Somatic TP53 Mutations Are Detectable in Circulating Tumor DNA from Children with Anaplastic Wilms Tumors

Abstract

Background: Diffuse anaplastic Wilms tumor (DAWT) is a rare, high-risk subtype that is often missed on diagnostic needle biopsy. Somatic mutations in TP53 are associated with the development of anaplasia and with poorer survival, particularly in advanced-stage disease. Early identification of DAWT harboring TP53 abnormalities could improve risk stratification of initial therapy and monitoring for recurrence.

Methods: Droplet digital polymerase chain reaction (ddPCR) was used to evaluate 21 samples from 4 patients with DAWT. For each patient, we assessed TP53 status in frozen tumor, matched germline DNA, and circulating tumor DNA (ctDNA) from plasma, serum, and urine collected throughout treatment.

Results: Mutant TP53 was detectable in ctDNA from plasma and serum in all patients. We did not detect variant TP53 in the same volume (200 μl) of urine. One patient displayed heterogeneity of TP53 in the tumor despite both histological sections displaying anaplasia. Concentration of ctDNA from plasma/serum taken prenephrectomy varied significantly between patients, ranging from 0.44 (0.05-0.90) to 125.25 (109.75-140.25) copies/μl. We observed variation in ctDNA throughout treatment, and in all but one patient, ctDNA levels fell significantly following nephrectomy.

Conclusion: We demonstrate for the first time that ddPCR is an effective method for detection of mutant TP53 in ctDNA from children with DAWT even when there is intratumoral somatic heterogeneity. This should be further explored in a larger cohort of patients, as early detection of circulating variant TP53 may have significant clinical impact on future risk stratification and surveillance.

Figure 1

MAF from serum and plasma for each case varies during treatment. Time points as follows: 1, diagnosis; 2, mid-chemotherapy; 3, pre-operative; 4, post-operative; and 5, end of treatment. MAF is calculated as mutant allele concentration divided by total concentration. Error bars represent the range of MAFs produced within the confidence intervals of the mutant and wild-type allele concentrations. Urine ctDNA is not shown.

Supplementary Figure 1

Contamination from germline cell-free DNA in serum versus plasma for case 6 at three different time points throughout treatment. Time points as follows: Dx, diagnosis; MC, midchemotherapy; PsO, postoperative.

Supplementary Figure 2

Comparison of ctDNA yield extracted by Qiagen and Norgen from plasma for case 9. Time points as follows: Dx, diagnosis; MC, midchemotherapy; PrO, preoperative; PsO, postoperative.