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Among pts with CLL/SLL, the median number of prior lines of therapy was 4 (range, 2-18), all pts received prior BTKi therapy, 27 (47%) received prior BTKi and BCL2i therapy, and 36 (63%) had C481S-mutated BTK. B Background: b Bruton tyrosine kinase inhibitors (BTKis) are standard of care for chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), but resistance can develop, most commonly because of BTK mutations. AbbVie, AstraZeneca, BeiGene, Century Therapeutics, Genentech, Genmab, Hutchison MediPharma, Iksuda Therapeutics, InnoCare Pharma, Janssen, Kite Pharma, MorphoSys, Myeloid Therapeutics, Novartis, Nurix Therapeutics, Pharmacyclics, Roche, Secura Bio, Servier Pharmaceuticals, Takeda, TG Therapeutics, Verastem, Vincerx Pharma, Xencor Research funding: Research Grants: All payments made to Sarah Cannon Research Institute, not to the physician. [Extracted from the article]

Abstract Background microRNAs (miRNAs) are small (~22 nt) non-coding RNAs that regulate cell movement, specification, and development. Expression of miRNAs is highly regulated, both spatially and temporally. Based on direct cloning, sequence conservation, and predicted secondary structures, a large number of miRNAs have been identified in higher eukaryotic genomes but whether these RNAs are simply a subset of a much larger number of noncoding RNA families is unknown. This is especially true in zebrafish where genome sequencing and annotation is not yet complete. Results We analyzed the zebrafish genome to identify the number and location of proven and predicted miRNAs resulting in the identification of 35 new miRNAs. We then grouped all 415 zebrafish miRNAs into families based on seed sequence identity as a means to identify possible functional redundancy. Based on genomic location and expression analysis, we also identified those miRNAs that are likely to be encoded as part of polycistronic transcripts. Lastly, as a resource, we compiled existing zebrafish miRNA expression data and, where possible, listed all experimentally proven mRNA targets. Conclusion Current analysis indicates the zebrafish genome encodes 415 miRNAs which can be grouped into 44 families. The largest of these families (the miR-430 family) contains 72 members largely clustered in two main locations along chromosome 4. Thus far, most zebrafish miRNAs exhibit tissue specific patterns of expression.

Purpose: The presence of MYC and BCL2 translocations (ie, double-hit lymphoma, DHL) in large B-cell lymphoma (LBCL) is associated with reduced chemosensitivity, but less is known on its impact on radiotherapy (RT) efficacy., Methods and Materials: Patients with LBCL who received their first course of RT for relapsed/refractory disease between 2008 and 2020 were eligible if there was adequate pathologic evaluation to be categorized as DHL versus non-DHL as per the World Health Organization (fifth edition). Separate analyses were conducted by treatment intent. Predictors for response (complete and partial) and local recurrence (LR) were evaluated using Cox regression analysis. LR analysis was restricted to curative-intent patients to ensure adequate follow-up., Results: Three hundred and eighty-three patients (102 DHL, 281 non-DHL, and 44% curative) were treated at 447 sites. Median time from diagnosis to RT was 11.6 months, with 38.7% of patients having primary chemorefractory disease, 37.4% having received >2 lines of systemic therapy, and 24% status post-stem cell transplant. Median biological equivalent dose (alpha/beta: 10) was 28 Gy (range: 3.2-60.0) for palliative and 46.9 Gy (range: 6.4-84.0) for curative-intent patients. With a median follow-up of 41.1 and 41.5 months among curative and palliative patients, respectively, the response was high (81.1% curative, 60.1% palliative). On univariate analysis, DHL pathology was not associated with RT response in either curative or palliative patients. Among curative patients, 2-year LR rate was 38.8%. On multivariable analysis, DHL pathology was associated with a 2 times higher risk of LR (95% CI: 1.05-3.67, P = .03), with a crude LR rate of 42.9% (DHL) versus 28.9% (non-DHL). RT was well tolerated with low rates of grade 3 or higher acute toxicity (1.8% curative, 2.9% palliative)., Conclusions: Relapsed/refractory LBCL remains radioresponsive with a 60%-80% response rate to RT. Although DHL pathology does not appear to influence RT response, its presence is associated with higher rates of LR, suggesting that it may be more radioresistant., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: To assess whether a radiation therapy (RT) dose affects response in bulky tumors in relapsed/refractory (r/r) diffuse large B-cell lymphoma (DLBCL)., Methods and Materials: Data from patients with r/r DLBCL treated with salvage- or palliative-intent RT (2008-2020) at a single institution were examined. Index lesion size ≥7.5 cm was defined as bulky. Equivalent doses in 2-Gy fractions (EQD2) were calculated to compare doses between conventional and hypofractionated (≥2.5 Gy/fraction) schemes. Objective response rates (ORRs) were compared using nonparametric Mann-Whitney U test or Kruskal-Wallis test with Dunn's multiple comparison corrections. Freedom from local progression (FFLP) was assessed using Kaplan-Meier and Cox proportional hazard regression analyses., Results: One hundred eighty-three courses of 151 unique patients were included (salvage: 37% and palliative: 63%). Nonbulky and bulky tumors were irradiated in 109 (60%) and 74 (40%) courses, respectively. Median EQD2 was 33 Gy (IQR, 23-39 Gy) with hypofractionation in 84 (46%) cases. Of those with post-RT imaging (80%), the ORR was 59%, with a trend toward worsened ORR in bulky tumors (50% vs 65%, P = .077). For bulky tumors, RT regimens with EQD2s >30 Gy were associated with better ORR (≤30 Gy vs >30 Gy: 27% vs 64%, P = .0073), whereas a lower EQD2 cutoff was sufficient for nonbulky tumors (≤20 Gy vs >20 Gy: 38% vs 75%, P = .0011). On multivariable regression analysis, bulky tumor size was associated with worsened FFLP (hazard ratio, 2.07; 95% CI, 1.16-3.68; P = .014), whereas high EQD2s >30 Gy were associated with better FFLP (hazard ratio, 0.48; 95% CI, 0.25-0.93; P = .031). Bulky tumors treated with EQD2s ≤30 Gy had the lowest median FFLP (4.0 months), whereas EQD2s >30 Gy had an unreached median FFLP (P = .0047)., Conclusions: Bulky r/r DLBCL tumors were associated with less favorable tumor control outcomes in the salvage and palliative settings. RT regimens with higher EQD2s (>30 Gy) should be considered if durable local control of bulky tumors is desired., Competing Interests: Disclosures J. Svoboda: TG Therapeutics: Research Funding; BMS: Consultancy, Research Funding; ADCT: Consultancy; SEAGEN: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; Adaptive: Consultancy, Research Funding; Atara: Consultancy; Incyte: Consultancy, Research Funding; Astra Zeneca: Consultancy, Research Funding; Merck: Research Funding; Genmab: Consultancy. All other authors have no conflicts of interest to disclose., (Copyright © 2024 American Society for Radiation Oncology. Published by Elsevier Inc. All rights reserved.)

Purpose: To identify the characteristics, indications, and toxicities among patients receiving proton beam therapy (PBT) in the final year of life at an academic medical center., Materials and Methods: A retrospective review of patients who received PBT within the final 12 months of life was performed. Electronic medical records were reviewed for patient and treatment details from 2010 to 2019. Patients were followed from the start of PBT until death or last follow-up. Acute (3 months) toxicities were graded using the Common Terminology Criteria for Adverse Events v5.0. Imaging response was assessed using the Response Evaluation Criteria in Solid Tumors v1.1. The χ 2 test was used to evaluate factors associated with palliative treatment. Simple logistic regression was used to evaluate factors associated with toxicity., Results: Bet299 patients were treated at the end of life (EOL) out of 5802 total patients treated with PBT (5.2%). Median age was 68 years (19-94 years), 58% male. The most common cancer was nonsmall cell lung cancer (27%). Patients were treated for symptom palliation alone (11%), durable control (57%), curative intent (16%), local recurrence (14%), or oligometastatic disease (2%). Forty-five percent received reirradiation. Median treatment time was 32 days (1-189 days). Acute toxicity was noted in 85% of the patients (31% G1, 53% G2, 15% G3). Thirteen patients (4%) experienced chronic toxicity. Breast and hematologic malignancy were associated with palliative intent χ 2 (1, N  = 14) = 17, P  = .013; (χ 2 (1, N  = 14) = 18, P  = .009)., Conclusion: The number of patients treated with PBT at the EOL was low compared to all comers. Many of these patients received treatment with definitive doses and concurrent systemic therapy. Some patients spent a large portion of their remaining days on treatment. A prognostic indicator may better optimize patient selection for PBT at the EOL., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Neil K Taunk reports a relationship with Varian Medical Systems Inc that includes: consulting or advisory and funding grants. Neil K Taunk reports a relationship with Therapanacea AI that includes: funding grants. Neil K Taunk reports a relationship with Radiological Society of North America that includes: funding grants. Neil K Taunk reports a relationship with POINT Biopharma Global Inc that includes: consulting or advisory. Neil K Taunk reports a relationship with Telix Pharmaceuticals Limited that includes: consulting or advisory. Neil K Taunk reports a relationship with Boston Scientific Corp that includes: consulting or advisory. James M Metz reports a relationship with Varian Medical Systems Inc that includes: speaking and lecture fees. James M Metz reports a relationship with IBA Dosimetry US that includes: speaking and lecture fees. Jeffrey D Bradley reports a relationship with Varian Medical Systems Inc that includes: consulting or advisory. Jeffrey D Bradley reports a relationship with AstraZeneca Pharmaceuticals LP that includes: board membership and consulting or advisory. Jeffrey D Bradley reports a relationship with Mevion Medical Systems that includes: board membership. Jeffrey D Bradley reports a relationship with Genentech that includes: board membership. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Author(s).)

Introduction: SBRT is an increasingly popular treatment for localized prostate cancer, though considerable variation in technical approach is common and optimal dose constraints are uncertain. In this study, we sought to identify dosimetric and patient-related predictors of acute rectal toxicity., Methods: Patients included in this study were treated with prostate SBRT on a prospective institutional protocol. Physician-graded toxicity and patient-reported outcomes were captured at one week, one month, and three months following SBRT. DVH data were extracted and converted into relative volume differential DVHs for NTCP modeling. Patient- and disease-related covariates along with NTCP model predictions were independently tested for significant association with physician-graded toxicity or a decline in bowel-related QoL. A multivariate model was constructed using forward selection, and significant parameter cutoff values were obtained with Fischer's exact test to group patients by risk of developing physician-graded toxicity or detriments in patient-reported QoL., Results: One hundred and three patients treated for localized prostate cancer with SBRT were included in our analysis. 52% of patients experienced a clinically significant decline in bowel-related QOL within 1 week of completion of treatment, while only 27.5% of patients developed grade 2+ physician-graded rectal toxicity. Sequential feature selection multivariate logistic regression identified rectal V22.5 Gy ( p  = 0.001) and D19% ( p  = 0.001) as independent predictors of clinically significant toxicity, while rectal V20Gy ( p  = 0.004) and D25.3% ( p  = 0.007) were independently correlated with physician-graded toxicity. Global multivariate step-wise logistic regression identified only D19% ( p  = 0.001) and V20Gy ( p  = 0.004) as independent predictors of acute bowel bother or physician-graded rectal toxicity respectively., Conclusions: Moderate doses to large rectal volumes, D19% and V20Gy, were associated with an increased incidence of a clinically significant decrease in patient-reported bowel QOL and physician-scored grade 2+ rectal toxicity, respectively. These dosimetric parameters may help practitioners mitigate acute toxicity in patients treated with prostate SBRT.

Background and Purpose: CD19-targeting chimeric antigen receptor T-cell (CART) therapy is a promising treatment for relapsed/refractory non-Hodgkin lymphoma, but most patients experience post-CART progression. We describe our institutional experience of salvage radiotherapy (SRT) in this setting., Materials and Methods: Of 94 patients who received CART therapy from 2018 to 2020, 21 received SRT for post-CART progression. Patients were divided into two groups: locoregional disease (n = 9 [43 %], all disease encompassable within an RT field) and advanced disease (n = 12 [57 %]). Patterns of failure, progression-free survival (PFS), overall survival (OS), and toxicity were assessed., Results: Median time from CART infusion to SRT was 4.0 months (range, 0.6-11.5 months). In the locoregional disease group, 8/9 patients (89 %) were treated with comprehensive SRT to a median dose of 37.5 Gy in a median of 15 fractions. In the advanced disease group, all patients (n = 12) were treated with focal SRT to a median dose of 20.8 Gy in a median of 5 fractions. Median follow-up post-SRT was 15.2 months. In-field response was observed in 8/9 (89 %) in the locoregional disease and 8/9 (89 %) evaluable patients in the advanced disease groups. 17/18 evaluable patients (94 %) patients experienced post-SRT progression, all with a distant component. Median OS was 7.4 months; 21 months for locoregional disease versus 2.4 months for advanced disease (p = 0.0002). Median PFS was 1.1 month, and similarly poor regardless of group. No grade ≥ 3 toxicities occurred., Conclusions: SRT post-CART therapy appears safe with encouraging in-field response but high rates of out-of-field progression, even for those presenting with locoregional disease, highlighting the need for integration of novel systemic agents., 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.)

Cherenkov images can be used for the quality assurance of dose homogeneity in total skin electron therapy (TSET). For the dose mapping purpose, this study reconstructed the patient model from 3D scans using registration algorithms and computer animation techniques. The Cherenkov light emission of the patient's surface was extracted from multi-view Cherenkov images, converted into dose distribution, and projected onto the patient's 3D model, allowing for dose cumulation and evaluation. The projected result from multiple Cherenkov cameras provides additional information about Cherenkov emission on the sides of the patients, which improves the agreement between the Cherenkov converted dose and the OSLD measurements.

Purpose: Combined modality therapy with multiagent chemotherapy and radiation therapy is a standard treatment option for aggressive mediastinal non-Hodgkin lymphomas (AMNHLs); however, concerns regarding acute and late radiation toxicities have fueled an effort to use systemic therapy alone. The use of proton therapy (PT) is a promising treatment option, but there are still limited data regarding clinical outcomes with this treatment modality. In this Particle Therapy Cooperative Group lymphoma subcommittee collaboration, we report outcomes of patients with AMNHL treated with pencil-beam scanning PT or double-scatter PT after chemotherapy., Methods and Materials: This was a multi-institutional retrospective observational cohort study of patients with AMNHL treated with PT following chemotherapy between 2011 and 2021. Progression-free survival (PFS), local recurrence-free survival (LRFS), and overall survival (OS) rates were estimated with the Kaplan-Meier method. PT toxicity was graded by the Common Terminology Criteria for Adverse Events version 5.0. A 2-tailed paired t test was used for dosimetric comparisons., Results: Twenty-nine patients were identified. With a median follow-up time of 4.2 years (range, 0.2-8.9 years), the estimated 5-year PFS for all patients was 93%, 5-year LRFS was 96%, and estimated 5-year OS was 87%. Maximum acute grade 1 (G1) toxicities occurred in 18 patients, and 7 patients had maximum G2 toxicities. No G3+ radiation-related toxicities were observed. Average mean lung dose and lung V20 Gy were lower for patients treated with pencil-beam scanning PT compared with double-scatter PT ( P  = .016 and .006, respectively), while patients with lower mediastinal disease had higher doses for all evaluated dosimetric heart parameters., Conclusions: PT after chemotherapy for patients with AMNHL resulted in excellent outcomes with respect to 5-year PFS, LRFS, and OS without high-grade toxicities. Future work with larger sample sizes is warranted to further elucidate the role of PT in the treatment of AMNHL., (© 2022 Published by Elsevier Inc. on behalf of American Society for Radiation Oncology.)

Purpose: To report objective response rates (ORR), time to local failure (TTLF), and overall survival (OS) among patients with relapsed or refractory diffuse large B-cell lymphoma after salvage- or palliative-intent radiation therapy (RT) and to investigate whether outcomes differed with conventional versus hypofractionated (≥2.5 Gy/fraction) RT., Methods and Materials: A single-institution observational cohort study was performed for patients who completed a course of RT for relapsed or refractory diffuse large B-cell lymphoma between January 1, 2008, and April 1, 2020. Predictors of ORR, TTLF, and OS were calculated using univariable and multivariable regression models. The Kaplan-Meier method was used to estimate TTLF and OS, and log-rank analysis was used to compare outcomes. Equivalent dose in 2 Gy fractions (EQD2) was calculated using an α/β of 10., Results: One-hundred and sixty-nine patients were treated with 205 RT courses (73 [36%] salvage, 132 [64%] palliative), and hypofractionated RT was used in 100 RT courses (49%). Median RT dose was 30 Gy (range, 8-60 Gy). ORR was 60% for the total cohort (53% and 69% for palliative and salvage cohorts, respectively). Over a median follow-up time of 4 months, median OS in all patients was 5 months (3 and 22 months for palliative and salvage cohorts, respectively). No statistically significant differences in ORR, TTLF, and OS were observed with hypofractionation compared with conventional fractionation. EQD2 ≥35 Gy was associated with improved ORR (odds ratio, 3.79 [1.19-12.03]; P  = .024) and prolonged TTLF (0.39 [0.18-0.87]; P  = .022), while double-hit receptor status (8.18 [1.08-62.05]; P  = .042), cell of origin (3.87 [1.17-8.74]; P  = .0012), and bulky disease (≥7.5 cm; 2.12 [1.18-3.81]; P  = .012) were associated with inferior TTLF. In the palliative-only cohort, a low-dose regimen of 8 Gy in 2 fractions was associated with similar ORR compared with other fractionation schema but trended towards inferior TTLF ( P  = .36)., Conclusions: Hypofractionation is not associated with differences in disease outcomes for patients with relapsed or refractory diffuse large B-cell lymphoma, while higher RT dose (EQD2 ≥35 Gy) may improve ORR and TTLF. Future work is warranted to elucidate the ideal dose and fractionation schema for such patients who will likely also undergo novel systemic agents and cellular therapies., (© 2022 The Author(s).)

Purpose: B-cell maturation antigen (BCMA)-targeted chimeric antigen receptor (CAR) T cells (CART-BCMA) are a promising treatment for relapsed/refractory multiple myeloma (r/rMM). We evaluated the safety and feasibility of bridging radiation (RT) in subjects treated on a phase I trial of CART-BCMA., Experimental Design: Twenty-five r/rMM subjects were treated in three cohorts with two doses of CART-BCMA cells ± cyclophosphamide. We retrospectively analyzed toxicity, response, and CART manufacturing data based on RT receipt., Results: Thirteen subjects received no RT <1 year before CART infusion (Group A). Eight subjects received RT <1 year before CART infusion (Group B) with median time from RT to apheresis of 114 days (range 40-301). Four subjects received bridging-RT (Group C) with a median dose of 22 Gy and time from RT to infusion of 25 days (range 18-35). Group C had qualitatively lower rates of grade 4 (G4) hematologic toxicities (25%) versus A (61.5%) and B (62.5%). G3-4 neurotoxicity occurred in 7.7%, 25%, and 25% in Group A, B, and C, respectively. G3-4 cytokine release syndrome was observed in 38.5%, 25%, and 25% in Group A, B, and C, respectively. Partial response or better was observed in 54%, 38%, and 50% of Group A, B, and C, respectively. RT administered <1 year ( P = 0.002) and <100 days ( P = 0.069) before apheresis was associated with lower in vitro proliferation during manufacturing; however, in vivo CART-BCMA expansion appeared similar across groups., Conclusions: Bridging-RT appeared safe and feasible with CART-BCMA therapy in our r/rMM patients, though larger future studies are needed to draw definitive conclusions., (©2021 American Association for Cancer Research.)

Recent improvements in chemoimmunotherapies, targeted agents, hematopoietic stem cell transplants, and cellular therapies have revolutionized treatment paradigms for patients with diffuse large B-cell lymphoma (DLBCL). Even in the relapsed or refractory setting, contemporary treatment options are delivered with curative intent and can lead to lasting remissions. Although such therapies have improved overall outcomes, they have increasingly led to a wide variety of presentations of recurrent tumors in need of palliation. Here, we review the use of radiotherapy (RT) in the palliation of DLBCL. We draw particular attention to the evolving role for hypofractionated RT and low-dose RT for DLBCL. We review the available literature on these topics and focus on commonly encountered clinical scenarios., (Copyright © 2021. Published by Elsevier Inc.)

Purpose: Radiation therapy (RT) with doses ranging from 24 Gray (Gy) to 40 Gy is a proven treatment modality for indolent orbital adnexal lymphoma (IOAL), but recently the use of low dose RT (LDRT, defined as 2 Gy x 2 fractions) has become a notable alternative. However, limited data exists comparing outcomes following LDRT to moderate-dose RT (MDRT, RT dose 4 - 36 Gy). We present a single institution retrospective analysis comparing outcomes of patients with IOALs following LDRT or MDRT., Methods: A total of 36 patients treated with 38 consecutive courses of RT were identified; LDRT was delivered for 14 courses and MDRT for 24 courses. Overall response rates (ORR) were recorded according to Deauville or RECIST criteria with a response characterized as a complete response (CR) or partial response. Local control (LC), orbital control (OC), and overall survival (OS) rates were estimated with the Kaplan-Meier method. RT toxicity was graded per CTCAEv5 and compared with the Fisher's exact test., Results: Median follow-up time was 29 months (m) (range, 4-129m), and median MDRT dose used was 24 Gy (range 21-36 Gy). Overall response rates (ORR) were 100% (CR 50%) and 87.5% (CR 58.3%) following LDRT and MDRT, respectively. OS at 2 years was 100% and 95% for the LDRT and MDRT groups, respectively (p=0.36). LC rates at 2 years was 100% for both LDRT and MDRT groups and at 4 years was 100% and 89% for the LDRT and MDRT groups, respectively (p=0.56). The 4-year OC rate (including both ipsilateral and contralateral relapses) was 80% and 85% for the LDRT and MDRT groups, respectively (p=0.79). No patient required treatment with RT to a previously irradiated orbit. Acute toxicities were reported following 6 LDRT courses compared to 20 MDRT courses (p=.014). No Grade 3 or higher acute toxicities occurred in either group. Late toxicities were reported following 2 LDRT courses compared to 10 MDRT courses (p=0.147)., Conclusions: LDRT produced similar ORR, LC, OC, and OS rates compared to MDRT with fewer acute and minimal late toxicities reported. Future multi-center studies with larger patient numbers are warranted to show significant associations., Competing Interests: EC reports grants from Novartis Pharmaceuticals Corporation, during the conduct of the study; personal fees from Novartis Pharmaceuticals Corporation, personal fees from Juno Therapeutics, Inc., personal fees from Kite Pharma, Inc., outside the submitted work. 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 © 2021 Baron, Wright, Lee, Carpenter, Manjunath, Briceño, Chong, Maity, Plastaras and Paydar.)

Purpose: Myeloma lesions of the head can present with central nervous system (CNS) involvement (leptomeningeal disease or brain metastasis), cranial neuropathy (CN), or impending neurologic involvement (INI). We analyzed response and survival after palliative radiation therapy (RT) to the brain and/or skull for myeloma lesions to determine whether CNS involvement fared worse than other RT indications., Methods and Materials: We retrospectively analyzed 54 palliative RT courses administered at our institution from 2008 to 2019. Eleven courses were administered for CNS disease, 28 for CN, and 15 for INI. Demographic, disease, and RT variables were recorded as well as clinical response, radiographic response, and survival. Univariate analyses were performed for differences between groups, effects of clinical and RT treatment factors on response, as well as dose response. Survival was analyzed with the Kaplan-Meier method and compared by the log-rank test., Results: This heavily pretreated cohort received a median of 20 to 24 Gy, most often to the base of skull, orbit(s), calvarium, or whole brain. Any clinical response (partial or complete vs no response or progressive disease) was significantly more likely for patients with CN and INI when collectively compared with patients with CNS disease ( P < .001). Dose response was significant for doses ≥15 and 20 Gy for the whole cohort ( P  = .026 and .005, respectively) and patients with CN/INI ( P  = .023 and .002, respectively). Additionally, patients with high-risk cytogenetics were less likely to clinically respond ( P  = .009). Patients with CNS disease had worse survival ( P  = .005)., Conclusions: Patients with leptomeningeal disease/brain metastasis have poor clinical response and survival after RT and their responses do not demonstrate a dose response. Given these poor outcomes, the potential benefit of RT may be limited for some patients who may be alternatively managed by supportive care or short RT courses. Patients with CN/INI have longer survival and better response rates and may benefit from RT courses ≥15 to 20 Gy., (© 2021 The Authors.)

Gingival myeloid sarcoma (MS) refractory to induction chemotherapy is a rare clinical entity and can be treated with palliative radiation therapy (RT). However, there are few previously published reports of RT approaches for the treatment of gingival MS. We present a single institution retrospective observational study of adult patients treated with palliative RT for chemotherapy refractory gingival MS. A total of six patients diagnosed with gingival MS in the setting of relapsed or refractory acute myeloid leukemia treated with palliative RT were identified, with a median age of 66 (range 52-77). Patients were treated with radiation doses ranging from 5 to 20 Gy in 2-10 fractions. Two patients had adequate follow-up time to assess treatment response. One patient who was simulated with PET/CT experienced a local complete response, while the other patient required retreatment 2 months after initial treatment and experienced an eventual local partial response. Three patients experienced radiation mucositis, with one patient experiencing grade 5 toxicity attributed to concomitant treatment with the radiosensitizer hydroxyurea. We believe that this study can provide a practical reference point for other clinicians given the rarity of gingival MS requiring palliative radiation therapy as a clinical entity., Competing Interests: AP has been a consultant for and received research funding from Astellas Pharma outside this study. 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 © 2021 Lee, Baron, Wright, Plastaras, Perl and Paydar.)

Total Skin Electron Therapy (TSET) utilizes high-energy electrons to treat cancers on the entire body surface. The otherwise invisible radiation beam can be observed via the optical Cherenkov photons emitted from interaction between the high-energy electron beam and tissue. Cherenkov emission can be used to evaluate the dose uniformity on the surface of the patient in real-time using a time-gated intensified camera system. Each patient was monitored during TSET by in-vivo detectors (IVD) as well as Scintillators. Patients undergoing TSET in various conditions (whole body and half body) were imaged and analyzed. A rigorous methodology for converting Cherenkov intensity to surface dose as products of correction factors, including camera vignette correction factor, incident radiation correction factor, and tissue optical properties correction factor. A comprehensive study has been carried out by inspecting various positions on the patients such as vertex, chest, perineum, shins, and foot relative to the umbilicus point (the prescription point).

Context: Advanced gynecologic malignancies can cause significant vaginal bleeding. Radiotherapy (RT) is often used to palliate symptoms, but limited data exist concerning the optimal dose and expected time to bleeding hemostasis in this population., Objectives: 1) To investigate the overall hemostasis response and kinetics of hemostasis in women with gynecologic malignancies receiving palliative RT. 2) To compare the efficacy of short-course RT (SCRT, less than or equal to five fractions, >3.5 Gy per fraction) vs. conventionally fractionated long-course regimens (greater than five fractions)., Methods: We identified women receiving palliative RT for bleeding gynecologic malignancies. Initial and maximal hemostasis responses (IHR and MHR) were recorded and categorized as progressive bleeding (PD), stable disease (SD), partial response (PR), or complete response (CR). Clinical variables were correlated with response or toxicity using binary logistic regression statistical methods., Results: Thirty-three women (median age 63) were identified between 2010 and 2019. Median follow-up and survival after RT were 131 days. About 54.5% (18 of 33) received SCRT. Median time to IHR was five days (two-and-a-half days with SCRT) and 78.8% (26 of 33) responded during treatment. Median time to MHR was 13 days. About 100% achieved PR or CR at MHR. Rates of CR were similar between SCRT (83%) and conventionally fractionated schedules (87%). Average durability of hemostatic control was 5.4 months. Overall rate of rebleeding and Grade 3+ toxicity was 9.1% (3 of 33 each)., Conclusion: Women receiving SCRT for bleeding gynecologic malignancies achieved rapid symptom control (often during treatment) with minimal rebleeding. In a population whose median survival is four months, SCRT effectively addresses symptomatic disease while minimizing patient burden and toxicity., (Copyright © 2020 American Academy of Hospice and Palliative Medicine. Published by Elsevier Inc. All rights reserved.)

Purpose: Single extracranial metastases from ovarian and uterine malignancies have historically been treated with surgery or conventional radiation. We report mature local control (LC), overall survival (OS), progression free survival (PFS), and toxicity for patients who completed 5-fraction stereotactic body radiation therapy (SBRT)., Methods: Patients with biopsy-proven, single extracranial metastases from primary ovarian and uterine malignancies treated with 5-fraction SBRT were included. Patients were stratified based on tumor volume (small < 50 cc or large ≥ 50 cc) and dose (low dose < 35 Gy or high ≥ 35 Gy). Kaplan-Meier method was used to estimate LC, OS, and PFS., Results: Between July 2007 and July 2012, 20 patients underwent SBRT to a single extracranial metastasis. Primary site was divided evenly between ovarian and uterine (n = 10 each). Metastases involved the liver (30%), abdominal lymph nodes (25%), lung (20%), pelvic lymph nodes (10%), spine (10%), and extremity (5%). The median gross tumor volume (GTV) was 42.5 cc (range, 5-273 cc) and the median dose to the GTV was 35 Gy (range, 30-50 Gy). At a median follow-up of 56 months, the 5-year LC and OS estimates were 73 and 46%. When stratified by tumor volume, the 5-year LC and OS for small tumors were significantly better at 100% (p < 0.01) and 65% (p < 0.02). When stratified by dose, the 5-year LC was 87.5% with high dose and 53.6% with low dose (p = 0.035). The 5-year PFS for the entire cohort was 20%. Four patients with small metastases who had complete response remained disease free at study completion and were considered cured (median PFS > 10 years). Treatment was generally well tolerated, and only one patient experienced a late grade III musculoskeletal SBRT related toxicity., Conclusions: SBRT is a versatile, well-tolerated, and effective treatment option for single extracranial metastases from ovarian and uterine primary tumors. 35 Gy in five fractions appears to be a practical minimum effective dose. Four patients with small metastases were disease free at the study completion and considered cured. However, patients with larger metastases (≥50 cc) may require higher SBRT dosing or alternative treatments., Competing Interests: SC and BC serve as clinical consultants to Accuray Inc. The Department of Radiation Medicine at Georgetown University Hospital receives a grant from Accuray to support a research coordinator. 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 © 2020 Aghdam, Repka, McGunigal, Pepin, Paydar, Rudra, Paudel, Pernia Marin, Suy, Collins, Barnes and Collins.)

Purpose: CD19-targeting chimeric antigen receptor T-cell (CART) therapy has emerged as a promising treatment for relapsed/refractory aggressive B-cell lymphoma (r/rABL), culminating in 2 US Food and Drug Administration-approved therapies: tisagenlecleucel (tisa-cel) and axicabtagene ciloleucel (axi-cel). Following leukapheresis and in preparation for CART infusion, contemporary bridging and lymphodepletion regimens rely mostly on cytotoxic chemotherapy. Here, in a cohort of patients treated with commercial tisa-cel and axi-cel, we show that bridging-RT may offer a supplemental approach., Methods and Materials: Thirty-one patients receiving commercial tisa-cel (n = 13) or axi-cel (n = 18) between August 2018 and February 2019 for r/rABL were retrospectively reviewed. Patients were categorized into 2 groups: (1) bridging-RT within 30 days of CART infusion or (2) nonbridging-RT (NBRT), in which patients received either remote RT greater than 30 days before CART infusion or no prior RT., Results: Five patients received bridging-RT within 30 days of CART infusion. Median bridging-RT dose was 37.5 Gy and was completed a median of 13 days before infusion. No grade 3 (G3) or higher RT-toxicities occurred. No patients in the bridging-RT group experienced G3 or higher CART-related toxicities (CRS or neurotoxicity), and 23% (n = 6) and 15% (n = 4) experienced G3-5 CRS and G3-5 neurotoxicity in the NBRT group, respectively. Overall treatment response in the bridging-RT and NBRT groups was 80% and 64%, respectively. The axi-cel CART product was associated with CRS (odds ratio [OR] = 26.67, P = .001) and CRS correlated with neurotoxicity (OR = 12.22, P = .028). There was a trend toward an association for CRS with metabolic tumor volume (OR = 1.06/mL, P = .141) and TLG (OR = 1.01/mL x standard uptake value, P = .099)., Conclusions: Bridging-RT before commercial CART does not appear to increase the risk for CART-related toxicities or negatively affect outcomes in r/rABL patients. No G3 or higher RT-toxicities occurred in this series. Pretreatment metabolic tumor burden may be associated with CART-associated CRS; however, larger patient numbers are required to elucidate significant associations. Future work to prospectively assess the value of bridging-RT is warranted., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose: To investigate the survival endpoints in women with International Federation of Gynecology and Obstetrics (FIGO) stage II endometrial cancer who received adjuvant vaginal brachytherapy (VBT) alone using multi-institutional pooled data., Methods and Materials: We performed a multi-institutional analysis of surgically staged patients with FIGO stage II endometrioid-type endometrial cancer treated with VBT alone. Patient, tumor, and treatment characteristics were collected and analyzed. Univariable and multivariable frailty survival models were performed to assess clinicopathologic risk factors for recurrence and death., Results: One hundred six patients were included (92 VBT alone and 14 VBT with chemotherapy) with median follow-up of 39.0 months. Pelvic node dissection was performed in 89.6% of patients. One hundred four patients (98.1%) and 2 patients (1.9%) had microscopic and macroscopic cervical stromal invasion, respectively. Grade 1 or 2 disease occurred in 88.6% of patients. For patients treated with VBT without chemotherapy, the 5-year estimates of vaginal failure, pelvic nodal failure, and distant metastases were 2.6%, 4.2%, and 7.2%, respectively. Five-year progression-free survival and overall survival were 74.0% and 76.2%, respectively. On univariable and multivariable models for progression-free survival, increasing age and lack of pelvic node resection were hazardous (P < .05)., Conclusions: Vaginal and pelvic failure rates were low in this selected population of stage II patients receiving adjuvant VBT without external beam radiation therapy. It is reasonable to consider adjuvant VBT alone in selected patients with grade 1 or 2 disease and microscopic cervical stromal invasion who underwent pelvic lymphadenectomy., (Published by Elsevier Inc.)

Background: Defining prognostic factors is a crucial initial step for determining the management of patients with brain metastases. Randomized trials assessing radiosurgery have commonly limited inclusion criteria to 1 to 4 brain metastases, in part due to multiple retrospective studies reporting on the number of brain metastases as a prognostic indicator. The present study reports on the survival of patients with 1 to 4 versus ≥5 brain metastases treated with radiosurgery., Methods: We evaluated a retrospective multi-institutional database of 1523 brain metastases in 507 patients who were treated with radiosurgery (Gamma Knife or Cyberknife) between 2001 and 2014. A total of 243 patients were included in the analysis. Patients with 1 to 4 brain metastases were compared with patients with ≥5 brain metastases using a standard statistical analysis. Cox hazard regression was used to construct a multivariable model of overall survival (OS). To find covariates that best separate the data at each split, a machine learning technique Chi-squared Automated Interaction Detection tree was utilized., Results: On Pearson correlation, systemic disease status, number of intracranial metastases, and overall burden of disease (number of major involved organ systems) were found to be highly correlated (P<0.001). Patients with 1 to 4 metastases had a median OS of 10.8 months (95% confidence interval, 6.1-15.6 mo), compared with a median OS of 8.5 months (95% confidence interval, 4.4-12.6 mo) for patients with ≥5 metastases (P=0.143). The actuarial 6 month local failure rate was 5% for patients with 1 to 4 metastases versus 3.2% for patients with ≥5 metastases (P=0.404). There was a significant difference in systemic disease status between the 2 groups; 30% of patients had controlled systemic disease in the <5 lesions group, versus 8% controlled systemic disease in the ≥5 lesions group (P=0.005). Patients with 1 to 4 metastases did not have significantly improved OS in a multivariable model adjusting for systemic disease status, systemic extracranial metastases, and other key variables. The Chi-squared Automated Interaction Detection tree (machine learning technique) algorithm consistently identified performance status and systemic disease status as key to disease classification, but not intracranial metastases., Conclusions: Although the number of brain metastases has previously been accepted as an independent prognostic indicator, our multicenter analysis demonstrates that the number of intracranial metastases is highly correlated with overall disease burden and clinical status. Proper matching and controlling for these other determinants of survival demonstrates that the number of intracranial metastases alone is not an independent predictive factor, but rather a surrogate for other clinical factors.

Background: Recent data suggest that intensity-modulated radiation therapy (IMRT) plus brachytherapy boost for unfavorable prostate cancer provides improved biochemical relapse-free survival over IMRT alone. Stereotactic body radiation therapy (SBRT) may be a less invasive alternative to brachytherapy boost. Here, we report the 3-year gastrointestinal (GI) and genitourinary (GU) toxicities of IMRT plus SBRT boost., Materials and Methods: Between March 2008 and September 2012, patients with prostate cancer were treated with robotic SBRT (19.5 Gy in three fractions) followed by fiducial-guided IMRT (45-50.4 Gy) on an institutional protocol. Toxicity was prospectively graded using the common terminology criteria for adverse events version 4.0 (CTCAEv.4) at the start of and at 1- to 6-month intervals after therapy. Rectal telangiectasias were graded using the Vienna Rectoscopy Score (VRS)., Results: At a median follow-up of 4.2 years (2.4-7.5), 108 patients (4 low-, 45 intermediate-, and 59 high-risk) with a median age of 74 years (55-92) were treated with SBRT plus IMRT, with 8% on anticoagulation and an additional 48% on antiplatelet therapy at the start of therapy. The cumulative incidence of late ≥grade 2 GI toxicity was 12%. Of these, 7% were due to late rectal bleeding, with six patients requiring up to two coagulation procedures. One patient with rectal telangiectasias was treated with hyperbaric oxygen (grade 3 toxicity). No rectal fistulas or stenoses were observed. Ten patients had multiple non-confluent telangiectasias (VRS grade 2), and three patients had multiple confluent telangiectasias (VRS grade 3). The cumulative incidence of late grade 3 GU toxicity was 6%. Most late toxicities were due to hematuria requiring bladder fulguration. There were no late ≥grade 4 GU toxicities., Conclusion: Rates of clinically significant GI and GU toxicities are modest following IMRT plus SBRT boost. Future studies should compare cancer control, quality of life, and toxicity with other treatment modalities for patients with high-risk prostate cancer.

Background: Proctitis following prostate cancer radiation therapy is a primary determinant of quality of life (QOL). While previous studies have assessed acute rectal morbidity at 1 month after stereotactic body radiotherapy (SBRT), little data exist on the prevalence and severity of rectal morbidity within the first week following treatment. This study reports the acute bowel morbidity 1 week following prostate SBRT., Materials and Methods: Between May 2013 and August 2014, 103 patients with clinically localized prostate cancer were treated with 35-36.25 Gy in five fractions using robotic SBRT delivered on a prospective clinical trial. Bowel toxicity was graded using the Common Terminology Criteria for Adverse Events version 4.0 (CTCAEv.4). Bowel QOL was assessed using the EPIC-26 questionnaire bowel domain at baseline, 1 week, 1 month, and 3 months. Time-dependent changes in bowel symptoms were statistically compared using the Wilcoxon signed-rank test. Clinically significant change was assessed by the minimally important difference (MID) in EPIC score. This was defined as a change of 1/2 standard deviation (SD) from the baseline score., Results: One-hundred and three patients with a minimum of 3 months of follow-up were analyzed. The cumulative incidence of acute grade 2 gastrointestinal (GI) toxicity was 23%. There were no acute ≥ grade 3 bowel toxicities. EPIC bowel summary scores maximally declined at 1 week after SBRT (-13.9, p < 0.0001) before returning to baseline at 3 months after SBRT (+0.03, p = 0.94). Prior to treatment, 4.9% of men reported that their bowel bother was a moderate to big problem. This increased to 28.4% (p < 0.0001) 1 week after SBRT and returned to baseline at 3 months after SBRT (0.0%, p = 0.66). Only the bowel summary and bowel bother score declines at 1 week met the MID threshold for clinically significant change., Conclusion: The rate and severity of acute proctitis following prostate SBRT peaked at 1 week after treatment and returned to baseline by 3 months. Toxicity assessment at 1 week can therefore minimize recall bias and should aid in the design of future clinical trials focused on accurately capturing and minimizing acute morbidity following SBRT.

Background: For patients with brain metastases, systemic disease burden has historically been accepted as a major determinant of overall survival (OS). However, less research has focused on specific history and physical findings made by clinicians and how such findings pertain to patient outcomes at a given time point. The aim of this study is to determine how the initial clinical assessment of patients with brain metastases, as part of the history and physical at the time of consultation, correlates to patient prognosis., Methods: We evaluated a prospective, multi-institutional database of 1523 brain metastases in 507 patients who were treated with radiosurgery (Gamma Knife or CyberKnife) from 2001 to 2014. Relevant history of present illness (HPI) and past medical history (PMH) variables included comorbidities, Eastern Cooperative Oncology Group (ECOG) performance status, and seizure history. Physical exam findings included a sensory exam, motor exam, and cognitive function. Univariate and multivariate Cox regression analyses were used to identify predictors of OS., Results: Two hundred ninety-four patients were included in the final analysis with a median OS of 10.8 months (95% CI, 7.8-13.7 months). On univariate analysis, significant HPI predictors of OS included age, primary diagnosis, performance status, extracranial metastases, systemic disease status, and history of surgery. Significant predictors of OS from the PMH included cardiac, vascular, and infectious comorbidities. On a physical exam, findings consistent with cognitive deficits were predictive of worse OS. However, motor deficits or changes in vision were not predictive of worse OS. In the multivariate Cox regression analysis, predictors of worse OS were primary diagnosis (p = 0.002), ECOG performance status (OR 1.73, p < 0.001), and presence of extracranial metastases (OR 1.22, p = 0.009)., Conclusion: Neurological deficits and systemic comorbidities noted at presentation are not associated with worse overall prognosis for patients with brain metastases undergoing radiosurgery. When encountering new patients with brain metastases, the most informative patient-related characteristics that determine prognosis remain performance status, primary diagnosis, and extent of extracranial disease.

Background: Stereotactic body radiation therapy (SBRT) for stage I non-small cell lung cancer (NSCLC) is considered standard of care in the medically inoperable patient population. Multiple methods of SBRT delivery exist including fiducial-based tumor tracking, which allows for smaller treatment margins and avoidance of patient immobilization devices. We explore the long-term clinical outcomes of this novel fiducial-based SBRT method., Methods: In this single institutional retrospective review, we detail the outcomes of medically inoperable pathologically confirmed stage I NSCLC. Patients were treated with the Cyberknife SBRT system using a planning target volume (PTV) defined as a 5-mm expansion from gross tumor volume (GTV) without creation of an internal target volume (ITV). Dose was delivered in three or five equal fractions of 10 to 20 Gy. Pretreatment and posttreatment pulmonary function test (PFT) changes and evidence of late radiological rib fractures were analyzed for the majority of patients. Actuarial local control, locoregional control, distant control, and overall survival were calculated using the Kaplan-Meier method., Results: Sixty-one patients with a median age of 75 years were available for analysis. The majority (80 %) of patients were deemed to be medically inoperable due to underlying pulmonary dysfunction. Eleven patients (18 %) developed symptomatic pneumothoraces secondary to fiducial placement under CT guidance, which precipitously dropped to 0 % following transition to bronchoscopic fiducial placement. The 2-year rib fracture risk was 21.4 % with a median time to rib fracture of 2.9 years. PFTs averaged over all patients and parameters demonstrated small absolute declines, 5.7 % averaged PFT decline, at approximately 1 year of follow-up, but only the diffusing capacity of lung for carbon monoxide (DL CO ) demonstrated a statistically significant decline (10.29 vs. 9.01 mL/min/mmHg, p  = 0.01). Five-year local control, locoregional control, and overall survival were 87.6, 71.8, and 39.3 %, respectively., Conclusions: Despite reduced treatment margins and lack of patient immobilization, SBRT with fiducial-based tumor tracking achieves clinically comparable long-term outcomes to other linac-based SBRT approaches., Competing Interests: Compliance with ethical standards Funding source No funding support is associated with this study. Conflict of interests Brian T. Collins, MD, is a paid speaker for Accuray Inc. Jonathan W. Lischalk, MD; Stephanie M. Woo, BA; Shaan Kataria, MD; Nima Aghdam, MD; Ima Paydar, MD; Michael C. Repka, MD; and Eric D. Anderson, MD, have no competing interests. Ethical approval All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. The article does not contain any studies with human or animal subjects performed by any of the authors. The local Health Research Institutional Review Board (IRB) approved this retrospective analysis of an established departmental treatment approach. Malika Danner, MD contributed to the development and maintenance of the IRB documentation. Informed consent For this type of study, formal consent is not required.

Purpose: Magnetic resonance imaging (MRI)-directed stereotactic body radiation therapy (SBRT) has been established as a safe and effective treatment for prostate cancer. For patients with contraindications to MRI, CT-urethrogram is an alternative imaging approach to identify the location of the prostatic apex to guide treatment. This study sought to evaluate the safety of urethrogram-directed SBRT for prostate cancer., Methods: Between February 2009 and January 2014, 31 men with clinically localized prostate cancer were treated definitively with urethrogram-directed SBRT with or without supplemental intensity-modulated radiation therapy (IMRT) at Georgetown University Hospital. SBRT was delivered either as a primary treatment of 35-36.25 Gy in five fractions or as a boost of 19.5 Gy in three fractions followed by supplemental conventionally fractionated IMRT (45-50.4 Gy). Toxicities were recorded and scored using the Common Terminology Criteria for Adverse Events version 4.0 (CTCAE v.4.0)., Results: The median patient age was 70 years with a median prostate volume of 38 cc. The median follow-up was 3.7 years. The patients were elderly (Median age = 70), and comorbidities were common (Carlson comorbidity index ≥2 in 36%). Seventy-one percent of patients utilized alpha agonists prior to treatment, and 9.7% had prior procedures for benign prostatic hyperplasia. The 3-year actuarial incidence rates of ≥Grade 3 GU toxicity and ≥Grade 2 GI toxicity were 3.2 and 9.7%, respectively, and there were no Grade 4 or 5 toxicities., Conclusion: Magnetic resonance imaging is the preferred imaging modality to guide prostate SBRT treatment. However, urethrogram-directed SBRT is a safe alternative for the treatment of patients with prostate cancer who are unable to undergo MRI.

Objective: To evaluate recurrence patterns and overall survival in patients treated with adjuvant radiation after surgical staging for Stage II endometrial carcinoma. Secondary goals include identification of prognostic factors for recurrence and toxicity assessment., Methods/materials: The medical records of 41 patients treated with adjuvant radiotherapy at Washington University School of Medicine after surgical staging for endometrial cancer (total abdominal hysterectomy and bilateral salpingo-oophorectomy, peritoneal cytology, lymph node dissection) were reviewed. Nineteen were treated with a combination of external beam radiotherapy and vaginal brachytherapy (VB), and 22 patients were treated with postoperative VB alone. Median followup for all patients was 41 months., Results: Median patient age was 59 years (range, 42-87 years). All tumors were of endometrioid histology. There were 20 Grade 1 tumors, 13 Grade 2 tumors, and 8 Grade 3 tumors. For all patients, the 5-year overall survival was 69.8%, and the 5-year recurrence-free survival was 89.0%. There was no statistically significant difference in overall survival (p = 0.510) or freedom from vaginal (p = 0.840), distant (p = 0.133), or any recurrence (p = 0.275) with respect to modality of treatment (external beam radiotherapy and VB vs. VB alone). There were no pelvic lymph node recurrences. In the univariate analysis, there were no risk factors influencing overall survival or recurrences. One patient experienced a toxicity requiring hospital admission. She was treated with pelvic external beam radiation plus brachytherapy., Conclusions: VB alone results in excellent local control for patients with Stage II endometrial cancer after surgical staging. Long-term toxicities are rare and more common in the group of patients who were treated with pelvic external beam plus brachytherapy., (Copyright © 2015 American Brachytherapy Society. Published by Elsevier Inc. All rights reserved.)

A number of genes have been implicated in regeneration, but the regulation of these genes, particularly pertaining to regeneration in higher vertebrates, remains an interesting and mostly open question. We have studied microRNA (miRNA) regulation of regeneration and found that an intact miRNA pathway is essential for caudal fin regeneration in zebrafish. We also showed that miR-203 directly targets the Wnt signaling transcription factor Lef1 during this process. Repression of Lef1 by miR-203 blocks regeneration, whereas loss of miR-203 results in excess Lef1 levels and fin overgrowth. Expression of Lef1 from mRNAs lacking 3' UTR recognition elements can rescue the effects of excess miR-203, demonstrating that these effects are due to specific regulation of lef1 by miR-203. Our data support a model in which regulation of Lef1 protein levels by miR-203 is a key limiting step during regeneration.

Background: microRNAs (miRNAs) are small (~22 nt) non-coding RNAs that regulate cell movement, specification, and development. Expression of miRNAs is highly regulated, both spatially and temporally. Based on direct cloning, sequence conservation, and predicted secondary structures, a large number of miRNAs have been identified in higher eukaryotic genomes but whether these RNAs are simply a subset of a much larger number of noncoding RNA families is unknown. This is especially true in zebrafish where genome sequencing and annotation is not yet complete., Results: We analyzed the zebrafish genome to identify the number and location of proven and predicted miRNAs resulting in the identification of 35 new miRNAs. We then grouped all 415 zebrafish miRNAs into families based on seed sequence identity as a means to identify possible functional redundancy. Based on genomic location and expression analysis, we also identified those miRNAs that are likely to be encoded as part of polycistronic transcripts. Lastly, as a resource, we compiled existing zebrafish miRNA expression data and, where possible, listed all experimentally proven mRNA targets., Conclusion: Current analysis indicates the zebrafish genome encodes 415 miRNAs which can be grouped into 44 families. The largest of these families (the miR-430 family) contains 72 members largely clustered in two main locations along chromosome 4. Thus far, most zebrafish miRNAs exhibit tissue specific patterns of expression.