| 28 |
2025 |
Clinical utility and predictive value of cerebrospinal fluid cell-free DNA profiling in non-small cell lung cancer patients with leptomeningeal metastasis
Abstract
Leptomeningeal
metastasis (LM) is a challenging complication of non-small cell lung cancer
(NSCLC). Cerebrospinal fluid (CSF) cell-free DNA (cfDNA) analysis using
next-generation sequencing (NGS) offers insights into resistance mechanisms and
potential treatment strategies. We conducted a study fr|om February 2022 to
April 2023 involving patients fr|om five hospitals in Taiwan who had recurrent
or advanced NSCLC with LM. These patients underwent CSF cfDNA analysis using a
118-gene targeted panel for NGS, with comprehensive clinical data collected.
Among 25 enrolled patients, 22 (88.0 %) had EGFR mutations, while three (12.0
%) had EML4-ALK fusion, KIF5B-RET fusion, and ERBB2 A775_G776insSVMA. CSF cfDNA
sequencing of 27 samples (fr|om 25 patients) all confirmed their original driver
mutations. Of total cohort, 18 patients (72.0 %) underwent intrathecal
pemetrexed (ITP), with a median survival time of 7.4 months (95.0 % confidence
interval, 3.3–11.6) fr|om the initiation of ITP to death. Among them, ten
individuals (55.6 %) survived beyond 6 months. Notably, MET copy number gain
(CNG) correlated significantly with survival time exceeding 6 months after ITP
(p = 0.007). The coexistence of EGFR T790M and EGFR-independent resistance
alterations was associated with shorter survival times after ITP, with a median
survival time of 1.9 months compared to 9.9 months for those without EGFR T790M
(p = 0.010). Our results highlight CSF cfDNA NGS's potential in LM resistance
understanding and ITP efficacy prediction. MET CNG positively impacts survival for
ITP recipients, whereas the coexistence of EGFR T790M and EGFR-independent
resistance mechanisms leads to poor outcomes.
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Neoplasia |
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| 27 |
2025 |
Enhanced Detection of Actionable Mutations in NSCLC Through Pleural Effusion Cell-Free DNA Sequencing: A Prospective Study
Abstract
Background: Inadequate tumour samples often hinder
molecular testing in non-small cell lung cancer (NSCLC). Plasma-based cell-free
DNA (cfDNA) sequencing has shown promise in bypassing these tissue limitations.
Nevertheless, pleural effusion (PE) samples may offer a richer cfDNA source for
mutation detection in patients with malignant PE.
Methods: This prospective study enrolled newly
diagnosed advanced NSCLC patients with malignant PE. PE samples were collected
for cfDNA NGS analysis. Meanwhile, PE cell pellet RNA was extracted for reverse
transcription polymerase chain reaction (RT-PCR) for clinically relevant actionable
mutations and then confirmed by Sanger sequencing. The concordance between PE
cell pellet RT-PCR and PE cfDNA NGS analyses was analysed.
Results: Fifty patients were enrolled. The median
age was 68.5 years, and the female-to-male ratio was 29:21. Most patients (74%)
were non-smokers. Notably, 45/50 patients (90%) had actionable mutations,
including EGFR exon 19 deletions (24%), EGFR L858R mutations (36%), HER2 exon20
insertions (10%), ROS1 rearrangements (4%), EGFR exon20 insertions (2%), ALK rearrangements
(4%), RET rearrangements (2%), KRAS G12C mutations (2%), and CD74-NRG1 fusions
(2%). Among the 50 enrolled patients, actionable mutations were detected in 44
(88%) by PE cfDNA NGS, 39 (78%) by PE cell pellet Sanger sequencing, and 33
(66%) by clinical tissue genetic testing (P=0.031). The detection of actionable
mutations fr|om PE cfDNA NGS remained consistently high across M1a to M1c
stages.
Conclusions: PE cfDNA genotyping has clinical
applicability for NSCLC patients and can serve as an additional source for
molecular testing. Incorporating PE NGS cfDNA analysis into genetic testing
enhances diagnostic yield and aids in identifying actionable mutations in
clinical practice.
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European Journal of Cancer |
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| 26 |
2024 |
Personalized circulating tumor DNA assay precisely predicts the response of neoadjuvant chemotherapy in breast cancer patients
Abstract
Introduction
Circulating tumor
DNA (ctDNA), which is detected in the blood as cell free DNA fragment fr|om
tumor cells, could indicates genomic landscape, tumor burden and treatment
response during chemotherapy as a non-invasive method. Especially, personized
ctDNA assay by next-generation sequencing(NGS) is able to detect a trace amount
of ctDNA and precisely figure out the change of the amount of ctDNA during NAC
and follow up period after curative surgery. Here, we performed serial ctDNA
evaluations in EBC patients who diagnosed as TNBC or HER2-positive BC and
received NAC followed by curative surgery. We aimed to predict NAC response and
detect minimal residual disease (MRD) using personalized ctDNA assay.
Methods
CtDNA was detected
by AlphaLiquid®Detect,
a tumor-informed personalized MRD detection assay exploring most of the
mutations in tumor. Whole exome
sequencing (WES) of tumor tissue and peripheral blood mononuclear cells (PBMCs)
was performed. Patient-specific somatic mutations were selected using a
proprietary algorithm. In brief, clonal variants were prioritized using
integrated information including variant allele frequency, population allele
frequency databases, somatic variant databases, variant pathogenicity, and
genomic context. Up to 100 variants were selected for patient monitoring. A
hybridization capture panel consisting of a pool of 4 patients’ selected
variants was synthesized. These bespoke panels (BSPs) were used for ctDNA
detection. Patients with more than 2 tumor-derived mutations detected in plasma
were considered as ctDNA positive.
Inclusion criteria
included patient who diagnosed as stage IIA-IIIC BC planned to NAC followed by
curative surgery. Among BC subtypes, TNBC and HER2-positive BC were allowed.
Collection of specimens and associated clinical data used in this study was approved
by the Institutional Review Board of Samsung Medical Center (IRB File
No.2021-02-033), and we received informed consents for this study.
Results
fr|om May 2021 to
Sep 2022, 158 patients has been enrolled. Archival tissues were not available
in 47 patients and tissue WES had failed in six patients. Therefore, 105
patients were enrolled and analyzed their ctDNA at diagnosis based on tissue
WES data.
Of 105 patients,
Median age of BC patients was 49.3 years of age (range: 26.1, 67.8). Thirty-six
patients (34.3%) were post-menopausal status and others (65.7%) were
pre-menopausal status. In BC pathology at diagnosis, 56 (53.3%) were TNBC and
other 49 (46.7%) were HER2-positive BC. Among HER2-positive BC, hormone
receptor (HR)-positive were in 21 (20.0%). In clinical stage at diagnosis,
stage II were 46 (43.8%) and stage III in 59 (56.2%).
In 105 patients,
median number of somatic mutations was 60 (interquartile range [IQR]: 38.5,
91). CtDNA detection rate at BC diagnosis was 90.5% and all of BC which not be
detected ctDNA was HER2-positive BC with clinical stage II. Median amount of
ctDNA at diagnosis was 20 (IQR: 6, 44).
In the number of
somatic mutations, there was no difference according to BC subtypes (P=0.121)
and clinical stage (P=0.700) but the amounts of ctDNA at diagnosis was
different. CtDNA was much more detected in TNBC compared to HER2-positive BC
(Median: 34.5 vs. 15.9; P<0.001) and clinical stage III compared to clinical
stage II (Median 33.2 vs. 19.5; P=0.002). After NAC, ctDNA at curative surgery
was tested in 70 patients. In 70 patients, pathologic complete response(pCR)
was observed in 44 patients. CtDNA was detected in ten patients (14.3%) and
eight did not achieve pCR and two with pCR (Specificity of assay: 95.5%,
positive predictive value: 0.80, P=0.032).
Conclusions
Personalized ctDNA assay can precisely detected ctDNA at diagnosis and
their detection rate was associated with BC subtypes and clinical stage. In
addition, ctDNA at curative surgery also can predict NAC response in EBC
patients. Long term ctDNA follow up of MRD would be warranted.
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Cancer Research |
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| 25 |
2024 |
High circulating tumor DNA (ctDNA) concentration was associated with shorter progression free survival in patients with metastatic breast cancer
Abstract
Background
Metastatic breast cancer can be classified into different subtypes depending on hormone receptor (HR) and HER2 status. The subtype can change during tumor progression, and repeated biopsy is needed to deliver the most appropriate treatment every time a new lesion is found. It is not always possible to get a new biopsy fr|om metastatic sites, and therefore liquid biopsy using circulating tumor DNA (ctDNA) is suggested as an alternative method to replace conventional biopsy.
Methods
We performed a prospective serial collection of 65 ctDNA samples fr|om 17 patients with metastatic breast cancer (mBC) at Seoul National University Hospital fr|om October 2020 to March 2022. We used IMBdx AlphaLiquid®100 method to detect the genetic changes and analyzed the correlation with clinical outcomes.
Results
Median age was 45 (range 32 – 62). Fifteen patients (88.2%) were relapsed mBC and most of the patients (14/17: 82.4%) were HR-positive and HER2-negative. Most of the patients had their ctDNA examined at baseline and at the time of maximal response and/or at progression. Fifteen patients (88.2%) received systemic therapy including hormone therapy, anti-HER2 therapy, and cytotoxic chemotherapy. Eight patients (47.1%) were on the first-line treatment for mBC, and 7 patients (41.2%) were on the second or later lines for mBC at the time of baseline sampling. The concentration of ctDNA and the sum of mutated allelic frequency was calculated for each sample. The ctDNA concentration ranged fr|om 0.71 to 1386.00 ng/mL, and the median value was 5.37 ng/mL. We dichotomized these samples into two groups, with ctDNA concentration either higher or lower than the median value. Then we analyzed progression free survival (PFS) of each group. Patients with higher ctDNA concentration showed shorter PFS (7 mo. vs. not reached, p< 0.001). The sum of mutated allelic frequency ranged fr|om 0.00% to 223.46% and the median value was 6.36%. Patients with higher mutated allelic frequency showed shorter PFS (6 mo. vs. 22 mo., p< 0.001). In addition, the PFS was significantly worse in patients who had mutated PIK3CA (5 mo vs. 22 mo, p< 0.001). The patients with mutated TP53 also showed shorter PFS (6 mo vs. 17 mo, p< 0.001) in univariate analysis. High estrogen receptor positivity in immunohistochemistry was correlated with lower mutated allelic frequency in ctDNA (p=0.003) but had no impact on the concentration of ctDNA (p=0.165). The concentration of ctDNA differed by metastatic sites. Patients with metastases to bones (p=0.007), liver (p< 0.001), soft tissue or lymph nodes (p=0.002) were more likely to have higher concentrations of ctDNA, while patients with brain metastases had significantly lower ctDNA concentration (p=0.006). When the sum of mutated allelic frequency of ctDNA and metastatic sites was analyzed, bone (p=0.001), liver (p< 0.001), and soft tissue or lymph node (p< 0.001) metastases had a positive correlation, while brain had negative correlation (p=0.017). Lung or pleural metastases had no significant correlation with ctDNA, neither concentration (p=0.271) nor mutated allelic frequency (p=0.965).
Conclusion
Patients with mBC with higher concentrations of ctDNA or higher mutated allelic frequency of ctDNA at baseline showed significantly shorter PFS. PIK3CAmt and TP53mt detected by liquid biopsy could be used as a poor prognostic biomarkers for mBC patients.
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AACR |
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| 24 |
2024 |
Assessing the clinical value of ctDNA sequencing for initial tumor profiling in metastatic colorectal cancer patients with sufficient tumor tissue
Abstract
Introduction
Tumor profiling including RAS, BRAF, HER2,
and MSI/MMR status, is required to determine the treatment for patients with
metastatic colorectal cancer (mCRC) at the time of diagnosis. While
comprehensive tumor profiling using tissue-based next-generation sequencing
(NGS) is increasingly adopted for initial profiling in mCRC, the role of
circulating tumor DNA (ctDNA) NGS as initial testing in patients with
sufficient tumor tissue is not clearly understood. We assessed the clinical
value of ctDNA sequencing compared to tumor NGS in patients with newly
diagnosed mCRC who have sufficient tumor specimens.
Methods
We prospectively enrolled consecutive
patients with newly diagnosed mCRC at the National Cancer Center Korea. As per
the institutional protocol for mCRC, initial tumor profiling was performed on
primary tumor tissue using an in-house NGS panel (NCC PCP ver.3), which
included 525 genes. For ctDNA sequencing, patients were evaluated using the
AlphaLiquid®100 comprehensive cancer panel (IMBdx, Inc.), which included 118
genes, before the initiation of chemotherapy. Additionally, immunohistochemical
(IHC) testing for HER2 and polymerase chain reaction (PCR)/IHC testing for MSI
and/or MMR were performed to assess the accuracy of HER2 and MSI/MMR status.
Results
A total of 188 patients were enrolled. In
139 eligible patients, 275 potentially actionable mutations were found in 12
selected CRC-related genes (APC, TP53, KRAS, NRAS, BRAF, FBXW7, PIK3CA, ERBB2,
SMAD4, NF1, EGFR, MET). Of these, 32% were found both in ctDNA and tissue; 54%
were found in ctDNA only; 12% in tissue only, and 2% were discordant. For
RAS/BRAF mutations, which are required for anti-EGFR treatment decisions, the
concordance rate between ctDNA and tissue NGS was 83.1%, and the concordance was
higher in patients with higher ctDNA concentrations. For 9 patients with
potentially actionable copy number variations (CNV) in EGFR, HER2, MET, and
FGFR1, 3 cases were found by both assays; 4 were found by ctDNA only, and 2
were found by tissue only. ctDNA NGS correctly predicted MSI/MMR status in 2
out of 4 patients with MSI-H/dMMR; in the 2 other patients, the MSI and MMR
statuses were different, suggesting potential false positivity. In addition,
the fold changes in ctDNA dynamics during treatment significantly correlated
with changes in tumor size and CEA levels, as well as with droplet digital PCR
copy number fold changes. In a patient with MET amplification, ctDNA NGS
identified MET Y1230H as a potential acquired resistance mutation after
crizotinib treatment, which responded to cabozantinib but not to capmatinib.
Conclusions
Initial tumor profiling using ctDNA NGS
yielded outcomes comparable to those of tumor tissue NGS in guiding treatment
for patients with newly diagnosed mCRC, thereby suggesting its utility as an
initial profiling method in mCRC.
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AACR |
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