Measurement of radiosensitivity in cervical tumours on the basis of the comet assay

PurposeThe aim of the study was the radiosensitivity assesment in squamous cell carcinoma (SCC) of the cervix on the basis of the comet assay in which the number of primary and residual DNA damage after 2 Gy dose of the radiation was measured.Material19 SCC were studied. The patients were not treated with chemo-or radiotherapy before biopsy.MethodSingle cell suspension from a biopsy was made by digesting with collagenase. The cell suspension was irradiated with doses 0–4 Gy. After the irradiation (initial DNA damage), or after 15 and 60 minuts of incubation at 37°C (residual DNA damage) cell suspension was mixed with poliakrylamide gel. Smears were made and cells were lysed with alkalic solution. Then electraphoresis was performed. The amount of damaged DNA stained with DAPI was measured with image analysis and Comet 3.0 pragramme. The measure of the DNA damage was tail moment, that is the length of comet tail and intesitivity of its fluorescence.ResultsThe differences in the number of primary (0 Gy), initial and residual DNA damage in the examined tumours were shown. Linear ralationship between number of initial DNA damage and radiation dose was obtained. Taxonomic analysis of initial DNA damage allowed for identification of 3 groups of patients of statisticaly different sensitivity. After 2 Gy dose of radiation, statisticaly differences in residual DNA damage after 0 and 15 minuts and 0 and 60 minuts were shown. The differences between patients were shown on the basis of the efficacy of the DNA damage repair (range 8.66%–91.73%).ConclusionThe comet assay seems to have the potential to be used as a predictive assay of individual radiosensitivity

Androgen receptor condensates as drug targets

Transcription factors are among the most attractive therapeutic targets, but are considered largely undruggable. Here we provide evidence that small molecule-mediated partitioning of the androgen receptor, an oncogenic transcription factor, into phase-separated condensates has therapeutic effect in prostate cancer models. We show that the phase separation capacity of the androgen receptor is driven by aromatic residues and short unstable helices in its intrinsically disordered activation domain. Based on this knowledge, we developed tool compounds that covalently attach aromatic moieties to cysteines in the receptors’ activation domain. The compounds enhanced partitioning of the receptor into condensates, facilitated degradation of the receptor, inhibited androgen receptor-dependent transcriptional programs, and had antitumorigenic effect in models of prostate cancer and castration-resistant prostate cancer in vitro and in vivo. These results establish a generalizable framework to target the phase- separation capacity of intrinsically disordered regions in oncogenic transcription factors and other disease-associated proteins with therapeutic intent

A glutamine-based single α-helix scaffold to target globular proteins

The binding of intrinsically disordered proteins to globular ones can require the folding of motifs into α-helices. These interactions offer opportunities for therapeutic intervention but their modulation with small molecules is challenging because they bury large surfaces. Linear peptides that display the residues that are key for binding can be targeted to globular proteins when they form stable helices, which in most cases requires their chemical modification. Here we present rules to design peptides that fold into single α-helices by instead concatenating glutamine side chain to main chain hydrogen bonds recently discovered in polyglutamine helices. The resulting peptides are uncharged, contain only natural amino acids, and their sequences can be optimized to interact with specific targets. Our results provide design rules to obtain single α-helices for a wide range of applications in protein engineering and drug design.We thank Luis Serrano for help with the Agadir predictions and helpful discussions, Ben Lehner and Ernest Giralt for helpful discussions and the ICTS NMR facility, managed by the scientific and technological centers of the University of Barcelona (CCiT UB), for their help in NMR. B.M. acknowledges funding from the Asociación Española contra el Cáncer (FCAECC project #POSTD211371MATE). C.G. acknowledges a graduate fellowship from MINECO (PRE2018-084684). M.S.-N. acknowledges funding from MINECO (PID2020-119810RB-I00). M.S.-N. holds a Ramón y Cajal contract (RYC2018-024759-I) from the Spanish Ministry of Science, Innovation, and Universities. X.S. acknowledges funding from AGAUR (2017 SGR 324), MINECO (BIO2015-70092-R and PID2019-110198RB-I00), and the European Research Council (CONCERT, contract number 648201). B.B.K acknowledges funding from the Novo Nordisk Foundation (#NNF18OC0033926). M.O. acknowledges funding from the Instituto Nacional de Bioinformática, The EU BioExcel Centre of Excellence for HPC and the Spanish Ministry of Science (PID2021-122478NB-I00) and the Instituto de Salud Carlos III–Instituto Nacional de Bioinformatica (ISCIII PT 17/0009/0007 co-funded by the Fondo Europeo de Desarrollo Regional). M.O. is an ICREA Academy scholar and J.A. is a Juan de la Cierva fellow. M.C. was supported by institutional funds of the Max Planck Society. This project has been carried out using the resources of CSUC. IRB Barcelona is the recipient of a Severo Ochoa Award of Excellence from MINECO (Government of Spain).Peer reviewe

Bromodeoxyuridine Labeling Index as an Indicator of Early Tumor Response to Preoperative Radiotherapy in Patients with Rectal Cancer

PURPOSE: Assessment of tumor proliferation rate using Bromodeoxyuridine labeling index (BrdUrdLI) as a possible predictor of rectal cancer response to preoperative radiotherapy (RT). METHODS AND MATERIAL: Ninety-two patients were qualified either to short RT (5 Gy/fraction/5 days) and surgery about 1 week after RT (schedule I), or to short RT and 4–5 weeks interval before surgery (schedule II). Tumor samples were taken twice from each patient: before RT and at the time of surgery. The samples were incubated with BrdUrd for 1 h at 37°C, and the BrdUrdLI was calculated as a percentage of BrdUrd-labeled cells. RESULTS: Thirty-eight patients were treated according to schedule I and 54 patients according to schedule II. Mean BrdUrdLI before RT was 8.5% and its value did not differ between the patients in the two compared groups. After RT tumors showed statistically significant growth inhibition (reduction of BrdUrdLI). As the pretreatment BrdUrd LI was not predictive for early clinical and pathologic tumor response, prognostic role of the ratio of BrdUrdLI after to BrdUrdLI before RT was considered. The ratios were calculated separately for fast (BrdUrd LI > 8.5%) and slowly (BrdUrd LI ≤ 8.5%) proliferating tumors and correlated with overall treatment time (OTT, i.e., time from the first day of RT to surgery). One month after RT, accelerated proliferation was observed only in slowly proliferating tumors. CONCLUSIONS: Pretreatment BrdUrdLI was not predictive for early clinical and pathologic tumor response. The ratio after/before RT BrdUrdLI was correlated to inhibition of proliferation in responsive tumors