Background: Allogeneic stem cell transplant (allo-SCT) is a mainstay of treatment for acute myeloid leukemia (AML). Its success depends largely on response of donor T lymphocytes against leukemia cells, known as graft-vs-leukemia (GvL) effect. A key potential driver of GvL is immune response to mutation-derived neoantigens. Previous studies in solid tumors have demonstrated enhanced immunogenicity of frameshift (FS)-derived peptides vs. those from non- synonymous single nucleotide variants (SNVs). We therefore hypothesized that AML cases bearing FS mutations in leukemia-associated genes would be more immunogenic than those with only other types of mutations (non-FS), and thus benefit more from allo-SCT via more robust GvL. Methods: We identified AML patients who had undergone allo-SCT between 2010 and 2022 and had next-generation sequencing data available on diagnostic specimens using a 42-gene hot spot panel. We compared the impact of tumor mutations present at diagnosis on overall survival and relapse-free survival based on FS versus non-FS status. Results: Ninety-five AML allo-SCT patients were identified. We observed superior relapse-free survival (P = 0.038, hazard ratio (HR): 0.24) and borderline superior overall survival (P = 0.058, HR: 0.55) post-transplant in de novo AML patients, who had at least one FS mutation (other than NPM1) ) in one of the 42 assessed genes versus those with only non-FS mutations. Conclusions: Our findings suggest that FS-mutated AML cases may benefit more from allo-SCT than those with only non-FS mutations, possibly due to increased generation of immunogenic neoepitopes. If validated in an expanded study, incorporation of somatic FS mutation status in AML could improve patient selection algorithms for bone marrow transplant and thereby lead to superior outcomes.

Supplementary Figures 1-3 from Evidence for the Direct Binding of Phosphorylated p53 to Sites of DNA Breaks In vivo

Supplementary Figure Legends from Evidence for the Direct Binding of Phosphorylated p53 to Sites of DNA Breaks In vivo

Response profile of subcutaneous PCa xenografts to RT

Abstract The proliferative activity of undifferentiated cells in intestinal crypts is essential for maintaining intestinal homeostasis and facilitating regeneration in response to both pathophysiological conditions and stress. DAB2IP (Disabled homolog 2 interacting protein), a tumor suppressor, plays a crucial role in regulating various oncogenic pathways. Our previous study unveiled that DAB2IP serves as a cell cycle regulator by facilitating PLK1-mediated mitosis progression. In this study, we uncovered a novel DAB2IP function as it mediates the maintenance and regeneration of intestinal epithelial homeostasis in normal conditions and following irradiation (IR)-induced gastrointestinal (GI) injury. Mechanistically, we identified that DAB2IP interacts with the histone acetyltransferase HBO1 and promotes PLK1-mediated phosphorylation of HBO1. This phosphorylation event leads to HBO-directed lysine 14 acetylation on histone 3 (H3K14Ac), the loading of the minichromosome maintenance protein (MCM) complex onto chromatin, progression of DNA replication, and maintenance of genome integrity. Furthermore, we found that ATR regulates Cdk1-mediated phosphorylation of DAB2IP and that phosphorylation of DAB2IP is crucial for the formation and activation of the HBO1-PLK1 complex. Finally, ablating DAB2IP phosphorylation results in increased genomic instability due to incomplete replication of genomic DNA, as shown via the accumulation of anaphase ultrafine bridges and 53BP1 nuclear bodies in the G1 cell cycle phase. Collectively, our findings highlight the regulatory role of DAP2IP in DNA replication-an essential process for maintaining genomic stability and intestinal homeostasis.

Supplementary Figures S1-S2 from Somatic Mutations in the Tyrosine Kinase Domain of Epidermal Growth Factor Receptor (EGFR) Abrogate EGFR-Mediated Radioprotection in Non–Small Cell Lung Carcinoma

ARV mediates DDR and clonogenic survival.

PDF file - 544K, Proximity ligation assay (PLA): Effect of DNA-Pkcs and ATM inhibition on EGFR-DNA-PKcs binding.NSCLCs or HBEC-3KT cells expressing wild type or gammaE746-E750 forms of EGFR following a 2 h pretreatment with vehicle, 10 muM NU7441 or 10 muM KU55933 were mock irradiated or exposed to 4 Gy IR and fixed at 60 minutes. Representative images of nuclei (blue) showing EGFR-DNA-PKcs complexes (intense red dots). Cropped images shown in Figure 4C

PDF file - 774K, Proximity ligation assay (PLA): To detect radiation-induced PP2A-DNA-PKcs binding. NSCLCs and HBEC cells expressing WT or gammaE746-E750 EGFR fixed 1 h following 0 Gy or 4 Gy IR. Left panel: representative images of nuclei (blue) and EGFR-DNA-PKcs complexes (red: PLA fluorescence). Square boxes represent cropped images shown in Figure 7C

Small cell lung cancers and squamous cell cancers can be epigenetically subtyped (Kong R et al 2022, Sato T et al, 2019) with focus on super-enhancer (SE) signals within or adjacent to transcription factors (TF). We reasoned that specific epigenomic subsets in lung adenocarcinomas (LUAD) operate with similar cellular programming showing TF dependency for lineage commitment and survival. We hypothesize that these dependencies on lineage programming are associated with a specific epigenomic landscape. Therefore, we investigated enrichment of SEs across the spectrum of LUADs to identify potential regulators for lineage commitment and survival. We obtained data for genome-wide H3K27ac enrichment by chromatin immunoprecipitation followed by sequencing (ChIP-seq) on a total of 53 LUAD cell lines. We performed unsupervised hierarchical clustering of signals on SEs at gene loci annotated with transcriptional regulation identified in two or more cell lines. To Uncover 3D contact information, we employed HiChIP, using antibody against SMC1, a cohesin complex, on NCI-H460. Three LUAD cell lines with distinct enhancer profiles were transfected with five enhancer segments of the SE at the PBX1 gene cloned into a Firefly luciferase plasmid. Lastly, four LUAD cell lines were treated with a PBX1 inhibitor, T417 for 72 hours in varying concentrations ranging from 20uM to 10nM and evaluated for sensitivity to the compound. Clustering analysis identified LUADs with differential enhancer enrichment for TFs. Among those, we focused on the PBX1 gene locus in LUADs harboring SEs proximal to its promoter. SMC1 HiChIP analysis on NCI-H460 suggest all enhancer segments are within a local topologically-associated domain with the PBX1 promoter. Luciferase reporter assay demonstrate activity of enhancer segments within the PBX1 SE, unique to each cell line (NCI-H460, PC-9 and NCI-H1437), with enhancers e4 and e5 rendered the highest level of activity while e2 being the most active in PC-9. The effects of these enhancer segments are congruent to H3K27ac enrichment patterns. Treatment with T417 showed reduction in cell proliferation at an IC50 of 2.5 uM and 0.97 uM for NCI-H460 and PC-9, respectively, whereas T417 did not affect cell viability in NCI-H1792 and NCI-H125. Data indicate a distinct subset of LUADs, enriched for the enhancer segments within the SE at the PBX1 gene locus, at which the HiChIP data displayed the enhancer segments lie within the same topologically-associated domain. The enhancer activity of enhancer segments correlated with H3K27ac enrichment pattern for each cell line, suggesting enrichment correlates with impact. Data with PBX1 T417 exposure suggests dependency of PBX1 in maintaining lineage state and survival for unique subsets of LUADs. Further investigation is needed to determine if T417 dependent inhibition is uniform to all PBX1 active LUADs and if other LUAD subsets are dependent on other TFs. Citation Format: Douglas M. Mansell, Maya Fridrich, Feng Jiang, Benjamin Chen, Ayushi Patel, Tian Li-Wang, Charles A. Powell, Hideo Watanabe. Epigenomic profiling uncovers distinct enrichment patterns of PBX1 super-enhancer among lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2231.

PDF file - 869K, Effect of DNA-PKcs or ATM inhibition on p-T2609 in NSCLCs. h pretreatment with 10 muM NU7441 or 10 muM KU55933. Square boxes represent cropped images shown in Figure 6C

Ataxia-telangiectasia mutated (ATM) drives the DNA damage response via modulation of multiple signal transduction and DNA repair pathways. Previously, ATM activity was implicated in promoting the non-homologous end joining (NHEJ) pathway to repair a subset of DNA double-stranded breaks (DSBs), but how ATM performs this function is still unclear. In this study, we identified that ATM phosphorylates the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), a core NHEJ factor, at its extreme C-terminus at threonine 4102 (T4102) in response to DSBs. Ablating phosphorylation at T4102 attenuates DNA-PKcs kinase activity and this destabilizes the interaction between DNA-PKcs and the Ku-DNA complex, resulting in decreased assembly and stabilization of the NHEJ machinery at DSBs. Phosphorylation at T4102 promotes NHEJ, radioresistance, and increases genomic stability following DSB induction. Collectively, these findings establish a key role for ATM in NHEJ-dependent repair of DSBs through positive regulation of DNA-PKcs.

Supplementary Figure 4 from Suppression of Nonhomologous End Joining Repair by Overexpression of HMGA2

PDF file - 729K, Proximity ligation assay (PLA): Effect of T2609A and S2056A on EGFR-DNA-PKcs binding. V3-WT DNA-PKCS, V3-S2056A and V3-T2609A CHO cells expressing wild type EGFR which were either mock-irradiated or exposed to 4 Gy IR. Left panel: representative images of nuclei (blue) and EGFR-DNA-PKcs complexes (red PLA fluorescence dots) after processing for PLA. Square boxes represent cropped images shown in Figure 5C.

The COVID-19 global pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has infected hundreds of millions of individuals. Following COVID-19 infection, a subset can develop a wide range of chronic symptoms affecting diverse organ systems referred to as post-acute sequelae of SARS-CoV-2 infection (PASC), also known as long COVID. A National Institutes of Health-sponsored initiative, RECOVER: Researching COVID to Enhance Recovery, has sought to understand the basis of long COVID in a large cohort. Given the range of symptoms that occur in long COVID, the mechanisms that may underlie these diverse symptoms may also be diverse. In this review, we focus on the emerging literature supporting the role(s) that viral persistence or reactivation of viruses may play in PASC. Persistence of SARS-CoV-2 RNA or antigens is reported in some organs, yet the mechanism by which they do so and how they may be associated with pathogenic immune responses is unclear. Understanding the mechanisms of persistence of RNA, antigen or other reactivated viruses and how they may relate to specific inflammatory responses that drive symptoms of PASC may provide a rationale for treatment.

Figure S1. DAB2IP expression does not affect key repair factors in NHEJ (Ku70, Ku80) and HR (RAD51, BRCA1) and PARP-1 mRNA expression. Figure S2. RANBP2, TRIP12, and RNF40 interact with both DAB2IP and PARP-1. Figure S3. Radiotherapy of RCC xenograft models.

ARVs are recruited to DNA damage sites.

PDF file - 210K, API inhibits ionizing radiation-induced Akt phosphorylation/activation