The Relationship Between Circulating Tumor Cell And The Characteristics Of Primary Tumor And The Response To Neoadjuvant Chemotherapy In Patients With Breast Cancer: Preliminary Results Of A Plot Study

Breast cancer is the most common malignant tumor in women. Owing to the development of combined therapy, the disease-free survival and overall survival of breast cancer have been rising obviously. However, recurrence and metastasis are still the prime causes of patients'deaths, in which micrometastasis plays an extremely important role. Circulating tumor cell (CTC), micrometastasis in peripheral blood, is defined as the presence of tumor cell in peripheral blood that is not detectable by the current routine clinicopathological and imaging methods. Recent clinical researches reveal that detection of CTC plays a vital role in the management of breast cancer patients. It's indicated that CTC≥5/7.5ml for metastatic breast cancer patients and the presence of CTC for non-metastatic patients newly diagnosed were associated with poor outcome. In addition, CTC increasing to or remaining in≥5/7.5ml after the first cycle of chemotherapy for the metastatic setting suggests the current treatment was ineffective or unsatisfactory. It can be said that changes of CTC during the adjuvant therapy for the non-metastatic setting provide useful information about the efficacy of chemotherapy and endocrinic treatment. What's more, Herceptin is beneficial to patients whose HER-2 was negative in primary tumor cell but positive in CTC. Therefore, CTC can be used not only for estimating and predicting prognosis, but also for monitoring and assessing responsiveness to therapy as well as stratifying patients for adjuvant therapy. However, still some problems need solving: firstly, what clinicopathologic characteristics tend to be detected with CTC? secondly, is there difference between the potencies of hematogenous dissemination in different molecular subtypes? thirdly, does CTC play a role in assessing and predicting the clinical response to neoadjuvant chemotherapy in patients with breast cancer? With these questions we performed this preliminary study and tried to get answers. Methods & ResultsPart I. Analysis of Relationship between the Presence of Circulating Tumor Cell and Clinicopathological Characteristics of Primary Tumor and Molecular Subtypes of Breast CancerMethods: Samples of peripheral blood (5ml) from 128 cases of patients with breast cancer and 30 cases of patients with breast fibroadenoma hospitalized for the first time were enriched by density gradient centrifugation for mononuclear cells and then detected by immunofluorescence with anti-cytokeratin8/18 antibody and DAPI for CTC. The expression of ER, PR, HER-2 was detected by immunohistochemistry. The relations between the positive rate of CTC and clinicopathological characteristics of primary tumor and molecular subtypes of breast cancer patients were analyzed.Results: The positive rate of CTC in breast cancer patients was 40.6% (52/128), while no CTC was detected in all patients with breast fibroadenoma. The positive rates of CTC in patients at pathologic lymph node stages (pN0, pN1, pN2 and pN3) were 15.4% (6/39), 27.0% (10/37), 64.7% (11/17) and 66.7% (12/18), respectively. There was significant difference in positive rate of CTC between different pN stages (P=0.0006). The positive rate of CTC in patients at stageⅠ,Ⅱ,ⅢandⅣwas 12.5% (3/24), 32.0% (16/50), 50.0% (17/34) and 80.0% (16/20), respectively. There was significant difference in positive rate of CTC between different TNM stages (P=0.0003). The positive rate of CTC in patients at stageⅣ(80.0%) was significantly higher than patients at stageⅠ～Ⅲ(33.3%, 36/108) (P=0.0009). Luminal, HER-2 and Basle-like subtype accounts for 56.2%(72/128),14.1%(18/18)和39.7% (38/128), respectively. The positive rate of CTC in patients with Luminal, HER-2 and Basle-like subtype was 30.6% (22/72), 61.1% (11/18) and 50.0% (19/38), respectively. There was significant difference in positive rate of CTC between different molecular subtypes (P=0.0230). The positive rate of CTC in patients with HER-2 and Basle-like subtype was significantly higher than patients with Luminal subtype (P=0.0161, P=0.0449) , respectively.Part II. Analysis of the Role of Circulating Tumor Cell Playing in Assessing the Clinical Response to Neoadjuvant Chemotherapy in Patients with Breast CancerMethods: Samples of peripheral blood (5ml) from 50 cases of patients with breast cancer and 15 cases of patients with breast fibroadenoma hospitalized for the first time and 10 healthy volunteers were enriched by density gradient centrifugation for mononuclear cells and then detected by real time quantitative RT-PCR for cytokeratin 19 mRNA-positive CTC. The effects of neoadjuvant chemotherapy (NCT) were evaluated according to the Response Evaluation Criteria in Solid Tumors (RECIST). The relations between status and changes of CTC before and after NCT (TE regimen: TAX 175 mg/m2,EPI 60 mg/m2,dl,q3w) and clinical response to NCT in breast cancer patients were analyzed.Results: Among 50 cases of breast cancer, 3 cases (6.0%) achieved clinical complete response (cCR), 35 cases (70.0%) achieved clinical partial response (cPR), 10 cases (20.0%) achieved stable disease (SD) and 2 cases (4.0%) achieved progress disease (PD). The response rate (RR) was 76.0%. The positive rate of CTC before NCT was 34.0% (17/50) while decreased to 14.0% (7/50) after NCT. There was significant difference in positive rate of CTC before and after NCT (P=0.0063). After NCT, RR of patients with CTC detected was 28.6% (2/7) while 83.7% (36/43) in patient with no CTC detected. There was significant difference in RR between two groups (P=0.0059). The RR of patients whose CTC status was positive/positive, positive/negative, negative/positive, negative/negative before and after NCT was 33.3% (2/6),72.7% (8/11),0% (0/1)和84.4% (27/32), respectively. There was significant difference in RR between four groups (P=0.0164).Conclusions1. CTC tends to be detected in patients at high pN and TNM stage as well as M1 stage. CTC detection is useful for evaluating and identifying patients at high risks of recurrence and metastasis. Moreover,CTC can also be detected in patients even at N0 and I stage , indicating that CTC detection is useful for more accurate risk assessment of patients with breast cancer.2. Among the three breast cancer molecular subtypes, CTC tends to be detected in patients with HER-2 and Basle-like subtype, which may be the main factor leading to their high risks of recurrence and metastasis as well as poor prognosis.3. The positive rate of CTC decreases significantly after NCT and the changes of CTC before and after NCT correlate closely with efficacy of NCT, indicating that CTC can be used to assess the response to NCT.