Combination radiofrequency ablation and intravenous radiolabeled liposomal Doxorubicin: imaging and quantification of increased drug delivery to tumors.

PURPOSE: To identify, with noninvasive imaging, the zone of radiopharmaceutical uptake after combination therapy with radiofrequency (RF) ablation and intravenous administration of technetium 99m ((99m)Tc) liposomal doxorubicin in a small-animal tumor model, and to quantify and correlate the uptake by using imaging and tissue counting of intratumoral doxorubicin accumulation. MATERIALS AND METHODS: This study was approved by the animal care committee. Two phases of animal experiments were performed. In the first experiment, a single human head-and-neck squamous cell carcinoma tumor was grown in each of 10 male nude rats. Seven of these animals were treated with intravenous (99m)Tc-liposomal doxorubicin followed by RF tumor ablation at a mean temperature of 70 degrees C + or - 2 for 5 minutes, and three were treated with intravenous (99m)Tc-liposomal doxorubicin only. Combination single photon emission computed tomography-computed tomography (SPECT/CT) was performed at 15 minutes, 4 hours, and 20 hours after therapy. In the second experiment, two tumors each were grown in 11 rats, but only one of the tumors was ablated after intravenous administration of (99m)Tc-liposomal doxorubicin. SPECT/CT and planar scintigraphy were performed at the same posttreatment intervals applied in the first experiment, with additional planar imaging performed at 44 hours. After imaging, tissue counting in the excised tumors was performed. Radiotracer uptake, as determined with imaging and tissue counting, was quantified and compared. In a subset of three animals, intratumoral doxorubicin accumulation was determined with fluorimetry and correlated with the imaging and tissue-counting data. RESULTS: At both SPECT/CT and planar scintigraphy, increased uptake of (99m)Tc-liposomal doxorubicin was visibly apparent in the ablated tumors. Results of quantitative analysis with both imaging and tissue counting confirmed significantly greater uptake in the RF ablation-treated tumors (P < .001). Intratumoral doxorubicin accumulation correlated closely with imaging (r = 0.9185-0.9871) and tissue-counting (r = 0.995) results. CONCLUSION: Study results show that increased delivery of intravenous liposomal doxorubicin to tumors combined with RF ablation can be depicted and quantified with noninvasive imaging.

Interventional therapy of head and neck cancer with lipid nanoparticle-carried rhenium 186 radionuclide.

PURPOSE: Minimally invasive interventional cancer therapy with drug-carrying lipid nanoparticles (ie, liposomes) via convection-enhanced delivery by an infusion pump can increase intratumoral drug concentration and retention while facilitating broad distribution throughout solid tumors. The authors investigated the utility of liposome-carrying beta-emitting radionuclides to treat head and neck cancer by direct intratumoral infusion in nude rats. MATERIALS AND METHODS: Four groups of nude rats were subcutaneously inoculated with human tongue cancer cells. After tumors reached an average size of 1.6 cm(3), the treatment group received an intratumoral infusion of liposomal rhenium-186 ((186)Re) (185 MBq [5 mCi]/cm(3) tumor). Three control groups were intratumorally infused with unlabeled liposomes, unencapsulated (186)Re-perrhenate, or unencapsulated intermediate (186)Re compound ((186)Re-N,N-bis[2-mercaptoethyl]-N',N'-diethyl-ethylenediamine [BMEDA]). In vivo distribution of (186)Re activity was measured by planar gamma-camera imaging. Tumor therapy and toxicity were assessed by tumor size, body weight, and hematology. RESULTS: Average tumor volume in the (186)Re-liposome group on posttreatment day 14 decreased to 87.7% +/- 20.1%, whereas tumor volumes increased to 395.0%-514.4% on average in the other three groups (P< .001 vs (186)Re-liposome). The (186)Re-liposomes provided much higher intratumoral retention of (186)Re activity, resulting in an average tumor radiation absorbed dose of 526.3 Gy +/- 93.3, whereas (186)Re-perrhenate and (186)Re-BMEDA groups had only 3.3 Gy +/- 1.2 and 13.4 Gy +/- 9.2 tumor doses, respectively. No systemic toxicity was observed. CONCLUSIONS: Liposomal (186)Re effectively treated head and neck cancer with minimal side effects after convection-enhanced interventional delivery. These results suggest the potential of liposomal (186)Re for clinical application in interventional therapy of cancer.

Type X collagen gene regulation by Runx2 contributes directly to its hypertrophic chondrocyte-specific expression in vivo.

The alpha1(X) collagen gene (Col10a1) is the only known hypertrophic chondrocyte-specific molecular marker. Until recently, few transcriptional factors specifying its tissue-specific expression have been identified. We show here that a 4-kb murine Col10a1 promoter can drive beta-galactosidase expression in lower hypertrophic chondrocytes in transgenic mice. Comparative genomic analysis revealed multiple Runx2 (Runt domain transcription factor) binding sites within the proximal human, mouse, and chick Col10a1 promoters. In vitro transfection studies and chromatin immunoprecipitation analysis using hypertrophic MCT cells showed that Runx2 contributes to the transactivation of this promoter via its conserved Runx2 binding sites. When the 4-kb Col10a1 promoter transgene was bred onto a Runx2(+/-) background, the reporter was expressed at lower levels. Moreover, decreased Col10a1 expression and altered chondrocyte hypertrophy was also observed in Runx2 heterozygote mice, whereas Col10a1 was barely detectable in Runx2-null mice. Together, these data suggest that Col10a1 is a direct transcriptional target of Runx2 during chondrogenesis.

Three-dimensional quantitative assessment of ablation margins based on registration of pre- and post-procedural MRI and distance map.

PURPOSE: Contrast-enhanced MR images are widely used to confirm the adequacy of ablation margin after liver ablation for early prediction of local recurrence. However, quantitative assessment of the ablation margin by comparing pre- and post-procedural images remains challenging. We developed and tested a novel method for three-dimensional quantitative assessment of ablation margin based on non-rigid image registration and 3D distance map. METHODS: Our method was tested with pre- and post-procedural MR images acquired in 21 patients who underwent image-guided percutaneous liver ablation. The two images were co-registered using non-rigid intensity-based registration. After the tumor and ablation volumes were segmented, target volume coverage, percent of tumor coverage, and Dice similarity coefficient were calculated as metrics representing overall adequacy of ablation. In addition, 3D distance map around the tumor was computed and superimposed on the ablation volume to identify the area with insufficient margins. For patients with local recurrences, the follow-up images were registered to the post-procedural image. Three-dimensional minimum distance between the recurrence and the areas with insufficient margins was quantified. RESULTS: The percent tumor coverage for all nonrecurrent cases was 100 %. Five cases had tumor recurrences, and the 3D distance map revealed insufficient tumor coverage or a 0-mm margin. It also showed that two recurrences were remote to the insufficient margin. CONCLUSIONS: Non-rigid registration and 3D distance map allow us to quantitatively evaluate the adequacy of the ablation margin after percutaneous liver ablation. The method may be useful to predict local recurrences immediately following ablation procedure.

Mutations and promoter SNPs in RUNX2, a transcriptional regulator of bone formation.

Cleidocranial dysplasia (CCD) is a dominantly inherited skeletal malformation syndrome with high penetrance and variable expressivity. It is caused by loss of function mutations in the RUNX2 gene that encodes for a transcription factor essential for osteoblast differentiation and chondrocyte maturation. To identify new pathogenic mutations associated with CCD we screened 38 CCD patients for mutations in the RUNX2 coding sequence. We also report the mutation screening of the "bone-related" RUNX2 promoter in CCD patients without mutation in the RUNX2 coding region. We identify eight new and three previously described mutations in the RUNX2 gene. Additionally, a total of five sequence variants in the RUNX2 promoter were detected. Three of them occur within putative zinc finger transcription factor binding sites. DHPLC analysis of chromosomes from the control population and CCD patients showed that two promoter sequence variants were unique for CCD families. Electrophoretic mobility shift assay (EMSA) with protein extracts from ROS17/2.8 and C3H10T1/2 cell lines demonstrated that the promoter sequence variants altered DNA-protein binding specificity. Moreover, one of the variants significantly decreased the expression of a RUNX2 reporter gene in osteoblastic ROS17/2.8 cells, but not in multipotent, mesenchymal C3H10T1/2 cells. Interestingly, one of these sites bound the TRPS1 transcription factor and we demonstrated that TRPS1 is able to repress the RUNX2 promoter. The in vitro functional studies in conjunction with analysis of clinical phenotype of CCD patients suggest that these promoter sequence variants may affect transcriptional activity of the RUNX2 gene. Analysis of the promoter variants and RUNX2-interacting proteins may help to identify important cis-elements and trans-factors that regulate the RUNX2 transcriptional network and identify new susceptibility markers for more common bone disorders.