OBJECTIVETo investigate whether α-hemoglobin stabilizing protein (AHSP), the α-globin-specific molecular chaperone, is regulated by erythroid transcription factor NF-E2.

METHODSWe established the stable cell line with NF-E2p45 (the larger subunit of NF-E2) short hairpin RNA to silence its expression. Western blot, real-time polymerase chain reaction, and chromatin immunoprecipitation (ChIP) analysis were performed to detect the expression of AHSP, the histone modifications at AHSP gene locus, and the binding of GATA-1 at the AHSP promoter with NF-E2p45 deficiency. ChIP was also carried out in dimethyl sulfoxide (DMSO)-induced DS19 cells and estrogen-induced G1E-ER4 cells to examine NF-E2 binding to the AHSP gene locus and its changes during cell erythroid differentiation. Finally, luciferase assay was applied in HeLa cells transfected with AHSP promoter fragments to examine AHSP promoter activity in the presence of exogenous NF-E2p45.

RESULTSWe found that AHSP expression was highly dependent on NF-E2p45. NF-E2 bound to the regions across AHSP gene locus in vivo, and the transcription of AHSP was transactivated by exogenous NF-E2p45. In addition, we observed the decrease of H3K4 trimethylation and GATA-1 occupancy at the AHSP gene locus in NF-E2p45-deficient cells. Restoration of GATA-1 in G1E-ER4 cells in turn led to increased DNA binding of NF-E2p45.

CONCLUSIONNF-E2 may play an important role in AHSP gene regulation, providing new insights into the molecular mechanisms underlying the erythroid-specific expression of AHSP as well as new possibilities for β-thalassemia treatment.

Base Sequence ; Blood Proteins ; genetics ; DNA Primers ; GATA1 Transcription Factor ; physiology ; Gene Expression Regulation ; physiology ; Gene Silencing ; HeLa Cells ; Humans ; Methylation ; Molecular Chaperones ; genetics ; NF-E2 Transcription Factor, p45 Subunit ; physiology ; Promoter Regions, Genetic ; Reverse Transcriptase Polymerase Chain Reaction

Purpose: To report on the local control and toxicity of 5-fraction, high-conformal ultrafractionated radiation therapy (RT) for primary tumors in patients with metastatic breast cancer (MBC) who did not undergo planned surgical intervention. Methods: We retrospectively reviewed 27 patients with MBC who underwent 5-fraction high-dose ultrafractionated intensity-modulated RT for their primary tumors between 2017 and 2022 at our institution. A median dose of 66.8 Gy (range, 51.8–83.6 Gy) was prescribed to the gross tumor, calculated in 2-Gy equivalents using an α/β ratio of 3.5, along with a simultaneous integrated boost of 81.5%. The primary endpoint of this study was local control. Results: The median tumor size and volume were 5.1 cm and 112.4 cm3 , respectively. Treatment was generally well tolerated, with only 15% of the patients experiencing mild acute skin toxicity, which resolved spontaneously. The best infield response rate was 82%, with the objective response observed at a median time of 10.8 months post-RT (range, 1.4–29.2), until local progression or the last follow-up. At a median follow-up of 18.3 months, the 2-year local control rate was 77%. A higher number of prior lines of systemic therapy was significantly associated with poorer 2-year local control (one–two lines, 94% vs three or more lines, 34%; p = 0.004). Post-RT, 67% of the patients transitioned to the next line of systemic therapy, and the median duration of maintaining the same systemic therapy post-RT was 16.3 months (range, 1.9–40.3). Conclusion In our small dataset, 5-fraction, high-conformal ultrahypofractionated breast RT offered promising 2-year local control with minimal toxicity. Further studies are warranted to investigate the optimal dose and role in this setting.

Aged ; Aged, 80 and over ; Carcinoma, Hepatocellular ; diagnosis ; pathology ; surgery ; Humans ; Keratin-18 ; analysis ; Keratin-19 ; analysis ; Male ; Pancreatic Neoplasms ; diagnosis ; pathology ; surgery ; alpha-Fetoproteins ; analysis

BACKGROUNDWWOX and FHIT are two candidate tumor suppressor genes located in active fragile sites, the damage of which has been associated with the development of breast cancer. The association of the expression of these genes and the development of breast cancer has not been fully explored. We evaluated mRNA and protein expression of WWOX and FHIT in breast tissue with normal histological appearances, atypical ductal hyperplasia, ductal carcinoma in situ, and invasive cancer to see if a progressive decline in expression was present.

METHODSReverse transcription-polymerase chain reaction and Western blotting were used to evaluate the specimens for mRNA and protein expression, including 28 specimens with normal tissue, 28 specimens with atypical ductal hyperplasia, 33 specimens with ductal carcinoma in situ, and 51 specimens with invasive ductal carcinoma.

RESULTSCompared with in situ and invasive cancer specimens, both normal and atypical hyperplasia specimens had greater rates of detectable mRNA (WWOX rate ratio = 2.95, 95% CI 1.24 - 7.08; FHIT rate ratio = 4.58, 95% CI 1.82 - 11.81) and Western blotting detectable protein (WWOX rate ratio = 4.12, 95% CI 1.63 - 10.73; FHIT rate ratio = 3.76, 95% CI 1.44 - 10.06). For both proteins, differences between normal and atypical hyperplasia specimens and between in situ and invasive carcinoma specimens were explainable by chance (P > 0.05 for each analysis). Within each histological category, differences among fractions of specimens showed that FHIT and WWOX mRNA and protein expression were explainable by chance (P > 0.05 for each analysis).

CONCLUSIONExpression of FHIT and WWOX decreases along with breast tissue progress from a normal histological appearance to atypical ductal hyperplasia, in situ cancer, and the final invasive cancer.

Acid Anhydride Hydrolases ; analysis ; genetics ; Breast ; pathology ; Breast Neoplasms ; genetics ; Chromosome Fragile Sites ; Female ; Genes, Tumor Suppressor ; Humans ; Hyperplasia ; Neoplasm Proteins ; analysis ; genetics ; Oxidoreductases ; analysis ; genetics ; Tumor Suppressor Proteins ; analysis ; genetics ; WW Domain-Containing Oxidoreductase