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Methods for detection of donor-derived cell-free DNA

A source and cell technology, applied in the direction of biochemical equipment and methods, microbial measurement/testing, etc., can solve the problems of invasive costs, transplant damage, lack of specificity, etc.

Pending Publication Date: 2021-05-04
NATERA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Previous attempts to use serum creatinine to determine kidney transplant status lacked specificity, and biopsy transplants were invasive and expensive and could lead to delayed diagnosis of graft injury and / or rejection

Method used

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  • Methods for detection of donor-derived cell-free DNA
  • Methods for detection of donor-derived cell-free DNA
  • Methods for detection of donor-derived cell-free DNA

Examples

Experimental program
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Effect test

example 1

[0769] Blood samples from kidney transplant recipients were analyzed retrospectively (292 plasma samples from 187 unique patients, of which 8 samples were excluded) after assessment of the patients' graft status by biopsy. Biopsies were graded by the Banff classification of T-cell and antibody-mediated acute rejection (AR) or non-AR (borderline, stable, or other injury). The samples analyzed for biopsy included 52 acute rejection (AR) samples and 240 non-acute rejection (non-AR) samples, including those in borderline rejection, with other injuries or stable.

[0770] Circulating cell-free DNA was extracted from 2 mL of plasma per sample by Qiagen cfDNA kit. The amount of cfDNA was then quantified using a LapChip. Library preparation was accomplished using the standard protocol using the Natera Panorama Library Prep Kit, except that the library was amplified by 18 cycles of PCR (as opposed to the standard 9 cycles). The amplified library was then purified using Ampure beads (...

example 2

[0782] Example 2. Optimization of detection of kidney transplant injury by assessment of donor-derived cell-free DNA via large-scale multiplex PCR.

[0783] introduce

[0784] Precision medicine and personalized customization of immunosuppressive drug regimens could improve the current state of organ transplant management. Invasive biopsies should be avoided where possible considering patient experience, and graft damage is usually detected late. Although advances in immunosuppressive drugs, organ harvesting methods, and human leukocyte antigen typing have reduced the number of clinically and biopsy-proven acute rejection events, subclinical acute rejection of kidney transplantation remains a significant risk. Kidney transplant management is particularly challenging due to the redundancy of serum creatinine measurements, which makes immunosuppressant dosing and adjustment far from individualized, in addition to late detection of graft injury. Therefore, rapid and non-invasiv...

example 3

[0847] Example 3. Evaluation of donor-derived cell-free DNA by large-scale multiplex PCR and next-generation sequencing to validate the detection of kidney transplant injury.

[0848] introduce

[0849] Kidney transplantation is the best option for patients with end-stage renal disease. According to the United Network for OrganSharing, more than 19,000 kidneys were transplanted in the United States in 2016 (cen.acs.org), and approximately 200,000 patients survived a functional kidney transplant (NIH Medline plus). Although life-long immunosuppressive maintenance regimens are designed to optimize treatment outcomes, approximately 20-30% of patients experience overall kidney transplant failure within the first 5 years, and only 55% of transplanted kidneys survive to 10 years (cen.acs.org ). Therefore, early intervention strategies are urgently needed to avoid or minimize acute / subclinical rejection episodes, nephrotoxicity, and enable management and monitoring of comorbidities...

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Abstract

The present disclosure provides methods for determining the status of an allograft within a transplant recipient from genotypic data measured from a mixed sample of DNA comprising DNA from both the transplant recipient and from the donor. The mixed sample of DNA may be preferentially enriched at a plurality of polymorphic loci in a way that minimizes the allelic bias, for example using massively multiplexed targeted PCR.

Description

[0001] Cross References to Related Applications [0002] This application claims U.S. Provisional Application No. 62 / 693,833, filed July 3, 2018; U.S. Provisional Application No. 62 / 715,178, filed August 6, 2018; U.S. Provisional Application No. 62, filed December 19, 2018 / 781,882; and priority of U.S. Provisional Application No. 62 / 834,315, filed April 15, 2019. Each of these applications cited above is hereby incorporated by reference in its entirety. technical field [0003] The present disclosure generally relates to methods for detecting donor-derived DNA in transplant recipients. Background technique [0004] There are currently approximately 190,000 living kidney recipients in the United States, and approximately 20,000 kidney transplants occur each year. Rapid detection of renal allograft injury and / or rejection remains a challenge. Previous attempts to use serum creatinine to determine renal transplant status lacked specificity, and biopsy transplants were invas...

Claims

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Application Information

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IPC IPC(8): C12Q1/686C12Q1/6883
CPCC12Q1/6883C12Q1/686C12Q2535/122C12Q2537/143C12Q2545/101C12Q2565/514C12Q2600/156
Inventor 所罗门·莫什克维奇伯恩哈德·齐默尔曼图多尔·蓬皮柳·康斯坦丁侯赛因·埃塞·基尔基兹拉尔阿利森·赖恩斯蒂米尔·西于尔永松费利佩·阿科斯塔·阿奇拉赖恩·斯韦纳顿
Owner NATERA
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