Internalizing antibody-based targeted gene delivery for human cancer cells

Gene therapy, a potential solution to hereditary and nonhereditary diseases, faces the challenges of safe and specific gene delivery. Cationic carrier molecules (e.g., liposome and polymers) that form noncovalent complexes with negatively charged DNA have been in use as nonviral gene delivery vectors. Although they tend to be relatively less efficient than viral systems, they have inherent advantages of flexibility and safety. Their derivatives, in conjugation with functional molecules such as peptides, proteins, growth factors, and antibodies, have been focused on to generate nanocarriers with low toxicity, high stability, high efficiency, and cell-specific targeting features. Here we describe internalizing polyclonal and monoclonal antibodies against a stress chaperone, mortalin/mtHsp70. We demonstrate that these internalizing anti-mortalin antibodies (i-mot Ab) could be employed for (1) internalization of nanoparticles (quantum dots, Qdots) and the generation of illuminating cells and (2) gene delivery. By using cancer and normal human cells in parallel, we further demonstrate that gene delivery can be specifically enhanced in human cancer cells if cationic polymer polyethylenimine (PEI) and i-mot Ab complex are used and may provide a novel cancer-targeting nanocarrier.     

Relationship between mitral regurgitation and left ventricular outflow obstruction in hypertrophic cardiomyopathy

To clarify the relationship between mitral regurgitation and left ventricular outflow obstruction, Doppler and two-dimensional echocardiographic studies were performed in 62 patients with hypertrophic cardiomyopathy (22 with and 40 without obstruction caused by mitral systolic anterior motion with septal contact). Pulsed Doppler echocardiography with color Doppler flow imaging demostrated that in 20 of the 22 patients with obstruction, mitral regurgitation occurred mainly during midsystole from the onset to the end of mitral-septal contact. Such midsystolic mitral regurgitation was not observed in patients without obstruction, except in three of 25 patients with mild mitral systolic anterior motion without septal contact. Furthermore, that regurgitation developed or disappeared together with the obstruction during follow-up periods or pharmacologic interventions. Two-dimensional echocardiography showed that in 21 of the 22 patients with obstruction, a distal residual portion of the "anterior" mitral leaflet moved anteriorly in early systole and protruded into the outflow tract during midsystole to cause the obstruction. In the other patient with obstruction, who had only early systolic mitral regurgitation, a distal residual "posterior" leaflet moved similary. These results may indicate that the midsystolic mitral regurgitation is hydrodynamically induced by the midsystolic pressure gradient across the protruding distal residual anterior mitral leaflet.     

Three-dimensional sonographic features of a fetus with Wolf-Hirschhorn syndrome

We present a case of fetal Wolf-Hirschhorn syndrome diagnosed by conventional two-dimensional and three-dimensional ultrasonography. Conventional two-dimensional ultrasonography revealed a diaphragmatic hernia, nuchal edema, and suspected hypospadias. Three-dimensional ultrasonography clearly showed a flattening of the face, a high forehead, a broad nasal bridge continuing to the forehead, exophthalmos, and micrognathia (resembling the appearance of a Greek warrior helmet), but conventional two-dimensional ultrasonography did not depict these findings. Prenatal chromosomal analysis confirmed the diagnosis of Wolf-Hirschhorn syndrome [46XY, del(4)(p15.2)]. Here we demonstrate how three-dimensional ultrasonography provided a novel visual depiction of the facial dysmorphism, which helped substantially in prenatal counseling.     

Ultrasonic vocalization impairment of Foxp2 (R552H) knockin mice related to speech-language disorder and abnormality of Purkinje cells

Previous studies have demonstrated that mutation in the forkhead domain of the forkhead box P2 (FOXP2) protein (R553H) causes speech-language disorders. To further analyze FOXP2 function in speech learning, we generated a knockin (KI) mouse for Foxp2 (R552H) [Foxp2 (R552H)-KI], corresponding to the human FOXP2 (R553H) mutation, by homologous recombination. Homozygous Foxp2 (R552H)-KI mice showed reduced weight, immature development of the cerebellum with incompletely folded folia, Purkinje cells with poor dendritic arbors and less synaptophysin immunoreactivity, and achieved crisis stage for survival 3 weeks after birth. At postnatal day 10, these mice also showed severe ultrasonic vocalization (USV) and motor impairment, whereas the heterozygous Foxp2 (R552H)-KI mice exhibited modest impairments. Similar to the wild-type protein, Foxp2 (R552H) localized in the nuclei of the Purkinje cells and the thalamus, striatum, cortex, and hippocampus (CA1) neurons of the homozygous Foxp2 (R552H)-KI mice (postnatal day 10), and some of the neurons showed nuclear aggregates of Foxp2 (R552H). In addition to the immature development of the cerebellum, Foxp2 (R552H) nuclear aggregates may further compromise the function of the Purkinje cells and cerebral neurons of the homozygous mice, resulting in their death. In contrast, heterozygous Foxp2 (R552H)-KI mice, which showed modest impairment of USVs with different USV qualities and which did not exhibit nuclear aggregates, should provide insights into the common molecular mechanisms between the mouse USV and human speech learning and the relationship between the USV and motor neural systems.     

Characterization of chronic idiopathic thrombocytopenic purpura in Japanese children: a retrospective multi-center study

The objectives of this study are to clarify (1) the difference in demographic and clinical variables at initial presentation between acute and chronic idiopathic thrombocytopenic purpura (ITP), and (2) the prognostic factors of patients with chronic ITP. We conducted a retrospective analysis of 247 children with newly diagnosed ITP between April 1991 and March 2006 who visited one of the 12 hospitals belonging to the Kyoto University Pediatric Hematologic Study Group. 180 and 67 cases were classified as the acute type and as the chronic type, respectively. Older age, higher initial platelet count, positive medical history or concomitant medical diagnosis, the absence of preceding infection or vaccination, and the absence of an increase in immunoglobulin were risk factors for the chronicity. The prognostic factors in chronic ITP were evaluated in 53 patients after excluding patients receiving splenectomy or having insufficient follow-up data. The overall time required for 50% resolution in patients with chronic ITP was approximately 5.6 years. Age at presentation of less than 3 years and higher platelet counts at the time of chronic ITP diagnosis were good prognostic factors. On the other hand, gender, initial platelet counts, and preceding infection or vaccination were not associated with the prognosis.     

The significance of strong histone deacetylase 1 expression in the progression of prostate cancer

Song Y H, Shiota M, Tamiya S, Kuroiwa K, Naito S & Tsuneyoshi M
(2011) Histopathology 58 , 773–780 The significance of strong histone deacetylase 1 expression in the progression of prostate cancer Aims: Histone deacetylases (HDACs) play important roles in many types of cancer. Recently, it has been reported that HDAC1 expression in prostate cancer is significantly higher than in benign prostate cell lines and tissues. The expression of HDAC1 in association with the clinicopathological data was investigated to define its functional and pathological roles in prostate cancer. Methods and results: HDAC1 expression was examined immunohistochemically in 148 patients with prostate cancer. Strong expression of HDAC1 in benign prostate glands, high‐grade prostatic intraepithelial neoplasia (PIN) and prostate cancer was observed in 17/148 (11%), 19/71 (27%) and 69/148 (47%) patients. Strong HDAC1 expression was correlated with high Gleason score (P = 0.025) and high pT stage (P = 0.012). Patients with strong HDAC1 expression had higher biochemical recurrence rates (P = 0.0010). Furthermore, strong HDAC1 expression had a significant impact on patient biochemical recurrence rates in multivariate analysis (P = 0.004). Conclusions: These results indicate that overexpression of HDAC1 contributes to progression and poor prognosis in prostate cancer. The findings may play an important role in the emergence of effective new approaches for therapy and prognostic markers of prostate cancer.     

Significance of urinary excretion of 8-hydroxy-2'-deoxyguanosine in healthy subjects and liver disease patients

BACKGROUND/AIMS: Although the importance of reactive oxygen species (ROS) in the pathogenesis of various diseases is stressed, clinical significance of the markers reflecting DNA oxidation such as 8-hydroxy-2'-deoxyguanosine (8-OHdG) remains to be clarified. METHODOLOGY: To examine clinical usefulness of 8-OHdG in healthy individuals in comparison with liver disease patients, urinary excretion of 8-OHdG was measured in 336 healthy individuals and 110 patients with liver disease. RESULTS: In healthy persons, the 8-OHdG excretion was increased in an age-dependent manner. It was positively correlated with cigarettes smoked a day and negatively correlated with body mass index (BMI) (P < 0.05, each). Age, smoking and BMI were independent predictors of urinary 8-OHdG excretion (P < 0.01, P < 0.01 and P < 0.05, respectively). In liver disease, the excretion of 8-OHdG was not changed, as compared with healthy individuals. However, the liver disease patients under the age of 40 had higher values of 8-OHdG than healthy persons. In addition, the urinary excretion of 8-OHdG was higher in patients with hepatitis C virus (HCV) infection than those with hepatitis B virus (HBV) infection. CONCLUSIONS: The results of the present study suggest that measurement of urinary 8-OHdG excretion is useful in assessing DNA oxidation caused by aging, smoking, body composition and liver disease.     

Aberrant DNA methylation of T-cell leukemia, homeobox 3 modulates cisplatin sensitivity in bladder cancer

The development of resistance to cisplatin during treatment of bladder cancer constitutes a major obstacle to curing bladder cancer. The identification of epigenetic biomarker molecules for cisplatin resistance and the elucidation of the function of the identified genes in bladder cancer will provide useful information. We found that the candidate gene TLX3 was unmethylated in cisplatin sensitive cells and methylated in resistant cells. The suppression of TLX3 expression using TLX3-specific shRNA in parental cells increased cisplatin resistance. Contrarily, overexpression of TLX3 in resistant cells induced increased sensitivity to cisplatin. We found that 22 (21%) out of 110 clinical samples of bladder cancer showed the methylated pattern using the COBRA assay in TLX3. We found a correlation between TLX3 methylation and the sensitivity to cisplatin in the clinical samples by SDI test. Cisplatin sensitivity was closely associated with the methylation status of TLX3. These findings showed that the TLX3 methylation may be useful as a novel biomarker for cisplatin resistance and can be used to design therapies to counteract the resistance against cisplatin in bladder cancer.