Citations

Frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17) is caused by mutations in the gene encoding the microtubule-associated protein, tau. Some FTDP-17 mutations affect exon 10 splicing. To correct aberrant exon 10 splicing while retaining endogenous transcriptional control, we evaluated the feasibility of using spliceosome-mediated RNA trans-splicing (SMaRT) to reprogram tau mRNA. We designed a pre-trans-splicing molecule containing human tau exons 10 to 13 and a binding domain complementary to the 3′ end of tau intron 9. A minigene comprising tau exons 9, 10, and 11 and minimal flanking intronic sequences was used as a target. RT-PCR analysis of SH-SY5Y cells or COS cells cotransfected with a minigene and a pre-trans-splicing molecule using primers to opposite sides of the predicted splice junction generated products containing exons 9 to 13. Sequencing of the chimeric products showed that an exact exon 9–exon 10 junction had been created, thus demonstrating that tau RNA can be reprogrammed by trans-splicing. Furthermore, by using the same paradigm with a minigene containing full-length intronic sequences, we show that cis-splicing exclusion of exon 10 can be by-passed by trans-splicing and that conversion of exon 10 – tau RNA into exon 10 + tau RNA could be achieved with ≈34% efficiency. Our results demonstrate that an alternatively spliced exon can be replaced by trans-splicing and open the way to novel therapeutic applications of SMaRT for tauopathies and other disorders linked to aberrant alternative splicing.

Abstract Purpose: Akt is a signal transduction protein that plays a central role in inhibiting apoptosis in a variety of cell types including human cancer cells. In cell lines derived from human non–small cell lung cancers (NSCLCs), Akt has been shown to confer chemoresistance by inhibition of apoptosis in response to different chemotherapeutic agents including platinum-based agents, which are often the first-line therapy for NSCLCs. Only 20% to 30% of patients with NSCLC treated with chemotherapy have clinical evidence of response. The purpose of this study is to determine whether or not overexpression of activated Akt [i.e., phosphorylated Akt (pAkt)] is correlated with survival. Experimental Design: We studied tumors from 61 patients with NSCLC in three tissue microarrays. All patients were followed for a period of 10 years or until death. The arrays were studied immunohistochemically with antibodies against pAkt, p53, and Ki-67. Results: There was a statistically significant difference in survival between the 14 patients with strong pAkt staining and the 47 patients with weak to absent pAkt staining both by log-rank (P = 0.0416) and Breslow analysis (P = 0.0446). Difference in survival time with respect to pAkt status was also statistically significant even after accounting for stage at diagnosis (P = 0.004). Neither p53 nor Ki-67 was a statistically significant prognostic factor. Conclusions: Overexpression of pAkt is an independent prognostic factor. Additional studies of human NSCLCs are warranted to drive the development of targeted tumor-specific antineoplastic therapies.

Abstract Purpose: To define the toxicities, pharmacodynamics, and clinical activity of the proteasome inhibitor, PS-341 (bortezomib), in patients with advanced malignancies. Patients and Methods: Twenty-eight patients (14 male and 14 female) received PS-341 twice weekly for 4 of 6 weeks (schedule I). Because toxicity necessitated dose omissions on this schedule, 16 additional patients (12 male and 4 female) received PS-341 twice weekly for 2 of every 3 weeks (schedule II). A total of 73 courses of treatment was given (median, 2; range, 1-4). Inhibition of 20S proteasome activity in peripheral blood mononuclear cells (PBMC) and accumulation of proteasome-targeted polypeptides in tumor tissue were evaluated as pharmacodynamic markers of PS-341 activity. Results: The most common toxicity was thrombocytopenia, which was dose limiting at 1.7 mg/m2 (schedule I) and 1.6 mg/m2 (schedule II), respectively. Sensory neuropathy was dose-limiting in a patient in schedule I. Grade ≥3 toxicities for schedule I were constipation, fatigue, myalgia, and sensory neuropathy. Grade ≥3 toxicities for schedule II were dehydration resulting from diarrhea, nausea and vomiting, fatigue, hypoglycemia, and hypotension. The maximum tolerated dose was 1.5 mg/m2 for both schedules. Reversible dose-dependent decreases in 20S proteasome activity in PBMCs were observed, with 36% inhibition at 0.5 mg/m2, 52% at 0.9 mg/m2, and 75% at 1.25 mg/m2. Accumulation of proteasome-targeted polypeptides was detected in tumor samples after treatment with PS-341. A patient with multiple myeloma had a partial response. Conclusion: PS-341 given 1.5 mg/m2 twice weekly for 2 of every 3 weeks is well tolerated and should be further studied.

Abstract Purpose: This phase I study was conducted to determine the toxicities, pharmacokinetics, and pharmacodynamics of BMS-214662, a farnesyl transferase inhibitor, in combination with paclitaxel and carboplatin, in patients with advanced solid tumors. Experimental Design: Patients with solid tumors received one of six escalating dose levels of BMS-214662 infused over 1 hour given following paclitaxel and carboplatin on the first day of a 21-day cycle. Toxicities were graded by the National Cancer Institute common toxicity criteria and recorded as maximum grade per patient for each treatment cycle. Inhibition of farnesyl transferase activity in peripheral blood mononuclear cells (PBMCs) was evaluated. Accumulation of unfarnesylated HDJ-2 in PBMCs of patients was evaluated as a marker of farnesyl transferase inhibition by BMS-214662. Results: Thirty patients received 141 cycles of treatment through six dose levels. Dose-limiting toxicities were neutropenia, thrombocytopenia, nausea, and vomiting. There was no pharmacokinetic interaction between BMS-214662 and paclitaxel. The maximum tolerated dose was established as BMS-214662 (160 mg/m2), paclitaxel (225 mg/m2) and carboplatin (area under the curve = 6 on day 1), every 21 days. Inhibition of HDJ-2 farnesylation in PBMCs of patients was shown. One measurable partial response was observed in a patient with taxane-resistant esophageal cancer. There was partial regression of evaluable disease in two other patients (endometrial and ovarian cancer). Stable disease (> 4 cycles) occurred in eight other patients. Conclusions: The combination of BMS-214662 with paclitaxel and carboplatin was well tolerated, with broad activity in solid tumors. There was no correlation between dose level and accumulation of unfarnesylated HDJ-2 in PBMCs nor tumor response.

Background— Sinus venosus atrial septal defect (SVASD) differs from secundum atrial septal defect by its atrial septal location and its association with anomalous pulmonary venous connection (APVC). Data on long-term outcome after surgical repair are limited. Methods and Results— We reviewed outcomes of 115 patients (mean age±SD 34±23 years) with SVASD who had repair from 1972 through 1996. APVC was present in 112 patients (97%). Early mortality was 0.9%. Complete follow-up was obtained for 108 patients (95%) at 144±99 months. Symptomatic improvement was noted in 83 patients (77%), and deterioration was noted in 17 patients (16%). At follow-up, 7 (6%) of 108 patients had sinus node dysfunction, a permanent pacemaker, or both, and 15 (14%) of 108 patients had atrial fibrillation. Older age at repair was predictive of postoperative atrial fibrillation ( P =0.033). No reoperations were required during follow-up. Survival was not different from expected for an age- and sex-matched population. Clinical improvement was more common with older age at surgery ( P =0.014). Older age at repair ( P =0.008) and preoperative New York Heart Association class III or IV ( P =0.038) were independent predictors of late mortality. Conclusions— Operation for SVASD is associated with low morbidity and mortality, and postoperative subjective clinical improvement occurs irrespective of age at surgery. Postoperative atrial fibrillation appears to be related to older age at operation. SVASD repair achieves survival similar to that of a matched population and should be considered whenever repair may impact survival or symptoms.

Purpose To determine the role of myeloablative conditioning and unrelated donor (URD) bone marrow transplantation in the treatment of patients with advanced B-cell chronic lymphocytic leukemia (CLL). Patients and Methods A total of 38 CLL patients received a matched URD transplant using bone marrow procured by the National Marrow Donor Program. The median age was 45 years (range, 26 to 57 years), the median time from diagnosis was 51 months, and the median number of prior chemotherapy regimens was three. Fifty-five percent of patients were chemotherapy refractory and 89% had received fludarabine. Conditioning included total-body irradiation in 92% of patients. Graft-versus-host disease (GVHD) prophylaxis consisted of methotrexate with cyclosporine or tacrolimus for 82% of patients. Results Twenty-one patients (58%) achieved complete response and six (17%) achieved partial response. Incidences of grades 2 to 4 acute GVHD were 45% at 100 days and incidences of chronic GVHD were 85% at 5 years. Eleven patients are alive and disease free at a median of 6 years (range, 3.0 to 9.0 years). Five-year overall survival, failure-free survival, disease progression rates, and treatment-related mortality (TRM) were 33%, 30%, 32%, and 38% respectively. Conclusion These data demonstrate that lasting remissions can be achieved after URD transplantation in patients with advanced CLL. High TRM suggest that myeloablative conditioning and HLA-mismatched donors should be avoided in future protocols, and it is mandatory to investigate transplant strategies with a lower morbidity and mortality, including the use of nonmyeloablative regimens.

The corpus cavernosum of the penis is one of the few vascular beds in which there is a change in oxygen tension with function (blood PO225-40mm Hg in the flaccid state, and 90-100mm Hg in the erect state). This change in oxygen tension exposes the components of the corpus cavernosum to a variety of cytokines, humoral, vasoactive, and growth factors which may affect the structure and function of the endothelial cells, smooth muscle cells, neurons and extracellular matrix. Among these cell types, endothelial cells are the first line of defense to blood-borne stress and can affect the underlying smooth muscle via paracrine mechanisms. Impotence is defined as the inability to obtain or sustain an erection sufficient for vaginal penetration and can result from a variety of pathological conditions, vascular disease, endocrine disease, neurological disease, and psychogenic disorders. The penis is a vascular organ and as such is susceptible to the effects of vascular diseases. This review will discuss the basic etiology of erection and vasculogenic erectile dysfunction and explore the role oxygen tension in regulating various cellular and humoral factors as well as trabecular structure and function.