCR) rates were observed in these patients. Treg decline after NAC was associated with better pathological response rates. Lower intratumoral infiltration of FOXP3+ Tregs after NAC (<3.4/HPF) was significantly associated with higher PCR rates for breast, and close to the significance limit for total (or both for breast and axillary) PCR rates (PCR for breast: 25 vs. 2.9 % for low vs. high Treg, p = 0.001; PCR for breast + axillary tissue: 13.9 vs. 0 %, p = 0.05). Despite better PCR rates, patients with high intratumoral Treg infiltrates (≥11.5/HPF) before chemotherapy had significantly shorter overall survival than patients with low Treg infiltrates (<11.5/HPF). Cox multivariate regression analyses demonstrated that the density of Treg infiltration before chemotherapy was the strongest predictor for survival. This study established the predictive and prognostic effect of intratumoral FOXP3+ Tregs in LABC patients. To predict clinical outcome, evaluation of FOXP3+ Tregs in tumoral tissues before and after NAC should be considered for these high-risk patients.     

昼夜节律蛋白Cry2作为结直肠癌预后和化疗敏感性预测因子的研究

目的:检测Cry2在结直肠癌组织中的表达,探讨Cry2在结直肠癌预后和辅助化疗敏感性预测中的作用。方法:收集16名规律接受包含5-氟尿嘧啶的新辅助化疗的结直肠癌切除术患者,使用实体肿瘤疗效评估标准评估化疗疗效,并根据疗效分为完全反应(CR)/部分反应(PR)和病灶稳定(SD)/病灶进展(PD)组,免疫组化检测Cry2表达,评分后进行双侧x²检验。收集307名在2001年至2005年间接受结直肠癌切除术患者的肿瘤组织,制作组织芯片,免疫组化检测Cry2表达,用log-rank法进行单因素分析,用Cox回归模型进行多因素分析,用Kaplan-Meier法绘制生存曲线。结果:CR/PR组Cry2表达较低,SD/PD组Cry2表达较高,差异显著(P<0.05)。307例中Cry2高表达者84人,平均生存83.46月,Cry2低表达者223人,平均生存100.10月,两组间差异显著(P<0.05)。Cry2低表达者预后较好。307例结直肠癌切除术患者Cry2表达高低风险比为1.70,其95%置信区间为1.09~2.66,差异显著(P<0.05),Cry2的表达是影响预后的独立因素。16例接受化疗的结直肠癌切除术患者Cry2表达高低风险比为2.88,其95%置信区间为1.03~8.06,差异显著(P<0.05),Cry2的表达是影响化疗预后的独立因素。结论:根据Cry2表达高低可预测结直肠癌新辅助化疗的疗效,也可预测结直肠癌患者的预后。     

Lymphocyte-rich gastric cancer: associations with Epstein-Barr virus, microsatellite instability, histology, and survival.

Lymphocyte-rich gastric carcinomas may have a better prognosis than cancers without a pronounced host inflammatory response. Two subsets of gastric cancer-Epstein-Barr virus-positive and microsatellite instability high-have been associated with a lymphocyte-rich phenotype. We assessed relationships between tumor-infiltrating lymphocytes, Epstein-Barr virus status, microsatellite instability status, and cancer-specific survival in 110 resected gastric cancers. Seven patients had Epstein-Barr virus-positive cancer, including 4 (3.7%) of 107 consecutive patients. Tumors from 17 patients (16%) were designated microsatellite instability high on the basis of negative immunohistochemical staining for MLH1; all tumors had intact expression of MSH2 and MSH6. Epstein-Barr virus-positive cancers had increased tumor-infiltrating lymphocytes compared with Epstein-Barr virus-negative cancers (median 450/10 HPF versus 21/10 HPF, P <.001). Microsatellite instability-high cancers also had increased tumor-infiltrating lymphocytes compared with non-microsatellite instability-high cancers (median 150/10 HPF versus 20/HPF, P <.001). Microsatellite instability-high cancers affected older patients and were more likely to be intestinal in the Lauren classification and expanding in the Ming classification. By univariate analysis, decreased risk of death from gastric cancer was significantly associated with low tumor stage, expanding growth pattern, increasing tumor-infiltrating lymphocyte count, and microsatellite instability-high status. High tumor-infiltrating lymphocyte count and microsatellite instability-high status retained statistical significance as favorable prognostic factors after adjustment for tumor stage in multivariate analysis. Tumor-infiltrating lymphocyte count retained statistical significance as a favorable prognostic factor after adjustment for microsatellite instability-high status; but microsatellite instability-high status did not remain a significant independent prognosticator after adjustment for tumor-infiltrating lymphocyte count. The association between microsatellite instability-high cancers and high tumor-infiltrating lymphocyte counts may account for the association of microsatellite instability-high gastric cancers with improved survival.     

Identification of HER-2/neu immunogenic epitopes presented by renal cell carcinoma and other human epithelial tumors.

HER-2/neu is a tumor-associated antigen overexpressed in a large variety of human tumors. Eight HER-2/neu peptides displaying HLA-A*0201 anchoring motifs were selected and tested for their binding affinity to HLA-A*0201 and their capacity to elicit cytotoxic T lymphocyte (CTL) responses in both HLA-A*0201 transgenic mice and in HLA-A*0201(+) healthy donors. Two high-affinity (p5 and p48) and one intermediate-affinity (p1023) peptides triggered CTL responses in both transgenic mice and humans, comparable to those observed for the well-known HER2/neu dominant peptide p369. CTL induced in transgenic mice lysed HLA-A*0201(+) RMA cells infected with recombinant HER-2/neu but not cells infected with wild-type vaccinia virus. Human CTL lysed HLA-A*0201(+) HER-2/neu(+) tumor cells of different origins (breast, colon, lung and renal cancer) irrespective of the expression levels of HER-2/neu. Importantly, primed CTL specific for these epitopes were detected in freshly isolated tumor-infiltrating lymphocytes from three renal cell carcinoma patients. Therefore, the HER-2/neu peptides p5, p48 and p1023 may be good candidates for immunotherapy of a broad spectrum of tumors, including renal cell carcinoma.     

Analysis of the relevance between molecular subtypes and efficacy of neoadjuvant chemotherapy in breast cancer as well as its prognostic factors

Background

Accurate pathological assessment of breast specimens after NACT is crucial. It is beneficial to determine the treatment efficacy and predict prognosis. So we should explore the relevance between molecular subtypes and efficacy of neoadjuvant chemotherapy in breast cancer as well as its prognostic factors, which about survival analysis and disease free survival involved, which was one part of contributing for evaluating in terms of global survival and disease free survival.

Expression of Inhibitory Receptors in Natural Killer (CD3CD56) Cells and CD3CD56 Cells in the Peripheral Blood Lymphocytes and Tumor Infiltrating Lymphocytes in Patients with Primary Hepatocellular Carcinoma

The cytolytic responses of NK (CD3(-)CD56(+)) and CD3(+)CD56(+) cells are inhibited by the engagement of the killer inhibitory receptors (p58.1, p58.2, and CD94) with respective ligands on the target cell. The expression of these receptors in peripheral blood lymphocytes (PBLs) (n = 18) and tumor infiltrating lymphocytes (TILs) (n = 7) was examined in patients with primary hepatocellular carcinoma (HCC). There were no differences in the expression of the three inhibitory receptors by both NK and CD3(+)CD56(+) PBLs in patients with HCC compared to that of control NK and CD3(+)CD56(+) PBLs, respectively (all P = NS). However, the expression of p58.1 by NK TILs and by CD3(+)CD56(+) TILs in patients with HCC was significantly decreased compared to that of hepatic lymphocytes of the control subjects (8.9% vs 37.85%, P = 0.047; 4.1% vs 25.2%, P = 0.049, respectively). The expression of p58.2 by CD3(+)CD56(+) TILs and CD94 by NK TILs was also decreased compared to that of hepatic lymphocytes of the control subjects (16.9% vs 73.1%, P = 0.047; 21% vs 49.95%, P = 0.037, respectively). These changes were limited to hepatic TILs, and this observation may reflect an adaptive anti-tumor phenomenon occurring in the microenvironment of HCC.     

Flow cytometric determination of cellular sources and frequencies of key cytokine-producing lymphocytes directed against recombinant LACK and soluble Leishmania antigen in human cutaneous leishmaniasis

Leishmaniasis, caused by infection with the protozoan parasite Leishmania, affects millions of individuals worldwide, causing serious morbidity and mortality. This study directly determined the frequency of cells producing key immunoregulatory cytokines in response to the recombinant antigen Leishmania homolog of receptors for activated kinase C (LACK) and soluble leishmania antigen (SLA), and it determined relative contributions of these antigens to the overall cytokine profile in individuals infected for the first time with Leishmania braziliensis. All individuals presented with the cutaneous clinical form of leishmaniasis and were analyzed for proliferative responses to LACK antigen and SLA, frequency of lymphocyte subpopulations (analyzed ex vivo), and antigen-induced (LACK and SLA) cytokine production at the single-cell level (determined by flow cytometry). The following were determined. (i) The Th1-type response previously seen in patients with cutaneous leishmaniasis is due to gamma interferon (IFN-gamma) production by several different sources, listed in order of contribution: CD4(+) T lymphocytes, CD4(-), CD8(-) lymphocytes, and CD8(+) T lymphocytes. (ii) SLA induced a higher frequency of lymphocytes producing IFN-gamma and tumor necrosis factor alpha (TNF-alpha) than did LACK. (iii) LACK induced an activation of monocyte populations as reflected by an increased percentage of CD14-positive cells. (iv) Neither SLA nor LACK induced detectable frequencies of cells producing interleukin-4 (IL-4) or IL-5. These data demonstrated a multifaceted immune response to SLA in human leishmaniasis involving Th1 CD4(+) T lymphocytes (IFN-gamma(+) and IL-10(-)/IL-4(-)), Tc1 CD8(+) T cells (IFN-gamma(+), and IL-10(-)/IL-4(-)), and a high frequency of TNF-alpha-producing lymphocytes. Moreover, it was determined that the recombinant antigen LACK acts as a weak inducer of Th1-type lymphocyte responses compared to SLA.     

Aldehyde dehydrogenase 1A1 expression in breast cancer is associated with stage, triple negativity, and outcome to neoadjuvant chemotherapy

Studies have shown that ALDH1A1 expression in the breast is associated with worse clinical outcome. ALDH1A1 inactivates cyclophosphamide, which is an integral agent in breast cancer chemotherapy regimens. The purposes of this study were to verify these results, to correlate ALDH1A1 expression with clinical outcome in patients treated with cyclophosphamide as part of the chemotherapy (adjuvant or neoadjuvant), and to evaluate ALDH1A1 as a useful marker to predict the clinical outcome of breast cancer subsets. A total of 513 primary breast cancers were studied. Tissue microarrays of the studied cases were stained with ALDH1A1. Key clinicopathological information was obtained. Disease-free survival and overall survival were calculated. Patients with neoadjuvant therapy who had substantial residual cancer burden (RCB) were included in the study. Fisher's exact test and Kaplan-Meier methods were used for statistical analysis. ALDH1A1 was expressed in 53 (10%) patients, with a higher frequency in triple negative, followed by HER2+, and finally hormonal receptor+/HER2- (P<0.0001). Tumors with advanced stage, node-positive, or larger tumor size were correlated with ALDH1A1 expression (P=0.006, P<0.0001, and P=0.05, respectively). ALDH1A1 expression was also correlated with worse disease-free survival (P<0.006) and overall survival (P<0.01) in patients who were treated with neoadjuvant chemotherapy. In all, 8 of 22 (36%) received neoadjuvant chemotherapy and died of disease-expressed ALDH1A1 (P=0.008). Similarly, 8 of 23 (35%) who received neoadjuvant chemotherapy and had tumor recurrence expressed this marker (P=0.002). The risk of recurrence was fivefold greater than negative ALDH1A1 tumors. The risk of recurrence became 11-fold greater when cyclophosphamide but not trastuzumab was part of the regimen. Our results are consistent with previous studies. Moreover, we found that ALDH1A1 could be a useful marker to predict worse clinical outcome after chemotherapy in the neoadjuvant setting with substantial RCB. However, a larger cohort is required to verify our results.     

乳腺癌新辅助化疗后临床评价与病理评价的差异分析

目的 探讨乳腺癌新辅助化疗后,临床疗效评价与病理评价之间存在差异的病理学基础.方法 收集中国医学科学院肿瘤医院2005年6月至2007年12月施行乳腺癌新辅助化疗的209例.新辅助化疗前均行核芯针穿刺活检.化疗结束后4周内实施乳腺癌根治术.新辅助化疗前后均对乳腺原发灶进行临床体检、乳腺X线检查和(或)超声检查.实施新辅助化疗后,依实体瘤的疗效评价标准(RECIST,1.1版)对乳腺癌原发灶进行临床疗效评价,依Miller和Payne(MP)分级系统进行病理评价.应用SPSS 15.0软件分析临床评价与病理评价的相关性.结果 (1)新辅助化疗后依临床体检结果进行临床评价:完全缓解33例,部分缓解124例,疾病稳定41例,疾病进展11例.(2)新辅助化疗前后均行乳腺X线检查87例,依乳腺X线检查进行临床评价:完全缓解8例,部分缓解42例,疾病稳定37例.(3)新辅助化疗后MP分级病理评价:1级14例,2级35例,3级106例,4级36例,5级18例.(4)临床体检相关的临床评价与病理评价存在统计学相关性(x2=33.668,P=0.001),乳腺X线检查相关的临床评价与病理评价存在统计学相关性(x2=22.404,P=0.004).(5)新辅助化疗病理评价与X线检查相关临床评价存在差异的病理学改变有:残存浸润癌以脉管瘤栓为主要表现形式;伴有大片黏液湖形成的黏液腺癌;导管内癌残存,伴明显沙砾样钙化及周围组织的沙砾样钙化;间质结节状纤维化等.结论 乳腺癌新辅助化疗的临床评价与病理评价存在统计学相关性.两者之间的差异有相应的病理学基础.伴有大片黏液湖形成的黏液腺癌、导管内癌的残存伴沙砾样钙化及间质结节状纤维化可能是临床评价低估治疗疗效的原因之一;而残存癌表现为脉管瘤栓可能是临床评价高估治疗疗效的原因之一.     

Combination of lentivector immunization and low-dose chemotherapy or PD-1/PD-L1 blocking primes self-reactive T cells and induces anti-tumor immunity

In the last two decades, anti-cancer vaccines have yielded disappointing clinical results despite the fact that high numbers of self/tumor-specific T cells can be elicited in immunized patients. Understanding the reasons behind this lack of efficacy is critical in order to design better treatment regimes. Recombinant lentivectors (rLVs) have been successfully used to induce antigen-specific T cells to foreign or mutated tumor antigens. Here, we show that rLV expressing a murine nonmutated self/tumor antigen efficiently primes large numbers of self/tumor-specific CD8(+) T cells. In spite of the large number of tumor-specific T cells, however, no anti-tumor activity could be measured in a therapeutic setting, in mice vaccinated with rLV. Accumulating evidence shows that, in the presence of malignancies, inhibition of T-cell activity may predominate overstimulation. Analysis of tumor-infiltrating lymphocytes revealed that specific anti-tumor CD8(+) T cells fail to produce cytokines and express high levels of inhibitory receptors such as programmed death (PD)-1. Association of active immunization with chemotherapy or antibodies that block inhibitory pathways often leads to better anti-tumor effects. We show here that combining rLV vaccination with either cyclophosphamide or PD-1 and PD-L1 blocking antibodies enhances rLV vaccination efficacy and improves anti-tumor immunity.     

HER2/<Emphasis Type="Italic">neu</Emphasis>, p53, Ki67, and hormone receptors do not change during neoadjuvant chemotherapy in breast cancer

The selection of a systemic breast cancer therapy is based on the expression pattern of immunohistochemical prognostic markers. In our study we sought to determine whether neoadjuvant chemotherapy may alter these expression patterns within the tumors. Our hypothesis was that the expression of the immunohistochemical prognostic markers does not differ between tissue specimens before and after neoadjuvant chemotherapy. We determined the protein expression levels of estrogen receptor, progesterone receptor, Ki67, p53 and HER2/neu in the core biopsy and the resected tumor sample from 25 patients receiving neoadjuvant chemotherapy. As a control group, we analyzed sample pairs from 30 patients who did not receive neoadjuvant chemotherapy. Additionally, we determined the relative HER2/neu gene copy number by FISH and/or real-time PCR. There were no significant differences in the changes in expression patterns from the core biopsy to the treated resected tumor between those who had received neoadjuvant chemotherapy and the control group. We suggest that it is sufficient to analyze the prognostic factors from either the core biopsy prior to chemotherapy or the treated tumor sample instead of investigating both samples. This would markedly reduce the costs.