TEST:

Idylla™ KRAS Mutation Test

KRAS mutation testing on liquid-based cytology using the Idylla or equivalent technique, combined with the PDAC EUS-FNB sample, should become a standard for diagnosis to avoid delaying treatment by doing another biopsy. Furthermore, knowledge of the KRAS status from treatment initiation could be used to isolate mutations requiring targeted treatments or inclusion in clinical research trials, especially for wild-type KRAS PDAC.

"Biocartis...announces that it will host a free corporate workshop at the Annual Meeting of the ‘Association for Molecular Pathology’ (AMP), a leading molecular diagnostics conference, taking place between 14-18 November 2023 in Salt Lake City, Utah (US)."

The Idylla Rapid IDH1/2 Mutation Assay had 92% concordance with NGS for FFPE sections. Compared with Idylla EGFR and KRAS assays (previously published) both control and target Cqs were considerably higher using the same number of sections or amount of DNA. A large DNA input requires stewardship for effective tissue utilization for molecular testing, especially in glioma biopsy samples, which may be limited.

P2, N=27, Recruiting, M.D. Anderson Cancer Center | Trial completion date: Oct 2023 --> Oct 2025 | Trial primary completion date: Oct 2023 --> Oct 2025

We report the largest North African analysis of NRAS and KRAS status in colorectal metastatic patients. This study showed the ability in low middle income countries to perform a high rate of valid tests and the unusual trend towards older patients for NRAS mutations.

In summary, our data show that high Ki67 expression in CRCs is associated with good clinical outcome. High Ki67 seems to be linked to CRC clinicopathological parameters and to the proliferation molecular pathway RAS/MAPK and indicates a cellular state of high proliferative activity. Finally, our findings argue for a clinical utility of Ki67 immunostaining as an independent prognostic biomarker in CRC and as a prediction marker for response to chemotherapy.

The significant association of high MVD-CD34 and mutated RAS status, suggesting that mutation of the RAS gene may increase angiogenic activity in CRC.

In our study, we found that the prognosis of KRAS-mutated mCRC patients differs according to the point mutation detected. Larger studies are needed to define clinical and molecular characteristics and prognosis of patients with KRAS-mutated mCRC in the era of personalised oncology.

Our study found that MSI/RAS/RAF mutated status is rare (9.5%) meanwhile both MSS and RAS/RAF wild cases, representing the worst prognostic group, represent 40% of the cases.

The first-line therapy consisted of anti-EGFR cetuximab in combination with irinotecan or oxaliplatin-based chemotherapy. PFS2 was statistically significantly shorter for RAS MT patients resulting in statistically non-significantly shorter OS in RAS MT patients. Our findings indicate that a change in RAS status is a negative prognostic factor for the second-line treatment, but this needs to be confirmed in large-scale studies. Testing RAS status analysing ctDNA might help to stratify patients during the metastatic treatment and to personalize the ongoing therapy.

Our results highlighted a difference in the molecular profile of KRAS, NRAS,and BRAF mutations' distribution between metastatic colorectal cancer and non-metastatic ones: NRAS and BRAF mutations were associated with metastasis advent; moreover, the G13D mutation of KRAS was correlated to a better prognosis than mutations in codon 12 of KRAS: G12A and G12S alleles gave the worst prognosis.

Patients with mCRC were treated with a targeted therapy (Bevacizumab or Cituximab) associated to chemotherapy as a first-line therapy according to the RAS status... Our study revealed that RAS muations were correlated with metastatic disease and worst OS and PFS.

The patient was given folforinox + avastin during 3 months, then Avastin+Xeloda as maintenance treatment. The question is: "Are RAS and BRAF mutations really exclusive to each other"? It seems difficult to answer: even if it's a rare event, it is important to search for BRAF mutations even if RAS is mutatedin specific patients.

Even if high concordance is noted between primary and metastatic tumors for the KRAS mutation according to literature, passenger mutations could change the deal, and primary tumor tissue will not be sufficient for a therapeutic decision.

Patients RASmut on liquid biopsy at baseline are randomized to receive FOLFIRI plus Cetuximab or FOLFIRI plus Bevacizumab... These preliminary data suggest that liquid biopsy might better recapitulate the heterogeneity of mCRC and might be useful in clinical practice to complement tissue testing. The clinical relevance of RAS variants detected in cfDNA will be determined in the LIBImAb trial.

P=N/A | N=55 | Completed | Sponsor: Royal Cornwall Hospitals Trust | Recruiting ➔ Completed | N=200 ➔ 55

P2 | "This is the first prospective randomized trial which evaluated anti-EGFR monoclonal antibody (panitumumab) in addition to standard of care (trifluridine-tipiracil) as third-line rechallenge therapy in chemo-refractory RASWT mCRC patients. Baseline liquid biopsy allows selection of RAS/BRAF WT ctDNA patients who could have a relevant clinical benefit. Clinical trial information: NCT05468892."

Beside established anti-CRC treatment, better understanding of the causality of CRC can be established by combining epidemiology and genetic/epigenetic on CRC etiology in Morocco. This approach may be able to significantly reduce the burden of disease in the country. Moreover, the Moroccan government should develop policy on CRC prevention and public health programs which may serve as a feasible setting to increase public awareness on lifestyle risk factors.

In this study, we challenged OPA with QNS samples, and OPA successfully detected SNVs and CNVs with an at least 83% concordance rate, compared to reference methods. We demonstrated that OPA can be a valuable, rapid method for molecular testing of advanced NSCLC cases with scant tissue.

P2, N=27, Recruiting, M.D. Anderson Cancer Center | Initiation date: Mar 2022 --> Mar 2023

Analytical concordance, repeatability, reproduc-ibility, and a robust limit of detection was documented in this assessment of the RNAseq approach. This data supports the RNAseq first approach, using FusionPlex® research assay (Invi-tae), and a decision has been made to adopt this workflow in our laboratory.

Ras/Braf mutation frequency in our population approaches occidental series.

Conclusions The Idylla TM KRAS mutation test shows high concordance and agreement with NGS testing to detect KRAS mutations in lung cancer samples. KRAS G12C mutations were found in about 18% of tumors without EGFR, BRAF, ALK, and ROS1 alterations.

Editorial acknowledgement The authors wish to thank Irene Mansilla-Núñez from TFS HealthScience for the medical writing support. NGS analysis suggested that resistance is promoted by a complex mutational process. In our study, RASmt by itself did not predict the ORR or PFS for P+CT, although they tended to be higher in RASwt pt.

In conclusion, this study showed that the frequency of KRAS mutations in CRC has been increasing in recent decades in the specific population of Greenland. The results of this study may be used in initiatives related to targeted therapy of CRC in specific ethnicities and in investigations focusing on the environmental factors of cancer-related somatic mutations.

Additionally, the cfDNA of 8 pts of C2 that progressed after 1L chemotherapy + panitumumab (C-P) was analyzed using next-generation sequencing (NGS) (AVENIO ® ctDNA expanded panel and Illumina NextSeq 550) containing 77 genes Results and Discussions A total of 65/64/58 (MAF ≥1/≥0.1/≥0.02% [C1]) and 45 (C2) pts were evaluable ( RAS wt at baseline as per solid and liquid biopsies and blood sample available at DP), of which 58/57/51 (C1) and 40 (C2) received C-P...However, a complex mutational process driving resistance is suggested based on NGS-cfDNA analysis. Further investigation in larger numbers of pts is warranted Study supported by Amgen S.A.

Secondary resistance is nearly universal among NSCLC patients who receive TKIs for actionable mutations. Solvent front secondary mutations in RET (G810X) as well as activation of KRAS or MET molecular pathways have been reported to confer resistance to RET inhibitors. To our knowledge, this is the first case of a RET fusion lung cancer with resistance to a RET inhibitor mediated by transformation to small cell carcinoma.

The results of this study do not reveal a clear correlation between mutation and response to immunotherapy. The mechanism regulating immune system activity in the tumor microenvironment remains unclear.

It is a minimally invasive method with a high degree of sensitivity and is representative of tumour heterogeneity. Future work is needed to evaluate cases with tissue-plasma discordance and the emerging monitoring of acquired mutations in RAS and BRAF as mechanisms of resistance.

"This result demonstrates a high concordance between the Idylla KRAS and NRAS-BRAF Mutation Tests and an existing IVD kit. In this manner, the Idylla mutation tests were validated for the detection of clinically significant KRAS, NRAS, and BRAF mutations in FFPE samples from colorectal cancer patients."

P2; Not yet recruiting --> Recruiting

Analysis of ctDNA is a viable strategy for clinical management of mCRC patients. Although the OncoBEAM assay sensitivity is somewhat higher, the fully automated Idylla platform also has good performance, while being cheaper and much less labor-intensive, for the detection of RAS mutations in plasma, either at diagnosis or after progression when considering anti-EGFR treatment rechallenge.

In this article, we review the data about the fast and fully automated real-time PCR platform Idylla™ (Biocartis, Mechelen, Belgium) permitting the mutational analyses of BRAF, KRAS, NRAS, EGFR and microsatellite instability notably in melanoma, non-small-cell lung cancer and colorectal cancer samples. Future applications as well as the implementation of Idylla™ in the workflow of pathology and/or molecular biology laboratories are also discussed.

Papillary renal cell neoplasm with reverse polarity represents 3.57% of PRCC in our study. PRNRP is a distinct and rare entity with characteristic morphological features and frequent mutation in KRAS genes. The positivity of GATA3 and L1CAM support that this tumor arises from the distal renal ducts.

"Out of 60 total BRAF-mutated cases (NGS+ IBMA), 14 (23%) received targeted therapy as first line, and 32 (53%) received immunotherapy, indicating that immunotherapy is the most common first-line systemic therapy in our cohort of patients with BRAF-mutated melanoma. The TAT of the testing platform did not factor into these decisions. NGS provides a more comprehensive assessment and may be the preferred choice with adequate samples."

The aim of our study was to investigate the use of TPs on the Idylla system to reduce TATs, such that results would be available with routine surgical pathology reports. Our approach of collecting TP samples directly from tissue and testing them via the Idylla system significantly reduced potential TATs. Samples processed with rapid intent often provided results the same day as or before the pathology report release.

"Performing KRAS testing from peripheral blood for patients, who have no elevated tumor markers, might be a novel option for treatment monitoring complementing routine imaging techniques."

"Our preliminary results show promise for the detection of mutant KRAS in NSCLC LBC and peripheral blood samples. Such ap- plication provides a means of rapid, non-invasive diagnostic turnover at local centres. Furthermore, given the reported higher prevalence of the KRAS mutation to the mutually exclusive molecular mutations (EGFR, ALK and ROS-1) we propose testing for the KRAS mutation first."

Our study shows rapid pre-screening for KRAS mutations in NSCLC can be achieved in a matter of hours from initial histological diagnosis along with PD-L1 status. It can eliminate the need for further single gene or panel testing in almost 30% of cases. These patients can begin a definitive treatment plan within days of diagnosis with no addi- tional costs for testing and a potential reduction in clinical appointments.

The study has shown a 46% mutation rate for KRAS muta- tions in NSCLC samples, and with a KRAS G12C inhibitor on the hori- zon we have shown that it is feasible to use KRAS testing as a screening tool and for treatment stratification purposes in parallel. Results from the BRAF validation demonstrate the ability to detect mutations in the BRAF gene in a number of tissue types.

Our results confirm the conclusion of recent reports that the Idylla™EGFR assay is not suitable in a resistance to EGFR TKI setting, also not in our cytological NSCLC samples prepared as cytoblocks using AGAR and paraffin embedding. KRAS and primary EGFR mutations were detected using the Idylla™ assays in virtually all cytological NSCLC samples. This analysis was rapid and time-saving compared to other mutation detection assays and may be useful if the amount of material is insufficient to perform a full set of molecular tests.

We demonstrated through liquid biopsy that KRAS G12C mutation can be acquired or lost at the time of PD due to cancer clonal evolution. Our underline the limitation to restrict biomarker studies to the analysis of primary tumor tissue for clinical trial stratification of cancer patients at PD, particularly if the tissue biopsy used for biomarker evaluation has been performed long before PD.