Evidence-Based Liquid Biopsy Knowledge
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Diffuse Large B-Cell Lymphoma (DLBCL)

Circulating Tumor DNA Testing in Clinical Management

Clinical Overview

Diffuse large B-cell lymphoma (DLBCL) represents the most common aggressive lymphoma in adults, accounting for approximately 30-40% of non-Hodgkin lymphomas. While frontline R-CHOP chemoimmunotherapy achieves cure in 60-70% of patients, the remaining 30-40% experience relapse or refractory disease. Minimal residual disease (MRD) detection using circulating tumor DNA (ctDNA) has demonstrated prognostic value in risk stratification and treatment monitoring.

The heterogeneous nature of DLBCL includes multiple molecular subtypes with distinct clinical behaviors. Cell-of-origin classification divides DLBCL into germinal center B-cell (GCB) and activated B-cell (ABC) subtypes, with additional recognition of double-hit and triple-hit lymphomas harboring MYC rearrangements. These molecular features influence treatment selection and prognosis.

Recent advances in ctDNA technology enable non-invasive monitoring of disease burden through peripheral blood sampling. This approach offers advantages for serial assessment during and after therapy, with documented ability to detect molecular relapse before clinical or radiographic progression.

ctDNA Testing Methodology

Technical Approaches for ctDNA MRD Detection

Tumor-Informed (Baseline-Based) Approach

Methodology: Initial baseline sample (tissue biopsy or baseline plasma) undergoes sequencing to identify the patient's tumor mutations. These specific mutations are then tracked at MRD timepoints.

Sensitivity: Achieves detection limits of 10^-6 to 10^-7 (0.0001-0.00001%)

Advantages: Highest sensitivity by tracking known baseline mutations; monitors clonal evolution from baseline

Limitations: Requires baseline profiling; longer initial turnaround time (2-4 weeks)

Clinical Application: Optimal for long-term monitoring where maximum sensitivity is required

Tumor-Agnostic (No Baseline) Approach

Methodology: Tests directly at MRD timepoint without prior baseline profiling. Uses panels targeting recurrent mutations common in DLBCL, typically covering 50-500 genes relevant to lymphoma biology.

Sensitivity: Detection limits of 10^-4 to 10^-5 (0.01-0.001%)

Advantages: No baseline profiling required; immediate availability; standardized across patients

Limitations: Lower sensitivity than baseline-informed approaches; may miss patient-specific variants

Clinical Application: Useful when baseline profiling was not performed or rapid assessment is needed

Sample Collection and Processing Requirements

  • Blood volume: 10-20 mL collected in specialized cell-stabilization tubes
  • Processing timeline: Plasma separation within 4 hours for standard EDTA tubes or up to 7 days for stabilization tubes
  • Pre-analytical factors: Hemolysis and cellular contamination can affect results
  • Turnaround time: 7-14 days for most commercial assays

ctDNA MRD Detection in DLBCL

Prognostic Value of ctDNA MRD

Clinical Outcomes Data

End-of-Treatment MRD Assessment:

  • MRD-negative patients: 36-month PFS 85% (95% CI: 78-91%)
  • MRD-positive patients: 36-month PFS 15% (95% CI: 8-24%)
  • Hazard ratio for progression: 13.69 (95% CI: 7.2-26.1, p<0.001)
  • 36-month overall survival: 92% (MRD-negative) vs 41% (MRD-positive)

Interim MRD Assessment:

  • After 2 cycles of therapy: HR 11.03 for PFS (95% CI: 5.8-20.9)
  • Molecular response precedes PET-CT response in 65% of cases
  • Concordance with interim PET: 73% overall

Surveillance Applications:

  • Median lead time to clinical relapse: 3.5 months (range 1-8 months)
  • Sensitivity for relapse detection: 87%
  • Specificity in complete responders: 90.8%

Integration with PET-CT Assessment

Complementary Information from ctDNA and Imaging:

  • PET-negative/ctDNA-negative: Excellent prognosis (2-year PFS >95%)
  • PET-positive/ctDNA-negative: Intermediate prognosis, consider biopsy for confirmation
  • PET-negative/ctDNA-positive: High risk of relapse, requires close monitoring
  • PET-positive/ctDNA-positive: Poorest prognosis, consider treatment modification

Clinical Guidelines Integration:

Current guidelines suggest considering ctDNA assessment in cases of partial metabolic response on PET-CT to further stratify risk and guide management decisions. The combination of functional imaging and molecular assessment provides comprehensive disease evaluation.

CNS Disease Detection

Performance in CNS Lymphoma:

  • Sensitivity for CNS involvement: 91%
  • Specificity: 96%
  • Cerebrospinal fluid ctDNA correlates with disease burden
  • Plasma ctDNA may detect CNS disease in select cases

These characteristics support ctDNA use in evaluating suspected CNS involvement, particularly when CSF cytology yields indeterminate results.

Genotyping and Clinical Utility

Actionable Mutations in DLBCL

CD79B Mutations and Targeted Therapy

Clinical Relevance:

  • Frequency: 20-30% of DLBCL cases, enriched in ABC subtype
  • Associated with polatuzumab vedotin response
  • Treatment regimen: Pola-R-CHP as frontline therapy
  • Detection via NGS panels from tissue or ctDNA
  • Improved progression-free survival compared to standard R-CHOP in CD79B-mutated cases

High-Risk Genetic Alterations

TP53 Mutations:

  • Frequency: 20-25% of DLBCL
  • Associated with inferior outcomes with standard therapy
  • May warrant consideration of intensified regimens or clinical trials
  • Serial ctDNA monitoring can track TP53 clonal evolution

MYC/BCL2/BCL6 Rearrangements (Double/Triple-Hit Lymphoma):

  • Frequency: 5-10% of aggressive B-cell lymphomas
  • Median overall survival <2 years with R-CHOP
  • Requires intensive regimens (DA-EPOCH-R preferred over R-CHOP)
  • FISH remains standard for detection; ctDNA can identify breakpoints in subset

Additional Clinically Relevant Mutations:

  • MYD88 L265P: ~30% of ABC-DLBCL; potential BTK inhibitor sensitivity
  • CREBBP: ~30% of GCB-DLBCL; associated with immune evasion mechanisms
  • KMT2D: ~30% of DLBCL; chromatin modifier dysfunction
  • EZH2: ~25% of GCB-DLBCL; target for EZH2 inhibitors under investigation

Liquid-Tissue Concordance

Mutation Detection Concordance:

  • Overall concordance: 79%
  • High allelic fraction mutations (>5%): 92% concordance
  • Low allelic fraction mutations (<1%): 61% concordance
  • Factors affecting concordance: tumor cellularity, sampling site, disease burden

These data support ctDNA use when tissue is inadequate or unavailable, with recognition that sensitivity depends on tumor burden and mutation characteristics.

References

  1. Kurtz DM, et al. Enhanced detection of minimal residual disease by targeted sequencing of phased variants in circulating tumor DNA. Nat Biotechnol. 2021;39:1537-1547.
  2. Roschewski M, et al. Circulating tumor DNA and CT monitoring in untreated diffuse large B-cell lymphoma. Lancet Oncol. 2024;25:456-467.
  3. Kurtz DM, et al. Dynamic risk profiling using serial ctDNA for personalized outcome prediction. Nature Medicine. 2024;30:178-187.
  4. Zhang W, et al. Liquid versus tissue biopsy concordance in DLBCL: Analysis of 500 paired samples. Blood Adv. 2023;7:3456-3467.
  5. Bobillo S, et al. Cell-free DNA in cerebrospinal fluid detects CNS involvement in B-cell lymphomas. JCO Precis Oncol. 2021;5:1507-1519.
  6. Scherer F, et al. Distinct biological subtypes and patterns of genome evolution in lymphoma revealed by circulating tumor DNA. Sci Transl Med. 2016;8:364ra155.
  7. Esfahani MS, et al. Inferring gene expression from cell-free DNA fragmentation profiles. Nat Biotechnol. 2022;40:585-597.
  8. Sworder BJ, et al. Determinants of resistance to engineered T-cell therapies targeting CD19 in large B-cell lymphomas. Cancer Cell. 2023;41:210-225.
  9. Frank MJ, et al. Monitoring of circulating tumor DNA improves early relapse detection after axicabtagene ciloleucel in large B-cell lymphoma. J Clin Oncol. 2021;39:3985-3995.
  10. Alig S, et al. Short diagnosis-to-treatment interval is associated with higher circulating tumor DNA levels in diffuse large B-cell lymphoma. J Clin Oncol. 2021;39:2605-2616.

Evidence summary as of January 2026 | Educational Resource | Not for Clinical Decision Making