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»: Optimal care for pathologic fractures centers on the use of a multidisciplinary team; thus, whenever there is a concern for pathologic fracture and proper workup is unable to be performed, prompt referral to a center equipped to manage these injuries should occur., »: Fixation strategies for pathologic fractures must take into account patient characteristics, cancer subtypes, and overall goals of treatment., »: As the treatments of cancers improve, patient life expectancy with disease will improve as well. This will lead to an increase in the incidence of impending or completed pathologic fractures. The broader subspecialties of orthopaedics must be aware of general principles in the diagnosis and management of these injuries., Competing Interests: Disclosure: The Disclosure of Potential Conflicts of Interest forms are provided with the online version of the article (http://links.lww.com/JBJSREV/A892)., (Copyright © 2023 by The Journal of Bone and Joint Surgery, Incorporated.)

Personalized treatment approaches for patients with limited liver metastases from colorectal cancer are critically needed. By leveraging three large, independent cohorts of patients with colorectal liver metastases (n = 336), we found that a proliferative subtype associated with elevated CIN70 scores is linked to immune exclusion, increased metastatic proclivity, and inferior overall survival in colorectal liver metastases; however, high CIN70 scores generate a therapeutic vulnerability to DNA-damaging therapies leading to improved treatment responses. We propose CIN70 as a candidate biomarker to personalize systemic treatment options for patients with metastatic colorectal cancer. These findings are potentially broadly applicable to other human cancers., (© 2022. The Author(s).)

Homologous recombination (HR) serves multiple roles in DNA repair that are essential for maintaining genomic stability, including double-strand DNA break (DSB) repair. The central HR protein, RAD51, is frequently overexpressed in human malignancies, thereby elevating HR proficiency and promoting resistance to DNA-damaging therapies. Here, we find that the non-canonical NF-κB factors p100/52, but not RelB, control the expression of RAD51 in various human cancer subtypes. While p100/p52 depletion inhibits HR function in human tumor cells, it does not significantly influence the proficiency of non-homologous end joining, the other key mechanism of DSB repair. Clonogenic survival assays were performed using a pair DLD-1 cell lines that differ only in their expression of the key HR protein BRCA2. Targeted silencing of p100/p52 sensitizes the HR-competent cells to camptothecin, while sensitization is absent in HR-deficient control cells. These results suggest that p100/p52-dependent signaling specifically controls HR activity in cancer cells. Since non-canonical NF-κB signaling is known to be activated after various forms of genomic crisis, compensatory HR upregulation may represent a natural consequence of DNA damage. We propose that p100/p52-dependent signaling represents a promising oncologic target in combination with DNA-damaging treatments., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Purpose: Cumulative evidence suggests that the addition of platinum agents as neoadjuvant chemotherapy (NACT) could improve the pathologic complete response (pCR) rate in triple-negative breast cancer (TNBC). We aimed to develop a DNA homologous recombination (HR)-associated gene expression score to predict tumor sensitivity to platinum-based NACT in TNBC., Methods: A retrospective cohort of 127 patients who were diagnosed with TNBC and received platinum-based NACT in Fudan University Shanghai Cancer Center from 2012 to 2017 was included in this study. Using quantitative reverse transcription-polymerase chain reaction (qRT-PCR), the expression levels of eight HR-associated genes were analyzed from formalin-fixed paraffin-embedded core-needle biopsy samples obtained before NACT. A random forest model was built to estimate the weight of each gene expression level and clinicopathological factors. The training set was used to modulate parameters and select the best model. The performance of the final model was evaluated in the validation set., Results: A 4-gene (BRCA1, XRCC5, PARP1, and RAD51) scoring system was developed. TNBC patients with a higher score had a nearly fourfold likelihood of achieving pCR to platinum-based NACT compared with patients with a lower score [odds ratio (OR) = 3.878; P < 0.001]. At the cutoff value of - 2.644, the 4-gene scoring system showed high sensitivity in predicting pCR in the breast (93.0%) and pCR in the breast/axilla (91.8%), while at the cutoff value of - 1.969, the 4-gene score showed high specificity for pCR in the breast (85.7%) and pCR in the breast/axilla (80.8%)., Conclusion: The qRT-PCR-based 4-gene score has the potential to predict pCR to platinum-based NACT in TNBC., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Homologous recombination (HR) is a primary DNA double-strand breaks (DSBs) repair mechanism. The recombinases Rad51 and Dmc1 are highly conserved in the RecA family; Rad51 is mainly responsible for DNA repair in somatic cells during mitosis while Dmc1 only works during meiosis in germ cells. This spatiotemporal difference is probably due to their distinctive mismatch tolerance during HR: Rad51 does not permit HR in the presence of mismatches, whereas Dmc1 can tolerate certain mismatches. Here, the cryo-EM structures of Rad51-DNA and Dmc1-DNA complexes revealed that the major conformational differences between these two proteins are located in their Loop2 regions, which contain invading single-stranded DNA (ssDNA) binding residues and double-stranded DNA (dsDNA) complementary strand binding residues, stabilizing ssDNA and dsDNA in presynaptic and postsynaptic complexes, respectively. By combining molecular dynamic simulation and single-molecule FRET assays, we identified that V273 and D274 in the Loop2 region of human RAD51 (hRAD51), corresponding to P274 and G275 of human DMC1 (hDMC1), are the key residues regulating mismatch tolerance during strand exchange in HR. This HR accuracy control mechanism provides mechanistic insights into the specific roles of Rad51 and Dmc1 in DNA double-strand break repair and may shed light on the regulatory mechanism of genetic recombination in mitosis and meiosis., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Immune checkpoint blockade (ICB) improves outcomes in non-small cell lung cancer (NSCLC) though most patients progress. There are limited data regarding molecular predictors of progression. In particular, there is controversy regarding the role of CDKN2A loss-of-function (LOF) in ICB resistance. We analyzed 139 consecutive patients with advanced NSCLC who underwent NGS prior to ICB initiation to explore the association of CDKN2A LOF with clinical outcomes. 73% were PD-L1 positive (≥ 1%). 48% exhibited high TMB (≥ 10 mutations/megabase). CDKN2A LOF was present in 26% of patients and was associated with inferior PFS (multivariate hazard ratio [MVA-HR] 1.66, 95% CI 1.02-2.63, p = 0.041) and OS (MVA-HR 2.08, 95% CI 1.21-3.49, p = 0.0087) when compared to wild-type (WT) patients. These findings held in patients with high TMB (median OS, LOF vs. WT 10.5 vs. 22.3 months; p = 0.069) and PD-L1 ≥ 50% (median OS, LOF vs. WT 11.1 vs. 24.2 months; p = 0.020), as well as in an independent dataset. CDKN2A LOF vs. WT tumors were twice as likely to experience disease progression following ICB (46% vs. 21%; p = 0.021). CDKN2A LOF negatively impacts clinical outcomes in advanced NSCLC treated with ICB, even in high PD-L1 and high TMB tumors. This novel finding should be prospectively validated and presents a potential therapeutic target., (© 2021. The Author(s).)

Competing Interests: Competing interests: None declared.

Purpose: To determine the results of pars plana vitrectomy for giant retinal tear detachments using transscleral diode laser retinopexy and short-term postoperative tamponade with perfluoro-n-octane (PFnO)., Methods: Twenty consecutive patients with fresh giant retinal tears were enrolled in a single-arm prospective study. One case was withdrawn for technical reasons. The remainder all underwent pars plana vitrectomy, PFnO injection, transscleral diode laser retinopexy to the edge of the giant retinal tear, and short-term postoperative heavy liquid tamponade. None of the cases had scleral buckling or lensectomy., Results: Nineteen cases (18 male and 1 female) with a mean age of 41 years (range 10-69 years) were followed up for a period of 6 months. Postoperative tamponade with PFnO was maintained for a mean of 7.6 days (range 4-21 days), after which it was exchanged for sulfur hexafluoride (SF6), perfluoropropane (C3F8) gas, or balanced salt solution. Final reattachment rate was 100%, with 3 (15.7%) patients requiring additional surgery. Best-corrected visual acuity at final follow-up was 20/40 or better in 11 eyes (58%), between 20/60 and 20/200 in 7 (37%), and 20/400 in 1 (5%)., Conclusion: In this series of acute giant retinal tears, transscleral diode laser retinopexy together with the use of PFnO for short-term postoperative tamponade achieved excellent anatomical and visual results.

Background and Aims: To describe rhegmatogenous retinal detachment surgery in a Dublin tertiary referral centre over a 4-year period and to examine factors predictive of surgical and anatomical success., Methods: A retrospective study was performed by reviewing the surgical log and the charts of patients who underwent a primary retinal detachment repair in a specialist centre over a 4-year period between 2012 and 2015. We excluded exudative and tractional cases. Multiple parameters were recorded including macular status, visual acuity, type of procedure, complications and visual and anatomical outcomes at 6 months post operation. Primary reattachment rate was calculated alongside change in visual acuity. Chi-square testing and analysis of variance were utilized to determine the effect which lens status, macular status, extent of breaks and type of procedure had on outcome and visual success., Results: A total of 613 primary rhegmatogenous retinal detachment repairs were performed over the 4-year period. Our primary reattachment rate was calculated to be 88.58% (annual range 85.25-91.30%) with a perioperative complication rate of 2.94%. We noted a statistical significant improvement in VA with a median logMAR VA of 1.0 at presentation to 0.5 at 6 months post procedure. Macular status (chi-square test p = 0.15, X 2  = 2.072) and lens status (chi-square test p = 0.2974, X 2  = 1.086) had no statistical effect on the final anatomical outcome. However, eyes with giant retinal tears were more likely to redetach than those without (chi-square test p = 0.0069, X 2  = 7.3). There was no statistical significant difference in the proportion of eyes achieving visual success by surgery category (one-way ANOVA analysis p = 0.501)., Conclusions: This is the first study of its kind in Ireland and will help surgeons benchmark their results against international standards in the future. Accurate recording of logMAR acuity and intraoperative complications is imperative to assist with prospective studies.

Purpose: To report the outcomes of eyes receiving surgical management for traumatic macular holes. To describe the preoperative and postoperative optical coherence tomography features of traumatic macular holes and to explore associations between preoperative clinical and optical coherence tomography features, and visual outcome., Methods: Retrospective study of patients undergoing vitrectomy for traumatic macular hole and entered into the Australian and New Zealand Society of Retinal Specialists surgical registry. Preoperative clinical data, surgical details, and 3-month postoperative outcomes were recorded prospectively. Longer-term outcomes at 12 months were requested retrospectively, as were preoperative and postoperative optical coherence tomography scans., Results: Hole closure was achieved in 91% (21/23) of patients with a single procedure. The average preoperative visual acuity was 20/120. At 3 months postoperatively, the mean visual acuity had improved to 20/70 (P < 0.001), 11/23 (48%) of eyes improved ≥15 letters, and the number of eyes with 20/40 acuity or better increased from 1/23 to 7/23. Eyes with worse visual outcomes (visual acuity < 20/80) had larger holes, worse preoperative acuity, and a greater extent of preoperative ellipsoid band attenuation than those with better postoperative visual acuity., Conclusion: Eyes receiving surgical management for traumatic macular hole achieved good anatomical results and approximately half had a substantial improvement in acuity. Ellipsoid band attenuation on preoperative optical coherence tomography and worse preoperative acuity were associated with poorer visual outcomes.

Case: Heterotopic ossification (HO) is a pathological formation of bone in nonosseous tissue and is a common complication of orthopaedic procedures. Radiotherapy is often used to prevent HO despite the small risk of secondary malignancy. Here, we report a case of a patient who developed a periprosthetic, radiation-induced sarcoma after delivery of a single fraction of 7 Gy for HO prophylaxis. This sarcoma was found to lie entirely within the treatment field and occurred within 5 years of radiation., Conclusion: Appropriate counseling regarding radiation-induced sarcoma formation should be provided to patients considering radiotherapy for this HO prophylaxis.

RAD51 is the central protein in homologous recombination (HR) repair, where it first binds ssDNA and then catalyzes strand invasion via a D-loop intermediate. Additionally, RAD51 plays a role in faithful DNA replication by protecting stalled replication forks; this requires RAD51 to bind DNA but may not require the strand invasion activity of RAD51. We previously described a small-molecule inhibitor of RAD51 named RI(dl)-2 (RAD51 inhibitor of D-loop formation #2, hereafter called 2 h), which inhibits D-loop activity while sparing ssDNA binding. However, 2 h is limited in its ability to inhibit HR in vivo, preventing only about 50 % of total HR events in cells. We sought to improve upon this by performing a structure-activity relationship (SAR) campaign for more potent analogues of 2 h. Most compounds were prepared from 1-(2-aminophenyl)pyrroles by forming the quinoxaline moiety either by condensation with aldehydes, then dehydrogenation of the resulting 4,5-dihydro intermediates, or by condensation with N,N'-carbonyldiimidazole, chlorination, and installation of the 4-substituent through Suzuki-Miyaura coupling. Many analogues exhibited enhanced activity against human RAD51, but in several of these compounds the increased inhibition was due to the introduction of dsDNA intercalation activity. We developed a sensitive assay to measure dsDNA intercalation, and identified two analogues of 2 h that promote complete HR inhibition in cells while exerting minimal intercalation activity., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Maternal embryonic leucine zipper kinase (MELK) activates pathways that mediate aggressive tumor growth and therapy resistance in many types of adult cancers. Pharmacologic and genomic inhibition of MELK impairs tumor growth and increases sensitivity to radiation and chemotherapy. On the basis of these promising preclinical studies, early-phase adult clinical trials testing the MELK inhibitor OTS167 are ongoing. To investigate whether MELK is also a therapeutic target in neuroblastoma, we analyzed MELK expression in primary tumors and cell lines, and examined the effects of OTS167 on neuroblastoma growth. In primary tumors, high levels of MELK were associated with advanced stage disease and inferior survival. Higher levels of MELK were also detected in tumorigenic versus nontumorigenic neuroblastoma cell lines, and cells with higher levels of MELK expression were more sensitive to OTS167 than low-MELK expressing cells. OTS167 suppressed the growth of neuroblastoma xenografts, and in a preclinical model of minimal residual disease, survival was prolonged with MELK inhibition. OTS167 treatment downregulated MELK and its target enhancer of zeste homolog 2 (EZH2), a component of the polycomb repressive complex 2 (PRC2) that is known to modulate the DNA damage response. We also show that OTS167 reduced the formation of collapsed replication forks induced by camptothecin or radiation. Taken together, our results indicate that MELK indirectly mediates efficient processing of replication-associated DNA lesions in neuroblastoma, and that OTS167 sensitizes cells to DNA-damaging agents by abrogating this process. Further studies evaluating the activity of combination treatment regimens with OTS167 in neuroblastoma are warranted., (©2019 American Association for Cancer Research.)

Purpose: To present the visual and safety outcomes of surgery for primary idiopathic macular holes including predictors of visual acuity and the impact of combined phacovitrectomy surgery., Design: Registry-style, prospective, nonrandomized, observational cohort study., Participants: Patients with idiopathic macular holes undergoing primary surgery., Methods: Surgeons were invited to submit clinical details of all macular hole cases receiving surgery. Baseline demographic and clinical information, and details of surgical intervention were collected. Primary follow-up data were collected at 3 months postoperatively or before revision surgery, and surgeons were also asked to submit data at 12 and 24 months postoperatively., Main Outcome Measures: Visual acuity improvement ≥15 letters and ≥0 letters, change in mean visual acuity, visual acuity ≥70 letters (20/40), retinal detachment, and endophthalmitis., Results: A total of 2455 eyes of 2366 patients were included in the study, and hole closure was achieved in 95.6% of eyes with a single procedure. Mean baseline vision was 48.3 letters. The proportion of successful eyes improving ≥15 letters at 3, 12, and 24 months was 59.1%, 69.4%, and 68.2%, respectively. The mean improvement in acuity at 3, 12, and 24 months was 16.0, 19.2, and 23.6 letters, and 92.4%, 93.4%, and 95.8% improved ≥0 letters at 3, 12, and 24 months, respectively. Eyes receiving SF6 gas had better visual acuities at all time points postoperatively (adjusted effect 3.4, 3.1, and 4.6 letters better at 3, 12, and 24 months vs. longer-acting gas, respectively). Combined phacovitrectomy in phakic eyes was associated with better corrected visual acuity postoperatively (vs. vitrectomy surgery alone), a difference that vanished when eyes went on to have subsequent cataract surgery. The rate of retinal detachment postoperatively was 1.3%, and the odds of detachment were greater in eyes receiving longer-acting gases versus SF6 gas (adjusted odds ratio, 2.2; 95% confidence interval, 1.04-4.77; P = 0.039). There were no reported cases of endophthalmitis., Conclusions: Macular hole surgery substantially improved acuity in approximately two thirds of patients and was seldom associated with loss of vision. SF6 gas was associated with better visual outcomes, an effect that warrants further study., (Copyright © 2018 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

The human retina is a complex structure of organised layers of specialised cells that support the transmission of light signals to the visual cortex. The outermost layer of the retina, the retinal pigment epithelium (RPE), forms part of the blood retina barrier and is implicated in many retinal diseases. Lysophosphatidic acid (LPA) is a bioactive lipid exerting pleiotropic effects in various cell types, during development, normal physiology and disease. Its producing enzyme, AUTOTAXIN (ATX), is highly expressed by the pigmented epithelia of the human eye, including the RPE. Using human pluripotent stem cell (hPSC)-derived retinal cells, we interrogated the role of LPA in the human RPE and photoreceptors. hPSC-derived RPE cells express and synthesize functional ATX, which is predominantly secreted apically of the RPE, suggesting it acts in a paracrine manner to regulate photoreceptor function. In RPE cells, LPA regulates tight junctions, in a receptor-dependent mechanism, with an increase in OCCLUDIN and ZONULA OCCLUDENS (ZO)-1 expression at the cell membrane, accompanied by an increase in the transepithelial resistance of the epithelium. High concentration of LPA decreases phagocytosis of photoreceptor outer segments by the RPE. In hPSC-derived photoreceptors, LPA induces morphological rearrangements by modulating the actin myosin cytoskeleton, as evidenced by Myosin Light Chain l membrane relocation. Collectively, our data suggests an important role of LPA in the integrity and functionality of the healthy retina and blood retina barrier., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: In this study we sought to determine if staging endoscopic bronchial ultrasound (EBUS) improves outcomes in stage I non-small-cell lung cancer (NSCLC) patients who received hypofractionated radiation therapy (HFRT)., Patients and Methods: Patients with stage I NSCLC treated with HFRT from 2008 to 2015 were retrospectively identified from 3 affiliated institutions. All patients underwent positron emission tomography/computed tomography staging and a subset of patients received pretreatment EBUS. Patients with and without pre-radiation therapy EBUS were compared for baseline characteristics. The log rank test was used to compare Kaplan-Meier estimates. Univariate analysis (UVA) and multivariable analysis (MVA) were used to analyze the effect of factors on disease-free survival (DFS) and overall survival (OS)., Results: Ninety-two patients met study criteria. Median follow-up for the entire cohort was 21 months. Two-year DFS and OS were 63% and 81%, respectively. Two-year freedom from local, regional, and distant failure were 93%, 87%, and 87%, respectively. Thirty-seven of 92 patients (40%) received pretreatment EBUS. There were no statistically significant differences in 2-year freedom from regional failure rates, DFS, or OS for EBUS-staged versus non-EBUS-staged patients. On UVA, smaller tumor size (P = .03) and higher performance status (P = .05) were associated with improved OS. On MVA, tumor size retained significance for improved OS (hazard ratio [HR], 0.44; 95% confidence interval [CI], 0.19-0.97; P = .04) and higher performance status showed a trend toward improved OS (HR, 0.51; 95% CI, 0.23-1.11; P = .09)., Conclusion: In this retrospective series, we did not detect a difference in regional failure or survival outcomes among stage I NSCLC patients who received invasive staging with EBUS before HFRT., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Through an Irish Health Service Executive (HSE) initiative to tackle excessive hospital outpatient waiting times, 996 patients referred to the Ophthalmology Outpatient Department (OPD) of the Mater Misericordiae University Hospital (MMUH), Eccles Street, Dublin 7, Ireland, were outsourced to a community medical eye clinic (CMEC) for consultation with specialist-registered ophthalmologists., Aims: The study aims to determine if patients referred as routine to the OPD department could be managed in a community setting., Methods: 996 patients were reviewed in the CMEC, and their data was collected and placed into a spreadsheet for analysis., Results: 61.2% of patients referred to the OPD were fully managed in the community clinic, and 34.9% required ophthalmic surgery in hospital., Conclusions: By facilitating direct listing of some of the surgical patients to the hospital theatre list, 89.8% of the 996 referrals received treatment without needing to attend the hospital outpatients department.

Purpose: Molecular-based cancer tests have been developed to augment the standard clinical and pathologic features used to tailor treatments to individual breast cancer patients. Homologous recombination (HR) repairs double-stranded DNA breaks and promotes tolerance to lesions that disrupt DNA replication. Recombination Proficiency Score (RPS) quantifies HR efficiency based on the expression of four genes involved in DNA damage repair. We hypothesized low RPS values can identify HR-deficient breast cancers most sensitive to DNA-damaging chemotherapy. Experimental Design: We collected pathologic tumor responses and tumor gene expression values for breast cancer patients that were prospectively enrolled on clinical trials involving preoperative chemotherapy followed by surgery ( N = 513). We developed an algorithm to calculate breast cancer-specific RPS (RPS b ) values on an individual sample basis. Results: Low RPS b tumors are approximately twice as likely to exhibit a complete pathologic response or minimal residual disease to preoperative anthracycline-based chemotherapy as compared with high RPS b tumors. Basal, HER2-enriched, and luminal B breast cancer subtypes exhibit low RPS b values. In addition, RPS b predicts treatment responsiveness after controlling for clinical and pathologic features, as well as intrinsic breast subtype. Conclusions: Overall, our findings indicate that low RPS breast cancers exhibit aggressive features at baseline, but they have heightened sensitivity to DNA-damaging chemotherapy. Low RPS b values in basal, HER2-enriched, and luminal B subtypes provide a mechanistic explanation for their clinical behaviors and genomic instability. RPS b augments standard clinical and pathologic features used to tailor treatments, thereby enabling more personalized treatment strategies for individual breast cancer patients. Clin Cancer Res; 23(15); 4493-500. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Homologous recombination (HR) is an evolutionarily conserved DNA repair process. Overexpression of the key HR protein RAD51 is a common feature of malignant cells. RAD51 plays two distinct genome-stabilizing roles, including HR-mediated repair of double-strand breaks (DSBs) and the promotion of replication fork stability during replication stress. Because upregulation of RAD51 in cancer cells can promote tumor resistance to DNA-damaging oncologic therapies, we and others have worked to develop cancer therapeutics that target various aspects of RAD51 protein function. Herein, we provide an overview of recent developments in this field, together with our perspectives on the challenges associated with these evolving anticancer strategies., (© 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Purpose: To determine the prevalence of glaucoma among patients referred to a glaucoma service with suspicious disc photographs from the diabetic retinopathy (DR) screening program., Methods: A clinical audit of all patients attending a single-center DR screening program in the Mater Misericordiae University Hospital, Dublin, between July 2010 and October 2011 was conducted with a minimum follow-up of 30 months. The DR screening service uses trained technician graders to assess 2-field color retinal photographs for the features of DR. Recently, the service was enhanced so that optic discs are also assessed for signs of glaucoma., Results: In the 16-month study period, 3,697 diabetic patients were photographed. Following photograph grading, 91 (2.46%) were judged to require referral for assessment at the glaucoma clinic. Of these, 63 (69.23%) presented for assessment. Thirteen patients (20.63%) were diagnosed with glaucoma, comprising 7 cases of low-tension glaucoma and 6 cases of primary open-angle glaucoma. Thirty-six patients (57.14%) were classified as glaucoma suspects and 14 patients (22.22%) were discharged back to the DR screening program following normal ocular examination. Only 6 (9.52%) of the 63 patients examined had an intraocular pressure greater than 21 mm Hg., Conclusions: The assessment of DR screening photographs for signs of glaucomatous optic nerve damage should be considered as part of a strategy to improve glaucoma case detection and to reduce the burden of this disease on society.

RAD51 is the central protein in homologous recombination (HR) DNA repair and represents a therapeutic target in oncology. Herein we report a novel class of RAD51 inhibitors that were identified by high throughput screening. In contrast to many previously reported RAD51 inhibitors, our lead compound 1 is capable of blocking RAD51-mediated D-loop formation (IC50 21.3 ± 7.8 μM) at concentrations that do not influence RAD51 binding to ssDNA. In human cells, 1 inhibits HR (IC50 13.1 ± 1.6 μM) without blocking RAD51's ability to assemble into subnuclear foci at sites of DNA damage. We determined that the active constituent of 1 is actually an oxidized derivative (termed RI(dl)-1 or 8) of the original screening compound. Our SAR campaign also yielded RI(dl)-2 (hereafter termed 9h), which effectively blocks RAD51's D-loop activity in biochemical systems (IC50 11.1 ± 1.3 μM) and inhibits HR activity in human cells (IC50 3.0 ± 1.8 μM).

Purpose: To determine whether sulfur hexafluoride (SF6) gas is noninferior to longer-acting gases in macular hole surgery and whether withholding postoperative face-down positioning (FDP) is noninferior to FDP., Design: Registry-style, prospective, nonrandomized, observational cohort study., Participants: Patients with idiopathic macular holes undergoing primary surgery., Methods: Surgeons were invited to submit clinical details of all macular hole cases receiving surgery. Baseline demographic and clinical information were collected, as well as details of surgical intervention and postoperative posturing advice. Primary follow-up data were collected 3 months postoperatively., Main Outcome Measures: Macular hole closure at 3 months. A noninferiority approach was used, with a noninferiority margin set at 5% decreased frequency of success., Results: A total of 2456 eyes of 2367 patients were included in the study. Outcomes were available in 94.9% of cases (2330/2456). The rate of macular hole closure was 95.0% (2214/2330). Sulfur hexafluoride gas was found to be noninferior to longer-acting gases (95% confidence interval [CI] for adjusted effect on success, -1.76 to +2.25), and noninferiority was demonstrated regardless of macular hole size. Although withholding FDP was found to be noninferior to FDP for the study population as a whole (95% CI for adjusted effect on success, -4.21 to +0.64), the result was inconclusive in holes >400 μm in diameter (95% CI, -9.31 to +1.04). Lack of internal limiting membrane (ILM) peel, increasing hole size, hole duration ≥9 months, increasing age, and 20-gauge surgery all were associated with lower odds of success. Vitreous attachment to the hole margin was not associated with outcome when corrected for hole size, and combined phacovitrectomy surgery was not observed to affect the odds of success in phakic eyes., Conclusions: Sulfur hexafluoride gas tamponade was noninferior to longer-acting gases in the surgical management of macular hole. Withholding FDP was noninferior to FDP in holes ≤400 μm in diameter. In holes >400 μm in diameter, noninferiority of withholding FDP could not be concluded. We would advise caution if posturing is withheld in this group on the basis of the results of this study and of others., (Copyright © 2016 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Most pediatric head and neck cancers are treated with radiotherapy, but the morbidity associated with radiotherapy has become a prominent issue. This article discusses the common long-term complications associated with head and neck radiotherapy for childhood cancers. It reviews approaches to minimize toxicity and details the toxicities that head and neck radiation inflicts on relevant functional measures. In addition, it discusses the risk of radiation-induced secondary cancers in childhood cancer survivors, as well as strategies to reduce them. Thus, this article addresses approaches to minimize long-term radiation toxicities in order to improve the quality of life for childhood cancer survivors., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The RAD54 family DNA translocases have several biochemical activities. One activity, demonstrated previously for the budding yeast translocases, is ATPase-dependent disruption of RAD51-dsDNA binding. This activity is thought to promote dissociation of RAD51 from heteroduplex DNA following strand exchange during homologous recombination. In addition, previous experiments in budding yeast have shown that the same activity of Rad54 removes Rad51 from undamaged sites on chromosomes; mutants lacking Rad54 accumulate nonrepair-associated complexes that can block growth and lead to chromosome loss. Here, we show that human RAD54 also promotes the dissociation of RAD51 from dsDNA and not ssDNA. We also show that translocase depletion in tumor cell lines leads to the accumulation of RAD51 on chromosomes, forming complexes that are not associated with markers of DNA damage. We further show that combined depletion of RAD54L and RAD54B and/or artificial induction of RAD51 overexpression blocks replication and promotes chromosome segregation defects. These results support a model in which RAD54L and RAD54B counteract genome-destabilizing effects of direct binding of RAD51 to dsDNA in human tumor cells. Thus, in addition to having genome-stabilizing DNA repair activity, human RAD51 has genome-destabilizing activity when expressed at high levels, as is the case in many human tumors., (© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: Vitrectomy for symptomatic vitreous floaters carries significant risks. Justification of surgery is difficult, particularly in healthy eyes with normal visual acuity and without a posterior vitreous detachment. This is the first reported case of optical coherence tomography being utilized to objectively assess the impact of a vitreous opacity on the macula., Case Presentation: A 37-year-old Caucasian female complained of the sudden onset of a ring-like floater in the central visual field of her left eye. Visual acuity was 20/20, there was no intraocular inflammation and the posterior vitreous was not detached. Complete blood count with differential, serology screen (including cysticercosis and echinococcus), chest x-ray and abdominal ultrasound found no evidence of systemic infective or cystic disease. A color photograph and B-scan ultrasound confirmed a 4.31 mm free-floating semi-translucent vitreous cyst with a hyperechogenic, pigmented surface and faint internal strands suspended in the mid-vitreous cavity, in the visual axis. The cyst moved with ocular movements, but only within the vitreous lacuna it resided in. Humphrey and Goldmann visual fields were normal. However, spectral domain optical coherence tomography (OCT) demonstrated shadowing on either side of the fovea, consistent with the ring-like scotoma described by the patient. Removing the retinal layers from the 3D-reconstructed macular cube OCT revealed a circular shadow on the macula. The patient elected for conservative management and at 3-month follow-up her symptoms had almost fully resolved as the cyst migrated to the inferior vitreous cavity, no longer casting a shadow on the macula., Conclusion: To our knowledge, this is the first description of using OCT as an objective, qualitative assessment of symptoms caused by large vitreous opacities and may provide a simple yet useful adjunctive tool in evaluating the risk-benefit ratio of vitrectomy in patients with large symptomatic vitreous floaters.

Background: The regulation of miR-126 by rs4636297 single nucleotide polymorphism (SNP) has been implicated in the pathogenesis of neovascularisation by promoting vascular endothelial growth factor, suggesting it could be associated with sight threatening diabetic retinopathy (STDR), but has not been previously investigated or reported., Materials and Methods: A case control study of 531 individuals with diabetes was genotyped for the rs4636297 SNP, using the Sequenom iPLEX Gold chemistry. STDR included people with severe non-proliferative diabetic retinopathy (NPDR) or proliferative diabetic retinopathy (PDR). Association was tested using logistic regression analysis, adjusting for confounding variables., Results: In an additive model, the A allele of rs4636297 SNP is significantly associated with STDR compared to people with none or mild diabetic retinopathy (DR) (odds ratio (OR) = 2.02, 95% confidence interval (CI) = 1.22-3.35, p = 0.006)., Conclusion: The A allele of rs4636297, known to be the non-functional allele for post-translational regulation of miR-126, is associated with STDR. This finding suggests that this locus would be a potential therapeutic target for inhibiting the development of DR., (© The Author(s) 2015.)

Background: Microbial keratitis (MK) is a sight-threatening emergency. Delayed diagnosis and treatment may exacerbate the condition and infection may spread to the posterior segment with resultant endophthalmitis. We describe the presentation, management, visual outcomes and microbial profiles of MK-associated endophthalmitis presenting to a tertiary referral centre., Methods: Prospective collection of data on all patients presenting with presumed MK-associated endophthalmitis from 1997 to 2007, to the Royal Victorian Eye and Ear Hospital. Outcome measures included: visual acuity, microbial profiles, and management strategy., Results: Thirty-seven cases of MK-associated endophthalmitis were identified over the study period, with a mean age of 73 years and 19 were male. Presenting acuities ranged from Snellen 2/60 to no perception of light (NPL). Thrity-four (91.9%) patients had a prior history of ocular disease. Identifiable non-ocular risk factors were present in 31 (83.8%), including steroid use, dementia, nursing home care or relative systemic immunosuppression. A culture positivity rate of 83.8% was recorded. The most common organisms identified included: Streptococcal species in 12 (32.4%), Pseudomonas aeruginosa in 11 (29.7%), and Staphylococcus aureus in eight (21.6%). Final acuities ranged from 6/36 to NPL. Sixteen (43.2%) eyes were eviscerated/enucleated as primary treatment. Overall, 23 (62.2%) patients required evisceration/enucleation, of which nine (39.1%) were due to Pseudomonas aeruginosa and seven (30.4%) to Streptococcal species (Streptococcal pneumonia)., Conclusions: MK-associated endophthalmitis is a serious ocular condition occurring more frequently in elderly populations, and those with long standing severe pre-existing ocular disease. Visual outcomes are poor, often requiring evisceration/enucleation.

RAD51 is the central protein that catalyzes DNA repair via homologous recombination, a process that ensures genomic stability. RAD51 protein is commonly expressed at high levels in cancer cells relative to their noncancerous precursors. High levels of RAD51 expression can lead to the formation of genotoxic RAD51 protein complexes on undamaged chromatin. We developed a therapeutic approach that exploits this potentially toxic feature of malignancy, using compounds that stimulate the DNA-binding activity of RAD51 to promote cancer cell death. A panel of immortalized cell lines was challenged with the RAD51-stimulatory compound RS-1. Resistance to RS-1 tended to occur in cells with higher levels of RAD54L and RAD54B, which are Swi2/Snf2-related translocases known to dissociate RAD51 filaments from dsDNA. In PC3 prostate cancer cells, RS-1-induced lethality was accompanied by the formation of microscopically visible RAD51 nuclear protein foci occurring in the absence of any DNA-damaging treatment. Treatment with RS-1 promoted significant antitumor responses in a mouse model, providing proof-of-principle for this novel therapeutic strategy., (©2014 American Association for Cancer Research.)

The aim of this study was to perform a systematic meta-analysis of biomarkers investigated with diabetic retinopathy (DR) in the vitreous, and to explore the molecular pathway interactions of these markers found to be consistently associated with DR. Relevant databases [PubMed and ISI web of science] were searched for all published articles investigating molecular biomarkers of the vitreous associated with DR. Based on set exclusion/inclusion criteria available data from studies with human vitreous samples were extracted and used for our meta-analysis. The interactions of significant biomarkers in DR were investigated via STRING and KEGG pathway analysis. Our meta-analysis of DR identifies eleven biomarkers as potential therapeutic candidates alternate to current anti-VEGF therapy. Four of these are deemed viable therapeutic targets for PDR; ET receptors (ET A and ET B), anti-PDGF-BB, blocking TGF-β using cell therapy and PEDF. The identification of supplementary or synergistic therapeutic candidates to anti VEGF in the treatment of DR may aid in the development of future treatment trials., (© 2014.)

Mutagenesis is a hallmark of malignancy, and many oncologic treatments function by generating additional DNA damage. Therefore, DNA damage repair is centrally important in both carcinogenesis and cancer treatment. Homologous recombination (HR) and nonhomologous end joining are alternative pathways of double-strand DNA break repair. We developed a method to quantify the efficiency of DNA repair pathways in the context of cancer therapy. The recombination proficiency score (RPS) is based on the expression levels for four genes involved in DNA repair pathway preference (Rif1, PARI, RAD51, and Ku80), such that high expression of these genes yields a low RPS. Carcinoma cells with low RPS exhibit HR suppression and frequent DNA copy number alterations, which are characteristic of error-prone repair processes that arise in HR-deficient backgrounds. The RPS system was clinically validated in patients with breast or non-small cell lung carcinomas (NSCLCs). Tumors with low RPS were associated with greater mutagenesis, adverse clinical features, and inferior patient survival rates, suggesting that HR suppression contributes to the genomic instability that fuels malignant progression. This adverse prognosis associated with low RPS was diminished if NSCLC patients received adjuvant chemotherapy, suggesting that HR suppression and associated sensitivity to platinum-based drugs counteract the adverse prognosis associated with low RPS. Therefore, RPS may help oncologists select which therapies will be effective for individual patients, thereby enabling more personalized care.

Purpose: Recent genome-wide association studies for diabetic retinopathy (DR) have identified novel single nucleotide polymorphisms (SNPs) associated with this potentially blinding disease. These markers could prove useful in risk profiling, if the association are validated by replication. To date, these associations have not been well assessed in independent cohorts. The objective of this study was to ascertain any association of these polymorphisms with advanced stages of DR., Methods: A total of 463 patients who had either type 1 (n = 46) or 2 (n = 417) diabetes were genotyped for 24 SNPs previously implicated in DR. Cases (n = 163) were defined as people with severe nonproliferative DR or proliferative DR. Control participants (n = 300) with a confirmed duration of diabetes of at least 5 years had either no evidence of DR or only mild DR., Results: Two SNPs (rs1073203 and rs4838605) were found to be significantly associated with DR in patients with diabetes after adjusting for covariants; rs1073203-G (P = 0.012, odds ratio [OR] = 0.317, 95% confidence interval [95% CI]: 0.129-0.778), rs1073203 in a dominant model (P = 0.005, OR = 0.251, 95% CI: 0.096-0.655), rs4838605 in an additive model (P = 0.047, OR = 1.650, 95% CI: 1.007-2.703), In a dominant model rs1073203 (P = 0.027, OR = 1.400, 95% CI: 0.101-0.857), was significantly associated with DR in type 2 diabetes after adjustment for covariants. This study was sufficiently powered to replicate previous findings., Conclusions: This study confirmed that two variants (rs1073203 and rs4838605) are associated with advanced stages of DR in our cohort. The underlying genes in these candidate regions provide interesting future gene association targets for understanding the pathogenesis of DR.

RAD51 is the central strand exchange recombinase in somatic homologous recombination, providing genomic stability and promoting resistance to DNA damage. An important tool for mechanistic studies of RAD51 is the D-loop or strand assimilation assay, which measures the ability of RAD51-coated single-stranded DNA (ssDNA) to search for, invade and exchange ssDNA strands with a homologous duplex DNA target. As cancer cells generally overexpress RAD51, the D-loop assay has also emerged as an important tool in oncologic drug design programs for targeting RAD51. Previous studies have adapted the traditional gel-based D-loop assay by using fluorescence-based substrates, which in principle allow for use in high-throughput screening platforms. However, these existing D-loop methods depend on linear oligonucleotide DNA duplex targets, and these substrates enable recombinase-independent ssDNA annealing that can obscure the recombinase-dependent strand assimilation signal. This compelled us to fundamentally re-design this assay, using a fluorescent target substrate that consists of a covalently closed linear double-hairpin dsDNA. This new microplate-based method represents a fast, inexpensive and non-radioactive alternative to existing D-loop assays. It provides accurate kinetic analysis of strand assimilation in high-throughput and performs well with human RAD51 and Escherichia coli RecA protein. This advance will aid in both mechanistic studies of homologous recombination and drug screening programs.

Radiation oncology exploits the biological interaction of radiation within tissue to promote tumor death while minimizing damage to surrounding normal tissue. The clinical delivery of radiation relies on principles of radiation physics that define how radiation energy is deposited in the body, as well as technology that facilitates accurate tumor targeting. This review will summarize the current landscape of recent biological and technological advances in radiation oncology, describe the challenges that exist, and offer potential avenues for improvement.

Homologous recombination (HR) is an essential process in cells that provides repair of DNA double-strand breaks and lesions that block DNA replication. RAD51 is an evolutionarily conserved protein that is central to HR. Overexpression of RAD51 protein is common in cancer cells and represents a potential therapeutic target in oncology. We previously described a chemical inhibitor of RAD51, called RI-1 (referred to as compound 1 in this report). The chloromaleimide group of this compound is thought to act as a Michael acceptor and react with the thiol group on C319 of RAD51, using a conjugate addition-elimination mechanism. In order to reduce the likelihood of off-target effects and to improve compound stability in biological systems, we developed an analogue of compound 1 that lacks maleimide-based reactivity but retains RAD51 inhibitory activity. This compound, 1-(3,4-dichlorophenyl)-3-(4-methoxyphenyl)-4-morpholino-1H-pyrrole-2,5-dione, named RI-2 (referred to as compound 7a in this report), appears to bind reversibly to the same site on the RAD51 protein as does compound 1. Like compound 1, compound 7a specifically inhibits HR repair in human cells.

Homologous recombination serves multiple roles in DNA repair that are essential for maintaining genomic stability. We here describe RI-1, a small molecule that inhibits the central recombination protein RAD51. RI-1 specifically reduces gene conversion in human cells while stimulating single strand annealing. RI-1 binds covalently to the surface of RAD51 protein at cysteine 319 that likely destabilizes an interface used by RAD51 monomers to oligomerize into filaments on DNA. Correspondingly, the molecule inhibits the formation of subnuclear RAD51 foci in cells following DNA damage, while leaving replication protein A focus formation unaffected. Finally, it potentiates the lethal effects of a DNA cross-linking drug in human cells. Given that this inhibitory activity is seen in multiple human tumor cell lines, RI-1 holds promise as an oncologic drug. Furthermore, RI-1 represents a unique tool to dissect the network of reaction pathways that contribute to DNA repair in cells.

Stable HIV-1 replication requires the DNA repair of the integration locus catalyzed by cellular factors. The human RAD51 (hRAD51) protein plays a major role in homologous recombination (HR) DNA repair and was previously shown to interact with HIV-1 integrase (IN) and inhibit its activity. Here we determined the molecular mechanism of inhibition of IN. Our standard in vitro integration assays performed under various conditions promoting or inhibiting hRAD51 activity demonstrated that the formation of an active hRAD51 nucleofilament is required for optimal inhibition involving an IN-DNA complex dissociation mechanism. Furthermore we show that this inhibition mechanism can be promoted in HIV-1-infected cells by chemical stimulation of the endogenous hRAD51 protein. This hRAD51 stimulation induced both an enhancement of the endogenous DNA repair process and the inhibition of the integration step. Elucidation of this molecular mechanism leading to the restriction of viral proliferation paves the way to a new concept of antiretroviral therapy based on the enhancement of endogenous hRAD51 recombination activity and highlights the functional interaction between HIV-1 IN and hRAD51.

Introduction: Treatment of technically operable, medically fit locoregionally advanced non-small cell lung cancer (NSCLC) patients is a controversial therapeutic challenge. Our group routinely uses a trimodality approach. Recent advances in radiotherapy allow for improved tumor targeting and daily patient positioning. We hypothesized that these technologies would improve pathologic response rates. We analyzed consecutively treated stage IIIA/IIIB NSCLC patients undergoing chemoradiotherapy before major lung resection, with particular attention paid to the impact of advanced technologies., Methods: Locoregionally advanced NSCLC patients (N2) staged in a multidisciplinary forum with mediastinoscopy were planned to receive platinum-based chemotherapy and 60Gy and major lung resection. Four-dimensional CT (4DCT) and image-guided radiotherapy (IGRT) were used as available. Survival endpoints were estimated using the Kaplan-Meier method and compared using the log-rank test. Multivariate analysis was performed using Cox proportional hazards models., Results: We identified 53 patients from 2/1999 to 2/2010. Median RT dose was 59Gy. 68% underwent lobectomy. Forty-three patients were downstaged pathologically (81%), 38 experienced mediastinal sterilization (72%), and 21 (40%) had complete pathologic response (pCR). 1 and 2 year OS were 85.5% and 61.6%. Superior OS and DFS were associated with nodal downstaging and mediastinal sterilization (pN0). Treatment with IGRT/4DCT in 10 patients resulted in high rates of nodal downstaging (100% vs 77%, p=0.0452), mediastinal sterilization (90% vs 67%, p=0.0769), and pCR (60% vs 35%, p=0.0728)., Conclusions: In selected patients, definitive dose CRT followed by major lung resection results in promising DFS and OS. The use of advanced radiotherapy techniques (4DCT and IGRT) appears to result in promising pathologic response rates., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Background: To evaluate the effect of perioperative anticoagulation and antiplatelet therapy on postoperative vitreous cavity haemorrhage following pars plana vitrectomy for diabetic eye disease., Design: Retrospective chart review., Participants: 139 patients., Methods: Retrospective collection of demographic, medical, surgical and postoperative data of all patients undergoing vitrectomy for diabetic eye disease at The Royal Victorian Eye and Ear Hospital., Main Outcome Measure: Correlation of the rates of persistent vitreous cavity haemorrhage and anticoagulation or antiplatelet treatment., Results: Sixty-eight of 155 (43.9%) eyes of 139 patients were on anticoagulation or antiplatelet therapy prior to surgery. At the time of surgery, 29 (42.6%) were on therapy. Eight of 29 (27.6%) patients had significant persistent vitreous cavity haemorrhage in the postoperative period, with four (13.8%) requiring secondary surgery. Thirty-nine (57.4%) patients had discontinued therapy prior to surgery. Among these, four (10.3%) had persistent bleeding, of which three (7.7%) required additional surgery. Six of 87 (6.9%) patients not on any anticoagulation/antiplatelet therapy had persistent postoperative vitreous cavity haemorrhage, with none requiring further surgery. Patients on anticoagulation/antiplatelet therapy at the time of surgery were more likely to experience persistent haemorrhage and subsequent reoperation (OR = 4.8, P = 0.0045 and OR = 6.6, P = 0.024, respectively)., Conclusion: Perioperative continuation of anticoagulation or antiplatelet treatment appears to increase the risk of persistent postoperative vitreous cavity haemorrhage and the necessity for vitreous cavity washout in this diabetic cohort. Appropriate preoperative cessation of treatment appeared to reduce this risk; however, caution must be taken with regard to the systemic risk associated with cessation of therapy., (© 2011 The Authors. Clinical and Experimental Ophthalmology © 2011 Royal Australian and New Zealand College of Ophthalmologists.)

Introduction: Endophthalmitis is a sight-threatening condition defined as any inflammation of the internal ocular spaces. It is classified as either endogenous or exogenous depending on the route of infection. Exogenous endophthalmitis results from direct inoculation as a complication of intra-ocular surgery, penetrating ocular trauma, intra-ocular foreign bodies, corneal ulceration and following a breach of ocular barriers from a periocular infection. We report a rare case of exogenous endophthalmitis with both unusual etiology and microbiology., Case Presentation: A 41-year-old Caucasian man with a history of depressive illness presented to our eye department with painful acute visual loss on a background history of chronic uveitis. Ocular examination revealed a dense fibrinous panuveitis with a suspicion of a focal lesion in the posterior segment. Microbiological sampling from his anterior chamber and posterior segment revealed a culture of Elizabethkingia meningoseptica. On closer questioning, he volunteered the occurrence of multiple episodes of deliberate needle ocular penetration. Following vitrectomy for associated retinal detachment, a final Snellen visual acuity of 6/60 was obtained., Conclusions: Elizabethkingia meningoseptica endophthalmitis is a rare condition, and visual results to date are poor.