Early Circulating Tumour DNA Variations Predict Tumour Response in Melanoma Patients Treated with Immunotherapy.

Antibodies targeting immune checkpoints were recently approved for metastatic melanoma. However, not all patients will respond to the treatment and some will experience grade III-IV immune-related adverse events. Therefore, early identification of non-responder patients would greatly aid clinical practice. Detection of circulating tumour DNA (ctDNA) is a non-invasive approach to monitor tumour response. Digital droplet PCR was used to quantify BRAF and NRAS mutations in the plasma of patients with metastatic melanoma treated with immunotherapy. In 16 patients, ctDNA variations mirrored tumour response (p = 0.034) and ctDNA augmentation during follow-up detected tumour progression with 100% specificity. In 13 patients, early ctDNA variation was associated with clinician decision at first evaluation (p = 0.0046), and early ctDNA increase with shorter progression-free survival (median 21 vs. 145 days; p = 0.001). Monitoring ctDNA variations early during immunotherapy may help clinicians rapidly to discriminate non-responder patients, allow early adaptation of therapeutic strategies, and reduce exposure to ineffective, expensive treatment.

Monitoring of KRAS-mutated ctDNA to discriminate pseudo-progression from true progression during anti-PD-1 treatment of lung adenocarcinoma.

OBJECTIVES: Pseudo-progression is a rare but worrying situation for both clinicians and patients during immunotherapy. Dedicated ir-RECIST criteria have been established to improve this situation. However, this can be sometimes considered inadequate and patients experiencing true progression may then receive inefficient treatments. Additional reliable tools to discriminate pseudo from true progression are thus needed. So far, no biomarker has been identified to distinguish pseudo from true progression. We hypothesize that biomarkers associated with the molecular characteristics of the tumor may be of interest. To avoid a tumor re-biopsy, circulating markers appear to be a less invasive and reproducible procedure. As ctDNA kinetics correlate with the response to treatment in KRAS-mutated adenocarcinoma, we anticipated that this analysis could be of interest. MATERIALS AND METHODS: We monitored the level of KRAS-mutated ctDNA by digital droplet PCR in serial plasma samples from two patients who had experienced pseudo-progression and compared the variations with those from of a patient that had true progression. RESULTS: ctDNA showed rapid and dramatic decreases in pseudo-progressive patients, whereas it was strongly increased in the progressive patient. CONCLUSIONS: ddPCR of ctDNA may thus be an additional tool to discriminate pseudo-progression from true progression for tumors that harbor an oncogenic addiction.

The RAS-related GTPase RHOB confers resistance to EGFR-tyrosine kinase inhibitors in non-small-cell lung cancer via an AKT-dependent mechanism.

Although lung cancer patients harboring EGFR mutations benefit from treatment with EGFR-tyrosine kinase inhibitors (EGFR-TKI), most of them rapidly relapse. RHOB GTPase is a critical player in both lung carcinogenesis and the EGFR signaling pathway; therefore, we hypothesized that it could play a role in the response to EGFR-TKI In a series of samples from EGFR-mutated patients, we found that low RHOB expression correlated with a good response to EGFR-TKI treatment while a poor response correlated with high RHOB expression (15.3 versus 5.6 months of progression-free survival). Moreover, a better response to EGFR-TKI was associated with low RHOB levels in a panel of lung tumor cell lines and in a lung-specific tetracycline-inducible EGFRL858R transgenic mouse model. High RHOB expression was also found to prevent erlotinib-induced AKT inhibition in vitro and in vivo Furthermore, a combination of the new-generation AKT inhibitor G594 with erlotinib induced tumor cell death in vitro and tumor regression in vivo in RHOB-positive cells. Our results support a role for RHOB/AKT signaling in the resistance to EGFR-TKI and propose RHOB as a potential predictor of patient response to EGFR-TKI treatment.

Monitoring KRAS mutations in circulating DNA and tumor cells using digital droplet PCR during treatment of KRAS-mutated lung adenocarcinoma.

Liquid biopsies are a new non-invasive strategy to detect and monitor the biology of non-small-cell lung cancer (NSCLC) in the era of personalized medicine. KRAS is an oncogenic driver that is mutated in 30% of NSCLCs and is associated with a poor prognosis. 62 samples from 32 patients, treated for metastatic KRAS-mutated lung adenocarcinoma, had DNA extracted from plasma and circulating tumor cells (CTCs) prospectively tested for the presence of KRAS mutations using droplet digital PCR. A KRAS mutation was detected in 82% of patients. Sensitivity was 78% for circulating free DNA (cfDNA) and 34% for CTCs. The presence of a KRAS mutation in cfDNA was correlated with a poor response to chemotherapy or targeted therapy. When a KRAS-mutated-DNA was detected and then monitored in cfDNA, its variation during targeted or conventional therapy was correlated with response, according to RECIST criteria, in 87.5% of cases (n=14/16), whereas this correlation was observed in 37.5% of cases for CTCs (n=3/8). We report the usefulness of using digital droplet PCR on liquid biopsies to predict and monitor responses to treatment of KRAS-mutated lung adenocarcinoma. ctDNA was much more sensitive than CTCs in this context.