Characterization of design grammar of peptides for regulating liquid droplets and aggregates of FUS.

Liquid droplets of aggregation-prone proteins, which become hydrogels or form amyloid fibrils, are a potential target for drug discovery. In this study, we proposed an experiment-guided protocol for characterizing the design grammar of peptides that can regulate droplet formation and aggregation. The protocol essentially involves investigation of 19 amino acid additives and polymerization of the identified amino acids. As a proof of concept, we applied this protocol to fused in sarcoma (FUS). First, we evaluated 19 amino acid additives for an FUS solution and identified Arg and Tyr as suppressors of droplet formation. Molecular dynamics simulations suggested that the Arg additive interacts with specific residues of FUS, thereby inhibiting the cation-π and electrostatic interactions between the FUS molecules. Second, we observed that Arg polymers promote FUS droplet formation, unlike Arg monomers, by bridging the FUS molecules. Third, we found that the Arg additive suppressed solid aggregate formation of FUS, while Arg polymer enhanced it. Finally, we observed that amyloid-forming peptides induced the conversion of FUS droplets to solid aggregates of FUS. The developed protocol could be used for the primary design of peptides controlling liquid droplets and aggregates of proteins.

Allelic loss of 3p25 associated with alterations of 5q22.3 approximately q23.2 may affect the prognosis of conventional renal cell carcinoma.

Little is known about the clinical significance at the frequent association of 3p loss with 5q gain/loss in conventional renal cell carcinoma (RCC). We analyzed the clinical significance of copy number gain and loss at 5q21 approximately q23 combined with allelic loss of 3p25 (including the VHL gene). Fifty RCCs were examined by dual-color fluorescence in situ hybridization with DNA probes for D3Z1 (3cen), cCI3-865 (3p25.1 approximately p25.3), D5S23 (5p15.2), cCI5-243 (5q21.2 approximately q21.3), and cCI5-215 (5q22.3 approximately q23.2). In patients who had 3p loss, there was a significant association of loss at 5q22.3 approximately q23.2 with large tumors (>7 cm) and high-grade tumors (both P < 0.05), whereas gain at 5q22.3 approximately q23.2 was associated with low-grade tumors (P < 0.05). There was also a significant association loss at 5q21.2 approximately q21.3 high-grade tumors in patients with 3p loss (P < 0.05). Patients with 3p loss and gain at 5q22.3 approximately q23.2 had a significantly better disease-specific survival than those who had 3p loss without such gain (P < 0.05). Allelic loss of 3p25 including the VHL gene is thought to be an immediate event in the development of conventional RCC. Copy number gains or losses of 5q21 approximately q23 are thought to be events that lead to tumor progression although the clinical significance of either gains or losses is not well known.

Renal cell carcinoma: allelic loss at chromosome 9 using the fluorescent multiplex-polymerase chain reaction technique.

Loss of tumor-suppressor genes on both arms of chromosome 9 appears to be common in many types of cancer. Chromosome 9q is often partially deleted in bladder cancer, lung cancer, and basal cell carcinoma. However, little data are available on allelic loss on chromosome 9 in renal cell carcinoma (RCC). One hundred and nine nonpapillary RCCs were studied for loss of heterozygosity (LOH) at 13 loci on chromosome 9 by using the fluorescent multiplex-polymerase chain reaction method to compare DNA from tumor samples and peripheral blood lymphocytes. At the loci tested, LOH was found in from 2.3% (9q31, D9S938) to 17% (9q22, 1AJL) of informative cases, and 27 (24.8%) of the 109 RCCs had LOH at 1 or more loci of chromosome 9. LOH was more often detected at 9q22 within the PTCH gene (17%) when compared with LOH at the other 12 loci (P = 0.0172). Regarding the relationship with clinical parameters, however, there were no statistically significant associations between this LOH and tumor stage or grade. Among the 109 tumors, 6 (5.5%) showed replication errors. Our results suggest that LOH of the PTCH gene may be related to the development of nonpapillary RCC, although the clinical relevance has not been not clarified.

Bax to Bcl-2 ratio and Ki-67 index are useful predictors of neoadjuvant chemoradiation therapy in bladder cancer.

OBJECTIVE: In this study, locally advanced bladder cancer was treated by radiation combined with cisplatin therapy and a retrospective analysis was conducted to predict the clinical response to chemoradiotherapy (CRT) based on the immunohistochemistry of apoptosis-related proteins. METHODS: Sixty-two patients (median age, 68 years; range, 45-89 years) with transitional cell carcinoma of the bladder (pT1G3-pT4M0) treated with CRT (median dose: 40.5 Gy of radiation and 230 mg of cisplatin) were studied. Mucosal biopsy was performed before and after CRT. Paraffin-embedded tumor specimens were examined with TUNEL and were immunostained for Ki-67, p53, Bcl-2 and Bax; the Bax/Bcl-2 ratio and apoptosis index (AI) were calculated. Clinical features of the patients and response to CRT were compared with data obtained from examination of the tumors. RESULTS: The 62 patients had a median follow-up period of 34 months (range, 3-84 months). Responses to CRT were as follows: CR, 34%; PR, 45%; NC, 21%. The survival rate of patients with Ki-67-positive tumors was significantly lower than those of patients with Ki-67-negative tumors (P < 0.05). No significant correlation was observed between the expression of any protein, the AI and the clinical response. However, the Bax/Bcl-2 ratio showed a significant association with the CR rate (P = 0.0289). CONCLUSIONS: The results of this study suggest that the combined assessment of Bcl-2 and Bax protein expression may be used to predict a clinical response to CRT based on the Bax/Bcl-2 ratio determined before therapy. The Ki-67 index may be a useful predictor of prognosis in patients treated by CRT.