A Phase II Study Evaluating the Effect of Intensity-Modulated Post-Mastectomy Radiation Therapy on Implant Failure Rates in Breast Cancer Patients with Immediate, 2-Stage Implant Reconstruction, with an MRI imaging correlative substudy.

BACKGROUND: Postmastectomy radiation therapy (PMRT) is a mainstay in the adjuvant treatment of node-positive breast cancer, but it poses risks for women with breast reconstruction. Multi-beam intensity-modulated radiation therapy (IMRT) improves dose conformality and homogeneity, potentially reducing complications in breast cancer patients with implant-based reconstruction. To investigate this hypothesis, we conducted a single-arm phase II clinical trial of breast cancer patients who underwent mastectomy/axillary dissection and prosthesis-based reconstruction. METHODS: The primary endpoint was the rate of implant failure within 24 months of permanent implant placement, which would be considered an improvement over historical controls if below 16%. Implant failure (IF) was defined as removal leading to a flat chest wall or replacement with another reconstruction. Patients were analyzed in two cohorts. Cohort 1 (RT-PI) received RT to the permanent implant (PI). Cohort 2 (RT-TE) received RT to the TE. Implant failure rates, adverse events, and quality of life (QOL) were analyzed. Follow-up/post-RT assessments were compared to the baseline/pre-RT assessments at 3-10 weeks after exchange surgery. A subgroup underwent serial MRIs to explore the association of MRI-detected changes with capsular contracture, a known adverse effect of RT. RESULTS: Between June 2014 and March 2017, 119 women were enrolled. Cohort 1 included 45 patients, and Cohort 2 had 74 patients. Among 100 evaluable participants, 25 experienced IF during the study period. IF occured in 8/42 (19%) and 17/58 (29%) in Cohorts 1 and 2, respectively. Among the IFs, the majority were due to capsular contracture (13), infection (7), exposure (3), and other reasons (2). Morphological shape features observed in longitudinal MRI images were associated with the development of Baker grade 3-4 contracture. CONCLUSIONS: The rate of implant failure in reconstructed breast cancer patients treated with IMRT was similar to, but not improved over, conventional, 3D-conformal methods. MRI imaging features show promise for predicting capsular contracture but require validation in larger studies.

Anthracyclines induce cardiotoxicity through a shared gene expression response signature.

TOP2 inhibitors (TOP2i) are effective drugs for breast cancer treatment. However, they can cause cardiotoxicity in some women. The most widely used TOP2i include anthracyclines (AC) Doxorubicin (DOX), Daunorubicin (DNR), Epirubicin (EPI), and the anthraquinone Mitoxantrone (MTX). It is unclear whether women would experience the same adverse effects from all drugs in this class, or if specific drugs would be preferable for certain individuals based on their cardiotoxicity risk profile. To investigate this, we studied the effects of treatment of DOX, DNR, EPI, MTX, and an unrelated monoclonal antibody Trastuzumab (TRZ) on iPSC-derived cardiomyocytes (iPSC-CMs) from six healthy females. All TOP2i induce cell death at concentrations observed in cancer patient serum, while TRZ does not. A sub-lethal dose of all TOP2i induces limited cellular stress but affects calcium handling, a function critical for cardiomyocyte contraction. TOP2i induce thousands of gene expression changes over time, giving rise to four distinct gene expression response signatures, denoted as TOP2i early-acute, early-sustained, and late response genes, and non-response genes. There is no drug- or AC-specific signature. TOP2i early response genes are enriched in chromatin regulators, which mediate AC sensitivity across breast cancer patients. However, there is increased transcriptional variability between individuals following AC treatments. To investigate potential genetic effects on response variability, we first identified a reported set of expression quantitative trait loci (eQTLs) uncovered following DOX treatment in iPSC-CMs. Indeed, DOX response eQTLs are enriched in genes that respond to all TOP2i. Next, we identified 38 genes in loci associated with AC toxicity by GWAS or TWAS. Two thirds of the genes that respond to at least one TOP2i, respond to all ACs with the same direction of effect. Our data demonstrate that TOP2i induce thousands of shared gene expression changes in cardiomyocytes, including genes near SNPs associated with inter-individual variation in response to DOX treatment and AC-induced cardiotoxicity.

Comparative Evaluation of Proton Therapy and Volumetric Modulated Arc Therapy for Brachial Plexus Sparing in the Comprehensive Reirradiation of High-Risk Recurrent Breast Cancer.

Purpose: Recurrent or new primary breast cancer requiring comprehensive regional nodal irradiation after prior radiation therapy (RT) to the supraclavicular area and upper axilla is challenging due to cumulative brachial plexus (BP) dose tolerance. We assessed BP dose sparing achieved with pencil beam scanning proton therapy (PBS-PT) and photon volumetric modulated arc therapy (VMAT). Methods and Materials: In an institutional review board-approved planning study, all patients with ipsilateral recurrent breast cancer treated with PBS-PT re-RT (PBT1) with at least partial BP overlap from prior photon RT were identified. Comparative VMAT plans (XRT1) using matched BP dose constraints were developed. A second pair of proton (PBT2) and VMAT (XRT2) plans using standardized target volumes were created, applying uniform prescription dose of 50.4 per 1.8 Gy and a maximum BP constraint <25 Gy. Incidence of brachial plexopathy was also assessed. Results: Ten consecutive patients were identified. Median time between RT courses was 48 months (15-276). Median first, second, and cumulative RT doses were 50.4 Gy (range, 42.6-60.0), 50.4 Gy relative biologic effectiveness (RBE) (45.0-64.4), and 102.4 Gy (RBE) (95.0-120.0), respectively. Median follow-up was 15 months (5-33) and 18 months for living patients (11-33) Mean BP max was 37.5 Gy (RBE) for PBT1 and 36.9 Gy for XRT1. Target volume coverage of V85% (volume receiving 85% of prescription dose), V90%, and V95% were numerically lower for XRT1 versus PBT1. Similarly, axilla I-III and supraclavicular area coverage were significantly higher for PBT2 than XRT2 at dose levels of V55%, V65%, V75%, V85%, and V95%. Only axilla I V55% did not reach significance (P = .06) favoring PBS-PT. Two patients with high cumulative BPmax (95.2 Gy [RBE], 101.6 Gy [RBE]) developed brachial plexopathy symptoms with ulnar nerve distribution neuropathy without pain or weakness (1 of 2 had symptom resolution after 6 months without intervention). Conclusions: PBS-PT improved BP sparing and target volume coverage versus VMAT. For patients requiring comprehensive re-RT for high-risk, nonmetastatic breast cancer recurrence with BP overlap and reasonable expectation for prolonged life expectancy, PBT may be the preferred treatment modality.

Hypoxia-Directed Treatment of Human Papillomavirus-Related Oropharyngeal Carcinoma.

PURPOSE: Standard curative-intent chemoradiotherapy for human papillomavirus (HPV)-related oropharyngeal carcinoma results in significant toxicity. Since hypoxic tumors are radioresistant, we posited that the aerobic state of a tumor could identify patients eligible for de-escalation of chemoradiotherapy while maintaining treatment efficacy. METHODS: We enrolled patients with HPV-related oropharyngeal carcinoma to receive de-escalated definitive chemoradiotherapy in a phase II study (ClinicalTrials.gov identifier: NCT03323463). Patients first underwent surgical removal of disease at their primary site, but not of gross disease in the neck. A baseline 18F-fluoromisonidazole positron emission tomography scan was used to measure tumor hypoxia and was repeated 1-2 weeks intratreatment. Patients with nonhypoxic tumors received 30 Gy (3 weeks) with chemotherapy, whereas those with hypoxic tumors received standard chemoradiotherapy to 70 Gy (7 weeks). The primary objective was achieving a 2-year locoregional control (LRC) of 95% with a 7% noninferiority margin. RESULTS: One hundred fifty-eight patients with T0-2/N1-N2c were enrolled, of which 152 patients were eligible for analyses. Of these, 128 patients met criteria for 30 Gy and 24 patients received 70 Gy. The 2-year LRC was 94.7% (95% CI, 89.8 to 97.7), meeting our primary objective. With a median follow-up time of 38.3 (range, 22.1-58.4) months, the 2-year progression-free survival (PFS) and overall survival (OS) rates were 94% and 100%, respectively, for the 30-Gy cohort. The 70-Gy cohort had similar 2-year PFS and OS rates at 96% and 96%, respectively. Acute grade 3-4 adverse events were more common in 70 Gy versus 30 Gy (58.3% v 32%; P = .02). Late grade 3-4 adverse events only occurred in the 70-Gy cohort, in which 4.5% complained of late dysphagia. CONCLUSION: Tumor hypoxia is a promising approach to direct dosing of curative-intent chemoradiotherapy for HPV-related carcinomas with preserved efficacy and substantially reduced toxicity that requires further investigation.

Omission of Radiotherapy After Breast-Conserving Surgery for Women With Breast Cancer With Low Clinical and Genomic Risk: 5-Year Outcomes of IDEA.

PURPOSE: Multiple studies have shown a low risk of ipsilateral breast events (IBEs) or other recurrences for selected patients age 65-70 years or older with stage I breast cancers treated with breast-conserving surgery (BCS) and endocrine therapy (ET) without adjuvant radiotherapy. We sought to evaluate whether younger postmenopausal patients could also be successfully treated without radiation therapy, adding a genomic assay to classic selection factors. METHODS: Postmenopausal patients age 50-69 years with pT1N0 unifocal invasive breast cancer with margins ≥2 mm after BCS whose tumors were estrogen receptor-positive, progesterone receptor-positive, and human epidermal growth factor receptor 2-negative with Oncotype DX 21-gene recurrence score ≤18 were prospectively enrolled in a single-arm trial of radiotherapy omission if they consented to take at least 5 years of ET. The primary end point was the rate of locoregional recurrence 5 years after BCS. RESULTS: Between June 2015 and October 2018, 200 eligible patients were enrolled. Among the 186 patients with clinical follow-up of at least 56 months, overall and breast cancer-specific survival rates at 5 years were both 100%. The 5-year freedom from any recurrence was 99% (95% CI, 96 to 100). Crude rates of IBEs for the entire follow-up period for patients age 50-59 years and age 60-69 years were 3.3% (2/60) and 3.6% (5/140), respectively; crude rates of overall recurrence were 5.0% (3/60) and 3.6% (5/140), respectively. CONCLUSION: This trial achieved a very low risk of recurrence using a genomic assay in combination with classic clinical and biologic features for treatment selection, including postmenopausal patients younger than 60 years. Long-term follow-up of this trial and others will help determine whether the option of avoiding initial radiotherapy can be offered to a broader group of women than current guidelines recommend.

Prophylactic Radiation Therapy Versus Standard of Care for Patients With High-Risk Asymptomatic Bone Metastases: A Multicenter, Randomized Phase II Clinical Trial.

PURPOSE: External-beam radiation therapy (RT) is standard of care (SOC) for pain relief of symptomatic bone metastases. We aimed to evaluate the efficacy of radiation to asymptomatic bone metastases in preventing skeletal-related events (SRE). METHODS: In a multicenter randomized controlled trial, adult patients with widely metastatic solid tumor malignancies were stratified by histology and planned SOC (systemic therapy or observation) and randomly assigned in a 1:1 ratio to receive RT to asymptomatic high-risk bone metastases or SOC alone. The primary outcome of the trial was SRE. Secondary outcomes included hospitalizations for SRE and overall survival (OS). RESULTS: A total of 78 patients with 122 high-risk bone metastases were enrolled between May 8, 2018, and August 9, 2021, at three institutions across an affiliated cancer network in the United States. Seventy-three patients were evaluable for the primary end point. The most common primary cancer types were lung (27%), breast (24%), and prostate (22%). At 1 year, SRE occurred in one of 62 bone metastases (1.6%) in the RT arm and 14 of 49 bone metastases (29%) in the SOC arm (P < .001). There were significantly fewer patients hospitalized for SRE in the RT arm compared with the SOC arm (0 v 4, P = .045). At a median follow-up of 2.5 years, OS was significantly longer in the RT arm (hazard ratio [HR], 0.49; 95% CI, 0.27 to 0.89; P = .018), which persisted on multivariable Cox regression analysis (HR, 0.46; 95% CI, 0.23 to 0.85; P = .01). CONCLUSION: Radiation delivered prophylactically to asymptomatic, high-risk bone metastases reduced SRE and hospitalizations. We also observed an improvement in OS with prophylactic radiation, although a confirmatory phase III trial is warranted.

Accelerated Partial Breast Irradiation for HER2-Positive Early-Stage Breast Cancer.

PURPOSE: HER2-positive (HER2+) breast cancer historically exhibited the most adverse local outcomes and, consequently, has had limited representation among trials of partial breast irradiation (PBI). We hypothesized that with contemporary HER2-targeted agents, patients may now exhibit excellent disease control outcomes after adjuvant PBI. METHODS AND MATERIALS: Using a prospectively maintained institutional database, we identified patients with HER2+ breast cancer treated with breast conserving surgery (BCS) and PBI from 2000 to 2022. Salient clinicopathologic and treatment parameters were collected. All patients received external beam PBI to 40 Gy in 10 daily fractions. Cumulative incidence functions were calculated to estimate the incidence of local recurrence (LR) with the competing risk of death. RESULTS: We identified 1248 patients who underwent PBI at our institution, of whom the study cohort comprised 52 (4%) with HER2+ breast cancer (median age, 64 years; range, 44-87). Nearly all had T1 tumors (98%; median size, 12 mm [range, 1-21 mm]). Most had estrogen receptor positive disease (88%), and all patients had negative surgical margins. Nearly all underwent sentinel node biopsy (94%), with the remainder undergoing no surgical axillary evaluation. Forty-two (81%) received chemotherapy; 40 (77%), hormone therapy; and 42 (81%), HER2-directed therapy, most commonly trastuzumab. At 143.8 person-years of follow-up (range, 7-226 months for each patient), we observed 2 LR events at 14 and 26 months, respectively, yielding a 2-year LR rate of 3.8%. No regional or distant recurrences were observed, nor were any breast-specific mortality events. Two deaths were observed, both without evidence of disease. CONCLUSIONS: Among a cohort of patients with HER2+ early-stage breast cancer managed with BCS and PBI, we observed a 2-year LR rate of 3.8%, with no regional or distant recurrences and excellent overall survival. These findings require confirmation with longer follow-up among larger cohorts but appear consistent with the excellent results of contemporary randomized trials of PBI unselected for HER2 status.

Standard-of-care systemic therapy with or without stereotactic body radiotherapy in patients with oligoprogressive breast cancer or non-small-cell lung cancer (Consolidative Use of Radiotherapy to Block [CURB] oligoprogression): an open-label, randomised, controlled, phase 2 study.

BACKGROUND: Most patients with metastatic cancer eventually develop resistance to systemic therapy, with some having limited disease progression (ie, oligoprogression). We aimed to assess whether stereotactic body radiotherapy (SBRT) targeting oligoprogressive sites could improve patient outcomes. METHODS: We did a phase 2, open-label, randomised controlled trial of SBRT in patients with oligoprogressive metastatic breast cancer or non-small-cell lung cancer (NSCLC) after having received at least first-line systemic therapy, with oligoprogression defined as five or less progressive lesions on PET-CT or CT. Patients aged 18 years or older were enrolled from a tertiary cancer centre in New York, NY, USA, and six affiliated regional centres in the states of New York and New Jersey, with a 1:1 randomisation between standard of care (standard-of-care group) and SBRT plus standard of care (SBRT group). Randomisation was done with a computer-based algorithm with stratification by number of progressive sites of metastasis, receptor or driver genetic alteration status, primary site, and type of systemic therapy previously received. Patients and investigators were not masked to treatment allocation. The primary endpoint was progression-free survival, measured up to 12 months. We did a prespecified subgroup analysis of the primary endpoint by disease site. All analyses were done in the intention-to-treat population. The study is registered with ClinicalTrials.gov, NCT03808662, and is complete. FINDINGS: From Jan 1, 2019, to July 31, 2021, 106 patients were randomly assigned to standard of care (n=51; 23 patients with breast cancer and 28 patients with NSCLC) or SBRT plus standard of care (n=55; 24 patients with breast cancer and 31 patients with NSCLC). 16 (34%) of 47 patients with breast cancer had triple-negative disease, and 51 (86%) of 59 patients with NSCLC had no actionable driver mutation. The study was closed to accrual before reaching the targeted sample size, after the primary efficacy endpoint was met during a preplanned interim analysis. The median follow-up was 11·6 months for patients in the standard-of-care group and 12·1 months for patients in the SBRT group. The median progression-free survival was 3·2 months (95% CI 2·0-4·5) for patients in the standard-of-care group versus 7·2 months (4·5-10·0) for patients in the SBRT group (hazard ratio [HR] 0·53, 95% CI 0·35-0·81; p=0·0035). The median progression-free survival was higher for patients with NSCLC in the SBRT group than for those with NSCLC in the standard-of-care group (10·0 months [7·2-not reached] vs 2·2 months [95% CI 2·0-4·5]; HR 0·41, 95% CI 0·22-0·75; p=0·0039), but no difference was found for patients with breast cancer (4·4 months [2·5-8·7] vs 4·2 months [1·8-5·5]; 0·78, 0·43-1·43; p=0·43). Grade 2 or worse adverse events occurred in 21 (41%) patients in the standard-of-care group and 34 (62%) patients in the SBRT group. Nine (16%) patients in the SBRT group had grade 2 or worse toxicities related to SBRT, including gastrointestinal reflux disease, pain exacerbation, radiation pneumonitis, brachial plexopathy, and low blood counts. INTERPRETATION: The trial showed that progression-free survival was increased in the SBRT plus standard-of-care group compared with standard of care only. Oligoprogression in patients with metastatic NSCLC could be effectively treated with SBRT plus standard of care, leading to more than a four-times increase in progression-free survival compared with standard of care only. By contrast, no benefit was observed in patients with oligoprogressive breast cancer. Further studies to validate these findings and understand the differential benefits are warranted. FUNDING: National Cancer Institute.

A Potential Pitfall and Clinical Solutions in Surface-Guided Deep Inspiration Breath Hold Radiation Therapy for Left-Sided Breast Cancer.

Purpose: Deep inspiration breath hold (DIBH) is an effective technique to spare the heart in treating left-sided breast cancer. Surface-guided radiation therapy (SGRT) is increasingly applied in DIBH setup and motion monitoring. Patient-specific breathing behavior, either thoracically driven or abdominally driven (A-DIBH), should be unaltered, online identified, and monitored accordingly to ensure reproducible heart-sparing treatment. Methods and Materials: Sixty patients with left-sided breast cancer treated with SGRT were analyzed: 20 A-DIBH patients with vertical chest elevation (VCE ≤ 5 mm) were prospectively identified, and 40 control patients were retrospectively and randomly selected for comparison. At simulation, both free-breathing (FB) and DIBH computed tomography (CT) were acquired, guided by a motion surrogate placed around the xiphoid process. For SGRT treatment setups, the region of interest (ROI) was defined on the CT chest surface, and the surrogate-based setup was a backup. For all 60 patients, the VCE was measured as the average of the FB-to-DIBH elevations at the breast and xiphoid process, together with abdominal elevation. In the 40-patient control group, A-DIBH patients (VCE ≤ 5 mm) were identified. Of the 20 A-DIBH patients, 10 were treated with volumetric modulated arc therapy plans, and 10 patients were treated with tangent plans. Clinical DIBH plans were recalculated on FB CT to compare maximum dose (DMax), 5% of the maximum dose (D5%), mean dose (DMean), and V30Gy, V20Gy, and V5Gy of the heart and lungs and their significance. Results: In the 20 A-DIBH patients, VCE = 3 ± 2 mm, surrogate motion (9 ± 6 mm), and abdomen motion of 14 ± 5 mm are found. Heart dose reduction from FB to DIBH is significant (P < .01): ∆DMax = -8.4 ± 9.8 Gy, ∆D5% = -2.4 ± 4.4 Gy, and ∆DMean = -0.6 ± 0.9 Gy. Six out of 40 control patients (15%) are found to have VCE ≤ 5 mm. Conclusions: A-DIBH (VCE ≤ 5 mm) patient population is significant (15%), and they should be identified in the SGRT workflow and monitored accordingly. A new abdominal ROI or an abdominal surrogate should be used instead of the conventional chest-only ROI. Patient-specific DIBH should be preserved for higher reproducibility to ensure heart sparing.

Proton Reirradiation for High-Risk Recurrent or New Primary Breast Cancer.

Radiotherapy is an integral component of multidisciplinary breast cancer care. Given how commonly radiotherapy is used in the treatment of breast cancer, many patients with recurrences have received previous radiotherapy. Patients with new primary breast cancer may also have received previous radiotherapy to the thoracic region. Curative doses and comprehensive field photon reirradiation (reRT) have often been avoided in these patients due to concerns for severe toxicities to organs-at-risk (OARs), such as the heart, lungs, brachial plexus, and soft tissue. However, many patients may benefit from definitive-intent reRT, such as patients with high-risk disease features such as lymph node involvement and dermal/epidermal invasion. Proton therapy is a potentially advantageous treatment option for delivery of reRT due to its lack of exit dose and greater conformality that allow for enhanced non-target tissue sparing of previously irradiated tissues. In this review, we discuss the clinical applications of proton therapy for patients with breast cancer requiring reRT, the currently available literature and how it compares to historical photon reRT outcomes, treatment planning considerations, and questions in this area warranting further study. Given the dosimetric advantages of protons and the data reported to date, proton therapy is a promising option for patients who would benefit from the added locoregional disease control provided by reRT for recurrent or new primary breast cancer.

Most large structural variants in cancer genomes can be detected without long reads.

Short-read sequencing is the workhorse of cancer genomics yet is thought to miss many structural variants (SVs), particularly large chromosomal alterations. To characterize missing SVs in short-read whole genomes, we analyzed 'loose ends'-local violations of mass balance between adjacent DNA segments. In the landscape of loose ends across 1,330 high-purity cancer whole genomes, most large (>10-kb) clonal SVs were fully resolved by short reads in the 87% of the human genome where copy number could be reliably measured. Some loose ends represent neotelomeres, which we propose as a hallmark of the alternative lengthening of telomeres phenotype. These pan-cancer findings were confirmed by long-molecule profiles of 38 breast cancer and melanoma cases. Our results indicate that aberrant homologous recombination is unlikely to drive the majority of large cancer SVs. Furthermore, analysis of mass balance in short-read whole genome data provides a surprisingly complete picture of cancer chromosomal structure.

What is the optimal timing of surgery after short-course radiotherapy for rectal cancer?

BACKGROUND: Short-course neoadjuvant radiotherapy is a valuable tool in managing rectal cancers and has improved local recurrence rates. However, limited and conflicting data has resulted in variable usage and a lack of consensus on the optimal timing of surgery following short-course radiotherapy. This review aims to provide a contemporary summation of the available evidence regarding the optimal time interval between short-course neoadjuvant radiotherapy and surgery. METHODS: A focused literature search was undertaken using the PubMed and Embase databases from January 1980 until January 2023. Randomised control trials, large observational studies and systematic reviews focusing on the use of short-course preoperative radiotherapy for localised rectal cancers, with a focus on the timing of surgery, were included. Primary outcomes were overall survival, disease-free survival and perioperative complications. RESULTS: Five randomised control trials, two meta-analyses, and two large, population-based studies were included for their eligibility and relevance. Increased downstaging and fewer postoperative complications are demonstrated in patients receiving delayed surgery (> four weeks), but more recent data raise concerns regarding increased rates of local recurrence in this cohort. Studies directly comparing different time intervals to surgery following short-course radiotherapy have failed to demonstrate an effect on overall survival. CONCLUSIONS: This review highlights the complexities and relative shortcomings of the available data with few high-quality studies and randomised control trials directly comparing different time intervals to surgery following short-course radiotherapy. Continuing research is needed to confirm existing findings and explore gaps in the current literature.

Genomic analysis and clinical correlations of non-small cell lung cancer brain metastasis.

Up to 50% of patients with non-small cell lung cancer (NSCLC) develop brain metastasis (BM), yet the study of BM genomics has been limited by tissue access, incomplete clinical data, and a lack of comparison with paired extracranial specimens. Here we report a cohort of 233 patients with resected and sequenced (MSK-IMPACT) NSCLC BM and comprehensive clinical data. With matched samples (47 primary tumor, 42 extracranial metastatic), we show CDKN2A/B deletions and cell cycle pathway alterations to be enriched in the BM samples. Meaningful clinico-genomic correlations are noted, namely EGFR alterations in leptomeningeal disease (LMD) and MYC amplifications in multifocal regional brain progression. Patients who developed early LMD frequently have had uncommon, multiple, and persistently detectable EGFR driver mutations. The distinct mutational patterns identified in BM specimens compared to other tissue sites suggest specific biologic underpinnings of intracranial progression.

A local ATR-dependent checkpoint pathway is activated by a site-specific replication fork block in human cells.

When replication forks encounter DNA lesions that cause polymerase stalling, a checkpoint pathway is activated. The ATR-dependent intra-S checkpoint pathway mediates detection and processing of sites of replication fork stalling to maintain genomic integrity. Several factors involved in the global checkpoint pathway have been identified, but the response to a single replication fork barrier (RFB) is poorly understood. We utilized the Escherichia coli-based Tus-Ter system in human MCF7 cells and showed that the Tus protein binding to TerB sequences creates an efficient site-specific RFB. The single fork RFB was sufficient to activate a local, but not global, ATR-dependent checkpoint response that leads to phosphorylation and accumulation of DNA damage sensor protein γH2AX, confined locally to within a kilobase of the site of stalling. These data support a model of local management of fork stalling, which allows global replication at sites other than the RFB to continue to progress without delay.

Long-molecule scars of backup DNA repair in BRCA1- and BRCA2-deficient cancers.

Homologous recombination (HR) deficiency is associated with DNA rearrangements and cytogenetic aberrations1. Paradoxically, the types of DNA rearrangements that are specifically associated with HR-deficient cancers only minimally affect chromosomal structure2. Here, to address this apparent contradiction, we combined genome-graph analysis of short-read whole-genome sequencing (WGS) profiles across thousands of tumours with deep linked-read WGS of 46 BRCA1- or BRCA2-mutant breast cancers. These data revealed a distinct class of HR-deficiency-enriched rearrangements called reciprocal pairs. Linked-read WGS showed that reciprocal pairs with identical rearrangement orientations gave rise to one of two distinct chromosomal outcomes, distinguishable only with long-molecule data. Whereas one (cis) outcome corresponded to the copying and pasting of a small segment to a distant site, a second (trans) outcome was a quasi-balanced translocation or multi-megabase inversion with substantial (10 kb) duplications at each junction. We propose an HR-independent replication-restart repair mechanism to explain the full spectrum of reciprocal pair outcomes. Linked-read WGS also identified single-strand annealing as a repair pathway that is specific to BRCA2 deficiency in human cancers. Integrating these features in a classifier improved discrimination between BRCA1- and BRCA2-deficient genomes. In conclusion, our data reveal classes of rearrangements that are specific to BRCA1 or BRCA2 deficiency as a source of cytogenetic aberrations in HR-deficient cells.

Non-cell-autonomous cancer progression from chromosomal instability.

Chromosomal instability (CIN) is a driver of cancer metastasis1-4, yet the extent to which this effect depends on the immune system remains unknown. Using ContactTracing-a newly developed, validated and benchmarked tool to infer the nature and conditional dependence of cell-cell interactions from single-cell transcriptomic data-we show that CIN-induced chronic activation of the cGAS-STING pathway promotes downstream signal re-wiring in cancer cells, leading to a pro-metastatic tumour microenvironment. This re-wiring is manifested by type I interferon tachyphylaxis selectively downstream of STING and a corresponding increase in cancer cell-derived endoplasmic reticulum (ER) stress response. Reversal of CIN, depletion of cancer cell STING or inhibition of ER stress response signalling abrogates CIN-dependent effects on the tumour microenvironment and suppresses metastasis in immune competent, but not severely immune compromised, settings. Treatment with STING inhibitors reduces CIN-driven metastasis in melanoma, breast and colorectal cancers in a manner dependent on tumour cell-intrinsic STING. Finally, we show that CIN and pervasive cGAS activation in micronuclei are associated with ER stress signalling, immune suppression and metastasis in human triple-negative breast cancer, highlighting a viable strategy to identify and therapeutically intervene in tumours spurred by CIN-induced inflammation.

Comparison of setup accuracy of optical surface image versus orthogonal x-ray images for VMAT of the left breast using deep-inspiration breath-hold.

To compare the setup accuracy of optical surface image (OSI) versus orthogonal x-ray images (2DkV) using cone beam computed tomography (CBCT) as ground truth for radiotherapy of left breast cancer in deep-inspiration breath-hold (DIBH). Ten left breast DIBH patients treated with volumetric modulated arc therapy (VMAT) were studied retrospectively. OSI, 2DkV, and CBCT were acquired weekly at treatment setup. OSI, 2DkV, and CBCT were registered to planning CT or planning DRR based on a breast surface region of interest (ROI), bony anatomy (chestwall and sternum), and both bony anatomy and breast surface, respectively. These registrations provided couch shifts for each imaging system. The setup errors, or the difference in couch shifts between OSI and CBCT were compared to those between 2DkV and CBCT. A second OSI was acquired during last beam delivery to evaluate intrafraction motion. The median absolute setup errors were (0.21, 0.27, 0.23 cm, 0.6°, 1.3°, 1.0°) for OSI, and (0.26, 0.24, 0.18 cm, 0.9°, 1.0°, 0.6°) for 2DkV in vertical, longitudinal and lateral translations, and in rotation, roll and pitch, respectively. None of the setup errors was significantly different between OSI and 2DkV. For both systems, the systematic and random setup errors were ≤0.6 cm and ≤1.5° in all directions. Nevertheless, larger setup errors were observed in some sessions in both systems. There was no correlation between OSI and CBCT whereas there was modest correlation between 2DkV and CBCT. The intrafraction motion in DIBH detected by OSI was small with median absolute translations <0.2 cm, and rotations ≤0.4°. Though OSI showed comparable and small setup errors as 2DkV, it showed no correlation with CBCT. We concluded that to achieve accurate setup for both bony anatomy and breast surface, daily 2DkV can't be omitted following OSI for left breast patients treated with DIBH VMAT.

The role of iron in the pathogenesis of endometriosis: a systematic review.

STUDY QUESTION: What is the role of iron in the pathophysiology of endometriosis? SUMMARY ANSWER: Iron excess is demonstrated wherever endometriotic tissues are found and is associated with oxidative stress, an inflammatory micro-environment, and cell damage; the iron-mediated oxidative stress is independently linked to subfertility, symptom severity, and malignant transformation. WHAT IS KNOWN ALREADY: Iron is found in excess in endometriotic tissues, and multiple mechanisms have been studied and posited to explain this. It is clear that iron excess plays a vital role in promoting oxidative stress and cell damage. The evidence base is large, but no comprehensive reviews exist to summarize our understanding and highlight the overarching themes to further our understanding and suggest future directions of study for the field. STUDY DESIGN SIZE DURATION: This systematic review with a thematic analysis retrieved studies from the PubMed, Embase, Web of Science, and Cochrane Library databases and searches were conducted from inception through to August 2022. Human and animal studies published in the English language were included and identified using a combination of exploded MeSH terms ('Iron' and 'Endometriosis') and free-text search terms ('Iron', 'Ferric', 'Ferrous', 'Endometriosis', 'Endometrioma'). PARTICIPANTS/MATERIALS SETTING METHODS: This review was reported in accordance with the PRISMA guidelines. All studies reporting original data concerning the role of iron or iron complexes in the pathophysiology of endometriosis were included. Studies that did not report original data or provided a review of the field were excluded. Bias analysis was completed for each included study by using the Newcastle-Ottawa scoring system. MAIN RESULTS AND THE ROLE OF CHANCE: There were 776 records identified and these were screened down to 53 studies which met the eligibility criteria, including 6 animal and 47 human studies, with 3556 individual participants. Iron excess is demonstrated in various tissues and fluids, including ovarian endometriomas, ovarian follicles, ectopic endometriotic lesions, and peritoneal fluid. Markers of oxidative stress are strongly associated with high iron levels, and aberrant expression of iron-transport proteins has been demonstrated. Abnormal resistance to ferroptosis is likely. Iron-mediated oxidative stress is responsible for a pro-inflammatory micro-environment and is linked to subfertility, symptom severity, and, possibly, malignant transformation. LIMITATIONS REASONS FOR CAUTION: A minority of the included studies were of objectively low quality with a high risk of bias and may lead to misleading conclusions. Additionally, multiple studies failed to appropriately characterize the included patients by known confounding variables, such as menstrual cycle phase, which may introduce bias to the findings. WIDER IMPLICATIONS OF THE FINDINGS: Current literature depicts a central role of aberrant iron mechanics and subsequent oxidative stress in endometriosis. It is likely that iron excess is at least partly responsible for the persistence and proliferation of ectopic endometriotic lesions. As such, iron mechanics represent an attractive target for novel therapeutics, including iron chelators or effectors of the iron-oxidative stress pathway. There are significant gaps in our current understanding, and this review highlights and recommends several topics for further research. These include the role of iron chelation, resistance to ferroptosis, the relationship between iron excess and localized hypoxia, systemic iron pathophysiology in endometriosis, and the role of oxidative stress in malignant transformation. STUDY FUNDING/COMPETING INTERESTS: J.W. and S.G.P. are supported by clinical fellowships at Liverpool University Hospital NHS Foundation trust. No additional funding was requested or required for the completion of this work. C.J.H. is supported by a Wellbeing of Women project grant (RG2137). D.K.H. is supported by a Wellbeing of Women project grant (RG2137) and an MRC clinical research training fellowship (MR/V007238/1). The authors have no conflicts of interest to declare. REGISTRATION NUMBER: A protocol was prospectively registered with the PROSPERO database in August 2021 (CRD42021272818).

Accelerated Partial Breast Irradiation: Association of Dosimetric Parameters With Patient-Reported Outcomes.

Purpose: Accelerated partial breast irradiation (APBI) after breast-conserving surgery offers a well-tolerated adjuvant radiation therapy option for patients with breast cancer. We sought to describe patient-reported acute toxicity as a function of salient dosimetric parameters during and after an APBI regimen of 40 Gy in 10 once-daily fractions. Methods and Materials: From June 2019 to July 2020, patients undergoing APBI were assigned a weekly, response-adapted, patient reported outcomes-common terminology criteria for adverse events-based acute toxicity assessment. Patients reported acute toxicity during treatment and for up to 8 weeks after treatment. Dosimetric treatment parameters were collected. Descriptive statistics and univariable analyses were used to summarize patient-reported outcomes and their correlation to corresponding dosimetric measures, respectively. Results: Overall, 55 patients who received APBI completed a total of 351 assessments. Median planning target volume was 210 cc (range, 64-580 cc), and median planning target volume:ipsilateral breast volume ratio was 0.17 (range, 0.05-0.44). Overall, 22% of patients reported moderate breast enlargement and 27% reported maximum skin toxicity as severe or very severe. Furthermore, 35% of patients reported fatigue, and 44% of patients reported pain in the radiated area as moderate to very severe. Median time to first report of any moderate to very severe symptom was 10 days (interquartile range, 6-27 days). By 8 weeks after APBI, most patients reported resolution of symptoms, with 16% reporting residual moderate symptoms. Upon univariable analysis, none of the ascertained salient dosimetric parameters were associated with maximum symptoms or with the presence of moderate to very severe toxicity. Conclusions: Weekly assessments during and after APBI showed that patients experienced moderate to very severe toxicities, most commonly skin toxicity, but that these typically resolved by 8 weeks after radiation therapy. More comprehensive evaluations among larger cohorts are warranted to define the precise dosimetric parameters that correspond to outcomes of interest.