Normal Tissue Damage Research Articles

Analysis of Peri-Implantitis Photothermal Therapy Effect According to Laser Irradiation Location and Angle: A Numerical Approach.

In recent years, dental implants have become increasingly popular around the world. However, if the implant is not properly managed, inflammation may occur, and the implant itself may need to be removed. Peri-implantitis is a common inflammation that occurs in dental implants, and various laser treatments have recently been studied to eliminate it. In this study, the situation of removing peri-implantitis using photothermal therapy, one of the various laser treatments, was analyzed theoretically and numerically. The temperature distribution in the tissue for various laser irradiation locations, angles, and power was calculated based on heat transfer theory, and the degree of thermal damage to tissue was analyzed using the Arrhenius damage integral. In addition, the thermally damaged region ratio of inflamed and normal tissue was analyzed using the Arrhenius thermal damage ratio and normal tissue Arrhenius thermal damage ratio to confirm the trend of treatment results for each treatment condition. The results of the study showed that if only the thermal damage to the inflamed tissue is considered, the laser should be angled vertically, and the laser should be applied to the center of the inflamed tissue rather than close to the implant. However, if the thermal damage to the surrounding normal tissue is also considered, it was found that the laser should be applied at 1.0 mm from the right end of the inflamed tissue for maximum effect. This will allow for more accurate clinical treatment of peri-implantitis in the future.

Hypoxia-responsive synthetic gene circuits to improve safety and potency of CAR T cell therapy for solid tumors.

37 Background: Engineered cell therapies expressing a chimeric antigen receptor (CAR) in human T cells have demonstrated great promise for the treatment of hematological malignancies. However, unique challenges remain in translating the success of CAR T cell therapies to solid tumors, namely: (1) specificity: there are few truly unique antigens present on solid tumors, leading to off-target effects that damage normal tissue and pose safety risks; and (2) suppression: solid tumors develop microenvironments that are immunosuppressive and decrease T cell survival and tumor-directed cytotoxicity. To address these challenges and improve the safety and potency of solid tumor cell therapy, we engineered cells with synthetic genetic circuits containing a previously validated hypoxia biosensor (HBS) that responds to upregulation of the native HIF1α and HIF2α transcription factors by hypoxia, enabling the cells to sense and respond to the hypoxic tumor microenvironment ubiquitous among solid tumors. Methods: We evaluated CAR expression from circuit architectures containing the HBS promoter driving expression of both CAR and native HIF1α, HIF2α, as well as from circuit architectures containing the HBS promoter and our toolkit of synthetic transcription factors and promoters. Circuits were integrated into HEK293 cells via PiggyBac transposon vector to generate stable cell lines. Cells were subsequently cultured in normoxia (21% O2) and hypoxia (1% O2), and harvested for flow cytometric analysis to evaluate CAR surface expression across a three day time frame. Results: We identified novel circuits providing fast reporter output with minimal background signal under hypoxic conditions, and elucidated design rules that enable or restrict amplification of transgene in native HIF1α and HIF2α-containing circuits. We observed differences in CAR transgene expression in hypoxia between circuits containing HIF1α and HIF2α-mediated feedback, suggesting mechanistic differences in regulation of HIF1α and HIF2α when overexpressed. Additionally, we observed greatly amplified transgene expression in circuit topologies containing feedback mediated by our synthetic transcription factor and promoter toolkit compared to native HIF1α and HIF2α-containing circuits and cells constitutively expressing CAR. Conclusions: We present a new therapeutic technology that senses hypoxic environments and enables high-output sense and respond behaviors with therapeutic outputs. We envision that this technology will enable development of safer, more effective cell therapies for solid tumors, as well as provide new biological and mechanistic insights that can inform the future development of hypoxia-responsive cell therapies.

Hypoxia-Responsive Prodrug of ATR Inhibitor, AZD6738, Selectively Eradicates Treatment-Resistant Cancer Cells.

Targeted therapy remains the future of anti-cancer drug development, owing to the lack of specificity of current treatments which lead to damage in healthy normal tissues. ATR inhibitors have in recent times demonstrated promising clinical potential, and are currently being evaluated in the clinic. However, despite the considerable optimism for clinical success of these inhibitors, reports of associated normal tissues toxicities remain a concern and can compromise their utility. Here, ICT10336 is reported, a newly developed hypoxia-responsive prodrug of ATR inhibitor, AZD6738, which is hypoxia-activated and specifically releases AZD6738 only in hypoxic conditions, in vitro. This hypoxia-selective release of AZD6738 inhibited ATR activation (T1989 and S428 phosphorylation) and subsequently abrogated HIF1a-mediated adaptation of hypoxic cancers cells, thus selectively inducing cell death in 2D and 3D cancer models. Importantly, in normal tissues, ICT10336 is demonstrated to be metabolically stable and less toxic to normal cells than its active parent agent, AZD6738. In addition, ICT10336 exhibited a superior and efficient multicellular penetration ability in 3D tumor models, and selectively eradicated cells at the hypoxic core compared to AZD6738. In summary, the preclinical data demonstrate a new strategy of tumor-targeted delivery of ATR inhibitors with significant potential of enhancing the therapeutic index.

Injectable Therapeutic Hydrogel with H2O2 Self-Supplying and GSH Consumption for Synergistic Chemodynamic/Low-Temperature Photothermal Inhibition of Postoperative Tumor Recurrence and Wound Infection.

Postoperative tumor recurrence and wound infection remain significant clinical challenges in surgery, often requiring adjuvant therapies. The combination treatment of photothermal therapy (PTT) and chemodynamic therapy (CDT) has proven to be effective in cancer treatment and wound infection. However, the hyperthermia during PTT increases the risk of normal tissue damage, severely impeding its application. Moreover, the efficacy of CDT is limited by insufficient hydrogen peroxide (H2O2) and excessive glutathione (GSH) levels at tumor or infection sites. Herein, an injectable and multifunctional CuO2@Au hydrogel system (CuO2@Au Gel) is developed for synergistic CDT and low-temperature PTT (LTPTT) to prevent tumor recurrence and bacterial wound infections. CuO2@Au Gel is constructed by embedding therapeutic CuO2@Au into low-melting point agarose hydrogel. In vitro and in vivo experiments confirm that the CuO2@Au in CuO2@Au Gel is capable of self-supplying H2O2 and depleting GSH, exhibiting effective CDT effect in acidic tumor or bacterial infected microenvironment. Additionally, it exhibits favorable photothermal conversion ability, inducing localized temperature elevation and synergistically enhancing CDT efficiency. The prepared CuO2@Au Gel demonstrates efficient tumor ablation capability in post-surgery recurrence mouse models and exhibits promising anti-infective efficiency in bacterial infection wound models, indicating significant potential in adjuvant therapy for post-surgical treatment and recovery.

Ti3C2Tx MXene nanosheet-based drug delivery/cascaded enzyme system for combination cancer therapy and anti-inflammation

Photothermal therapy (PTT)-based combination therapy is an effective approach to cancer treatment. However, there is a possibility of tumor development due to the inflammation caused by PTT. In addition, hyperthermia can cause damage to normal tissues. This work reported a drug delivery/cascade enzyme system based on Ti3C2Tx MXene nanosheets. Ultra-small, degradable mesoporous silica nanoparticles (DS-MSN) were prepared to load phloretin (Phl), which has anti-inflammatory, antioxidant, and anticancer properties. Subsequently, glucose oxidase (GOx) was adsorbed on the surface of DS-MSN. The nanoparticles were then modified on Ti3C2Tx MXene nanosheets and finally treated with polyethylene glycol (PEG) to obtain the multifunctional system named Ti3C2Tx-Phl@DS-MSN-GOx-PEG (TPDMGP). The high near-infrared photothermal conversion efficiency of Ti3C2Tx made it suitable for PTT. GOx and Ti3C2Tx formed a cascade enzyme system inside cancer cells: Ti3C2Tx exhibited catalase (CAT)-like activity that decomposed intracellular hydrogen peroxide (H2O2) into O2, which was provided to GOx for enhanced starvation therapy. GOx in the presence of O2 produces H2O2, for the oxygen production of Ti3C2Tx. Meanwhile, DS-MSN could be degraded in the reducing environment inside cancer cells as it contains disulfide bonds in its backbone, thus releasing Phl for chemotherapy. Such a combination therapy strategy could achieve a satisfactory therapeutic outcome by low-temperature PTT while evading normal tissue damage by high temperature. The anti-inflammatory effects of both Ti3C2Tx and Phl enabled TPDMGP to eliminate the inflammation caused by PTT via the scavenging of reactive oxygen species (ROS). The multifunctional system is expected to generate new paradigms for cancer PTT.

Potential of Ruthenium as a photosensitizer in radiation-activated Photodynamic Therapy (radioPDT)

In Photodynamic Therapy (PDT), light is used to activate photosensitizers (PS) to generate reactive oxygen species (ROS) as a cell-killing mechanism. However, PDT's application is limited to the surface and small size tumors because of the inefficiency of light in penetrating larger and deep-seated tumors. We have developed novel nanoparticles (NPs) that can be activated by radiation, a process known as the radiation-activated PDT (radioPDT). In our studies we developed a new variant of pegylated poly-lactic-co-glycolic (PEG-PLGA) encapsulated nanoscintillators (NSC) and ruthenium-based photosensitizer (Ru/rPDT) that exhibits minimal cell toxicity while having significant cell killing effect upon radiation activation. Our data suggest that radioPDT with Ru/rPDT can be a more effective strategy than previous protoporphyrin-based iterations in treating many deep-seated tumors.Introduction and Background: Toxicity and normal tissue damage are limiting factors for the application of radiotherapy (RT) where higher doses of radiation is required to cure deep-seated tumor such as prostate cancer. In Photodynamic Therapy (PDT), photosensitizers (PS) are activated by light to generate reactive oxygen species (ROS) to kill tumor cells. However, the main limitation of the current PDT is the limited tissue penetration of PS-activating light. Thus, PDT cannot be effectively used to treat deep-seated and larger size tumors. RT has higher tissue penetration depth, making the radiation-activated PDT (radioPDT) advantageous in treating deep seated tumors.We aim to develop novel radioPDT nanoparticles (NP) with more efficient anti-cancer therapeutic effect in vitro and in vivo to enhance targeted radiotherapy.Methods: LaF3:Ce3+ nanoscintillators (NSCs) were synthesized in a single step wet chemistry technique. The NSCs along with Ruthenium-based PS (Ru) with matching excitation/emission spectra were successfully encapsulated in PEG-PLGA. UV-Vis spectrometry and inductively coupled plasma mass spectrometry (ICP-MS) were used to confirm the presence of NSCs and Ru, and dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used to determine the size of NP. Singlet oxygen generation was measured by Singlet Oxygen Sensor Green (SOSG) and direct singlet oxygen fluorescence. RadioPDT therapeutic potential was evaluated in PC3 prostate cancer cell under radiation by Alamar Blue cell viability assay.Results: UV-Vis spectrometry and ICP-MS determined the successful PEG-PLGA encapsulation of NSC and Ru. The TEM imaging confirmed the encapsulation NSC inside the nanocarrier PEG-PLGA. TEM and DLS measurements showed the size of radioNP ranged from 90nm to 150nm with polydispersity index of <0.3. Singlet oxygen generation by Ru/rPDT NPs was greater than PPIX based radioPDT (PPIX/rPDT) NPs. Furthermore, the direct measurement of singlet oxygen generation with fluorescence spectroscopy of Ru/rPDT showed preserved singlet oxygen yield and no evidence of quenching. The Ru/rPDT and PPIX/rPDT showed minimal PC3 cell cytotoxicity in dark conditions. The PC3 cell killing efficiency was comparable between Ru/rPDT and PPIX/rPDT either under light or radiation activation.Conclusion: Our in vitro studies show Ru/rPDT significantly increases anti-cancer therapeutic effect of RT, with minimal toxicity. Future studies will refine in vivo NP biodistribution, dosing, and RT dose/fractionation schemes.

302 Diagnostic Evidence Gauge of Spatial Transcriptomics (DEGAS-ST): Using transfer learning to map clinical data to spatial transcriptomics in prostate cancer

OBJECTIVES/GOALS: The 'field effect' is a concept in pathology that pre-malignant tissue changes forecast health. Spatial transcriptomics could detect these changes earlier than histopathology, suggesting new early cancer screening methods. Knowing how normal tissue damage relates to cancer’s origin and progression may improve long-term outcomes. METHODS/STUDY POPULATION: We trained DEGAS, our machine learning framework, with prostate cancer data, combining both general cancer patterns and in-depth genetic information from individual tumors. The Tumor Cancer Genome Atlas (TCGA) shows how gene patterns in tumors relate to patient outcomes, emphasizing the differences between tumors from different patients (intertumor). On the other hand, spatial transcriptomics (ST) shows the genetic variety within a single tumor (intratumor) but has limited samples, making it hard to know which genetic differences are important for treatment. DEGAS bridges these areas by finding tissue sections that resemble those in TCGA profiles and are key indicators of patient survival. DEGAS serves as a valuable tool for generating clinically-important hypotheses. RESULTS/ANTICIPATED RESULTS: DEGAS identified benign-appearing glands in a normal prostate as being highly associated with poor progression-free survival. These glands have transcriptional signatures similar to high-grade prostate cancer. We confirmed this finding in a separate prostate cancer ST dataset. By integrating single cell (SC) data we demonstrated that cells annotated as cancerous in the SC data map to regions of benign glands in the ST dataset. We pinpoint several genes, chiefly Microseminoprotein-β (MSMB, PSP94), where reduced expression is highly correlated with poor progression-free survival. Cell type specific differential expression analysis further revealed that loss of MSMB expression associated with poor outcomes occurs specifically in luminal epithelia, the putative progenitor of prostate cancer. DISCUSSION/SIGNIFICANCE: DEGAS reveals that normal-appearing tissue can be highly-associated with tumor progression and underscores the importance of the 'field effect' in cancer research. Traditional analysis may miss such nuance, hiding key transitional cell states. Validating gene markers could boost early cancer detection and understanding of metastasis.

Abstract 1124: Prevention of radiation-induced malignancies in Li-Fraumeni syndrome

Abstract PURPOSE: Li-Fraumeni Syndrome (LFS) is a genetic disorder associated with a significant risk of early-onset cancer. This condition is driven by germline mutations in the TP53 gene which plays a primary role in the regulation of the radiation response. Aberrant TP53 function contributes to radiation vulnerability and a greater risk of secondary, radiation-induced malignancies (RIMs). As a result, therapeutic options for LFS patients are often limited to exclude radiotherapy (RT), which may otherwise be beneficial for the treatment of certain primary tumors. Data from our lab demonstrate an aberrant transcriptomic response to irradiation (IR) in mutant p53 patient skin fibroblasts compared to wildtype; however, it is unknown whether reprogramming this radiation response can decrease the risk of RIM in LFS. Metformin, a commonly prescribed anti-diabetic drug, is associated with lower cancer incidence and may decrease cancer-related mortality in murine LFS models. In addition to its potential anti-tumorigenicity, studies have shed light on the ability of metformin to prevent IR-induced damage in normal tissue; hence, we hypothesize that metformin can reprogram the radiation response to protect against radiation injury and delay the onset of RIM in LFS. METHODS: To establish a murine model of RIM in LFS, and to investigate whether metformin can delay tumor onset in this model, whole-body or localized IR were administered to mice harboring a hotspot Trp53R172H/+ mutation in the presence and absence of metformin. Serial MRI was conducted to monitor for tumor development. To understand the effect of metformin on the mutant p53 radiation response in vivo, a similar murine workflow was established and irradiated skin was collected longitudinally from untreated and metformin-treated cohorts for whole transcriptome sequencing. Sequencing data were functionally validated in a separate cohort of mice using flow cytometry. In parallel, we performed RNA sequencing on LFS patient fibroblasts to characterize the effect of metformin on the human radiation response. RESULTS: We demonstrate that IR decreases tumor latency in TP53R172H/+ mice, and that metformin significantly delays tumor onset within the radiation field. Moreover, transcriptomic data revealed that metformin upregulates apoptosis following IR in TP53R172H/+ mice. Flow cytometry analysis of the radiation response validated these findings, demonstrating that metformin promotes the apoptosis-driven clearance of damaged, potentially tumorigenic cells following IR. Transcriptomic data from patient cell lines will be presented. CONCLUSIONS: Overall, we show that metformin may delay RIM in LFS mice, and have begun to characterize the biology underpinning this reprogrammed response to IR. This study is the first to highlight metformin as a radioprotective agent in the context of germline mutant p53, with the potential to broaden RT treatment options for LFS patients. Citation Format: Pamela Psarianos, Nicholas Fischer, David Malkin. Prevention of radiation-induced malignancies in Li-Fraumeni syndrome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1124.

Abstract 7107: Clustered DNA double strand breaks for cancer treatment via multi-gRNA CRISPR system

Abstract Radiation therapy is one of the most common cancer treatments. Ionizing radiation generates DNA double-strand breaks (DSBs), which are the most lethal type of DNA damage. The dominant pathway for repairing DSBs is non-homologous end joining (NHEJ), but homologous recombination (HR) is also important. NHEJ is dominant pathway in all cell cycle phases while HR is largely restricted to S and G2 phases. Unrepaired or misrepaired DSBs can lead to cell death. KU70/80 and DNA-PKcs are essential for NHEJ. KU70/80 binds to broken ends, and DNA-PKcs binds to DNA-bound KU70/80, which activates DNA-PKs kinase allowing NHEJ to proceed. Clustered DSBs (two or more DSBs separated by &amp;lt;200 bp) generate small DNA fragments that are efficiently bound by KU70/80 and DNA-PKcs, but fail to activate DNA-PKcs kinase, inhibiting NHEJ and shifting repair toward HR. Thus, clustered DSBs are poorly repaired and more cytotoxic than dispersed DSBs. There are two types of ionization radiation: high LET and low LET. High LET radiation has a higher ionization density and therefore induces more clustered DSBs than low LET, which explains why high LET radiation is more cytotoxic at a given dose. A major limitation of radiotherapy is normal tissue damage. We hypothesize that mimicking high-LET-induced clustered DSBs using CRISPR/Cas9 targeted to cancer-specific DNA sequences (e.g., indel mutations) will effectively kill cancer cells but spare normal cells. In this approach, DSBs are not targeted to specific genes, nor is CRISPR/Cas9 used for gene editing. Instead, cancer cell killing is achieved by inducing poorly repaired DSBs anywhere in cancer cell DNA. Thus, this therapeutic approach is distinct from conventional targeted therapies that are oncogenic pathway-dependent, and this eliminates oncogenic pathway-dependent resistance mechanisms. However, types of resistance are possible, such as upregulated DSB repair. Our data indicate that the cytotoxicity of clustered DSBs depends on the number of small DNA fragments induced, thus we observe similar cytotoxicity with a localized, 4-DSB cluster and with three non-localized 2-DSB clusters, as each generates 3 small fragments per target chromosome. This finding significantly increases available targets as 2-DSB clusters can be induced at indels and flanking (wild-type) sites, generating clustered DSBs in cancer cells but efficiently repaired dispersed DSBs in normal cells lacking the cancer indel. Data also indicates that a single DSB targeted to each amplicon of highly amplified regions is also cytotoxic, and we are exploring the effectiveness of combination clustered DSBs targeted to amplified regions. We are also exploring how DSB repair inhibitors (like ATMi, DNA-PKcsi or RAD51i) can augment cytotoxic DSB effects, potentially mitigating that resistance pathway. These proof-of-principal studies represent the first approach to targeted cancer therapy that is oncogenic pathway-independent. Citation Format: Gamze Badakul, Neelam Sharma, Jac A. Nickoloff. Clustered DNA double strand breaks for cancer treatment via multi-gRNA CRISPR system [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7107.

Rosamultin ameliorates radiation injury via promoting DNA injury repair and suppressing oxidative stress in vitro and in vivo

Radiotherapy remains the preferred treatment option for cancer patients with the advantages of broad indications and significant therapeutic effects. However, ionizing radiation can also damage normal tissues. Unfortunately, there are few anti-radiation damage drugs available on the market for radiotherapy patients. Our previous study showed that rosamultin had antioxidant and hepatoprotective activities. However, its anti-radiation activity has not been evaluated. Irradiating small intestinal epithelial cells and mice with whole-body X-rays radiation were used to evaluate the in vitro and in vivo effects of rosamultin, respectively. Intragastric administration of rosamultin improved survival, limited leukocyte depletion, and reduced damage to the spleen and small intestine in irradiated mice. Rosamultin reversed the downregulation of the apoptotic protein BCL-2 and the upregulation of BAX in irradiated mouse small intestine tissue and in irradiation-induced small intestinal epithelial cells. DNA-PKcs antagonists reversed the promoting DNA repair effects of rosamulin. Detailed mechanistic studies revealed that rosamultin promoted Translin-associated protein X (TRAX) into the nucleus. Knockdown of TRAX reduced the protective effect of rosamultin against DNA damage. In addition, rosamultin reduced irradiation-induced oxidative stress through promoting Nrf2/HO-1 signaling pathway. To sum up, in vitro and in vivo experiments using genetic knockdown and pharmacological activation demonstrated that rosamultin exerts radioprotection via the TRAX/NHEJ and Nrf2/HO pathways.

FLASH Proton Radiation Therapy Mitigates Inflammatory and Fibrotic Pathways and Preserves Cardiac Function in a Preclinical Mouse Model of Radiation-Induced Heart Disease

PurposeStudies during the past 9 years suggest that delivering radiation at dose rates exceeding 40 Gy/s, known as “FLASH” radiotherapy, enhances the therapeutic index of radiation therapy (RT) by decreasing normal tissue damage while maintaining tumor response compared to conventional (or standard) RT. This study demonstrates the cardioprotective benefits of FLASH proton RT (F-PRT) compared to standard (conventional) proton RT (S-PRT), as evidenced by reduced acute and chronic cardiac toxicities. Methods and MaterialsMice were imaged using cone beam computed tomography to precisely determine the heart's apex as the beam isocenter. Irradiation was conducted utilizing a shoot-through technique with a 5mm diameter circular collimator. Bulk RNA-sequencing was performed on non-irradiated (NR) samples, as well as apexes treated with F-PRT or S-PRT, at 2 weeks following a single 40 Gy dose. Inflammatory responses were assessed through multiplex cytokine/chemokine microbead assay and immunofluorescence analyses. Levels of perivascular fibrosis were quantified using Masson's Trichrome and Picrosirius red staining. Additionally, cardiac tissue functionality was evaluated by 2D echocardiograms at 8 and 30 weeks post-PRT. ResultsRadiation damage was specifically localized to the heart's apex. RNA profiling of cardiac tissues treated with PRT revealed that S-PRT uniquely upregulated pathways associated with DNA damage response, induction of tumor necrosis factor superfamily and inflammatory response, while F-PRT primarily affected cytoplasmic translation, mitochondrion organization and ATP synthesis. Notably, F-PRT led to a milder inflammatory response, accompanied by significantly attenuated changes in TGF-β1 and αSMA levels. Critically, F-PRT decreased collagen deposition, and better preserved cardiac functionality compared to S-PRT. ConclusionsThis study demonstrated that F-PRT reduces the induction of an inflammatory environment with lower expression of inflammatory cytokines and profibrotic factors. Importantly, the results indicate that F-PRT better preserves cardiac functionality, as confirmed by echocardiography analysis, while also mitigating the development of long-term fibrosis.

High variability in short and long-term recovery kinetic of blood cell count and blood chemistry in a partial body irradiation mouse model

Purpose In the aftermath of a nuclear disaster or accident, survivors will suffer from radiation-induced normal tissue damage. Recovery after radiation exposure is dictated by several factors, one of which is degree of shielding at time of exposure. This study aims to characterize the short and late term changes in kinetics and magnitude of pancytopenia and blood chemistry in a model of heterogeneous radiation exposure, or partial body irradiation (PBI), compared to whole body irradiation (WBI). Materials and methods Male C57BL/6 mice, 8-10 weeks of age, were WBI at 6 different doses (6, 6.1. 6.15, 6.2, 6.5, and 7.5 Gy) to establish the LD50. To determine the effect of shielding on blood cell counts and chemistry, animals were either WBI at 6 Gy (LD2230) or 6 Gy PBI with one leg shielding (LD030). Complete blood counts and chemistry were measured at 1, 5-, 10-, 20-, 30- and 120-days post-irradiation. Results and conclusions Irradiated animals had significant depletion of white blood cells, red blood cells and platelets up to 10 days post-irradiation. Separation between PBI and WBI were observed at 10- and 20-days post-irradiation at which point PBI animals showed sign of recovery while overall cell count remains depleted in WBI animals up to 30 days post-irradiation. In addition, significant changes were found in parameters indicative of hematopoietic injury including hemoglobin count, hematocrit count and white blood cell population. Significant changes were observed in kidney function with changes to blood urea nitrogen and calcium concentration at 5-days post-irradiation. At 10-days post-irradiation. liver function changes differentiated WBI from PBI animals. Long-term, irradiated animal’s chemistry values and many blood counts were not significantly different from Sham. In conclusion, partial shielding ensured complete survival and demonstrated a different recovery kinetics of blood and chemistry parameters after irradiation compared to survivors of whole body irradiation and no single hemopoietic parameter was able to consistently differentiate irradiated from Sham animals. This seems to indicate that there is no single robust hemopoietic parameter to differentiate those exposed from those who were not due to the inherent variability in individual responses. Furthermore, there were no significant long-term effects on these blood parameters between survivors of WBI and PBI except that shielding accelerated recovery.

Embracing Personalized Strategies in Radiotherapy for Nasopharyngeal Carcinoma: Beyond the Conventional Bounds of Fields and Borders.

Radiotherapy is the primary treatment modality for non-metastatic nasopharyngeal carcinoma (NPC) across all TN-stages. Locoregional control rates have been impressive even from the 2D radiotherapy (RT) era, except when the ability to deliver optimal dose coverage to the tumor is compromised. However, short- and long-term complications following head and neck RT are potentially debilitating, and thus, there has been much research investigating technological advances in RT delivery over the past decades, with the primary goal of limiting normal tissue damage. On this note, with a plateau in gains of therapeutic ratio by modern RT techniques, future advances have to be focused on individualization of RT, both in terms of dose prescription and the delineation of target volumes. In this review, we analyzed the guidelines and evidence related to contouring methods, and dose prescription for early and locoregionally advanced (LA-) NPC. Next, with the preference for induction chemotherapy (IC) in patients with LA-NPC, we assessed the evidence concerning radiotherapy adaptations guided by IC response, as well as functional imaging and contour changes during treatment. Finally, we discussed on RT individualization that is guided by EBV DNA assessment, and its importance in the era of combinatorial immune checkpoint blockade therapy with RT.

GSH/pH Cascade-Responsive Nanoparticles Eliminate Methicillin-Resistant Staphylococcus aureus Biofilm via Synergistic Photo-Chemo Therapy.

Bacterial biofilm infection threatens public health, and efficient treatment strategies are urgently required. Phototherapy is a potential candidate, but it is limited because of the off-targeting property, vulnerable activity, and normal tissue damage. Herein, cascade-responsive nanoparticles (NPs) with a synergistic effect of phototherapy and chemotherapy are proposed for targeted elimination of biofilms. The NPs are fabricated by encapsulating IR780 in a polycarbonate-based polymer that contains disulfide bonds in the main chain and a Schiff-base bond connecting vancomycin (Van) pendants in the side chain (denoted as SP-Van@IR780 NPs). SP-Van@IR780 NPs specifically target bacterial biofilms in vitro and in vivo by the mediation of Van pendants. Subsequently, SP-Van@IR780 NPs are decomposed into small size and achieve deep biofilm penetration due to the cleavage of disulfide bonds in the presence of GSH. Thereafter, Van is then detached from the NPs because the Schiff base bonds are broken at low pH when SP@IR780 NPs penetrate into the interior of biofilm. The released Van and IR780 exhibit a robust synergistic effect of chemotherapy and phototherapy, strongly eliminate the biofilm both in vitro and in vivo. Therefore, these biocompatible SP-Van@IR780 NPs provide a new outlook for the therapy of bacterial biofilm infection.

Exploring Deep Learning for Estimating the Isoeffective Dose of FLASH Irradiation From Mouse Intestinal Histological Images

PurposeUltra-high dose rate (FLASH) irradiation has been reported to reduce normal tissue damage compared with conventional dose rate (CONV) irradiation without compromising tumor control. This proof-of-concept study aims to develop a deep learning (DL) approach to quantify the FLASH isoeffective dose (dose of CONV that would be required to produce the same effect as the given physical FLASH dose) with post-irradiation mouse intestinal histological images. Methods and materials84 healthy C57BL/6J female mice underwent 16 MeV electron CONV (0.12Gy/s; n=41) or FLASH (200Gy/s; n=43) single fraction whole abdominal irradiation. Physical dose ranged from 12 to 16Gy for FLASH and 11 to 15Gy for CONV in 1Gy increments. 4 days after irradiation, 9 jejunum cross-sections from each mouse were H&E stained and digitized for histological analysis. CONV dataset was randomly split into training (n=33) and testing (n=8) datasets. ResNet101-based DL models were retrained using the CONV training dataset to estimate the dose based on histological features. The classical manual crypt counting (CC) approach was implemented for model comparison. Cross-section-wise mean squared error (CS-MSE) was computed to evaluate the dose estimation accuracy of both approaches. The validated DL model was applied to the FLASH dataset to map the physical FLASH dose into the isoeffective dose. ResultsThe DL model achieved a CS-MSE of 0.20Gy2 on the CONV testing dataset compared with 0.40Gy2 of the CC approach. Isoeffective doses estimated by the DL model for FLASH doses of 12, 13, 14, 15, and 16 Gy were 12.19±0.46, 12.54±0.37, 12.69±0.26, 12.84±0.26, and 13.03±0.28 Gy, respectively. ConclusionsOur proposed DL model achieved accurate CONV dose estimation. The DL model results indicate that in the physical dose range of 13 to 16 Gy, the biological dose response of small intestinal tissue to FLASH irradiation is represented by a lower isoeffective dose compared to the physical dose. Our DL approach can be a tool for studying isoeffective doses of other radiation dose modifying interventions.

Microbial biosurfactant based nanoemulsions and their applications: A review

AbstractNanoemulsions exhibit a wide range of practical applications owing to their smaller particle size, stability, and enhanced bioavailability. Conventional surfactants widely used in nanoemulsion formulations namely, polysorbate 80, macrogolglycerol ricinoleate, and 4‐octylphenol polyethoxylate have been associated with anaphylactoid responses, alterations in normal flora, cytotoxicity and tissue damage. The use of less toxic, environment‐friendly microbial biosurfactants in the formulation of nanoemulsions have been demonstrated to manifest stability, slow drug release, enhanced skin permeability, and bioactivity. The type and composition of mixed microbial biosurfactants used in nanoemulsions can decrease the droplet size and synergistically increase the stability. Moreover, a lower concentration of drug‐loaded surfactin‐based nanoemulsion synergistically inhibited the proliferation of cancer cells by 50% (IC50). The scope of the current review is to focus on the prospect of substituting conventional surfactants with biosurfactants obtained from microbial origin for nanoemulsion formulations. It also aims to draw attention to the paucity of research in the combinatorial study of biosurfactants and functional drugs used in nanoemulsions. The review emphasizes the bioactivity of microbial biosurfactant‐based nanoemulsions and their applications in diverse sectors. The differences in emulsion stability and size of microbial biosurfactant and conventional surfactant‐based nanoemulsions have been highlighted. The study intends to promote comparative research between nanoemulsions prepared using conventional surfactants and microbial biosurfactants.

Dual Nucleosomal Double-Strand Breaks Are the Key Effectors of Curative Radiation Therapy

Most ionizing radiation produces δ-rays of ≈1 keV that can impart MGy doses to 100 nm3 volumes of DNA. These events can produce severe dual double-strand breaks (DDSBs) on nucleosomes, particularly in dense heterochromatic DNA. This is the most common multiply damaged site, and their probabilities determine the biological effectiveness of different types of radiation. We discuss their frequency, effect on cell survival, DNA repair, and imaging by gold nanoparticle tracers and electron microscopy. This new and valuable nanometer resolution information can be used for determining the optimal tumor cure by maximizing therapeutic effects on tumors and minimizing therapeutic effects on normal tissues. The production of DDSBs makes it important to deliver a rather high dose and LET to the tumor (&gt;2.5 Gy/Fr) and at the same time reach approximately 1.8–2.3 Gy of the lowest possible LET per fraction in TP53 intact normal tissues at risk. Therefore, their intrinsic low-dose hyper-sensitivity (LDHS)-related optimal daily fractionation window is utilized. Before full p53 activation of NHEJ and HR repair at ≈½ Gy, the low-dose apoptosis (LDA) and LDHS minimize normal tissue mutation probabilities. Ion therapy should thus ideally produce the lowest possible LET in normal tissues to avoid elevated DDSBs. Helium to boron ions can achieve this with higher-LET Bragg peaks, producing increased tumor DDSB densities. Interestingly, the highest probability of complication-free cure with boron or heavier ions requires a low LET round-up for the last 10–15 GyE, thereby steepening the dose response and further minimizing normal tissue damage. In conclusion, the new high-resolution DSB and DDSB diagnostic methods, and the new more accurate DNA-repair-based radiation biology, have been combined to increase our understanding of what is clinically important in curative radiation therapy. In fact, we must understand that we already passed the region of optimal LET and need to go back one step rather than forward, with oxygen being contemplated. As seen by the high overkill and severely high LET in the distal tumor and the increased LET to normal tissues (reminding of neutrons or neon ions), it is therefore preferable to use lithium–boron ions or combine carbon with an optimal 10–15 GyE photon, electron, or perhaps even a proton round-up, thus allowing optimized, fractionated, curative, almost complication-free treatments with photons, electrons, and light ions, introducing a real paradigm shift in curative radiation therapy with a potential 5 GyE tumor boost, 25% increase in complication-free cure and apoptotic–senescent Bragg Peak molecular light ion radiation therapy.

Targeted nuclear irradiation with a proton microbeam induces oxidative DNA base damage and triggers the recruitment of DNA glycosylases OGG1 and NTH1

DNA is the major target of radiation therapy of malignant tumors. Ionizing radiation (IR) induces a variety of DNA lesions, including chemically modified bases and strand breaks. The use of proton beam therapy for cancer treatment is ramping up, as it is expected to reduce normal tissue damage. Thus, it is important to understand the molecular mechanisms of recognition, signaling, and repair of DNA damage induced by protons in the perspective of assessing not only the risk associated with human exposure to IR but also the possibility to improve the efficacy of therapy. Here, we used targeted irradiation of nuclear regions of living cells with controlled number of protons at a high spatio-temporal resolution to detect the induced base lesions and characterize the recruitment kinetics of the specific DNA glycosylases to DNA damage sites. We show that localized irradiation with 4 MeV protons induces, in addition to DNA double strand breaks (DSBs), the oxidized bases 7,8-dihydro-8-oxoguanine (8-oxoG) and thymine glycol (TG) at the site of irradiation. Consistently, the DNA glycosylases OGG1 and NTH1, capable of excising 8-oxoG and TG, respectively, and initiating the base excision repair (BER) pathway, are recruited to the site of damage. To our knowledge, this is the first direct evidence indicating that proton microbeams induce oxidative base damage, and thus implicating BER in the repair of DNA lesions induced by protons.

Using SRIM code for calculating Bragg curve, projected range, straggling of 12C ion in biological materials

‘The Stopping and Range of Ions in Matter’ SRIM program, utilizing the built-in compound directory has been used to compute Bragg curve, projected range, longitudinal, lateral straggle of 12C ion with energies of 1 − 150 MeV in biological matter: liquid water (benchmark) human blood, skin, thyroid, and spleen are assessed. Results show that in the studied biological media for higher energy beam i): noticeable modification are seen in Bragg peak, shift in peak-position; ii) maximum projected range: 595.2 μm suggests effective use of 12C ions for cancer tumor therapy, especially for superficial cancer tumors, without much damage to healthier cells; iii) longitudinal straggling ~ 22.6 μm, low lateral straggling ~ 4.7 μm suggests potential application of 12C ions in treating cancer tumor nearby critical organs, causing minimal damage in nearby normal tissues.

Heterozygous variants of NOD2, IL10RA, PLA2G6 and COL7A1 correlate with Crohn's disease

To identify candidate pathogenic genes of early-stage Crohn's disease (CD) and predict potential roles of genetic factors in CD, we performed whole exome sequencing on a child with early-stage Crohn's disease (CD) and her parents (core family), found that the patient carried heterozygous variants of 4 genes: NOD2 c. 2257 C > T, IL10RA c. 301 C > T, PLA2G6 c. 2029 C > T, COL7A1 c. 3190 G > A. Heterozygous variants of NOD2, IL10RA, PLA2G6 and COL7A1, intestinal inflammatory response is triggered, normal intestinal wall tissue damage, leading to CD phenotype.