Detection Of Bone Marrow Micrometastasis In Breast Cancer And Tumor Specific CTLs Immunotherepy

Objective: Breast cancer is the most common malignant tumor among women, the incidence is becoming higher in recent years. There is no significant improvement of long-term survival rate though comprehensive measures including surgery are taken. The major reason is associated with metastasis to remote organs. It is showed that, by clinical statistics, bone marrow micrometastasis of breast cancer is the signal of distant metastasis. The detection of micrometastasis may be one of targets to predict the prognosis of breast cancer. It not only help us supervise metastasis dynamically, observe the treatment effect, but also has been an independent factor of predicting the prognosis of breast cancer. Therefore, improving the prognosis of breast cancer needs the treatment of bone marrow micrometastasis. In this research, we have detected bone marrow micrometastasis of breast cancer patients, then used tumor specific CTLs as the treatment method of bone marrow micrometastasis, and discussed the regulations of bone marrow micrometastasis. By this study, we want to find a suitable way to predict the prognosis and increase the long-term survival rate of breast cancer patients.Methods: 82 breast cancer cases in our hospital were taken between March and December in 2007. All of them were female, aged from 27 to 82, with a mean of 44.8, and have been proved by patheology. 10 were used as normal person negative control. According to the UICC breast cancer TNM, Detected the cell positive rate of CK18 and CK19 in bone marrow by FCM. Bone marrow puncture was done in 1-3Ddays before operation. In the 82 cases, 23 were detected to be CK18 and CK19 positive expression ones. 23 were divided into two groups: 17 had been treated with CTLs immunotherapy after operation, other therapies would be done in sequence. Therapies of the other 6 were same except the use of CTLs. Each patient with immunotherapy had been collected lymph nodes and peripheral blood. CTLs were cultivated from these two tissues, and injected into the same patient 10-14 days after operation. Then observed the effect of CTLs.Results1.From bone marrow specimen of 82 breast cancer patients, CK18.+,CK19~+/CD45~- cells were detected in 23 cases by FCM, breast cancer bone marrow micrometastasis positive detection rate was 28.05%.2. Bone marrow micrometastasis positive rate in breast cancer patients was increased by protopathic tumor size. The micrometastasis positive rate was 7.69%(1/13),14.58(7/48)%,71.43%(15/21) respective for the≤2cm,2~5cm and>5cm (in diameter). There are statistical significance among three groups, P=0.0007. 3. Breast cancer bone marrow micrometastasis positive rate was increased by the clinical TNM staging increased. Bone marrow micrometastasis positive rate was 12.50% (1/8), 17.65% (9/51), and 56.52% (13/23) respectively for StageⅠ,ⅡandⅢ. There are statistical significance among three groups, P=0.004.4 Breast cancer bone marrow micrometastasis positive rate was increased with histological grade increasing. Positive rate of the Grade III group was 68.42% (13/19), higher than the Grade II group 17.65% (9/51) and Grade I group 8.33% (1/12), There are statistical significance among three groups, P =0.003.5. There was not relationship between breast cancer bone marrow micrometastasis positive rate and their ages and menstruation, P>0.05.6. Bone marrow micrometastasis positive rate in breast cancer patients was decreased with ER and PR expressions in tumor tissues intensified. Bone marrow micrometastasis positive rate in ER and PR positive groups was 13.33%(4/30),10.34%(3/29) respectively, lower than ER and PR negative group 36.54%(19/54),37.74%(20/53), P<0.05. There was not relationship between bone marrow micrometastasis positive rate and C-erbB-2 expression in the tumor tissues(P>0.05)7. 23 breast cancer bone marrow micrometastasis positive patients had been divided into 2 groups: there were 17 patients in foscarnet group, and all had immunotherapy of tumor specific CTLs successfully. Only 2 patients had slight fever, anoresia or sleepiness, other patients had no discomfort feelings. Other 6 patients without CTLs treatment as control group. Each of the 23 cases had bone marrow micrometastasis detection 20 days later of operation. Then compared the micrometastasis positive detection rate with the one before operation, and predicted the effect. Among 17 patients of foscarnet group, there were 14 cases who could not be detected bone marrow micrometastasis, the immunotherapy clearance rate (negative rate) was 82.4%. Among 6 patients of control group, there were only 1 case who could not be detected bone marrow micrometastasis, the clearance rate (negative rate) was 16.7%. There are statistical significance between two groups (P =0.00028).Conclusion1. Breast cancer micrometastasis cell positive rate of CK18 and CK19 in bone marrow detecteted by FCM .is 28.05%.2 In this research, breast cancer bone marrow micrometastasis positive rate is increased by the clinical TNM staging increased. The clinical TNM staging in breast cancer relates to the size of the primary tumor and the axillary lymph nodes metastasis. The more invading ability the tumor has, the more chance to metastasis will be. Breast cancer patients of grade III group have more chances for bone marrow micrometastasis.3. Breast cancer bone marrow micrometastasis positive rate increases as histological grading going up, which indicates that low-differentiated cancer cells have stronger ability to reproduce, invade and transfer. It is necessary to use complex therapies to these patients and observe the distant metastasis.4. Breast cancer bone marrow micrometastasis positive rate relates to ER and PR protein expressions in tumor tissues. Breast cancer bone marrow micrometastasis positive rate decreases with ER and PR protain expressions intensified,which indicates that the lower tumor cells differentiate, the lower ER and PR express, and they are more capable to infiltrate and metastasis.5. No relationship is found between breast cancer bone marrow micrometastasis positive detection rate of the 82 cases and their ages, menstruation, pathological typing or the C-erbB-2 expression in the tumor tissues.6. The data has also shown that there is 12.50% (1/8) bone marrow micrometastasis in clinicalⅠ, which indicates that breast cancer is ageneral disease. Its metastasis and spread may occur in early phase of the disease. It is obvious that treatment to bone marrow metastasis must be done.7. Among the 23 breast cancer bone marrow micrometastasis positive patients, the immunotherapy clearance rate (negative rate) is 82.4%, the clearance rate (negative rate) of control group is 16.7%. There are statistical significance between two groups, P=0.00028. Tumor specific CTLs immunotherapy has the ability to clean the bone marrow micrometastasis of breast cancer. The side-effect is slight, and patients are easy to accept the therapy.