Establishment Of Canine Model For Lung Tumor And Pharmacokinetics Study Of Bronchial Artery Infusion Chemotherapy

BackgroundLung cancer is one of the most common malignant tumors. According to WHO statistics, more than 1 million people die from lung cancer every year (WHO,2002). In China, lung cancer is the No.1 leading cause of death among all types of malignancies. in the cities, and the morbidity and mortality have increased significantly in both urban and rural areas. Developed in the late 1970(?), (?)al arterial infusion chemotherapy (BAI) one of the effactive treatment modalities for patients with advanced lung cancer. BAI is a minimally invasive interventional therapy which delivers chemical drugs directly to the tumor via bronchial, with the aim to increase intratumoral drug concentration and prolong drug stasis within lesions while maintain the blood drug concentration at low level. Therefore, the advantages of BAI are higher efficacy and less side effects. But the clinical pharmacokinetic studies of BAI are not mature, with few papers in the literature. Some representative studies calculate the pharmacokinetic parameters by detecting the blood concentration of several chemotherapeutic drugs. There are main reaseons for this limitation. Firstly, it is difficult to establish a suitable large animal model of lung cancer. Secondly, it is also difficult to measure the real-time concentration of chemical durgs inside mass in the course of interventional treatment.Canine transmissible venereal tumor (CTVT) is a naturally occurring tumor, which has many characteristic such as stability, reliability and diversity in in-situ transplantation, is a good tumor strain for the canine lung cancer model.Microdialysis was developed in the 1970s. This technique provides a means of continuously sampling substances in the interstitial space fluid on the premise of not destroying environment in the organ. Principle of microdialysis is the same as conventional dialysis, with the exception that its sampling equipment (probe) is tiny which can be put into various kinds of organs and tissues. Since microdialysis has many advantages such as little damage, high authenticity and small deviation, it has been extensively applied in numerous fields of natural science.The aims of this study are:to explore the optimal method for the establishment of cTVT allograft lung cancer model, then try to discover the blood supply of the model, and finally to determine the intra-tumoral pharmacokinetics of BAI using microdialysis in order to better guide clinical BAI protocol and provide a theoretical frame of reference. By acompishing these work, we also hope to establish a set of mature methods to study drug metabolism after various vascular interventions in the futrue.Objective1. To explore the optimal method for the establishment of CTVT lung cancer model.2. To study the blood supply of CTVT lung cancer.3. To determine the intra-tumoral pharmacokinetics of BAI using microdialysis technique.4. To explore one series of pharmacokinetics study methods on vascular interventional treatment in vivo tumor.Methods1.18 beagle dogs were divided into 2 groups. Group A, cell suspension inoculation group, in which 12 dogs underwent percutaneous CT-guided puncture of lungs and innoculation of CTVT cell suspension (live cell concentration of 108/ml) into dog lungs. Group B, fresh tumor fragments inoculation group, in which 6 dogs underwent percutaneous CT-guided puncture of lungs and innoculation of 2～3 fresh tumor fragments (1.5～2.0mm). Chest CT was performed immediately after inoculation, and 4,6,,8 and 10 weeks after innoculation in order to observe the tumor growth and metastasis. Animals were observed daily for general state of health. Autopsy and pathological examinations were performed to observe whether tumor necrosis and micro-pathological changes until their natural death or at 10 weeks after furthur experiment. Repeated measure method was performed to analyse the tumor growth state, and determine the differences between the two groups.2. In 12 dogs with CTVT lung cancer, bronchial artery digital subtraction angiography (BA-DSA), pulmonary digital subtraction angiography (PA-DSA), trans-bronchial arterial contrast enhanced multi-slice CT angiography (BA-MSCTA) and trans-pulmonary arterial contrast enhaced multi-slice CT angiography (PA-MSCTA) were performed. Images were analyzed and enhancement of lung nodules were determined. CT after pulmonary arterial lipiodol injection was also performed. Finally, blood supply of CTVT lung cancer model was judged.3. Experimental dogs were divided into two groups:BAI treatment group (group B, n=3); Venous groups (group V, n=3). After the dogs were anaesthetized, percutaneous bronchial arterial angiography or other systemic arterial angiography was performed to clarify the blood supply to the lung tumors. Then chest CT scan was performed to decide puncture point, angle and route. Two 20G chiba needles were inserted to cross the semi-chest, one through the tumor and one through the adjacent lung parenchyma. A linear microdialysis probes were threaded through the canula, making sure the membrane (samping part) was within the tumor and the lung parenchyma of interest. For group B, carboplatin 16.5mg/kg dissolved in 250ml 5% GS was infused into the bronchial artery slowly within an hour using a micro-pomp. In groupⅤ, same dosage of carboplatin was administered through saphenous vein using the miaro-pump. Then dialysate samples were collected at the outlet. The drug concentration was determined by Mass Spectrometry Instrument. The time-curves of the drug concentration in extracellular fluid of tumor and lung tissue were drawn. Data were processed with Microsoft Excel and DAS2.0 software. Pharmacokinetic parameters were calculated.Results1. The technical success rates of both groups were 100%. Complications (pneumothorax, hemothorax, pulmonary hemorrhage, hemoptysis, etc.) after inoculation were almost equal. Tumor formation rate of group B (tumor fragment inoculation,100%,12/12) was significantly higher than group A(cell suspension inoculation,66.67%,9/15) (P=0.037<0.05). The mean largest diameter of Group A was 1.059±0.113,1.827±0.084 and 2.189±0.153cm at 6th,8th,10th week; while in Group B 1.716±0.102,2.392±0.076,2.734±0.138cm, respectively (p<0.05). Chest wall and subcutaneous plantings were found in 6 dogs in Group A, with pleural effusion appeared at 7th week and severe pleural effusion and mediastinal lymph node enlargement at 9th week in all dogs. All dogs in Group B showed cachexia status. In Group B, chest wall inoculatation occurred in 2 dogs, and lung metastasis was observed at 10th week. There were no pleural effusion and mediastinal lymph node enlargement. No metastases outside the chest were observed. Two dogs underwent pathological examination and no ecrosis was found.2. For 14 nodules larger than 2cm, hyperplastic bronchial arteries leading to the tumor were found in 12 nodules on BA-DSA. For 5 nodules 1～2cm in diameter, tumor staining were obvious in 2 nodules on BA-DSA. No tumor staining was observed in four dogs on PA-DSA. In 12 nodules more than 2cm, BA-MSCTA diaplayed chaotic tumor blood vessels and tumor staining within tumors. Chest CT scan was performed in 8 experimental dogs after pulmonary artery was embolized with lipiodol. Two lesions (less than 1cm) were shown to have lipiodol deposition (2/5). A droplet of lipiodol deposition was seen in one nodule larger than 2cm. Scattered dotted lipiodol deposits were found in two nodules larger than 2cm.3. Plasma carboplatin concentrations changed regularly after administration via both BAI or venous route. Some pharmacokinetic parameters between the two groups were significantly different. AUC values in BAI group and intravenous group were 58.66±4.768 and 76.992±8.873(mg/L*h), respectively. And Tmax value 1.053±0.046 and 1±0 (h), Cmax values 28.833±3.711 and 81.3±9.839 (mg/L), tl/2z values 51.471±22.165 and 9.188±12.687 (h), respectively, (p<0.05, for all). In both groups, drug concentrations in tumor and normal lung tissue changed regularly. The tumor pharmacokinetic parameters in two groups were:AUC values 1660.76±339.77 and 459.49±49.99 (mg/L* min), Cmax values 21500.39±4359.91 and 4627.27±722.676 (mg/L), MRT values 74.322±8.048 and 98.23±3.934 (min), in BAI groups andⅣgroup, respectively. Analysis of variance FAUC=17.336, P=0.001, FCmax=33.556, P=0, FMRT=9.091, P=0.006. (p<0.05).Conclusions1. It is feasible to establish a CTVT lung cancer model by percutaneous puncture. Tissue fragments inoculation method is simple, efficient, with few complications compared with cell suspension inoculation method. It is the ideal method to establish lung cancer model with CTVT.2. CTVT allograft lung tumor was mainly supplied by the bronchial artery.Tumor blood supply was related to tumor size, and the greater the tumor size was, the greater the likelihood of bronchial arterial supply was; Pulmonary artery may supply the tumor, especially in tumors smaller than 1cm. For larger tumors, blood supply was from the bronchial artery exclusively.3. Compared with intravenous administration, BAI could significantly increase the peak concentration within the tumor, prolong the local action time, increase drug bioavailability in local tumor and enhance tumor cell killing, while decreasing the blood concentration and reducing systemic side effects.4. The average residual time of carboplatin can be prolonged after dministered by BAI. But drug accumulation is prone to take place if administration is repeated in a short time compared with intravenous administration. 5. It is an effective way to study vascular interventional pharmacokinetics by establishing CTVT lung cancer model by percutaneous implantation, combined with microdialysis probe implanted by CT-guided puncture.