Supplementary Material for: ctDNA in Neuroendocrine Carcinoma of Gastroenteropancreatic Origin or of Unknown Primary: The CIRCAN-NEC Pilot Study

<b><i>Introduction:</i></b> Gastroenteropancreatic neuroendocrine carcinomas (GEPNEC) are characterized by a heterogeneous molecular profile and a poor prognosis. Circulating tumour DNA (ctDNA) analysis may be useful for NEC management. This study aimed at describing ctDNA mutations, to assess their predictive value for response to chemotherapies, and their change according to disease progression. <b><i>Methods:</i></b> The CIRCAN-NEC study included patients with GEPNEC or NEC from an unknown primary, scheduled to begin first- or second-line chemotherapy. Blood samples were collected prior to chemotherapy initiation, at first evaluation, and during disease progression. ctDNA was sequenced by next-generation sequencing (NGS). Molecular response was defined as a decrease of at least 30% of the mutant allele fraction. <b><i>Results:</i></b> All 24 patients included received platinum-etoposide first-line chemotherapy; 19 received a FOLFIRI-based post-first-line regimen. Twenty-two patients had at least one driver mutation: <i>TP53</i> (<i>n</i> = 21), <i>RB1</i> (<i>n</i> = 2), <i>KRAS</i> (<i>n</i> = 4), and <i>BRAF</i> (<i>n</i> = 3). Ten (42%) had an “adenocarcinoma-like” profile. Five of 6 patients with matching ctDNA/tissue NGS harboured at least one concordant mutation (44% concordance at the gene level). The concordance rate between ctDNA mutation/immunohistochemistry profile was 64% (7/11) for <i>TP53</i>/p53+ and 14% (1/7) for <i>RB1</i>/pRb−. In this pilot study including few patients by subgroups, patients with <i>KRAS</i> (HR = 3.60, 95% CI [1.06–12.04]) and <i>BRAF</i> (HR = 4.25, 95% CI [1.11–16.40]) mutations had shorter progression-free survival (PFS) under platinum-etoposide, while the 2 patients with <i>RB1</i> mutations had shorter PFS under FOLFIRI-based chemotherapy. Twenty-eight periods of treatment were assessed: 10 patients had a molecular response (7/10 had a morphological response), which was associated with longer PFS (HR = 0.37, 95% CI [0.15; 0.91]). <b><i>Conclusion:</i></b> This pilot study shows a high sensitivity of ctDNA assessment, which is encouraging for the future management of GEPNEC (tumour molecular diagnosis and evaluation of disease progression).

<b><i>引言：</i></b> 胃肠胰神经内分泌癌（Gastroenteropancreatic neuroendocrine carcinomas, GEPNEC）具有异质性分子特征且预后较差。循环肿瘤DNA（circulating tumour DNA, ctDNA）检测或可助力神经内分泌癌的临床管理。本研究旨在描述ctDNA的突变特征，评估其对化疗应答的预测价值，并分析其随疾病进展的动态变化。<b><i>方法：</i></b> 本CIRCAN-NEC研究纳入了拟接受一线或二线化疗的胃肠胰神经内分泌癌或原发灶不明神经内分泌癌患者。分别于化疗开始前、首次疗效评估时及疾病进展阶段采集血液样本。采用下一代测序（next-generation sequencing, NGS）技术对ctDNA进行测序。分子应答定义为突变等位基因分数至少降低30%。<b><i>结果：</i></b> 纳入的24例患者均接受了铂类依托泊苷一线化疗；其中19例后续接受了基于FOLFIRI的二线治疗方案。22例患者检出至少1种驱动突变，包括<i>TP53</i>（<i>n</i>=21）、<i>RB1</i>（<i>n</i>=2）、<i>KRAS</i>（<i>n</i>=4）及<i>BRAF</i>（<i>n</i>=3）。10例（42%）患者呈现“腺癌样”分子特征。6例匹配ctDNA与组织样本NGS检测的患者中，5例检出至少1种一致性突变，基因水平一致性为44%。ctDNA突变与免疫组化特征的一致性方面，<i>TP53</i>/p53阳性的一致性率为64%（7/11），<i>RB1</i>/pRb阴性的一致性率为14%（1/7）。在本亚组患者数量有限的先导研究中，携带<i>KRAS</i>（风险比=3.60，95%置信区间[1.06–12.04]）与<i>BRAF</i>（风险比=4.25，95%置信区间[1.11–16.40]）突变的患者，接受铂类依托泊苷治疗后的无进展生存期（progression-free survival, PFS）更短；而2例携带<i>RB1</i>突变的患者，接受基于FOLFIRI的化疗后的PFS更短。本研究共评估了28个治疗周期：10例患者出现分子应答，其中7/10同时出现形态学应答，分子应答与更长的PFS相关（风险比=0.37，95%置信区间[0.15; 0.91]）。<b><i>结论：</i></b> 本先导研究证实ctDNA检测具有较高的灵敏度，这为胃肠胰神经内分泌癌的未来临床管理（包括肿瘤分子诊断与疾病进展评估）提供了鼓舞人心的依据。