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How to determine HRD status?

 
When the homologous recombination (HR) pathway is disrupted
by gene mutations, promoter methylation, or undetermined causes, the HR pathway stops working, leading to Homologous Recombination Deficiency (HRD).

 
Tumors with HRD cannot repair themselves effectively after sustaining damage, contributing to genomic instability. There are limitations to determining HRD status when evaluating each 'cause'/ cause individually. Evaluating genomic instability allows
for the assessment of HRD regardless of the specific cause.

 
HRD status can be measured by “cause” through mutations in the HRR pathway (e.g., BRCA1 and BRCA2) and by the “effect” of the presence of genomic scars at a given threshold or functional assay. HRD testing can identify 30% more PARPi-effective population than BRCA testing. (BRCA~20% vs. HRD~50%).
 



The AmoyDx® HRD Focus Panel offers dependable and cost-effective solutions for evaluating PARP inhibitor (PARPi)-related biomarkers and advancing targeted treatment research in ovarian cancer. Leveraging proprietary HANDLE technology, the Genomic Scar Score (GSS) algorithm, and the AmoyDx NGS Data Analysis System (ANDAS), the HRD Focus assay can be seamlessly integrated into your laboratory workflow. Our methodologies enable streamlined end-to-end solutions, requiring less than 1 hour of hands-on operation, delivering high-performance assessment of genomic instability, and providing automated and secure data analysis.
 



Comprehensively Designed GSS Algorithm


The AmoyDx proprietary GSS algorithm is a machine learning-based model which assesses genomic instability by analyzing different types of copy number events across the genome.1




Longer PFS with PARPi Treatment for GSS-positive Group1


The study highlights the promising value of GSS in identifying patients 
who may respond favorably to PARPi treatment. 


 
 
1. Yuan W, Ni J, Wen H, et al. Genomic Scar Score: A robust model predicting homologous recombination deficiency based on genomic instability. BJOG. 2022;129 Suppl 2:14-22. doi:10.1111/1471-0528.17324.

Mutations( Hover over each mutational type to highlight genes covered )

All
SNV/INDELS
BRCA1

BRCA1

SNV/INDELS
BRCA2

BRCA2

SNV/INDELS

Cost-effective

Cost-effective

Decentralized assay solution with ease of use

Decentralized assay solution with ease of use

Fast turnaround time within 3 days

Fast turnaround time within
3 days

Detection of BRCA1/2 genes (SNVs/InDels) and Genomic Scar Score (GSS)

Detection of BRCA1/2 genes (SNVs/InDels) and Genomic Scar Score (GSS)

Specifications

Target regions
whole coding regions and intron/exon boundaries of BRCA1 and BRCA2 genes, and ~24,000 genome-wide SNPs
Alterations detected
BRCA1/2 genes (SNVs/InDels) and Genomic Scar Score (GSS)
Sample type
FFPE tissue
DNA input
Optimal 100ng (minimum 50ng)
Data output per sample
4 Gb
Sequencing type
PE150
Sequencer
Illumina NextSeq 550Dx
TAT for library prep
5 hours (hands-on time <1 hour )
TAT from sample to report
3 days

Technology

The AmoyDx HANDLE® technology is a proprietary amplicon-based method for NGS library preparation.
It offers a time-saving and cost-effective protocol that can be completed within 5 hours, requiring just about 1 hour of hands-on time.

During the library construction process, each individual DNA molecule is tagged with a unique molecular index (UMI) at both ends. This feature enables high sensitivity in variant detection by eliminating any library amplification and sequencing bias.

The probe consists of an extension arm and a ligation arm, both complementary to the target gene region.
Target-specific probes simultaneously hybridize to the DNA/cDNA fragment, forming a circular structure with the intended target captured between the probes. Subsequently, remaining linear probes, single-strand, and double-strand DNA are digested, leaving only the target circular DNA for PCR amplification to form the final library.

Publications

1. Fountzilas, E.; Papadopoulou, K.; Chatzikonstantinou, T.; Karakatsoulis, G.; Constantoulakis, P.; Tsantikidi, A.; Tsaousis, G.; Karageorgopoulou, S.; Koumarianou, A.; Mauri, D.; et al. Concordance between Three Homologous Recombination Deficiency (HRD) Assays in Patients with High-Grade Epithelial Ovarian Cancer. Cancers 2023, 15, 5525.

2. Kekeeva, T.; Andreeva, Y.; Tanas, A.; Kalinkin, A.; Khokhlova, S.; Tikhomirova, T.; Tyulyandina, A.; Popov, A.; Kuzmenko, M.; Volkonsky, M.; et al. HRD Testing of Ovarian Cancer in Routine Practice: What Are We Dealing With? Int. J. Mol. Sci. 2023, 24, 10497.

3. Magliacane, G.; Brunetto, E.; Calzavara, S.; Bergamini, A.; Pipitone, G.B.; Marra, G.; Redegalli, M.; Grassini, G.; Rabaiotti, E.; Taccagni, G.; et al. Locally Performed HRD Testing for Ovarian Cancer? Yes, We Can! Cancers 2023, 15, 43.

4. Doig, Kenneth D. et al. Homologous Recombination Repair Deficiency: An Overview for Pathologists. Modern Pathology, Volume 36, Issue 3, 100049

5. Fumagalli, C., Betella, I., Ranghiero, A., Guerini-Rocco, E., Bonaldo, G., Rappa, A., Vacirca, D., Colombo, N. and Barberis, M. 2022. In-house testing for homologous recombination repair deficiency (HRD) testing in ovarian carcinoma: a feasibility study comparing AmoyDx HRD Focus panel with Myriad myChoiceCDx assay. Pathologica - Journal of the Italian Society of Anatomic Pathology and Diagnostic Cytopathology. 114, 4 (Sep. 2022), 288-294.

6. Scheiter A, Hierl F, Winkel I, Keil F, Klier-Richter M, Coulouarn C, Lüke F, Kandulski A, Evert M, Dietmaier W, et al. Wnt/β-Catenin-Pathway Alterations and Homologous Recombination Deficiency in Cholangiocarcinoma Cell Lines and Clinical Samples: Towards Specific Vulnerabilities. Journal of Personalized Medicine. 2022; 12(8):1270.

7. Liu H, Zhang Z, Chen L, Pang J, Wu H and Liang Z (2022) Next-Generation Sequencing Reveals a Very Low Prevalence of Deleterious Mutations of Homologous Recombination Repair Genes and Homologous Recombination Deficiency in Ovarian Clear Cell Carcinoma. Front. Oncol. 11:798173.

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