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Purpose: Human papillomavirus (HPV) is a significant cause of cervical cancer. We hypothesized that detecting viral cell-free HPV DNA (cfDNA) before, during, and after chemoradiation (chemoRT) could provide insights into disease extent, clinical staging, and treatment response., Experimental Design: A total of 66 patients with locally advanced cervical cancer were enrolled between 2017 and 2023, with 49 receiving standard-of-care treatment and 17 participating in a clinical trial combining a therapeutic HPV vaccine (PDS0101; IMMUNOCERV). Plasma was collected at baseline, weeks 1, 3, and 5 of chemoRT, and 3 to 4 months after chemoRT. HPV cfDNA was quantified using droplet digital PCR targeting the HPV E6/E7 oncogenes of 13 high-risk types. MRI was performed at baseline and before brachytherapy., Results: The median follow-up was 23 months, with recurrence-free survival (RFS) of 78.4% at 2 years. Baseline nodal disease extent correlated with HPV cfDNA levels. HPV cfDNA levels peaked in week 1 of radiation and decreased through treatment. Patients receiving the PDS0101 vaccine had a higher rate of undetectable HPV type 16 cfDNA compared with those who received standard-of-care therapy. HPV cfDNA clearance correlated with better 2-year RFS (92.9% vs. 30%, log-rank; P = 0.0067). The strongest predictor of RFS was HPV cfDNA clearance in follow-up achieving a concordance index score of 0.83, which improved when combined with MRI response (concordance index, 0.88)., Conclusions: HPV cfDNA levels change dynamically during chemoRT. HPV cfDNA levels at follow-up predict RFS, and delivery of therapeutic HPV vaccine with chemoRT was linked to rapid HPV cfDNA decline. Monitoring HPV cfDNA during and after chemoRT may guide tailoring of personalized treatment., (©2024 American Association for Cancer Research.) more...

Herein, we present a protocol for culturing patient-derived organoids (PDOs) of cervical cancer that includes workflows for tumor biopsy/resection tissue and cytobrush-sampled cells. We describe steps for PDO culture initiation, including rinsing, gentle dissociation, Lymphoprep separation, and cell assessment, as well as seeding cells from surgical and cytobrush tissue digestion. We then provide guidance on PDO maintenance and passage and techniques for producing conditioned medium. Overall, this protocol serves as a valuable guide for establishing and maintaining cervical cancer PDOs. For complete details on the use and execution of this protocol, please refer to Colbert et al. 1 ., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.) more...

For decades, chemoradiosensitization with checkpoint kinase inhibitors has been proposed but largely unexplored. A recent study reports that the novel ataxia telangiectasia and Rad3-related kinase inhibitor RP-3500 synergizes with radiation to control Atm-/- tumors in vivo. RP-3500 did not radiosensitize wild-type or Brca-1-deficient tumors, highlighting the need for a genotype-tailored approach. See related article by Ng et al., p. 5643., (©2024 American Association for Cancer Research.) more...

Adenoid cystic carcinoma (ACC) is a rare, usually slow-growing yet aggressive head and neck malignancy. Despite its clinical significance, our understanding of the cellular evolution and microenvironment in ACC remains limited. We investigated the intratumoral microbiomes of 50 ACC tumor tissues and 33 adjacent normal tissues using 16S rRNA gene sequencing. This allowed us to characterize the bacterial communities within the ACC and explore potential associations between the bacterial community structure, patient clinical characteristics, and tumor molecular features obtained through RNA sequencing. The bacterial composition in the ACC was significantly different from that in adjacent normal salivary tissue, and the ACC exhibited diverse levels of species richness. We identified two main microbial subtypes within the ACC: oral-like and gut-like. Oral-like microbiomes, characterized by increased diversity and abundance of Neisseria, Leptotrichia, Actinomyces, Streptococcus, Rothia, and Veillonella (commonly found in healthy oral cavities), were associated with a less aggressive ACC-II molecular subtype and improved patient outcomes. Notably, we identified the same oral genera in oral cancer and head and neck squamous cell carcinomas. In both cancers, they were part of shared oral communities associated with a more diverse microbiome, less aggressive tumor phenotype, and better survival that reveal the genera as potential pancancer biomarkers for favorable microbiomes in ACC and other head and neck cancers. Conversely, gut-like intratumoral microbiomes, which feature low diversity and colonization by gut mucus layer-degrading species, such as Bacteroides, Akkermansia, Blautia, Bifidobacterium, and Enterococcus, were associated with poorer outcomes. Elevated levels of Bacteroides thetaiotaomicron were independently associated with significantly worse survival and positively correlated with tumor cell biosynthesis of glycan-based cell membrane components., (© 2024. The Author(s).) more...

Objectives: This study investigates retreatment rates in single-fraction radiation therapy (SFRT) for painful bone metastasis in patients with limited life expectancy. We compared retreatment-free survival (RFS) in patients from a rapid access bone metastases clinic (RABC) and non-RABC patients, identifying factors associated with retreatment., Methods: In this observational study, we analysed RABC patients who received SFRT between April 2018 and November 2019, using non-RABC SFRT patients as a comparison group. Patients with prior or perioperative radiation therapy (RT) were excluded. The primary endpoint was same-site and any-site retreatment with RT or surgery. Patient characteristics were compared using χ 2 and Student's t-tests, with RFS estimates based on a multistate model considering death as a competing risk using Aalen-Johansen estimates., Results: We identified 151 patients (79 RABC, 72 non-RABC) with 225 treatments (102 RABC, 123 non-RABC) meeting eligibility criteria. Of the 22 (10.8%) same-site retreatments, 5 (22.7%) received surgery, 14 (63.6%) received RT and 3 (13.6%) received both RT and surgery. We found no significant differences in any-site RFS (p=0.97) or same-site RFS (p=0.11)., Conclusions: RFS is high and similar comparable in the RABC and non-RABC cohorts. Retreatment rates are low, even in patients with low Eastern Cooperative Oncology Group scores., Competing Interests: Competing interests: PPLee reports personal fees and non-financial support from Viewray, Inc., grants, personal fees and non-financial support from AstraZeneca, Inc., personal fees and non-financial support from Varian, Inc, outside the submitted work. PPLin reports book royalties from Springer Nature, outside the submitted work., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.) more...

Background: Chemoradiation (CRT) may modulate the immune milieu as an in-situ vaccine. Rapid dose delivery of brachytherapy has unclear impact on T-cell repertoires. HPV-associated cancers express viral oncoproteins E6/E7, which enable tracking antigen/tumor-specific immunity during CRT., Methods: Thirteen cervical cancer patients on a multi-institutional prospective protocol from 1/2020-1/2023 underwent standard-of-care CRT with pulsed-dose-rate brachytherapy boost (2 fractions). Cervix swabs at various timepoints underwent multiplex DNA deep sequencing of the TCR-β/CDR3 region with immunoSEQ. Separately, HPV-responsive T-cell clones were also expanded ex vivo. Statistical analysis was via Mann-Whitney-U., Results: TCR productive clonality, templates, frequency, or rearrangements increased post-brachytherapy in 8 patients. Seven patients had E6/E7-responsive evolution over CRT with increased productive templates (ranges: 1.2-50.2 fold-increase from baseline), frequency (1.2-1.7), rearrangements (1.2-40.2), and clonality (1.2-15.4). Five patients had HPV-responsive clonal expansion post-brachytherapy, without changes in HPV non-responsive clones. Epitope mapping revealed VDJ rearrangements targeting cervical cancer-associated antigens in 5 patients. The only two patients with disease recurrence lacked response in all metrics. A lack of global TCR remodeling correlated with worse recurrence-free survival, p = 0.04., Conclusion: CRT and brachytherapy alters the cervical cancer microenvironment to facilitate the expansion of specific T-cell populations, which may contribute to treatment efficacy., (Copyright © 2023 American Brachytherapy Society. Published by Elsevier Inc. All rights reserved.) more...

Objective: To describe the long-term outcomes of patients with stage IVA cervical cancer, a rare and deadly disease for which long-term toxicity data are scarce, to guide clinician counseling and survivorship support., Methods: In a retrospective review of a prospectively maintained database, we identified 76 patients with stage IVA cervical cancer with biopsy- or MRI-proven bladder mucosal involvement who received definitive radiotherapy (external beam radiotherapy [EBRT] alone or EBRT plus brachytherapy) with or without chemotherapy at our institution between 2000 and 2020. We used Kaplan-Meier modeling to estimate recurrence-free survival (RFS) and overall survival (OS) and used proportional hazard modeling to identify clinical variables associated with recurrence or survival. We performed actuarial competing risk modeling for severe late toxicity (grades 3 to 5, occurring >6 months of follow-up) and vesicovaginal fistulae (VVF), censoring for pelvic recurrence and death, and made comparisons between potential predictors using Gray's test and binary logistic regression., Results: The median follow-up time was 76 months (interquartile range 58-91). The median OS duration was 35 months (range, 18-not reached), and the 2- and 5-year OS rates were 53.6% and 40.9%, respectively. OS and RFS did not differ significantly between patients who received EBRT alone (N = 18) or EBRT plus brachytherapy (N = 49). Current smoking was a strong predictor of severe late toxicity, whose incidence was 14% at 2 years and 17% at 10 years. The VVF incidence was 24% at 2 years and 32% at 10 years., Conclusion: Patients with stage IVA cervical cancer, even those who receive EBRT alone, can have long-term survival. These patients should be followed closely for late radiation-related toxicity., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023. Published by Elsevier Inc.) more...

Introduction: The objective of this study was to assess differences in long-term sexual and menopausal side effects after uterine cancer treatment among treatment modalities., Methods and Materials: This is a cross-sectional study that examined women treated for uterine cancer from 2006-2018. Eligible women included those who underwent a hysterectomy/bilateral salpino-oophorectemy alone (HS), with brachytherapy (BT), or with external beam radiation therapy (EBRT). A noncancer cohort of women who underwent a hysterectomy/BSO for benign indications were also identified (non-CA). To compare outcomes, we utilized a shortened form of the female sexual function index (FSFI) and the menopause survey, which consists of 3 subscales: hot flashes, vaginal symptoms, and urinary symptoms. Demographic, comorbidity, and other treatment variables were collected. Survey totals were compared across cohorts using ANOVA tests and logistic regression., Results: A total of 284 women completed the Menopause Survey (Non-CA 64, HS 60, BT 69, EBRT 91); 116 women reported sexual activity in the last 4 weeks and completed the FSFI (NC 32, HS 21, BT 31, EBRT 32). The mean FSFI score for the entire cohort was 11.4 (SD 4.16), which indicates poor sexual function. There was no significant difference between any cohort in the overall FSFI score (p = 0.708) or in any of the FSFI subscales (all p > 0.05). On univariate analysis, BT was associated with fewer menopausal hot flashes and vaginal symptoms compared to the non-CA cohort (p < 0.05), which did not persist on multivariable analysis., Conclusion: There was no significant difference in sexual dysfunction or menopausal symptoms in those treated for uterine cancer with or without adjuvant radiation. Most patients reported poor sexual function., (Copyright © 2023 American Brachytherapy Society. Published by Elsevier Inc. All rights reserved.) more...

Background: Stereotactic body radiotherapy (SBRT) has the potential to ablate localised pancreatic ductal adenocarcinoma. Selective dismutase mimetics sensitise tumours while reducing normal tissue toxicity. This trial was designed to establish the efficacy and toxicity afforded by the selective dismutase mimetic avasopasem manganese when combined with ablative SBRT for localised pancreatic ductal adenocarcinoma., Methods: In this adaptive, randomised, double-blind, placebo-controlled, phase 1b/2 trial, patients aged 18 years or older with borderline resectable or locally advanced pancreatic cancer who had received at least 3 months of chemotherapy and had an Eastern Cooperative Oncology Group performance status of 0-2 were enrolled at six academic sites in the USA. Eligible patients were randomly assigned (1:1), with block randomisation (block sizes of 6-12) with a maximum of 24 patients per group, to receive daily avasopasem (90 mg) or placebo intravenously directly before (ie, within 180 min) SBRT (50, 55, or 60 Gy in five fractions, adaptively assigned in real time by Bayesian estimates of 90-day safety and efficacy). Patients and physicians were masked to treatment group allocation, but not to SBRT dose. The primary objective was to find the optimal dose of SBRT with avasopasem or placebo as determined by the late onset EffTox method. All analyses were done on an intention-to-treat basis. This study is registered with ClinicalTrials.gov, NCT03340974, and is complete., Findings: Between Jan 25, 2018, and April 29, 2020, 47 patients were screened, of whom 42 were enrolled (median age was 71 years [IQR 63-75], 23 [55%] were male, 19 [45%] were female, 37 [88%] were White, three [7%] were Black, and one [2%] each were unknown or other races) and randomly assigned to avasopasem (n=24) or placebo (n=18); the placebo group was terminated early after failing to meet prespecified efficacy parameters. At data cutoff (June 28, 2021), the avasopasem group satisfied boundaries for both efficacy and toxicity. Late onset EffTox efficacy response was observed in 16 (89%) of 18 patients at 50 Gy and six (100%) of six patients at 55 Gy in the avasopasem group, and was observed in three (50%) of six patients at 50 Gy and nine (75%) of 12 patients at 55 Gy in the placebo group, and the Bayesian model recommended 50 Gy or 55 Gy in five fractions with avasopasem for further study. Serious adverse events of any cause were reported in three (17%) of 18 patients in the placebo group and six (25%) of 24 in the avasopasem group. In the placebo group, grade 3 adverse events within 90 days of SBRT were abdominal pain, acute cholangitis, pyrexia, increased blood lactic acid, and increased lipase (one [6%] each); no grade 4 events occurred. In the avasopasem group, grade 3-4 adverse events within 90 days of SBRT were acute kidney injury, increased blood alkaline phosphatase, haematoma, colitis, gastric obstruction, lung infection, abdominal abscess, post-surgical atrial fibrillation, and pneumonia leading to respiratory failure (one [4%] each).There were no treatment-related deaths but one late death in the avasopasem group due to sepsis in the setting of duodenal obstruction after off-study treatment was reported as potentially related to SBRT., Interpretation: SBRT that uses 50 or 55 Gy in five fractions can be considered for patients with localised pancreatic ductal adenocarcinoma. The addition of avasopasem might further enhance disease outcomes. A larger phase 2 trial (GRECO-2, NCT04698915) is underway to validate these results., Funding: Galera Therapeutics., Competing Interests: Declaration of interests CMT reports grant or contracts with US National Institutes of Health (NIH and CPRIT), licenses with Xerient Pharmaceuticals for licensed or individual patent for radioprotection of the upper gastrointestinal tract, and consulting fees with Phebry. JMF reports consultant and advisory board roles for Viewray, and payment for presentations for Boston Scientific. TAA reports an unfunded study steering committee role for Galera Therapeutics (Galera). MP reports research support to their institution from Merck and Varian, royalties to serve as section editor for cholangiocarcinoma, consulting fees from Voxelmetric and Syntactx, a speakers honorarium for Oakstone CME, advisory board participation for Varian, and an unpaid ASTRO Education Committee role. BC reports royalties from Springer, honoraria for Varian-Educational and Oakstone, and a leadership or fiduciary role with the National Comprehensive Cancer Network. MSB reports funding from Galera, institution grants or contracts from Nanobiotix, Silenseed, Trisalis, Artidis, SUC2, CPRIT, AstraZeneca, Merck, Siemens Medical, and Boston Scientific (Augmenix) and consulting fees from Oncosil and Strapax. SP reports consulting or advisory roles for Zymeworks, Ipsen, Novartis, Janssen, Boehringer Ingelheim, and AskGene Pharma, and institutional research funding from Mirati Therapeutics, Lilly, Xencor, Novartis, Rgenix, Bristol-Myers Squibb, Astellas Pharma, Framewave, 4D Pharma, Boehringer Ingelheim, NGM Biopharmaceuticals, Janssen, Arcus Biosciences, Elicio Therapeutics, Bionte, Ipsen, Zymeworks, Pfizer, ImmunoMET, Immuneering, and Amal Therapeutics. C-WDT reports an institution-sponsored research agreement with Sirtex, and individual and institutional honoraria from PanTher. DWK reports grants or contracts from Bold Therapeutics, and payment or honoraria from AstraZeneca. JC reports travel expense money from Galera. EJK reports research grants and contracts from NIH, US Department of Defense, Cancer Prevention and Research Institute of Texas, and Artidis, royalties or licenses from Taylor and Francis, and Kallisio, consulting fees from Kallisio, AstraZeneca, RenovoRx, MD Anderson Physician Network, MJH Life Sciences, and Quantum Aurea Capital, honoraria from Northwestern University and Amplity, a patent pending for 3D oral stents, scientific and medical advisory board roles for Cholangiocarcinoma Foundation, and stock or stock options with Quantum Aurea Capital. PD reports consulting fees from the American Society for Radiation Oncology, payment or honoraria from American Society for Clinical Oncology, Conveners, Physicians Education Resource, Imedex, and Bayer. RAW reports royalties from McGraw-Hill. GOS reports a research agreement with Artios Pharma. RSST reports institutional funding from Galera and NIH National Cancer Institute (NCI). YY reports personal fees from AbbVie, Amgen, Bexion Pharmaceuticals, BeyondSpring Pharmaceuticals, Boehringer Ingelheim Pharmaceuticals, Bristol Myers Squibb, Century Therapeutics, Enliven Therapeutics, Repare Therapeutics, Servier Pharmaceuticals, Starpax Pharmaceuticals, and Vertex Pharmaceuticals during the conduct of the study. PFT reports research funding from NIH NCI. RAB is an employee and officer of, reports receiving compensation from, and holds equity in the sponsor, Galera, residual royalties resulting from the acquisition of Confluence from Aclaris Therapeutics, stock or stock options with Galera, Euclises, Aclaris Therapeutics, and Novocure, and patents for methods for treatment of diseases, (US Pat 10,493,081, 2019 issued to Galera); a patent methods for treatment of diseases (US Pat 9,149,483, 2015 issued to Galera). JH was an employee and officer of, reports receiving compensation from, and holds equity in the sponsor, Galera. JMH reports research funding to their institution and support for attending a meeting from Canopy Cancer Collective, royalties or licenses from Springer, stock or stock options with Histosonics, and consulting fees from Histosonics and Boston Scientific. SEH is an employee of MyCareGorithm and has her own limited liability company, Beyond the White Coat. She has received travel assistance from ViewRay, stock options from Rittenhouse, and her institution received research funding from ViewRay and Galera. LEC, JAJ, VB, MM, GM, NR, EBL, MHGK, SB, and SM declare no competing interests., (Copyright © 2023 Elsevier Ltd. All rights reserved.) more...

Tumor microbiota can produce active metabolites that affect cancer and immune cell signaling, metabolism, and proliferation. Here, we explore tumor and gut microbiome features that affect chemoradiation response in patients with cervical cancer using a combined approach of deep microbiome sequencing, targeted bacterial culture, and in vitro assays. We identify that an obligate L-lactate-producing lactic acid bacterium found in tumors, Lactobacillus iners, is associated with decreased survival in patients, induces chemotherapy and radiation resistance in cervical cancer cells, and leads to metabolic rewiring, or alterations in multiple metabolic pathways, in tumors. Genomically similar L-lactate-producing lactic acid bacteria commensal to other body sites are also significantly associated with survival in colorectal, lung, head and neck, and skin cancers. Our findings demonstrate that lactic acid bacteria in the tumor microenvironment can alter tumor metabolism and lactate signaling pathways, causing therapeutic resistance. Lactic acid bacteria could be promising therapeutic targets across cancer types., Competing Interests: Declaration of interests L.E.C. reports grants from American Society for Clinical Oncology, Radiology Society of North America, National Institutes of Health, and MD Anderson Cancer Center during the conduct of the study. A.J. reports personal fees from Genentech during the conduct of the study, as well as personal fees from Genentech outside the submitted work. L.L. reports other support from AstraZeneca and Pfizer, and grants from NCI outside the submitted work. J.A.W. reports other support from Micronoma during the conduct of the study, as well as other support from Imedex, Dava Oncology, Illumina, and PeerView outside the submitted work. P.O. reports Faculty grant/research support from Merck Sharp and Dohme Corp, Deinove Pharmaceuticals, Summit Pharmaceuticals, Melinta Pharmaceuticals, and Napo Pharmaceutical, in addition to consultant work with Napo Pharmaceutical, Ferring Pharmaceutical, Summit Pharmaceutical, and SNIPR Biome Company, all outside of the submitted work., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.) more...

Purpose: Human papillomavirus (HPV) is the primary driver of cervical cancer. Although studies in other malignancies correlated peripheral blood DNA clearance with favorable outcomes, research on the prognostic value of HPV clearance in gynecologic cancers using intratumoral HPV is scarce. We aimed to quantify the intratumoral HPV virome in patients undergoing chemoradiation therapy (CRT) and associate this with clinical characteristics and outcomes., Methods and Materials: This prospective study enrolled 79 patients with stage IB-IVB cervical cancer undergoing definitive CRT. Cervical tumor swabs collected at baseline and week 5 (end of intensity modulated radiation therapy) were sent for shotgun metagenome sequencing and processed via VirMAP, a viral genome sequencing and identification tool for all known HPV types. The data were categorized into HPV groups (16, 18, high risk [HR], and low risk [LR]). We used independent t tests and Wilcoxon signed-rank to compare continuous variables and χ 2 and Fisher exact tests to compare categorical variables. Kaplan-Meier survival modeling was performed with log-rank testing. HPV genotyping was verified using quantitative polymerase chain reaction to validate VirMAP results using receiver operating characteristic curve and Cohen's kappa., Results: At baseline, 42%, 12%, 25%, and 16% of patients were positive for HPV 16, HPV 18, HPV HR, and HPV LR, respectively, and 8% were HPV negative. HPV type was associated with insurance status and CRT response. Patients with HPV 16+ and other HPV HR+ tumors were significantly more likely to have a complete response to CRT versus patients with HPV 18 and HPV LR/HPV-negative tumors. Overall HPV viral loads predominantly decreased throughout CRT, except for HPV LR viral load., Conclusions: Rarer, less well-studied HPV types in cervical tumors are clinically significant. HPV 18 and HPV LR/negative tumors are associated with poor CRT response. This feasibility study provides a framework for a larger study of intratumoral HPV profiling to predict outcomes in patients with cervical cancer., (Copyright © 2023. Published by Elsevier Inc.) more...

Background: Squamous cell carcinoma of the anus (SCCA) is a rare gastrointestinal cancer. Factors associated with progression of HPV infection to anal dysplasia and cancer are unclear and screening guidelines and approaches for anal dysplasia are less clear than for cervical dysplasia. One potential contributing factor is the anorectal microbiome. In this study, we aimed to identify differences in anal microbiome composition in the settings of HPV infection, anal dysplasia, and anal cancer in this rare disease., Methods: Patients were enrolled in two prospective studies. Patients with anal dysplasia were part of a cross-sectional cohort that enrolled women with high-grade lower genital tract dysplasia. Anorectal tumor swabs were prospectively collected from patients with biopsy-confirmed locally advanced SCCA prior to receiving standard-of-care chemoradiotherapy (CRT). Patients with high-grade lower genital tract dysplasia without anal dysplasia were considered high-risk (HR Normal). 16S V4 rRNA Microbiome sequencing was performed for anal swabs. Alpha and Beta Diversity and composition were compared for HR Normal, anal dysplasia, and anal cancer., Results: 60 patients with high-grade lower genital tract dysplasia were initially enrolled. Seven patients had concurrent anal dysplasia and 44 patients were considered HR Normal. Anorectal swabs from 21 patients with localized SCCA were included, sequenced, and analyzed in the study. Analysis of weighted and unweighted UniFrac distances demonstrated significant differences in microbial community composition between anal cancer and HR normal (p = 0.018). LEfSe identified that all three groups exhibited differential enrichment of specific taxa. Peptoniphilus (p=0.028), Fusobacteria (p=0.0295) , Porphyromonas (p=0.034) , and Prevotella (p=0.029) were enriched in anal cancer specimens when compared to HR normal., Conclusion: Although alpha diversity was similar between HR Normal, dysplasia and cancer patients, composition differed significantly between the three groups. Increased anorectal Peptoniphilus, Fusobacteria , Porphyromonas , and Prevotella abundance were associated with anal cancer. These organisms have been reported in various gastrointestinal cancers with roles in facilitating the proinflammatory microenvironment and neoplasia progression. Future work should investigate a potential role of microbiome analysis in screening for anal dysplasia and investigation into potential mechanisms of how these microbial imbalances influence the immune system and anal carcinogenesis., Competing Interests: CT is on the clinical advisory board of Accuracy, has a patent for oral amifostine as a radioprotectant of the upper gastrointestinal tract issued, licensed, and with royalties paid from Xerient Pharmaceuticals and a pending patent for prolyl hydroxylase inhibitors as a radioprotectant of the gastrointestinal tract, was the lead principal investigator of a multicenter trial testing the effects of high-dose stereotactic body radiation therapy with the radiomodulator, GC4419, and is a paid consultant for Phebra Pty, Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Elnaggar, Huynh, Lin, Hillman, Abana, El Alam, Tomasic, Karpinets, Kouzy, Phan, Wargo, Holliday, Das, Mezzari, Ajami, Lynn, Minsky, Morris, Milbourne, Messick, Klopp, Futreal, Taniguchi, Schmeler and Colbert.) more...

Although high-risk human papillomavirus infection is a well-established risk factor for cervical cancer, other co-factors within the local microenvironment may play an important role in the development of cervical cancer. The current study aimed to characterize the cervicovaginal microbiota in women with premalignant dysplasia or invasive cervical cancer compared with that of healthy women. The study comprised 120 Ethiopian women (60 cervical cancer patients who had not received any treatment, 25 patients with premalignant dysplasia, and 35 healthy women). Cervicovaginal specimens were collected using either an Isohelix DNA buccal swab or an Evalyn brush, and ribosomal RNA sequencing was used to characterize the cervicovaginal microbiota. Shannon and Simpson diversity indices were used to evaluate alpha diversity. Beta diversity was examined using principal coordinate analysis of weighted UniFrac distances. Alpha diversity was significantly higher in patients with cervical cancer than in patients with dysplasia and in healthy women ( p < 0.01). Beta diversity was also significantly different in cervical cancer patients compared with the other groups (weighted UniFrac Bray-Curtis, p < 0.01). Microbiota composition differed between the dysplasia and cervical cancer groups. Lactobacillus iners was particularly enriched in patients with cancer, and a high relative abundance of Lactobacillus species was identified in the dysplasia and healthy groups, whereas Porphyromonas , Prevotella , Bacteroides , and Anaerococcus species predominated in the cervical cancer group. In summary, we identified differences in cervicovaginal microbiota diversity, composition, and relative abundance between women with cervical cancer, women with dysplasia, and healthy women. Additional studies need to be carried out in Ethiopia and other regions to control for variation in sample collection. more...

Aims: Our understanding of the impact of adjuvant therapy on longitudinal quality of life (QoL) following surgery for patients with uterine cancer is limited. The purpose of this study was to compare QoL in patients who have undergone surgery with or without radiation therapy for uterine cancer., Materials and Methods: This was a cross-sectional cohort study that examined women treated for uterine cancer at MD Anderson Cancer Center from 2006 to 2017. Participants included those who underwent hysterectomy/bilateral salphingo-oophorectomy alone, with brachytherapy or external beam radiation therapy (EBRT). A non-cancer cohort of women who underwent a hysterectomy/bilateral salphingo-oophorectomy for benign indications was also identified (non-CA). To compare QoL we used the Functional Assessment of Cancer Therapy - Endometrial survey (FACT-En), a validated survey used to assess QoL. The survey has five subscales: physical, social, emotional, functional and an endometrial cancer-specific subscale. Cohorts were compared using ANOVA tests., Results: In total, 309 women responded to the questionnaire (hysterectomy/bilateral salphingo-oophorectomy 64, brachytherapy 77, EBRT 96, non-CA 72). The median time from surgery to survey completion was 6.7 years. The mean total FACT-En score for the entire cohort was 144 [standard deviation 22]. Overall QoL was different between cohorts, with the EBRT cohort reporting the lowest QoL (mean 139.4 [21.6]) and the brachytherapy cohort the highest (150.6 [18.2], P = 0.006). Among patients who had undergone cancer treatment, the EBRT cohort reported the worst endometrial-specific QoL (53.5 [8.6]), while again the brachytherapy group reported the highest score (57.5 [6.1], P = 0.007)., Conclusions: QoL differences in women who have undergone different treatments for uterine cancer may persist years after treatment. In women with endometrial cancer who require adjuvant therapy, brachytherapy does not appear to have any long-term detriments on QoL., (Copyright © 2022 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.) more...

Purpose: The immune system's role in mediating the cytotoxic effects of chemoradiotherapy remains not completely understood. The integration of immunotherapies into treatment will require insight into features and timing of the immune microenvironment associated with treatment response. Here, we investigated the role of circulating neutrophils and tumor-associated myeloid cells (TSAMs) as potential agents and biomarkers for disease-related outcomes in locally advanced cervical cancer (LACC)., Material and Methods: Hematologic parameters for two LACC patient cohorts, a retrospective clinical and a prospective translational cohort, were obtained at baseline, weekly during chemoradiotherapy for the retrospective cohort, biweekly during chemoradiotherapy for the prospective cohort, and at the first follow-up visit for both cohorts (mean 14.7 weeks, range 8.1-25.1 weeks for the prospective cohort and 5.3 weeks with a range of 2.7-9.0 weeks for the retrospective cohort). In both cohorts, baseline as well as mean and lowest on-treatment values for platelets, hemoglobin, absolute neutrophil count (ANC), and absolute lymphocyte count (ALC) were analyzed for correlations with disease-related outcomes. In the prospective cohort, circulating myeloid cells were isolated from peripheral blood mononuclear cells (PBMCs), and TSAMs were isolated from tumor tissue via a novel serial cytobrush sampling assay. The samples were analyzed by flow cytometry., Results: In both cohorts, the only hematologic parameter significantly associated with survival was elevated on-treatment mean ANC (mANC), which was associated with lower local failure-free and overall survival rates in the retrospective and prospective cohorts, respectively. mANC was not associated with a difference in distant metastases. CD11b + CD11c - TSAMs, which act as a surrogate marker for intratumoral neutrophils, steadily decreased during the course of chemoRT and nadier'd at week 5 of treatment. Conversely, circulating myeloid cells identified from PBMCs steadily increased through week 5 of treatment. Regression analysis confirmed an inverse relationship between circulating myeloid cells and TSAMs at this time point., Conclusions: These findings identify on-treatment mean neutrophil count as a predictor of disease-related outcomes, suggest that neutrophils contribute to chemoradiation treatment resistance, and demonstrate the importance of techniques to measure intratumoral immune activity., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 Published by Elsevier B.V. on behalf of European Society for Radiotherapy and Oncology.) more...

We evaluated the association of disease outcome with T cell immune-related characteristics and T cell receptor (TCR) repertoire in malignant ascites from patients with high-grade epithelial ovarian cancer. Ascitic fluid samples were collected from 47 high-grade epithelial ovarian cancer patients and analyzed using flow cytometry and TCR sequencing to characterize the complementarity determining region 3 TCR β-chain. TCR functions were analyzed using the McPAS-TCR and VDJ databases. TCR clustering was implemented using Grouping of Lymphocyte Interactions by Paratope Hotspots software. Patients with poor prognosis had ascites characterized by an increased ratio of CD8+ T cells to regulatory T cells, which correlated with an increased productive frequency of the top 100 clones and decreased productive entropy. TCRs enriched in patients with an excellent or good prognosis were more likely to recognize cancer antigens and contained more TCR reads predicted to recognize epithelial ovarian cancer antigens. In addition, a TCR motif that is predicted to bind the TP53 neoantigen was identified, and this motif was enriched in patients with an excellent or good prognosis. Ascitic fluid in high-grade epithelial ovarian cancer patients with an excellent or good prognosis is enriched with TCRs that may recognize ovarian cancer-specific neoantigens, including mutated TP53 and TEAD1. These results suggest that an effective antigen-specific immune response in ascites is vital for a good outcome in high-grade epithelial ovarian cancer., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Yoshida-Court et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.) more...

Purpose: Complicating factors such as time pressures, anatomic variants in the spine, and similarities in adjacent vertebrae are associated with incorrect level treatments of the spine. The purpose of this work was to mitigate such challenges by fully automating the treatment planning process for diagnostic and simulation computed tomography (CT) scans., Methods and Materials: Vertebral bodies are labeled on CT scans of any length using 2 intendent deep-learning models-mirroring 2 different experts labeling the spine. Then, a U-Net++ architecture was trained, validated, and tested to contour each vertebra (n = 220 CT scans). Features from the CT and auto-contours were input into a random forest classifier to predict whether vertebrae were correctly labeled. This classifier was trained using auto-contours from cone beam computed tomography, positron emission tomography/CT, simulation CT, and diagnostic CT images (n = 56 CT scans, 751 contours). Auto-plans were generated via scripting. Each model was combined into a framework to make a fully automated clinical tool. A retrospective planning study was conducted in which 3 radiation oncologists scored auto-plan quality on an unseen patient cohort (n = 60) on a 5-point scale. CT scans varied in scan length, presence of surgical implants, imaging protocol, and metastatic burden., Results: The results showed that the uniquely designed convolutional neural networks accurately labeled and segmented vertebral bodies C1-L5 regardless of imaging protocol or metastatic burden. Mean dice-similarity coefficient was 85.0% (cervical), 90.3% (thoracic), and 93.7% (lumbar). The random forest classifier predicted mislabeling across various CT scan types with an area under the curve of 0.82. All contouring and labeling errors within treatment regions (11 of 11), including errors from patient plans with atypical anatomy (eg, T13, L6) were detected. Radiation oncologists scored 98% of simulation CT-based plans and 92% of diagnostic CT-based plans as clinically acceptable or needing minor edits for patients with typical anatomy. On average, end-to-end treatment planning time of the clinical tool was less than 8 minutes., Conclusions: This novel method to automatically verify, contour, and plan palliative spine treatments is efficient and effective across various CT scan types. Furthermore, it is the first to create a clinical tool that can automatically verify vertebral level in CT images., (Published by Elsevier Inc.) more...

Background: Clinically relevant genetic predictors of radiation response for cervical cancer are understudied due to the morbidity of repeat invasive biopsies required to obtain genetic material. Thus, we aimed to demonstrate the feasibility of a novel noninvasive cervical swab technique to (1) collect tumor DNA with adequate throughput to (2) perform whole-exome sequencing (WES) at serial time points over the course of chemoradiation therapy (CRT)., Methods: Cervical cancer tumor samples from patients undergoing chemoradiation were collected at baseline, at week 1, week 3, and at the completion of CRT (week 5) using a noninvasive swab-based biopsy technique. Swab samples were analyzed with whole-exome sequencing (WES) with mutation calling using a custom pipeline optimized for shallow whole-exome sequencing with low tumor purity (TP). Tumor mutation changes over the course of treatment were profiled., Results: 216 samples were collected and successfully sequenced for 70 patients (94% of total number of tumor samples collected). A total of 33 patients had a complete set of samples at all four time points. The mean mapping rate was 98% for all samples, and the mean target coverage was 180. Estimated TP was greater than 5% for all samples. Overall mutation frequency decreased during CRT but mapping rate and mean target coverage remained at >98% and >180 reads at week 5., Conclusion: This study demonstrates the feasibility and application of a noninvasive swab-based technique for WES analysis which may be applied to investigate dynamic tumor mutational changes during treatment to identify novel genes which confer radiation resistance., Competing Interests: The authors have declared that no competing interests exist. more...

Purpose: We compared the magnitude of changes in bone mineral density (BMD), within and outside the radiation field, among women who received pelvic radiation therapy (RT) with or without chemotherapy for cervical cancer., Methods and Materials: In this secondary analysis of a prospective study, we analyzed serial computed tomography scans and dual-energy x-ray absorptiometry scans from 78 patients who received definitive RT or chemoradiation therapy (CRT) for cervical cancer at a single institution from 2008 to 2015. BMD values at L1, L2, L3, and L4 were measured. We compared changes in BMD within the radiation field (ie, at L4) with those outside the field (ie, at L1). Linear mixed models were also used to examine the effect of RT on changes in BMD over time and covariate adjustment., Results: The median age of the 78 patients was 45.5 years (range, 23-88 years); all received RT and 76 (97%) received concurrent CRT. Treatment was associated with significant declines in BMD in all 4 lumbar vertebral bodies over time (P < .05), with nadir at 3 months for L4 and at 1 year for L1. Pairwise comparisons at 3 months and 2 years after treatment indicated that BMD in L4 (within the RT field) had improved (P = .037), but BMD in L1 (outside the RT field) was no different at 3 months and 2 years., Conclusions: Significant BMD declines were observed in all lumbar vertebral bodies immediately after RT. However, in-field vertebral bodies reached nadir BMD earlier than those located outside the RT field. Our results suggest that treatment and patient-related factors other than RT may contribute to declines in BMD after treatment for cervical cancer. Routine bone density screening and post-RT therapy with hormones may be beneficial for selected patients who receive CRT for cervical cancer., (Copyright © 2022 American Society for Radiation Oncology. Published by Elsevier Inc. All rights reserved.) more...

Background: Gut microbiome community composition differs between cervical cancer (CC) patients and healthy controls, and increased gut diversity is associated with improved outcomes after treatment. We proposed that functions of specific microbial species adjoining the mucus layer may directly impact the biology of CC., Method: Metagenomes of rectal swabs in 41 CC patients were examined by whole-genome shotgun sequencing to link taxonomic structures, molecular functions, and metabolic pathway to patient's clinical characteristics., Results: Significant association of molecular functions encoded by the metagenomes was found with initial tumor size and stage. Profiling of the molecular function abundances and their distributions identified 2 microbial communities co-existing in each metagenome but having distinct metabolism and taxonomic structures. Community A (Clostridia and Proteobacteria predominant) was characterized by high activity of pathways involved in stress response, mucus glycan degradation and utilization of degradation byproducts. This community was prevalent in patients with larger, advanced stage tumors. Conversely, community B (Bacteroidia predominant) was characterized by fast growth, active oxidative phosphorylation, and production of vitamins. This community was prevalent in patients with smaller, early-stage tumors., Conclusions: In this study, enrichment of mucus degrading microbial communities in rectal metagenomes of CC patients was associated with larger, more advanced stage tumors., (© 2022. The Author(s).) more...

Purpose: Patients with localized squamous cell carcinoma of the anus (SCCA) who experience treatment toxicity or recurrences have few therapeutic options. Investigation into the microbiome's influence on treatment toxicity and its potential use as a predictive biomarker could improve these patients' outcomes. Our study presents the first longitudinal characterization of the SCCA tumor microbiome and its associations with treatment-related toxicities., Methods and Materials: This prospective cohort study included patients with nonmetastatic SCCA receiving standard-of-care chemoradiation therapy. Anorectal swabs of the tumor site were collected before, during, and after treatment. Patient-reported quality-of-life metrics were collected at similar time points. 16S rRNA gene sequencing was used to perform diversity and taxonomic characterization of the SCCA microbiome. Wilcoxon signed-rank tests were used to compare microbial diversity and abundance over time. Wilcoxon rank-sum tests were used to compare microbial profiles of high and low toxicity groups., Results: Twenty-two patients with SCCA were included in this study with a median age of 58.5 years (range, 39-77 years), and 18 (82%) were women. Alpha diversity remained relatively stable throughout chemoradiation therapy except for decreases in the Observed Features (P = .03) index at week 5 relative to baseline. Tumor microbial compositions changed significantly by the end of treatment (P = .03). Differential enrichment of bacteria at specific time points occurred during treatment, including the enrichment of Clostridia at follow-up (vs week 5, q = 0.019) and Corynebacterium at week 5 (vs baseline, q = 0.015; vs follow-up, q = 0.022). Patients experiencing high toxicity at week 5 had higher relative counts of Clostridia, Actinobacteria, and Clostridiales at baseline (P = .03 for all)., Conclusions: The tumor microbiome changes during and after chemoradiation therapy, and patient-reported toxicity levels are associated with patients' microbial profiles. Further studies into these microbial characterizations and toxicity associations will elucidate the tumor microbiome's role in predicting treatment-related outcomes for patients with SCCA., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.) more...

Objective: Radiation therapy (RT) may improve outcomes for patients with oligometastatic cancer. We sought to determine if there are long-term survivors treated with definitive RT for recurrent or oligometastatic gynecological cancer (ROMGC), and to evaluate the clinical and disease characteristics of these patients., Methods: We performed a landmark analysis in 48 patients with ROMGC who survived for ≥5 years following definitive RT of their metastasis. Patient characteristics were extracted from the medical record. DFS was modeled using the Kaplan-Meier method., Results: This cohort included 20 patients (42%) with ovarian cancer, 16 (33%) with endometrial cancer, 11 (23%) with cervical cancer, and one (2%) with vaginal cancer. The sites of ROMGC were the pelvic (46%), para-aortic (44%), supraclavicular (7%), mediastinal (4%), axillary (4%) lymph nodes and the lung (5.5%). Median total RT dose and fractionation were 62.1 Gy and 2.1 Gy/fraction; one patient was treated with SBRT. 32 patients (67%) received chemoradiation; these patients had higher rates of median DFS than those treated with RT alone (93 vs. 34 months, P = 0.05). At median follow-up of 11.7 years, 11 (23%) patients had progression of disease. 20 (42%) patients had died, 9 (19%) died from non-gynecologic cancer and 8 (17%) from gynecologic cancer (three were unknown). 25 (52%) patients were alive and disease-free (10 initially had endometrial cancer [63% of these patients], eight had cervical cancer [73%], six had ovarian cancer [30%], one had vaginal cancer [100%])., Conclusions: Long-term survival is possible for patients treated with definitive RT for ROMG, however randomized data are needed to identify which patients derive the most benefit., Competing Interests: Declaration of Competing Interest None., (Copyright © 2021 Elsevier Inc. All rights reserved.) more...