Clinical relevance of circulating CK-19 mRNA-positive cells detected during the adjuvant tamoxifen treatment in patients with early breast cancer.

BACKGROUND: The purpose of this study was to evaluate the effect of adjuvant treatment with tamoxifen on the CK-19 mRNA+ cells in patients with early-stage breast cancer. PATIENTS AND METHODS: CK-19 mRNA+ cells were prospectively and longitudinally detected using a specific real-time PCR assay for CK-19 mRNA in 119 patients with estrogen and/or progesterone receptor-positive tumors during the period of tamoxifen administration. RESULTS: Twenty-two (18.5%) patients had detectable CK-19 mRNA+ cells after the completion of adjuvant chemotherapy and in 15 (68.2%) of them adjuvant tamoxifen could not eliminate these cells (persistently positive). In 68 (57.1%) patients, no CK-19 mRNA+ cells could be detected throughout the follow-up period (persistently negative). Seven (46.7%) of the 15 persistently positive and six (8.8%) of the 68 persistently negative patients developed disease recurrence (P = 0.00026). Persistency of CK-19 mRNA+ cells was associated with a significantly lower median disease-free interval (P = 0.0001) and overall survival (P = 0.0005). Multivariate analysis revealed that the detection of CK-19 mRNA+ cells during the administration of tamoxifen was associated with an increased risk of relapse [hazard ratio (HR) = 22.318, P = 0.00006] and death (HR = 13.954, P < 0.00001). CONCLUSIONS: The detection of CK-19 mRNA+ cells throughout the period of adjuvant tamoxifen treatment is an independent poor prognostic factor in patients with early breast cancer.     

Assessment of the effect of surgery on the kinetics of circulating tumour cells in patients with operable breast cancer based on cytokeratin-19 mRNA detection

Aims

The aim of this study was to evaluate the effect of surgery on the kinetics of CTCs in breast cancer patients.

Methods

The detection of CK-19 mRNA-positive CTCs in the blood by RT-PCR was analysed in 104 stage 0-IIIA patients at 4 time-points: prior to surgery, upon completion, 24 h after surgery and 15 days after surgery. Furthermore, a late sample was assessed prior to initiation of adjuvant chemotherapy in a subgroup of 53 patients. As negative controls, peripheral blood was obtained from 50 female patients undergoing excision of benign breast lesions and from 11 female patients receiving surgery for early-stage colorectal cancer.

Results

A significant percentage of blood samples from breast cancer patients (14.4%) were negative for CK-19 preoperatively but turned transiently positive early postoperatively. However, no significant difference in CK-19 mRNA detection was noted among the first 4 examined time-points. There was no significant correlation between CK-19 mRNA-positive cells and classic prognostic factors. A significant increase in CK-19 mRNA-positivity (32.1%) was observed in a late sample of the subgroup of 53 patients before adjuvant chemotherapy after a median of 54 days, postoperatively.

Conclusions

Surgery may result in CTC detection in a small proportion of early breast cancer patients. There is no clear correlation to indicate which patients are expected to have detectable CTCs. Although CTCs are detected in a small proportion of patients during the perioperative period, the detection rate may increase over time and with longer follow-up.     

Intraoperative examination of sentinel lymph nodes by ultrarapid immunohistochemistry in breast cancer

BACKGROUND: The ultrarapid immunohistochemistry (IHC) technique was applied to the intraoperative examination of sentinel lymph nodes (SLNs) because routine SLN frozen section examinations sometimes produce false-negative results. The present study was undertaken to develop a reliable protocol for the ultrarapid IHC of SLNs. METHODS: SLNs from 79 breast cancer patients with clinically negative axillary node were examined intraoperatively by frozen hematoxylin-eosin (H&E) stain and by ultrarapid cytokeratin IHC assay. On the basis of the result of serially sectioned permanent study, the sensitivity and accuracy of each intraoperative technique were compared. RESULTS: The total number of dissected SLNs was 178 with a mean of 2.3 (1-5) per patient. The mean turnaround time for ultrarapid IHC was 20 min. The sensitivity rates of frozen H&E staining and ultrarapid IHC were 70.0 and 85.0%, respectively (P = 0.083). Each method had a specificity of 100%. The accuracy rates for frozen H&E staining and rapid IHC were 92.4 and 96.2%, respectively (P = 0.083). Ultrarapid IHC detected one additional patient with sentinel node micrometastasis and two additional patients with isolated tumor cells (ITCs). In those patients, two underwent completion axillary dissection simultaneously and could avoid a second operation. CONCLUSIONS: Ultrarapid cytokeratin IHC enhanced the intraoperative detection of sentinel node micrometastasis and ITCs in breast cancer without consuming much time. In patients who need completion axillary dissection after sentinel node biopsy, this technique could be helpful in avoiding a second operation.     

Using one-step nucleic acid amplification (OSNA) for intraoperative detection of lymph node metastasis in breast cancer patients avoids second surgery and accelerates initiation of adjuvant therapy

BackgroundSentinel lymph node (SLN) analysis is conventionally analyzed using immunohistochemistry and in the case of SLN involvement, justifies a second surgery for axillary lymph node (ALN) resection, thus delaying the initiation of adjuvant therapies.Patients and methodsThree hundred and eighty-one patients with early stage breast cancer (BC) were considered in this retrospective study. SLNs were detected using combined radioisotope and dye detection. SLN involvement was analyzed using routine intraoperative One-Step Nucleic Acid Amplification (OSNA) assay, in 100 patients and compared with the conventional histopathology carried out previously in 281 patients.ResultsConsidering positive SLNs as ‘++’ (CK19 mRNA copy number>5000), ‘+’ (250 < CK19 mRNA copy number <5000) and positive by inhibition in the OSNA group and macro-, micrometastases and isolated tumor cells in the histopathology group, no difference in SLN involvement rate was found between the two groups with 29.0% and 29.9% of positive SLNs, respectively. Using OSNA intraoperatively, the mean time to process the SLN was 42 min allowing immediate ALN resection, reduced significantly (P < 0.01) the re-intervention rate (9% versus 39%) and significantly (P < 0.01) accelerated the initiation of adjuvant therapy (6.2 versus 8.4 weeks).ConclusionsUsing OSNA for intraoperative SLN analysis avoids second surgery for ALN resection in most patients and accelerates initiation of adjuvant therapy.     

Prospective study found thatperipheral lymph node sampling reduced the false-negative rate ofsentinel lymph node biopsy forbreast cancer

Background:Although sentinel lymph node biopsy (SLNB) can accurately predict the status of axillary lymph node (ALN) metastasis, the high false?negative rate (FNR) of SLNB is still the main obstacle for the treatment of patients who receive SLNB instead of ALN dissection (ALND). The purpose of this study was to evaluate the clinical signiifcance of SLNB combined with peripheral lymph node (PLN) sampling for reducing the FNR for breast cancer and to discuss the effect of “skip metastasis” on the FNR of SLNB. Methods:At Shandong Cancer Hospital Affliated to Shandong University between March 1, 2012 and June 30, 2015, the sentinel lymph nodes (SLNs) of 596 patients with breast cancer were examined using radiocolloids with blue dye tracer. First, the SLNs were removed; then, the area surrounding the original SLNs was selected, and the visible lymph nodes in a ifeld of 3–5cm in diameter around the center (i.e., PLNs) were removed, avoiding damage to the structure of the breast. Finally, ALND was performed. The SLNs, PLNs, and remaining ALNs underwent pathologic examination, and the relationship between them was analyzed. Results:The identiifcation rate of SLNs in the 596 patients was 95.1% (567/596); the metastasis rate of ALNs was 33.7% (191/567); the FNR of pure SLNB was 9.9% (19/191); and after the SLNs and PLNs were eliminated, the FNR was 4.2% (8/191), which was signiifcantly decreased compared with the FNR before removal of PLNs (P=0.028). According to the detected number (N) of SLNs, the patients were divided into four groups of N=1, 2, 3, and≥4; the FNR in these groups was 19.6, 9.8, 7.3, and 2.3%, respectively. For the patients with≤2 or≤3 detected SLNs, the FNR after removal of PLNs was signiifcantly decreased compared with that before removal of PLNs (N≤2: 14.0% vs. 4.7%, P=0.019; N≤3: 12.2% vs. 4.7%,P=0.021), whereas for patients with≥4 detected SLNs, the decrease in FNR was not statistically signiifcant (P=1.000). In the entire cohorts, the “skip metastasis” rate was 2.5% (15/596); the FNR caused by “skip metastasis” was 2.1% (4/191). Conclusions:The FNR of SLNB was associated with the number of SLNs. For patients with≤3 detected SLNs, PLN sampling can reduce the FNR of SLNB to an acceptable level of less than 5%. Because of the existence of the “skip metastasis” and distinct metastasis patterns, the FNR of SLNB cannot be completely eliminated.     

Peripheral blood circulating cytokeratin-19 mRNA-positive cells after the completion of adjuvant chemotherapy in patients with operable breast cancer.

BACKGROUND: The purpose of this study was to evaluate the prognostic significance of the molecular detection of cytokeratin 19 (CK-19) mRNA-positive cells in the peripheral blood of women with operable breast cancer after the completion of adjuvant chemotherapy. PATIENTS AND METHODS: Blood from 161 patients with stage I and II breast cancer, obtained after the completion of adjuvant chemotherapy, was tested by nested RT-PCR for CK-19 mRNA detection. Using univariate and multivariate analyses possible interactions with other prognostic factors and association of CK-19 mRNA detection with risk of relapse, disease-free interval (DFI) and overall survival were investigated. RESULTS: After completion of adjuvant chemotherapy, 27.3% of patients had peripheral blood CK-19 mRNA-positive cells; there was no association of this finding with any other prognostic factors or the type of chemotherapy regimen used. For patients with less than four involved axillary lymph nodes the risk of relapse was 3.81 [95% confidence interval (CI) 1.06-13.71] times higher, and the DFI was significantly reduced (P = 0.028) if CK-19 mRNA-positive cells were detectable in the blood after the completion of adjuvant chemotherapy. In contrast, for patients with four or more involved lymph nodes, the presence of CK-19 mRNA-positive cells after adjuvant chemotherapy did not significantly affect the risk of relapse or DFI. Furthermore, the risk of relapse was higher (hazards ratio 3.70; 95% CI 1.09-13.89) and the DFI was reduced (P = 0.022) for patients with detectable CK-19 mRNA-positive cells following adjuvant cyclophosphamide, methotrexate and 5-fluorouracil (CMF) as compared with epirubicin, cyclophosphamide and 5-fluorouracil (FEC) or sequential taxotere-epirubicin and cyclophosphamide (T/EC) chemotherapy. CONCLUSIONS: The detection of CK-19 mRNA-positive cells in the peripheral blood after adjuvant chemotherapy may be of clinical relevance for patients with early breast cancer and less than four involved axillary lymph nodes.     

Detection of breast cancer micrometastases in bone marrow using reverse-transcriptase chain reaction and southern hybridization

Thirty-flveto40%ofallpatientswithbreastcarcinoma,includingupt024%ofpatientswithn0evidence0fmetastasesatthetimeofdiagnosis,willrelapseafterprimarytherapy.l"']Themostreliableprognosticindicesofaxillarylymphnodestatusandsizeofprimarytumorcannotpredictinwh0mthediseasewillrecur.Bonemarrowisafrequentandreadilyaccessiblesiteofmetastases.Inup8O%ofpatients,therelaPsedevelopsstariingfrombonemarrowmetastasesats0mepointinthepr0cessoftheirillness.l3]Currentmethodstodetectboneinvolvement,suchasX-rayandbon…     

Sentinel lymph node biopsy after neoadjuvant chemotherapy predicts pathological axillary lymph node status in breast cancer patients with clinically positive axillary lymph nodes at presentation

Background

It is still controversial whether axillary lymph node (ALN) dissection (ALND) can be omitted after negative sentinel lymph node (SLN) biopsy (SLNB) in breast cancer (BC) patients with clinically positive ALNs at presentation treated with neoadjuvant chemotherapy (NAC). The study aim was to analyze whether SLNB could be useful in these patients.

Methods

In a retrospective study, eligible patients were women with invasive BC with clinically positive ALNs at presentation, treated with NAC then a total or partial mastectomy, with an intraoperative histological examination of SLNs and non-SLNs suspicious for metastasis followed by ALND. Non-SLNs suspicious for metastasis were defined as hard or large nodes located in the same level of the axilla where clinically positive ALNs had been initially identified. The results of SLNB and clinicopathological characteristics were analyzed for correlation with pathological ALN status.

Results

In a consecutive series of 105 women with 107 BC cases, 81 (75.7 %) had at least 1 SLN, and the remaining 26 (24.3 %) had at least 1 non-SLN suspicious for metastasis. The intraoperative (or final) histological examination of these nodes revealed that the false-negative (FN) rate and accuracy were 8.2 (or 6.3) % and 95.1 (or 96.3) %, respectively. Estrogen receptor status at presentation, pathological tumor response, lymphovascular invasion after NAC, and NAC regimen were correlated with pathological ALN status.

Conclusion

The histological examination of SLNs and that of non-SLNs suspicious for metastasis are useful for predicting pathological ALN status in BC patients with clinically positive ALNs at presentation who are treated with NAC.     

乳腺癌前哨淋巴结临床病理的研究

目的：探讨乳腺癌前哨淋巴结能否准确反映腋窝淋巴结转移情况,方法对36例例早期乳腺癌,常规HE染色检测腋窝淋巴结,对HE染色阴性的前哨淋巴结及所属非前啉巴结,连续切片、免疫组化染色,检测其肿瘤的转移情况,结果常规病理检测36例中,10例腋窝有转移,其中2例仅哨淋巴结有肿瘤转移,连续切片,组化检测26例阴性的前哨淋巴结共61例,非前哨淋巴结406枚,2例（2枚）仅前哨淋巴结发现有肿瘤转移,余245例前哨淋巴结及非前哨淋巴结无肿瘤转移,HE染色的检测阴性的前哨淋巴结患者假阴性率为7．7％（2／26）。结论早期腺癌前哨淋巴结可以准确反映腋窝巴结转移情况。     

Clinical outcome of patients with lymph node-negative breast carcinoma who have sentinel lymph node micrometastases detected by immunohistochemistry

BACKGROUND: The ideal pathologic assessment of sentinel lymph nodes (SLNs) in patients with breast carcinoma remains controversial. The authors evaluated how detailed assessment of SLNs using immunohistochemistry (IHC) and serial sectioning would affect treatment decisions and outcomes in patients with breast carcinoma who had negative SLNs on standard hematoxylin and eosin staining. METHODS: The SLNs from patients who were treated between June 1998 and June, 1999 and who had negative lymph node status determined by hematoxylin and eosin staining (n = 84 patients) were evaluated further with serial sectioning and cytokeratin IHC. Patients were offered adjuvant therapy based on primary tumor factors. RESULTS: The median patient age was 57 years, and the median tumor size was 1.2 cm. At a median follow-up of 40.2 months, 81 patients (96%) were alive with no evidence of disease, 1 patient was alive with disease, 1 patient had died of disease, and 1 patient had died of other causes. Fifteen patients (18%) had micrometastases identified on IHC. Of the total 84 patients, information regarding adjuvant therapy was not available for 5 patients. Of the remaining 79 patients, 10 patients (13%) were not offered adjuvant chemotherapy but had positive SLN status determined by IHC. SLN status based on IHC evaluation did not correlate with age (P = 0.077), tumor size (P = 0.717), grade (P = 0.148), estrogen receptor status (P = 1.000), or lymphovascular invasion (P = 0.274). Furthermore, IHC-detected positive SLN status did not correlate with distant metastasis (P = 0.372) or overall or distant metastasis-free survival (P = 0.543 and P = 0.540, respectively). CONCLUSIONS: Although the finding of SLN micrometastases by IHC may change management in > 12% of patients, preliminary results suggested that such micrometastases do not affect outcomes significantly.     

乳腺癌前哨淋巴结活检40例报告

目的 :通过对乳腺癌前哨淋巴结 (SLN)活检 ,探讨其对腋淋巴结转移情况预测的准确性。方法 :采用亚甲蓝染料法对 4 0例乳腺癌行腋窝蓝染淋巴结活检 ,后行常规腋窝淋巴结清除 (ALND) ,两标本均送病理检查。结果 :全组 4 0例患者检出SLN 38例 ,2例未找到SLN ,检出率为 95 % ( 38/ 4 0 ) ,有 8例SLN为阳性 ,1例SLN为假阴性 ,腋窝淋巴结 (ALN)有 9例转移 ,SLN与ALN病理检查完全符合者 37例 ,准确率为 92 5 % ( 37/ 38) ;灵敏度为 88 9% ( 8/ 9) ;假阴性率为 11 1% ( 1/ 9)。结论 :亚甲蓝染色法能准确地鉴别SLN及预测乳腺癌腋窝淋巴结状态     

The value of immunohistochemistry in sentinel lymph node histopathology in breast cancer

The optimal protocol for the histopathological examination of sentinel lymph nodes (SLNs) in breast cancer has not been determined. The value of more detailed examination using immunohistochemistry (IHC) is controversial. A total of 476 SLNs from 216 patients were reviewed. Sentinel lymph nodes were sectioned at three levels at 100 mum intervals and stained with haematoxylin and eosin (H&E). If the H&E sections showed no evidence of metastasis, then the three serial sections were stained with a murine monoclonal anti-cytokeratin antibody (CAM 5.2). Metastatic deposits were classified as macrometastasis (> 2.0 mm), micrometastasis (0.2-2.0 mm) or isolated tumour cells (ITC, < 0.2 mm). Of the 216 patients, 56 (26%) had metastasis as identified by H&E. Immunohistochemistry detected metastatic deposits in a further nine patients (4%), of whom four (2%) had micrometastasis and five (2%) had ITC only. Those cases with micrometastases were all, on review, visible on the H&E sections. Immunohistochemistry detects only a small proportion of metastasis in SLNs. All metastatic deposits identified by IHC were either micrometastasis or ITC. Until the prognostic significance of these deposits has been determined, IHC may be of limited value in the histopathological examination of SLNs.     

应用RT-PCR检测乳腺癌患者外周血CK-19mRNA的临床意义

目的:探索取材于乳腺癌患者外周血诊断微转移的方法。方法:66例乳腺癌和37例乳腺良性疾病患者的外周血,分离其有核细胞后进行细胞总RNA的抽提,运用RT-PCR技术进行CK-19 mRNA的检测。结果:以RT-PCR终产物出现460bp条带定为阳性。66例乳腺癌患者CK-19mRNA有24例表达,总阳性率为36.36％。37例乳腺良性疾病者CK-19mRNA无一例表达。两组有统计学意义(x2=17.54,P<0.001)。结论:RT-PCR检测CK-19 mRNA可以用于临床诊断乳腺癌患者外周血中微转移。     

应用RT-PCR检测乳腺癌患者外周血CEA mRNA和CK-19 mRNA的研究及意义

目的：检测乳腺癌患者外周血中CEA mRNA和CK-19 mRNA的表达，建立取材于外周血早期诊断乳腺癌微转移的方法。方法：65例乳腺癌和37例乳腺良性疾病患者的外周血，分离其有核细胞后进行细胞总RNA的抽提，运用果式RT-PCR技术进行CEA mRNA和CK-19 mRNA的检测。结果：以     

hMAM和CK19 mRNA联合检测早期乳腺癌循环肿瘤细胞的临床价值

目的:评价hMAM和CK19 mRNA联合检测早期乳腺癌循环肿瘤细胞的临床价值.方法:采用RT-PCR检测早期乳腺癌患者CK19、hMAM mRNA阳性循环肿瘤细胞.结果:hMAM和CK19 mRNA联合检测早期乳腺癌患者循环肿瘤细胞阳性率(52.0%)均高于良性乳腺疾病患者(16.7%)和健康体检者(5.0%), P值分别为0.004和0.000;阳性率与癌组织HER-2过表达相关,P=0.049.联合检测的敏感度为57.5%,特异度为88.6%.26例联合检测阳性患者,13例随访期出现转移复发,P=0.001;中位无瘤生存期明显降低,P=0.000.结论:hMAM和CK19 mRNA是一组敏感度和特异度较好的诊断早期乳腺癌患者循环肿瘤细胞的基因标志,可能作为早期乳腺癌术后监测转移复发辅助指标.