Olaparib enhances sensitization of BRCA-proficient breast cancer cells to x-rays and protons.

PURPOSE: This study aimed to compare the radiosensitizing effect of the PARP inhibitor, Olaparib, between proton and X-rays irradiations in BRCA-proficient breast cancer (BC) cells. METHODS: Two BRCA-proficient BC cell lines, MDA-MB-231 and T47D BC, were used. Cell proliferation was assessed using the CCK-8 assay, and radiosensitivity was determined through the clonogenic survival assay. Flow cytometry was employed to analyze cell cycle distribution and apoptosis. The kinetics of DNA damage repair were evaluated using γH2AX immunofluorescence imaging and the comet assay. Tumor spheroid assays were conducted to test radiosensitivity in a three-dimensional culture condition. RESULTS: Olaparib sensitized both MDA-MB-231 and T47D cells to proton and X-ray irradiation in the clonogenic assay. MDA-MB-231 cells exhibited a higher dose enhancement factor for Olaparib than T47D cells. Olaparib increased radiation-induced G2/M cell cycle arrest and apoptosis specifically in MDA-MB-231 cells. γH2AX immunostaining and the comet assay showed Olaparib augmented radiation-induced DNA damage and apoptosis. The enhancement effect of Olaparib was more pronounced in proton irradiation than in X-ray irradiation, particularly in MDA-MB-231 cells than T47D cells. Both radiation and Olaparib dose-dependently inhibited spheroid growth in both cell lines. The synergy scores demonstrated that Olaparib interacted more strongly with protons than X-rays. The addition of an ATR inhibitor further enhanced Olaparib-induced proton radiosensitization in MDA-MB-231 cells. CONCLUSION: This study found that Olaparib enhanced radiation efficacy in BRCA-proficient breast cancer cells, with a more pronounced effect observed with proton irradiation compared to X-ray irradiation. Combining Olaparib with an ATR inhibitor increased the radiosensitizing effect of protons.

Multi spectral holographic ellipsometry for a complex 3D nanostructure.

We present an innovative ellipsometry technique called self-interferometric pupil ellipsometry (SIPE), which integrates self-interference and pupil microscopy techniques to provide the high metrology sensitivity required for metrology applications of advanced semiconductor devices. Due to its unique configuration, rich angle-resolved ellipsometric information from a single-shot hologram can be extracted, where the full spectral information corresponding to incident angles from 0° to 70° with azimuthal angles from 0° to 360° is obtained, simultaneously. The performance and capability of the SIPE system were fully validated for various samples including thin-film layers, complicated 3D structures, and on-cell overlay samples on the actual semiconductor wafers. The results show that the proposed SIPE system can achieve metrology sensitivity up to 0.123 nm. In addition, it provides small spot metrology capability by minimizing the illumination spot diameter up to 1 µm, while the typical spot diameter of the industry standard ellipsometry is around 30 µm. As a result of collecting a huge amount of angular spectral data, undesirable multiple parameter correlation can be significantly reduced, making SIPE ideally suited for solving several critical metrology challenges we are currently facing.

An Octopus-Derived Peptide with Antidiuretic Activity in Rats.

Discovering new drug candidates with high efficacy and few side effects is a major challenge in new drug development. The two evolutionarily related peptides oxytocin (OXT) and arginine vasopressin (AVP) are known to be associated with a variety of physiological and psychological processes via the association of OXT with three types of AVP receptors. Over decades, many synthetic analogs of these peptides have been designed and tested for therapeutic applications; however, only a few studies of their natural analogs have been performed. In this study, we investigated the bioactivity and usefulness of two natural OXT/AVP analogs that originate from the marine invertebrate Octopus vulgaris, named octopressin (OTP) and cephalotocin (CPT). By measuring the intracellular Ca2+ or cyclic AMP increase in each OXT/AVP receptor subtype-overexpressing cell, we found that CPT, but not OTP, acts as a selective agonist of human AVP type 1b and 2 receptors. This behavior is reminiscent of desmopressin, the most widely prescribed antidiuretic drug in the world. Similar to the case for desmopressin, a single intravenous tail injection of CPT into Sprague-Dawley rats reduced urine output and increased urinary osmolality. In conclusion, we suggest that CPT has a significant antidiuretic effect and that CPT might be beneficial for treating urological conditions such as nocturia, enuresis, and diabetes insipidus.

High-Throughput 3D Tumor Spheroid Array Platform for Evaluating Sensitivity of Proton-Drug Combinations.

Proton beam therapy (PBT) is a critical treatment modality for head and neck squamous cell carcinoma (HNSCC). However, not much is known about drug combinations that may improve the efficacy of PBT. This study aimed to test the feasibility of a three-dimensional (3D) tumor-spheroid-based high-throughput screening platform that could assess cellular sensitivity against PBT. Spheroids of two HNSCC cell lines-Fadu and Cal27-cultured with a mixture of Matrigel were arrayed on a 384-pillar/well plate, followed by exposure to graded doses of protons or targeted drugs including olaparib at various concentrations. Calcein staining of HNSCC spheroids revealed a dose-dependent decrease in cell viability for proton irradiation or multiple targeted drugs, and provided quantitative data that discriminated the sensitivity between the two HNSCC cell lines. The combined effect of protons and olaparib was assessed by calculating the combination index from the survival rates of 4 × 4 matrices, showing that Cal27 spheroids had greater synergy with olaparib than Fadu spheroids. In contrast, adavosertib did not synergize with protons in both spheroids. Taken together, we demonstrated that the 3D pillar/well array platform was a useful tool that provided rapid, quantitative data for evaluating sensitivity to PBT and drug combinations. Our results further supported that administration of the combination of PBT and olaparib may be an effective treatment strategy for HNSCC patients.

Immunomodulation of HDAC Inhibitor Entinostat Potentiates the Anticancer Effects of Radiation and PD-1 Blockade in the Murine Lewis Lung Carcinoma Model.

Although the combination of radiotherapy and immunotherapy has proven to be effective in lung cancer treatment, it may not be sufficient to fully activate the antitumor immune response. Here, we investigated whether entinostat, a histone deacetylase inhibitor, could improve the efficacy of radiotherapy and anti-PD-1 in a murine syngeneic LL/2 tumor model. A total of 12 Gy of X-rays administered in two fractions significantly delayed tumor growth in mice, which was further enhanced by oral entinostat administration. Flow cytometry-aided immune cell profiling revealed that entinostat increased radiation-induced infiltration of myeloid-derived suppressor cells and CD8+ T cells with decreased regulatory T-cells (Tregs). Transcriptomics-based immune phenotype prediction showed that entinostat potentiated radiation-activated pathways, such as JAK/STAT3/interferon-gamma (IFN-γ) and PD-1/PD-L1 signaling. Entinostat augmented the antitumor efficacy of radiation and anti-PD-1, which may be related to an increase in IFN-γ-producing CD8+ T-cells with a decrease in Treg cells. Comparative transcriptomic profiling predicted that entinostat increased the number of dendritic cells, B cells, and T cells in tumors treated with radiation and anti-PD-1 by inducing MHC-II genes. In conclusion, our findings provided insights into how entinostat improves the efficacy of ionizing radiation plus anti-PD-1 therapy and offered clues for developing new strategies for clinical trials.

Dual Oxidase 2 (DUOX2) as a Proteomic Biomarker for Predicting Treatment Response to Chemoradiation Therapy for Locally Advanced Rectal Cancer: Using High-Throughput Proteomic Analysis and Machine Learning Algorithm.

High-throughput mass-spectrometry-based quantitative proteomic analysis was performed using formalin-fixed, paraffin-embedded (FFPE) biopsy samples obtained before treatment from 13 patients with locally advanced rectal cancer (LARC), who were treated with concurrent chemoradiation therapy (CCRT) followed by surgery. Patients were divided into complete responder (CR) and non-complete responder (nCR) groups. Immunohistochemical (IHC) staining of 79 independent FFPE tissue samples was performed to validate the predictive ability of proteomic biomarker candidates. A total of 3637 proteins were identified, and the expression of 498 proteins was confirmed at significantly different levels (differentially expressed proteins-DEPs) between two groups. In Gene Ontology enrichment analyses, DEPs enriched in biological processes in the CR group included proteins linked to cytoskeletal organization, immune response processes, and vesicle-associated protein transport processes, whereas DEPs in the nCR group were associated with biosynthesis, transcription, and translation processes. Dual oxidase 2 (DUOX2) was selected as the most predictive biomarker in machine learning algorithm analysis. Further IHC validation ultimately confirmed DUOX2 as a potential biomarker for predicting the response of nCR to CCRT. In conclusion, this study suggests that the treatment response to RT may be affected by the pre-treatment tumor microenvironment. DUOX2 is a potential biomarker for the early prediction of nCR after CCRT.

Spatiotemporal characterisation of ischaemic lesions in transient stroke animal models using diffusion free water elimination and mapping MRI with echo time dependence.

BACKGROUND AND PURPOSE: The excess fluid as a result of vasogenic oedema and the subsequent tissue cavitation obscure the microstructural characterisation of ischaemic tissue by conventional diffusion and relaxometry MRI. They lead to a pseudo-normalisation of the water diffusivity and transverse relaxation time maps in the subacute and chronic phases of stroke. Within the context of diffusion MRI, the free water elimination and mapping method (FWE) with echo time dependence has been proposed as a promising approach to measure the amount of free fluid in brain tissue robustly and to eliminate its biasing effect on other biomarkers. In this longitudinal study of transient middle cerebral artery occlusion (MCAo) in the rat brain, we investigated the use of FWE MRI with echo time dependence for the characterisation of the tissue microstructure and explored the potential of the free water fraction as a novel biomarker of ischaemic tissue condition. METHODS: Adult rats received a transient MCAo. Diffusion- and transverse relaxation-weighted MRI experiments were performed longitudinally, pre-occlusion and on days 1, 3, 4, 5, 6, 7 and 10 after MCAo on four rats. Histology was performed for non-stroke and 1, 3 and 10 days after MCAo on three different rats at each time point. RESULTS: The free water fraction was homogeneously increased in the ischaemic cortex one day after stroke. Between three and ten days after stroke, the core of the ischaemic tissue showed a progressive normalisation in the amount of free water, whereas the inner and outer border zones of the ischaemic cortex depicted a large, monotonous increase with time. The specific lesions in brain sections were verified by H&E and immunostaining. The tissue-specific diffusion and relaxometry MRI metrics in the ischaemic cortex were significantly different compared to their conventional counterpart. CONCLUSIONS: Our results demonstrate that the free water fraction in FWE MRI with echo time dependence is a valuable biomarker, sensitive to the progressive degeneration in ischaemic tissue. We showed that part of the heterogeneity previously observed in conventional parameter maps can be accounted for by a heterogeneous distribution of free water in the tissue. Our results suggest that the temporal evolution of the free fluid fraction map at the core and inner border zone can be associated with the pathological changes linked to the evolution of vasogenic oedema. Namely, the homogeneous increase in free water one day after stroke and its tendency to normalise in the core of the ischaemic cortex starting three days after stroke, followed by a progressive increase in free water at the inner border zone from three to ten days after stroke. Finally, the monotonous increase in free fluid in the outer border zone of the cortex reflects the formation of fluid-filled cysts.

Magnetic resonance spectroscopy with transcranial direct current stimulation to explore the underlying biochemical and physiological mechanism of the human brain: A systematic review.

A large body of molecular and neurophysiological evidence connects synaptic plasticity to specific functions and energy metabolism in particular areas of the brain. Furthermore, altered plasticity and energy regulation has been associated with a number of neuropsychiatric disorders. A favourable approach enabling the modulation of neuronal excitability and energy in humans is to stimulate the brain using transcranial direct current stimulation (tDCS) and then to observe the effect on neurometabolites using magnetic resonance spectroscopy (MRS). In this way, a well-defined modulation of brain energy and excitability can be achieved using a dedicated tDCS protocol to a predetermined brain region. This systematic review was guided by the preferred reporting items for systematic reviews and meta-analysis and summarises recent literature studying the effect of tDCS on neurometabolites in the human brain as measured by proton or phosphorus MRS. Limitations and recommendations are discussed for future research. The findings of this review provide clear evidence for the potential of using tDCS and MRS to examine and understand the effect of neurometabolites in the in vivo human brain.

Incorporating sarcopenia and inflammation with radiation therapy in patients with hepatocellular carcinoma treated with nivolumab.

BACKGROUND: We investigated the combined effects of sarcopenia and inflammation on outcomes in patients with HCC treated with nivolumab. MATERIALS AND METHODS: We reviewed 102 patients treated with nivolumab between 2017 and 2018. Sarcopenia was diagnosed when the L3 skeletal muscle indices were < 42 cm2/m2 and < 38 cm2/m2 in men and women, respectively. Baseline neutrophil-to-lymphocyte ratio (NLR) and absolute lymphocyte count were used as surrogate markers of inflammation and immune cell reservoir. High NLR (hNLR) was defined as NLR ≥ 3, and severe lymphopenia (sLP) was defined as lymphocyte < 800/µL. The overall survival (OS) and progression-free survival (PFS) were analyzed. RESULTS: With a median follow-up of 21.9 (interquartile range, 8.3-58.3) months, patients with sarcopenia showed shorter OS than those without sarcopenia (median, 2.9 vs. 7.5 months, respectively). Patients with either hNLR or sLP exhibited inferior survival than those without risk factor (median OS, 2.8 vs. 14.5 months; median PFS, 1.3 vs. 3.7 months, respectively). Among 70 patients treated with RT, benefit of RT was observed in patients with sarcopenia or those without hNLR/sLP (all p < 0.05). After multivariable analysis, RT, hNLR/sLP, albumin-bilirubin (ALBI) grade, and alpha-fetoprotein were significantly associated with OS (all p < 0.05), and hNLR/sLP was also associated with decreased PFS together with ALBI grade, alpha-fetoprotein, and RT (all p < 0.05). CONCLUSION: The current study hypothetically demonstrated that the risk group stratified by hNLR/sLP outweighs the significance of sarcopenia in predicting outcomes after nivolumab. Furthermore, patients with sarcopenia might benefit from RT, especially those without risk factors of hNLR/sLP.

Isolation of Scalarane-Type Sesterterpenoids from the Marine Sponge Dysidea sp. and Stereochemical Reassignment of 12-epi-Phyllactone D/E.

The chemical investigation of the marine sponge Dysidea sp., which was collected from Bohol province in the Philippines, resulted in the identification of 15 new scalarane-type sesterterpenoids (1-14, 16), together with 15 known compounds. The chemical structures of the new compounds were elucidated based on NMR spectroscopy and HRMS. The structure of 12-epi-phyllactone D/E (15) isolated during this study was originally identified in 2007. However, careful inspection of our experimental 13C NMR spectrum revealed considerable discrepancies with the reported data at C-9, C-12, C-14, and C-23, leading to the correction of the reported compound to the C-12 epimer of 15, phyllactone D/E. The biological properties of compounds 1-16 were evaluated using the MDA-MB-231 cancer cell line. Compound 7, which bears a pentenone E-ring, exhibits significant cytotoxicity with a GI50 value of 4.21 µM.

Effect of photobiomodulation therapy on radiodermatitis in a mouse model: an experimental animal study.

This study aimed to evaluate the effect of photobiomodulation (PBM) for prevention of radiodermatitis in an irradiated mouse model and compare the efficacy of PBM using 633- or 830-nm wavelengths. Irradiated mice were randomly distributed into three groups: A (633 nm), B (830 nm), and C (without PBM). On post-irradiation days 7 and 21, we compared acute damage and recovery in treated skin samples to non-irradiated skin using H&E, Masson's trichrome, anti-CD45 and PCNA immunohistochemistry, and a TUNEL assay. Grade 3 radiodermatitis was evident only in group C. Compared with that in group C, the skin in groups A and B had significantly less epidermal hyperplasia, inflammatory cell infiltration, and thinner dermis on day 7 and less inflammatory cell infiltration, fewer apoptotic cells, and thinner dermis on day 21. However, there was no significant difference between groups A and B. This study indicates PBM could prevent severe radiodermatitis by reducing epidermal and dermal damage, inflammation, and apoptosis. There was no difference in PBM efficacy between the 633- and 830-nm wavelengths.

Manganese Ferrite Nanoparticles Enhance the Sensitivity of Hepa1-6 Hepatocellular Carcinoma to Radiation by Remodeling Tumor Microenvironments.

We evaluated the effect of manganese ferrite nanoparticles (MFN) on radiosensitization and immunologic responses using the murine hepatoma cell line Hepa1-6 and the syngeneic mouse model. The clonogenic survival of Hepa1-6 cells was increased by hypoxia, while being restricted by ionizing radiation (IR) and/or MFN. Although MFN suppressed HIF-1α under hypoxia, the combination of IR and MFN enhanced apoptosis and DNA damage in Hepa1-6 cells. In the Hepa1-6 syngeneic mouse model, the combination of IR and MFN notably limited the tumor growth compared to the single treatment with IR or MFN, and also triggered more frequent apoptosis in tumor tissues than that observed under other conditions. Increased expression of PD-L1 after IR was not observed with MFN alone or the combination of IR and MFN in vitro and in vivo, and the percentage of tumor-infiltrating T cells and cytotoxic T cells increased with MFN, regardless of IR, in the Hepa1-6 syngeneic mouse model, while IR alone led to T cell depletion. MFN might have the potential to overcome radioresistance by alleviating hypoxia and strengthening antitumor immunity in the tumor microenvironment.

Dedicated diffusion phantoms for the investigation of free water elimination and mapping: insights into the influence of T2 relaxation properties.

Conventional diffusion-weighted (DW) MRI suffers from free water contamination due to the finite voxel size. The most common case of free water contamination occurs with cerebrospinal fluid (CSF) in voxels located at the CSF-tissue interface, such as at the ventricles in the human brain. Another case refers to intra-tissue free water as in vasogenic oedema. In order to avoid the bias in diffusion metrics, several multi-compartment methods have been introduced, which explicitly model the presence of a free water compartment. However, fitting multi-compartment models in DW MRI represents a well known ill conditioned problem. Although during the last decade great effort has been devoted to mitigating this estimation problem, the research field remains active. The aim of this work is to introduce the design, characterise the NMR properties and demonstrate the use of two dedicated anisotropic diffusion fibre phantoms, useful for the study of free water elimination (FWE) and mapping models. In particular, we investigate the recently proposed FWE diffusion tensor imaging approach, which takes explicit account of differences in the transverse relaxation times between the free water and tissue compartments.

Therapeutic Potential of (-)-Agelamide D, a Diterpene Alkaloid from the Marine Sponge Agelas sp., as a Natural Radiosensitizer in Hepatocellular Carcinoma Models.

Radiation therapy (RT) is an effective local treatment for unresectable hepatocellular carcinoma (HCC), but there are currently no predictive biomarkers to guide treatment decision for RT or adjuvant systemic drugs to be combined with RT for HCC patients. Previously, we reported that extracts of the marine sponge Agelas sp. may contain a natural radiosensitizer for HCC treatment. In this study, we isolated (-)-agelamide D from Agelas extract and investigated the mechanism underlying its radiosensitization. (-)-Agelamide D enhanced radiation sensitivity of Hep3B cells with decreased clonogenic survival and increased apoptotic cell death. Furthermore, (-)-agelamide D increased the expression of protein kinase RNA-like endoplasmic reticulum kinase/inositol-requiring enzyme 1α/activating transcription factor 4 (PERK/eIF2α/ATF4), a key pathway of the unfolded protein response (UPR) in multiple HCC cell lines, and augmented radiation-induced UPR signaling. In vivo xenograft experiments confirmed that (-)-agelamide D enhanced tumor growth inhibition by radiation without systemic toxicity. Immunohistochemistry results showed that (-)-agelamide D further increased radiation-induced ATF4 expression and apoptotic cell death, which was consistent with our in vitro finding. Collectively, our results provide preclinical evidence that the use of UPR inducers such as (-)-agelamide D may enhance the efficacy of RT in HCC management.

Topical Application of Phlorotannins from Brown Seaweed Mitigates Radiation Dermatitis in a Mouse Model.

Radiation dermatitis (RD) is one of the most common side effects of radiotherapy; its symptoms progress from erythema to dry and moist desquamation, leading to the deterioration of the patients' quality of life. Active metabolites in brown seaweed, including phlorotannins (PTNs), show anti-inflammatory activities; however, their medical use is limited. Here, we investigated the effects of PTNs in a mouse model of RD in vivo. X-rays (36 Gy) were delivered in three fractions to the hind legs of BALB/c mice. Macroscopic RD scoring revealed that PTNs significantly mitigated RD compared with the vehicle control. Histopathological analyses of skin tissues revealed that PTNs decreased epidermal and dermal thickness compared with the vehicle control. Western blotting indicated that PTNs augmented nuclear factor erythroid 2-related factor 2 (NRF2)/heme oxygenase-1 (HO-1) pathway activation but attenuated radiation-induced NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) and inflammasome activation, suggesting the mitigation of acute inflammation in irradiated mouse skin. PTNs also facilitated fast recovery, as indicated by increased aquaporin 3 expression and decreased γH2AX (histone family member X) expression. Our results indicate that topical PTN application may alleviate RD symptoms by suppressing oxidative stress and inflammatory signaling and by promoting the healing process. Therefore, PTNs may show great potential as cosmeceuticals for patients with cancer suffering from radiation-induced inflammatory side effects such as RD.

Oxygenated Theonellastrols: Interpretation of Unusual Chemical Behaviors Using Quantum Mechanical Calculations and Stereochemical Reassignment of 7α-Hydroxytheonellasterol.

A total of eight new oxygenated 4-exo-methylene sterols, 1-8, together with one artifact 9 and six known sterols 11-16, were isolated from the marine sponge Theonella swinhoei collected from the Bohol province in Philippines. Structures of sterols 1-8 were determined from 1D and 2D NMR data. Among the sterols, 8α-hydroxytheonellasterol (4) spontaneously underwent an allylic 1,3-hydroxyl shift to produce 15α-hydroxytheonellasterol (9) as an artifact; this was rationalized by quantum mechanical calculations of the transition state. In addition, the 1,2-epoxy alcohol subunit of 8α-hydroxy-14,15-ß-epoxytheonellasterol (5) was assigned using the Gauge-Independent Atomic Orbital (GIAO) NMR chemical shift calculations and subsequent DP4+ analysis. Finally, comparison of the 13C chemical shifts of isolated 7α-hydroxytheonellasterol (6) with the reported values revealed significant discrepancies at C-6, C-7, C-8, and C-14, leading to reassignment of the C-7 stereochemistry in the known structure.

Checkpoint Kinase 1 (CHK1) Inhibition Enhances the Sensitivity of Triple-Negative Breast Cancer Cells to Proton Irradiation via Rad51 Downregulation.

Due to a superior dose conformity to the target, proton beam therapy (PBT) continues to rise in popularity. Recently, considerable efforts have been directed toward discovering treatment options for use in combination with PBT. This study aimed to investigate the targeting of checkpoint kinase 1 (CHK1), a critical player regulating the G2/M checkpoint, as a promising strategy to potentiate PBT in human triple-negative breast cancer (TNBC) cells. Protons induced cell-cycle arrest at the G2/M checkpoint more readily in response to increased CHK1 activation than X-rays. A clonogenic survival assay revealed that CHK1 inhibition using PF-477736 or small interfering RNA (siRNA) enhanced the sensitivity toward protons to a greater extent than toward X-rays. Western blotting demonstrated that PF-477736 treatment in the background of proton irradiation increased the pro-apoptotic signaling, which was further supported by flow cytometry using annexin V. Immunofluorescence revealed that proton-induced DNA double-strand breaks (DSBs) were further enhanced by PF-477736, which was linked to the downregulation of Rad51, essential for the homologous recombination repair of DSBs. Direct inactivation of Rad51 resulted in enhanced proton sensitization. Collectively, these data suggest that targeting CHK1 may be a promising approach for improving PBT efficacy in the treatment of TNBC.

Cyclin D1 is Associated with Radiosensitivity of Triple-Negative Breast Cancer Cells to Proton Beam Irradiation.

Proton therapy offers a distinct physical advantage over conventional X-ray therapy, but its biological advantages remain understudied. In this study, we aimed to identify genetic factors that contribute to proton sensitivity in breast cancer (BC). Therefore, we screened relative biological effectiveness (RBE) of 230 MeV protons, compared to 6 MV X-rays, in ten human BC cell lines, including five triple-negative breast cancer (TNBC) cell lines. Clonogenic survival assays revealed a wide range of proton RBE across the BC cell lines, with one out of ten BC cell lines having an RBE significantly different from the traditional generic RBE of 1.1. An abundance of cyclin D1 was associated with proton RBE. Downregulation of RB1 by siRNA or a CDK4/6 inhibitor increased proton sensitivity but not proton RBE. Instead, the depletion of cyclin D1 increased proton RBE in two TNBC cell lines, including MDA-MB-231 and Hs578T cells. Conversely, overexpression of cyclin D1 decreased the proton RBE in cyclin D1-deficient BT-549 cells. The depletion of cyclin D1 impaired proton-induced RAD51 foci formation in MDA-MB-231 cells. Taken together, this study provides important clues about the cyclin D1-CDK4-RB1 pathway as a potential target for proton beam therapy in TNBC.

Comparison of Proton and Photon Beam Irradiation in Radiation-Induced Intestinal Injury Using a Mouse Model.

When radiotherapy is applied to the abdomen or pelvis, normal tissue toxicity in the gastrointestinal (GI) tract is considered a major dose-limiting factor. Proton beam therapy has a specific advantage in terms of reduced doses to normal tissues. This study investigated the fundamental differences between proton- and X-ray-induced intestinal injuries in mouse models. C57BL/6J mice were irradiated with 6-MV X-rays or 230-MeV protons and were sacrificed after 84 h. The number of surviving crypts per circumference of the jejunum was identified using Hematoxylin and Eosin staining. Diverse intestinal stem cell (ISC) populations and apoptotic cells were analyzed using immunohistochemistry (IHC) and a terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay, respectively. The crypt microcolony assay revealed a radiation-dose-dependent decrease in the number of regenerative crypts in the mouse jejunum; proton irradiation was more effective than X-ray irradiation with a relative biological effectiveness of 1.14. The jejunum is the most sensitive to radiations, followed by the ileum and the colon. Both types of radiation therapy decreased the number of radiosensitive, active cycling ISC populations. However, a higher number of radioresistant, reserve ISC populations and Paneth cells were eradicated by proton irradiation than X-ray irradiation, as shown in the IHC analyses. The TUNEL assay revealed that proton irradiation was more effective in enhancing apoptotic cell death than X-ray irradiation. This study conducted a detailed analysis on the effects of proton irradiation versus X-ray irradiation on intestinal crypt regeneration in mouse models. Our findings revealed that proton irradiation has a direct effect on ISC populations, which may result in an increase in the risk of GI toxicity during proton beam therapy.