X-Ray Psoralen Activated Cancer Therapy (X-PACT).

This work investigates X-PACT (X-ray Psoralen Activated Cancer Therapy): a new approach for the treatment of solid cancer. X-PACT utilizes psoralen, a potent anti-cancer therapeutic with current application to proliferative disease and extracorporeal photopheresis (ECP) of cutaneous T Cell Lymphoma. An immunogenic role for light-activated psoralen has been reported, contributing to long-term clinical responses. Psoralen therapies have to-date been limited to superficial or extracorporeal scenarios due to the requirement for psoralen activation by UVA light, which has limited penetration in tissue. X-PACT solves this challenge by activating psoralen with UV light emitted from novel non-tethered phosphors (co-incubated with psoralen) that absorb x-rays and re-radiate (phosphoresce) at UV wavelengths. The efficacy of X-PACT was evaluated in both in-vitro and in-vivo settings. In-vitro studies utilized breast (4T1), glioma (CT2A) and sarcoma (KP-B) cell lines. Cells were exposed to X-PACT treatments where the concentrations of drug (psoralen and phosphor) and radiation parameters (energy, dose, and dose rate) were varied. Efficacy was evaluated primarily using flow cell cytometry in combination with complimentary assays, and the in-vivo mouse study. In an in-vitro study, we show that X-PACT induces significant tumor cell apoptosis and cytotoxicity, unlike psoralen or phosphor alone (p<0.0001). We also show that apoptosis increases as doses of phosphor, psoralen, or radiation increase. Finally, in an in-vivo pilot study of BALBc mice with syngeneic 4T1 tumors, we show that the rate of tumor growth is slower with X-PACT than with saline or AMT + X-ray (p<0.0001). Overall these studies demonstrate a potential therapeutic effect for X-PACT, and provide a foundation and rationale for future studies. In summary, X-PACT represents a novel treatment approach in which well-tolerated low doses of x-ray radiation are delivered to a specific tumor site to generate UVA light which in-turn unleashes both short- and potentially long-term antitumor activity of photo-active therapeutics like psoralen.

Quantitative ultrasound measurements of bone quality in female adolescents with idiopathic scoliosis compared to normal controls.

OBJECTIVE: The aims of this study were to compare the speed-of-sound (SOS) between adolescent idiopathic scoliosis (AIS) patients and controls using quantitative ultrasound examination and to further analyze the relationship between the SOS and curve type, curve magnitude, maturation status and Risser's sign in AIS patients compared to controls. METHODS: Seventy-eight female AIS patients and 58 healthy female controls 10 to 16 years of age were recruited to participate. Quantitative ultrasound measurements were performed at the non-dominant distal end of the radius. The standard method for estimating the SOS and z-score was used. Comparisons were made between the SOS values and z-score in AIS patients and age-matched Asian adolescents. RESULTS: The SOS values of the patients were significantly lower than the controls (P < .01). The percentage of cases with low bone quality was 25% in the entire AIS sample. The prevalence of low bone quality in AIS patients was 20.5%. However, there were no correlations between the SOS and types of scoliosis (P > .05). The SOS values among different severity groups were significant, particularly between the 10° to 19° and 20° to 39° groups as well as between 10° to 19° and ≥40° groups. However, there was no significant correlation between the SOS and Cobb angles. Significant correlations were also found between the pre- and post-menarchy status in patients. There was a significant difference in the SOS values for different Rissers' signs (P < .05). CONCLUSIONS: Compared to nonscoliotic controls, subjects with AIS had a generally lower SOS, indicating lower bone quality. The age, Risser's sign, or maturation status, may have an effect on the bone quality; however, the curve type and magnitude do not affect the bone quality. The results of this study indicate that slower bone maturation may affect the bone quality in adolescents with AIS.

Ligand-independent toll-like receptor signals generated by ectopic overexpression of MyD88 generate local and systemic antitumor immunity.

Although critical for initiating and regulating immune responses, the therapeutic use of individual cytokines as anticancer immunotherapeutic agents has achieved only modest clinical success. Consequently, many current strategies have focused on the use of specific immunotherapeutic agonists that engage individual receptors of innate immune networks, such as the Toll-like receptor (TLR) system, each resulting in specific patterns of gene expression, cytokine production, and inflammatory outcome. However, these immunotherapeutics are constrained by variable cellular TLR expression and responsiveness to particular TLR agonists, as well as the specific cellular context of different tumors. We hypothesized that overexpression of MyD88, a pivotal regulator of multiple TLR signaling pathways, could circumvent these constraints and mimic coordinated TLR signaling across all cell types in a ligand-independent fashion. To explore this hypothesis, we generated an adenoviral vector expressing MyD88 and show that Ad-MyD88 infection elicits extensive Th1-specific transcriptional and secreted cytokine signatures in all murine and human cell types tested in vitro and in vivo. Importantly, in vivo intratumoral injection of Ad-MyD88 into established tumor masses enhanced adaptive immune responses and inhibited local tumor immunosuppression, resulting in significantly inhibited local and systemic growth of multiple tumor types. Finally, Ad-MyD88 infection of primary human dendritic cells, tumor-associated fibroblasts, and colorectal carcinoma cells elicited significant Th1-type cytokine responses, resulting in enhanced tumor cell lysis and expansion of human tumor antigen-specific T cells. Thus, Ad-MyD88 initiated robust antitumor activity in established murine tumor microenvironments and in human contexts, suggesting its potential effectiveness as a clinical immunotherapeutic strategy.

Hyaluronan, CD44, and emmprin regulate lactate efflux and membrane localization of monocarboxylate transporters in human breast carcinoma cells.

Interactions of hyaluronan with CD44 in tumor cells play important cooperative roles in various aspects of malignancy and drug resistance. Emmprin (CD147; basigin) is a cell surface glycoprotein of the immunoglobulin superfamily that is highly up-regulated in malignant cancer cells and stimulates hyaluronan production, as well as several downstream signaling pathways. Emmprin also interacts with various monocarboxylate transporters (MCT). Malignant cancer cells use the glycolytic pathway and require MCTs to efflux lactate that results from glycolysis. Glycolysis and lactate secretion contribute to malignant cell behaviors and drug resistance in tumor cells. In the present study, we find that perturbation of endogenous hyaluronan, using small hyaluronan oligosaccharides, rapidly inhibits lactate efflux from breast carcinoma cells; down-regulation of emmprin, using emmprin small interfering RNA, also results in decreased efflux. In addition, we find that CD44 coimmunoprecipitates with MCT1, MCT4, and emmprin and colocalizes with these proteins at the plasma membrane. Moreover, after treatment of the cells with hyaluronan oligosaccharides, CD44, MCT1, and MCT4 become localized intracellularly whereas emmprin remains at the cell membrane. Together, these data indicate that constitutive interactions among hyaluronan, CD44, and emmprin contribute to regulation of MCT localization and function in the plasma membrane of breast carcinoma cells.

PTEN and rapamycin inhibiting the growth of K562 cells through regulating mTOR signaling pathway.

OBJECTIVE: To investigate, in vitro, the regulatory effects of tumor-suppressing gene PTEN on mTOR (mammalian target of rapamycin) signaling pathway, the effects of transfected PTEN and rapamycin on the growth inhibition, and apoptosis induction for human leukemia cell line K562 cells. METHODS: K562 cells were transfected with recombined adenovirus-PTEN vector containing green fluorescent protein (Ad-PTEN-GFP), followed by the treatment of the cells with or without rapamycin. The proliferation inhibition rate and apoptotic rate of these transfected and/or rapamycin treated K562 cells were measured by MTT assay and flow cytometry (FCM), the expression levels of PTEN-, mTOR-, cyclinD1- and P27kip1- mRNA were measured by real-time fluorescent relative-quantification reverse transcriptional PCR (FQ-PCR), the protein expression levels of PTEN, Akt, p-Akt were detected by western blotting. RESULTS: The proliferation of K562 cells was inhibited by PTEN gene transfection with/without the treatment of rapamycin. The expression levels of PTEN- and P27kip1- mRNA were up-regulated, and the mTOR- and cyclinD1- mRNA were down-regulated in K562 cells after the cells transfected with wild type PTEN gene and treated with rapamycin. CONCLUSION: PTEN and rapamycin inhibited mTOR expression by acting as an upstream regulator of mTOR. Low dose rapamycin in combination with over-expressed PTEN might have synergistic effects on inhibiting the proliferation and promoting apoptosis of K562 cells.

Adenovirus infection triggers a rapid, MyD88-regulated transcriptome response critical to acute-phase and adaptive immune responses in vivo.

Nearly 50 years ago, the discovery of interferon prompted the notion that host cells innately respond to viral invasion. Since that time, technological advances have allowed this response to be extensively characterized and dissected in vitro. However, these advances have only recently been applied to highly complex, in vivo biological systems. To this end, we exploited high-titer adenovirus (Ad) vectors to globally investigate the innate immune response to nonenveloped viral infection in vivo. Our results indicated a potent cellular transcriptome response shortly after infection, with global assessments revealing significant dysregulation in approximately 15% of the measured transcripts derived from Ad vector-transduced tissue. Bioinformatics-based transcriptome analysis revealed a complex innate response to Ad infection, with induction of proinflammatory responses (and suppression of metabolism and mitochondrial genes) akin to those observed when mice are challenged with lipopolysaccharide. Despite this commonality, there were many unique aspects of the Ad-dependent transcriptome response, including the upregulation of several RNA regulatory mechanisms and apoptosis-related pathways, accompanied by the suppression of lysosomal and endocytic genes. Our results also implicated the Toll-like receptors (TLRs) in these responses, prompting specific investigations into this pathway. By using MyD88KO mice, our results confirmed that Ad-induced dysregulation of five functionally related gene clusters are significantly dependent on this TLR adaptor gene. MyD88 deficiency also resulted in significantly diminished, although not abolished, adaptive and acute-phase immune responses to Ad, confirming the transcriptome data, as well as specifically identifying MyD88 as a significant Ad immunity amplifier and regulator in vivo.