Baseline VDJ clonotype detection using a targeted sequencing NGS assay: allowing for subsequent MRD assessment

Hultcrantz, Malin; Rustad, Even H.; Yellapantula, Venkata; Arcila, Maria E.; Ho, Caleb; Syed, Mustafa; Papaemmanuil, Elli; Zhang, Yanming; Maura, Francesco; Landgren, Ola

doi:10.1038/s41408-020-00343-w

Cited by 9 publications

(7 citation statements)

References 15 publications

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“…As MM is a plasma-cell malignancy with clonal Bcell origin, immunoglobulin heavy (IGH) and light chain V(D)J rearrangement sequences have been successfully exploited by next-generation sequencing, 45,46 which is moving towards clinical implementation as a minimal residual disease (MRD) tracking. 47,48 We have applied the scVDJ analysis based on 10x Genomics to confirm the clonotype of tumour cells and found that 27.8% (5/18) MM samples possessed more than one clonotype. According to a previous report of bulk sequencing works, sub-clones were rare and only found in 3.2% of MM patients by VDJ sequencing.…”

Section: Discussionmentioning

confidence: 99%

Single‐cell RNA‐seq reveals clonal diversity and prognostic genes of relapsed multiple myeloma

Lü

et al. 2022

Clinical & Translational Med

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Background Multiple myeloma (MM) is a clinically and biologically heterogeneous plasma‐cell malignancy. Despite extensive research, disease heterogeneity and relapse remain a big challenge in MM therapeutics. We tried to dissect this disease and identify novel biomarkers for patient stratification and treatment outcome prediction by applying single‐cell technology. Methods We performed single‐cell RNA sequencing (scRNA‐seq) and variable‐diversity‐joining regions‐targeted sequencing (scVDJ‐seq) concurrently on bone marrow samples from a cohort of 18 patients with newly diagnosed MM (NDMM; n = 12) or refractory/relapsed MM (RRMM; n = 6). We analysed the malignant clonotypes using scVDJ‐seq data and conducted data integration and cell‐type annotation through the CCA algorithm based on gene expression profiling. Furthermore, we identified disease status‐specific genes and modules by comparison of NDMM and RRMM datasets and explored the findings in a larger MM cohort from the MMRF CoMMpass study. Results We found that all the myeloma cells in either diagnosed or relapsed samples were dominated by a major clone, with a few subclones in several samples (n = 5). Next, we investigated the universal transcriptional features of myeloma cells and identified eight meta‐programs correlated with this disease, especially meta‐programs 1 and 8 (M1 and M8), which were the most significant and related to cell cycle and stress response, respectively. Furthermore, we classified the malignant plasma cells into eight clusters and found that the cell numbers in clusters 2/6/7 were exclusively higher in relapsed samples. Besides, we identified several attractive candidates for biomarkers (e.g. SMAD1 and STMN1) associated with disease progression and relapse in our dataset and related to overall survival in the CoMMpass dataset. Conclusions Our data provide insights into the heterogeneity of MM as well as highlight the relevance of intra‐tumour heterogeneity and discover novel biomarkers that might be a potent therapy.

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Section: Discussionmentioning

confidence: 99%

Single‐cell RNA‐seq reveals clonal diversity and prognostic genes of relapsed multiple myeloma

Lü

et al. 2022

Clinical & Translational Med

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“…The tumor cell-specific rearrangement in the immunoglobulin V(D)J gene region (particularly at the IGH locus) is already exploited as a target by NGS (both amplicon- and capture-based [ 18 ]) or allele specific oligonucleotide (ASO) PCR. Here, the identification of the clonotypic rearrangement is particularly worthwhile also in routine clinical practice, as it represents an extremely specific molecular marker that is useful for monitoring the disease burden along its various stages.…”

Section: Discussionmentioning

confidence: 99%

Single-Cell RNA Sequencing for the Detection of Clonotypic V(D)J Rearrangements in Multiple Myeloma

Matera

Marella

Maeda

et al. 2022

IJMS

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Multiple myeloma (MM) has a highly heterogeneous genetic background, which complicates its molecular tracking over time. Nevertheless, each MM patient’s malignant plasma cells (PCs) share unique V(D)J rearranged sequences at immunoglobulin loci, which represent ideal disease biomarkers. Because the tumor-specific V(D)J sequence is highly expressed in bulk RNA in MM patients, we wondered whether it can be identified by single-cell RNA sequencing (scRNA-seq). To this end we analyzed CD138+ cells purified from bone marrow aspirates of 19 samples with PC dyscrasias by both a standard method based on bulk DNA and by an implementation of the standard 10x Genomics protocol to detect expressed V(D)J sequences. A dominant clonotype was easily identified in each sample, accounting on average for 83.65% of V(D)J-rearranged cells. Compared with standard methods, scRNA-seq analysis proved highly concordant and even more effective in identifying clonal productive rearrangements, by-passing limitations related to the misannealing of consensus primers in hypermutated regions. We next validated its accuracy to track 5 clonal cells with absolute sensitivity in a virtual sample containing 3180 polyclonal cells. This shows that single-cell V(D)J analysis may be used to find rare clonal cells, laying the foundations for functional single-cell dissection of minimal residual disease.

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“…As an example, a targeted NGS assay (myTYPE) for detection of IGH translocations, copy number gains and losses, and somatic mutations in multiple myeloma was found to have a >99% sensitivity and specificity compared to FISH [50]. Using this targeted assay, the tumor-specific V(D)J sequences could be identified and were found to be identical to the V(D)J clonotype found using LymphoTrack in 93% of samples [40]. Furthermore, Rustad et al used bulk RNA sequencing to identify clonal CDR3 sequences that can be used for MRD tracking [44].…”

Section: Next-generation Sequencingmentioning

confidence: 97%

“…Next-generation sequencing (NGS) takes advantage of the unique V(D)J sequences created during B-cell development [36][37][38][39][40][41][42][43]. In the immunoglobulin heavy chain (IGH) variable region, rearrangement occurs in the variable (V), diversity (D), and joining (J) gene segments creating a unique V(D)J sequence (Figure 1) [36].…”

Section: Next-generation Sequencingmentioning

confidence: 99%

“…In a recent study by Ho et al using LymphoTrack, the baseline clonotype detection rate was below 80% in samples with <5% plasma cell content while it was as high as 98% in samples with 10% plasma cells [46]. The limiting factor in sensitivity is often sample quality and tumor content rather than technical limitations [30,39,40,46]. Provided that the baseline clonotype is identified, the clonotype can be detected in virtually 100% of follow-up samples provided that the tumor clone is present [45,46].…”

Section: Next-generation Sequencingmentioning

confidence: 99%

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Minimal Residual Disease in Multiple Myeloma—Current Approaches and Future Clinical Implications

Akhlaghi

Firestone

Hultcrantz

2022

Hemato

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The prognosis and clinical outcomes for patients with multiple myeloma have improved significantly over the past two decades. A substantial number of patients now achieve complete remission after induction therapy, and more sensitive methods are needed to assess response. Minimal or measurable residual disease (MRD) has been incorporated in many clinical trials as well as in clinical practice. The importance of MRD assessment and correlation between MRD negativity and prolonged progression-free and overall survival has been confirmed in numerous clinical trials and several meta-analyses. Recent studies have even suggested that MRD negativity can partly overcome the impact of the negative prognostic factors such as high-risk cytogenetics or adverse revised international scoring system (R-ISS) stage. MRD can be measured in the bone marrow via imaging and via emerging blood-based techniques. The most common methods are multicolor flow cytometry and next-generation sequencing of bone marrow samples. Using these methods in optimal settings, MRD negativity with a sensitivity level of 10−6 can be detected. In this review, we discuss the benefits and limitations of these techniques as well as the clinical implications.

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Baseline VDJ clonotype detection using a targeted sequencing NGS assay: allowing for subsequent MRD assessment

Cited by 9 publications

References 15 publications

Single‐cell RNA‐seq reveals clonal diversity and prognostic genes of relapsed multiple myeloma

Single‐cell RNA‐seq reveals clonal diversity and prognostic genes of relapsed multiple myeloma

Single-Cell RNA Sequencing for the Detection of Clonotypic V(D)J Rearrangements in Multiple Myeloma

Minimal Residual Disease in Multiple Myeloma—Current Approaches and Future Clinical Implications

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