背景：尽管转移性结直肠癌（mCRC）的分子分割取得了进展，但超越 RAS 状态的治疗作用仍然局限于 ERBB2 扩增、BRAF 突变和 MSI-H 患者的有限亚组。因此，有效治疗的优化是必要的。抗表皮生长因子受体单克隆抗体的再激发通常是经验性的，作为晚期治疗有一定的益处。我们以前发现在 EGFR 阻断期间出现在血液中的突变 RAS 和 EGFR 胞外克隆，在抗体退出后下降，从而恢复药物敏感性基于这一原理，我们设计了 CHRONOS，这是一个多中心的 II 期抗 EGFR 治疗试验，通过监测 RAS 的突变状态进行再激发，循环肿瘤DNA（CTDNA）中 bra 和 EGFR 的表达。据我们所知，这是第一次液体活检在 MCRC 中驱动抗 EGFR 再激发治疗的介入性临床试验。

Background: Despite advances in molecular segmentation of metastatic colorectal cancer (mCRC), beyond RAS status therapeutic actionability remains confined to the limited subgroups of ERBB2 amplified, BRAF mutated and MSI-H patients. Optimization of available treatments is therefore warranted. Rechallenge with anti-EGFR monoclonal antibodies is often empirically used with some benefit as late-line therapy. We previously found that mutant RAS and EGFR ectodomain clones, which emerge in blood during EGFR blockade, decline upon antibody withdrawal leading to regain drug sensitivity. Based on this rationale, we designed CHRONOS, a multicenter phase II trial of anti-EGFR therapy rechallenge guided by monitoring of the mutational status of RAS, BRAF and EGFR in circulating tumor DNA (ctDNA). To our knowledge, this is the first interventional clinical trial of liquid biopsy for driving anti-EGFR rechallenge therapy in mCRC.

方法：符合条件的患者为 PS ECOG 0-2 RAS/BRAF WT MCRC，首先达到客观反应，然后在任何含抗 EGFR 抗体的治疗方案中进展，显示 RAS，BRAF 和 EGFR 外域 WT 状态在进展到最后一个无 EGFR 方案后的分子筛选中增加 tDNA。通过 ddpcr 和下一代测序分析 CTDNA 的克隆进化。每两周静脉注射 6 毫克/公斤的帕尼托-穆玛，直至病情进展。主要终点是 RECIST 版本 1.1 的客观有效率（ORR），独立中心回顾共有 27 名患者和 6 名应答者被要求宣布研究阳性（幂=85%，I 型误差=0.05 ) .

Methods: Eligible patients were PS ECOG 0-2 RAS/BRAF WT mCRC having first achieved an objective response and then progression in any treatment line with an anti-EGFR antibody containing regimen, displaying RAS, BRAF and EGFR ectodomain WT status in ctDNA at molecular screening after progression to the last anti-EGFR-free regimen. Clonal evolution in ctDNA was analyzed by ddPCR and next generation sequencing. Panitumumab 6 mg/kg was administered IV every two weeks until progression. The primary endpoint was objective response rate (ORR) by RECIST version 1.1 with independent central review. 27 total patients and 6 responses were required to declare the study positive (power = 85%, type I error = 0.05).

结果：2019 年 8 月 19 日至 2020 年 11 月 6 日，52 例患者通过液体活检筛查，36 例（69%）中 CTDNA 的 RAS/BRAF/EGFR 突变阴性。其中，27 名患者在 4 个中心登记。中位年龄为 64 岁（42-80 岁）。PS ECOG 分别为 0/50%、1/46%、2/4%。既往 antEGFR 应用于一线的比例为 63%，二线的比例为 15%和大于 22%，既往治疗的中位数为 3。观察到 8/27 应答（2 例未确认）（ORR=30%，95%Cl:12-47%）11/27 稳定（40%，95%Cl:24-59%），8/11 持续>4个月。疾病控制率（PR+SD>4 个月）为 59%（95%Cl:41-78%）。中位无进展生存期为 16 周。中位反应持续时间为 1 周/周（我正在进行），最大毒性等级为 G3，仅限于 19%的皮肤病患者。

Results: Between Aug 19, 2019 and Nov 6, 2020 52 patients were screened by liquid biopsy and 36 (69%) were negative in ctDNA for RAS/BRAF/EGFR mutations. Of these, 27 patients were enrolled in 4 centers. Median age was 64 years (range: 42-80). PS ECOG was 0/50%, 1/46%, 2/4%. Previous anti-EGFR was administered in 1st line in 63%, 2nd in 15% and > 2nd in 22%. Median number of previous treatments was 3. The primary endpoint was met, with 8/27 partial responses (PR) observed (2 unconfirmed) (ORR = 30%, 95% CI: 12-47%). Stable disease (SD) was obtained in 11/27 (40%, 95% CI: 24-59%), lasting > 4 months in 8/11. Disease control rate (PR plus SD > 4 months) was therefore obtained in 16/27 (59%, 95% CI: 41-78%). Median progression-free survival was 16 weeks. Median duration of response was 17 weeks (1 ongoing). Maximal grade toxicity was G3, limited to dermatological and occurring in 19% of patients. ctDNA dynamics were studied in all patients.

结论：液体活检驱动抗 EGFR 抗体再激发可使三分之一的患者产生更客观的反应。对血液中肿瘤 DNA 进行基因分型以指导治疗可有效地应用于晚期 CRCS 的治疗。

Conclusions: Liquid biopsy-driven rechallenge with anti-EGFR antibodies leads to further objective responses in one third of patients. Genotyping tumor DNA in the blood to direct therapy can be effectively incorporated in the management of advanced CRCs.