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Sequence combination for detecting tumor mutational burden, and design method thereof

A technology of mutation load and design method, applied in genomics, proteomics, instruments, etc., to achieve high throughput, cost reduction, and wide clinical application

Active Publication Date: 2019-03-05
BEIJING GENEPLUS TECH +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, the calculation method and threshold of TMB have not formed a guiding specification. In this field, a sequence combination for detecting TMB and a method for calculating TMB are needed to predict the efficacy of immune checkpoint inhibitors

Method used

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  • Sequence combination for detecting tumor mutational burden, and design method thereof
  • Sequence combination for detecting tumor mutational burden, and design method thereof
  • Sequence combination for detecting tumor mutational burden, and design method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0092] Example 1 In this example, 18 cases of simultaneous detection of the whole exome chip and tissue samples of the chip of the present invention are taken as an example to illustrate the performance of the present invention for detecting TMB.

[0093] figure 1 Shows the sample detection implementation process of the present invention, including sample DNA extraction, library building, target area capture using liquid-phase probes, high-throughput sequencing, biological information analysis, identification of true somatic and germline mutations, and TMB calculation .

[0094] 1. DNA extraction: the scope of application of the sample includes surgically excised fresh pathological tissues, formaldehyde-fixed paraffin-embedded case tissues and paraffin sections; this example uses the sequencing results of blood cell gDNA as a control to exclude germline mutations. DNA extraction steps: samples such as blood cells and tissues are extracted according to the instructions of QIAamp DNA...

Embodiment 2

[0134] Example 2 Detection of tTMB for non-small cell lung cancer.

[0135] In this example, according to the TMB calculation method described in Example 1, retrospective statistics were performed using the same method as in Example 1 to detect the data of 1353 cases of non-small cell lung cancer.

[0136] Results: The tTMB distribution of non-small cell lung cancer was obtained as image 3 . According to literature reports, currently in clinical trials related to non-small cell lung cancer, the upper quartile or upper tertile of the total sample is used as the threshold to distinguish TMB-H or TMB-L. Therefore, in this embodiment, for non-small cell lung cancer, TMB≥9 mutation / Mb is defined as TMB-H, and TMB<9 mutation / Mb is defined as TMB-L.

Embodiment 3

[0137] Example 3 Determination of bTMB threshold for non-small cell lung cancer.

[0138] In this example, 100 samples of newly diagnosed non-small cell lung cancer (NSCLC) were used to assess the consistency of tissue plasma TMB, and 100 samples were tested for both untreated tissue samples and plasma samples.

[0139] Refer to Example 1 for the experimental analysis process of tissue samples in this embodiment.

[0140] The experimental analysis process of the plasma sample in this embodiment is as follows, using blood cell gDNA as the sample control.

[0141] 1. DNA extraction:

[0142] For whole blood, plasma / blood cell separation is required first: Collect 10 mL of peripheral blood, and perform plasma / blood cell separation in time (EDTA anticoagulation tube, within 4 hours; Streck tube within 72 hours). The separation steps are as follows:

[0143] Step 1: Centrifuge at 1600g for 10 minutes at 4°C. After centrifugation, divide the upper plasma into multiple 1.5mL or 2.0mL centrifuge...

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Abstract

The invention discloses a sequence combination for detecting tumor mutational burden, and a design method thereof. The method comprises the steps that (a) target areas requiring design probes are determined, and a database of tumor mutation data containing large sample determination; (b) in terms of each target area, sliding is conducted with a length of one window to obtain a plurality of windows, and each window is scored; (c) in terms of each target area, the window with the highest score is the target probe area. The invention also discloses a method of calculating TMB of organs and bloodplasma samples, and a quality control method of calculating TMB of organs and blood plasma samples.

Description

Technical field [0001] The present invention belongs to the field of biotechnology. More specifically, the present invention relates to sequence combinations and methods for detecting tumor mutation burden. Background technique [0002] In recent years, programmed death inhibitor-1 (PD-1) protein or its ligand (PD-L1) immune checkpoint inhibitors have created many miracles, among which many patients have been rescued from endangered conditions and have long-term survive. However, for patients without biomarker selection, the effective rate of PD-1 / PD-L1 immune checkpoint inhibitors is only -20%. Therefore, the efficacy predictor of this type of inhibitor has become a research hotspot. [0003] PD-L1 expression is a predictive indicator that has been included in the NCCN guidelines. According to clinical data, it can be seen that as the expression level of PD-L1 increases, the response rate, progression-free survival and median survival of patients to pembrolizumab are also signi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G16B20/00G16B25/00
Inventor 易玉婷管彦芳田超易鑫杨玲
Owner BEIJING GENEPLUS TECH