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Targeted chromatin ligation, a robust epigenetic profiling technique for small cell numbers.

ABSTRACT: The complexity and inefficiency of chromatin immunoprecipitation strategies restrict their sensitivity and application when examining rare cell populations. We developed a new technique that replaces immunoprecipitation with a simplified chromatin fragmentation and proximity ligation step that eliminates bead purification and washing steps. We present a simple single tube proximity ligation technique, targeted chromatin ligation, that captures histone modification patterns with only 200 cells. Our technique eliminates loss of material and sensitivity due to multiple inefficient steps, while simplifying the workflow to enhance sensitivity and create the potential for novel applications.

SUBMITTER: Zarnegar MA

PROVIDER: S-EPMC5622369 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Targeted chromatin ligation, a robust epigenetic profiling technique for small cell numbers.

Zarnegar Mark A MA Reinitz Felicia F Newman Aaron M AM Clarke Michael F MF

Nucleic acids research 20170901 17

The complexity and inefficiency of chromatin immunoprecipitation strategies restrict their sensitivity and application when examining rare cell populations. We developed a new technique that replaces immunoprecipitation with a simplified chromatin fragmentation and proximity ligation step that eliminates bead purification and washing steps. We present a simple single tube proximity ligation technique, targeted chromatin ligation, that captures histone modification patterns with only 200 cells. O ...[more]

PMID: 28973448

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Project description:PurposeTheoretical studies have shown that dose-painting-by-numbers (DPBN) could lead to large gains in tumor control probability (TCP) compared to conventional dose distributions. However, these gains may vary considerably among patients due to (a) variations in the overall radiosensitivity of the tumor, (b) variations in the 3D distribution of intra-tumor radiosensitivity within the tumor in combination with patient anatomy, (c) uncertainties of the 3D radiosensitivity maps, (d) geometrical uncertainties, and (e) temporal changes in radiosensitivity. The goal of this study was to investigate how much of the theoretical gains of DPBN remain when accounting for these factors. DPBN was compared to both a homogeneous reference dose distribution and to nonselective dose escalation (NSDE), that uses the same dose constraints as DPBN, but does not require 3D radiosensitivity maps.MethodsA fully automated DPBN treatment planning strategy was developed and implemented in our in-house developed treatment planning system (TPS) that is robust to uncertainties in radiosensitivity and patient positioning. The method optimized the expected TCP based on 3D maps of intra-tumor radiosensitivity, while accounting for normal tissue constraints, uncertainties in radiosensitivity, and setup uncertainties. Based on FDG-PETCT scans of 12 non-small cell lung cancer (NSCLC) patients, data of 324 virtual patients were created synthetically with large variations in the aforementioned parameters. DPBN was compared to both a uniform dose distribution of 60 Gy, and NSDE. In total, 360 DPBN and 24 NSDE treatment plans were optimized.ResultsThe average gain in TCP over all patients and radiosensitivity maps of DPBN was 0.54 ± 0.20 (range 0-0.97) compared to the 60 Gy uniform reference dose distribution, but only 0.03 ± 0.03 (range 0-0.22) compared to NSDE. The gains varied per patient depending on the radiosensitivity of the entire tumor and the 3D radiosensitivity maps. Uncertainty in radiosensitivity led to a considerable loss in TCP gain, which could be recovered almost completely by accounting for the uncertainty directly in the optimization.ConclusionsOur results suggest that the gains of DPBN can be considerable compared to a 60 Gy uniform reference dose distribution, but small compared to NSDE for most patients. Using the robust DPBN treatment planning system developed in this work, the optimal DPBN treatment plan could be derived for any patient for whom 3D intra-tumor radiosensitivity maps are known, and can be used to select patients that might benefit from DPBN. NSDE could be an effective strategy to increase TCP without requiring biological information of the tumor.

Dataset Information

Targeted chromatin ligation, a robust epigenetic profiling technique for small cell numbers.

Publications

Targeted chromatin ligation, a robust epigenetic profiling technique for small cell numbers.

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