Dados do Trabalho

Título

Detection of circulating tumor DNA (ctDNA) in plasma and urine of renal cell carcinoma patients for prognostic stratification

Introdução

Renal cell carcinoma (RCC) is the most common renal tumor in the adult population. Clear cell renal cell carcinoma (ccRCC) is the most prevalent subtype, accounting for about 75% of cases. Even though CCR prognosis is excellent when diagnosed at an early stage, the 5-year survival rate is 10% in metastatic cases. Thus, tools for early identification of high-risk patients are important for early therapeutic approach aiming greater treatment success. CEP 2397/17

Objetivo

1) To characterize the somatic mutations of RCC and use them as tumor marker for tracking ctDNA in plasma and urine of RCC patients;
2) To test the role of ctDNA in identifying patients with high-risk for disease progression.

Métodos

RCC patients were recruited at A.C.Camargo Cancer Center (2017-2019) from the Oncourology Department. Baseline urine and plasma samples were taken pre-surgery. For ccRCC patients, additional samples were collected at 6-8 weeks (M1), 6 months (M2), 18 months (M3), and 30 months (M4) post-surgery. Tumor tissue and leukocyte DNAs were assessed using NGS on the Illumina NextSeq platform to identify somatic mutations, using a custom panel with 28 RCC-related genes (RCC-28 panel). Cases without somatic mutation in genes of this panel were analyzed on the ION GeneStudio S5 Plus (S5) platform using the Comprehensive Cancer Panel (CCP). Tumor somatic mutations were tracked in cell free DNA (cfDNA) from urine and plasma by deep amplicon sequencing using the Ion S5 platform. The Fisher exact test was employed for statistical analysis.

Resultados

79 RCC patients participated in the study. 50 (63.3%) presented somatic mutations in genes of the RCC-28 panel, involving 76.1% of the ccRCC tumors. 23 of the remaining 29 RCC tumor samples were tested using the CCP panel. 4 were positive for somatic mutations. Overall, VHL was the most mutated gene (32.9%, 55.8% in ccRCC), followed by PBRM1 (17.7%, 30.2% in ccRCC).
For patients with identified somatic variants, ctDNA was positive in 13.04% of baseline plasma and 12.5% of urine samples. Max variant allelic frequency (VAF) was 10.7% in plasma and 11.6% in urine.
During monitoring of ccRCC patients, at 6 to 8 weeks after surgery (M1), 10% had ctDNA-positive plasma while all urine samples were negative. At 6 months (M2), all plasma and urine were ctDNA-negative. By 18 months (M3), 11.8% of plasma and 7.1% of urine samples were ctDNA-positive. At 30 months (M4), all plasma and urine samples were ctDNA-negative.
ctDNA-positive baseline plasma was more frequent in patients with disease progression (42.9%) than overall ccRCC patients (12.5%) (p=.0155) and patients positive for ctDNA in baseline plasma had a mean tumor size 4.2cm larger than ctDNA-negative patients (p=.027). Progression-free survival was reduced in patients that were positive for ctDNA in baseline plasma (p=.004) and overall survival was lower in cases with positive ctDNA in plasma samples collected in the 1st year after surgery (samples M1 or M2) (p=<.001).

Conclusões

The custom RCC-28 Panel shows a good capacity for identifying somatic variants in RCC tumors, specially the ccRCC subtype. RCC is a tumor that releases low amount of tumor DNA in urine and plasma. The presence of ctDNA in plasma before surgery seems to be informative for identifying patients at a higher risk of disease progression.

Palavras-chave

circulating tumor DNA, liquid biopsy, renal cell carcinoma

Financiador do resumo

PRONON - Ministério da Saúde