HRD Testing: A Crucial Tool in Cancer Diagnosis

Homologous recombination deficiency (HRD) testing is a valuable diagnostic and prognostic tool in the field of oncology. It is a molecular assay that plays a prominent role in identifying tumours with impaired DNA repair capabilities. Thereby, it guides personalised treatment strategies, especially in ovarian, breast and prostate cancers. HRD testing is especially important in the era of precision medicine, where therapy is increasingly personalised to a patient's genetic makeup.

What is HRD Testing?
The HRD’s full form is "Homologous Recombination Deficiency" testing. It is a genomic assessment designed to detect whether a tumour exhibits defects in the homologous recombination repair (HRR) pathway. The HRR pathway is essential for the accurate repair of DNA double-strand breaks.

When the repair mechanism fails due to mutations or epigenetic alterations, particularly in genes like BRCA1 and BRCA2, cells become susceptible to genomic instability. It leads to cancer progression.

HRD test evaluates several markers, including loss of heterozygosity (LOH), telomeric allelic imbalance (TAI), and large-scale state transitions (LSTs). These are collectively referred to as genomic scars. These markers form the basis for calculating a Genomic Instability Score (GIS), a surrogate for HRD status.

Only two HRD tests are currently FDA-approved, both relying on genomic scar detection and BRCA mutation testing. However, researchers are actively exploring new technologies, including functional assays like RAD51 foci formation, to provide real-time HRD assessment. These advancements aim to overcome limitations like outdated HRD status due to tumour evolution and limited predictive accuracy.

Why HRD Testing Matters in Oncology

Tumours with homologous recombination deficiency are often more sensitive to DNA-damaging agents like platinum-based chemotherapy and poly (ADP-ribose) polymerase (PARP) inhibitors. The mechanism of PARPi involves synthetic lethality—tumours deficient in HRR cannot repair the double-strand breaks that result when PARP activity is blocked, leading to tumour cell death.

HRD status becomes an important biomarker for predicting response to PARPi, especially in cancers with BRCA mutations or BRCA-like characteristics, collectively termed "BRCAness." Identifying this phenotype expands the therapeutic window for patients who do not carry BRCA mutations but exhibit similar vulnerabilities.

HRD Testing in Ovarian Cancer

The utility of HRD testing in ovarian cancer is particularly well-documented. High-grade serous ovarian carcinoma (HGSOC) exhibits homologous recombination deficiency.

According to current guidelines, HRD testing in ovarian cancer is now considered standard practice to determine eligibility for maintenance therapy with PARP inhibitors following platinum-based chemotherapy.

Several FDA-approved companion diagnostic tests are validated for ovarian cancer, e.g., Myriad’s MyChoice CDx and FoundationOne CDx. These tests calculate a genomic instability score based on LOH, TAI, and LST, with defined thresholds to classify tumours as HRD-positive or HRD-negative.

HRD Testing vs BRCA Testing

While both HRD testing and BRCA testing are related, they are not synonymous. BRCA testing focuses solely on identifying mutations in the BRCA1 and BRCA2 genes, either germline or somatic.

In contrast, HRD testing provides a broader evaluation of genomic instability and includes both BRCA and non-BRCA alterations affecting the HRR pathway.

It means HRD testing identifies a wider cohort of patients who might benefit from PARP inhibitors. All BRCA-mutated tumours are HRD-positive, but not all HRD-positive tumours have BRCA mutations.

Genomic Scars and Their Limitations

Genomic scars, LOH, TAI, and LST, reflect the cumulative DNA damage in a tumour over time due to deficient homologous recombination repair. Their combined measurement is known as the HRD score or Genomic Instability Score (GIS). The composite score is validated in breast, ovarian and prostate cancers.

However, genomic scars are static markers. They do not necessarily reflect the tumour’s current HRD status.

For example, a tumour initially classified as HRD-positive may acquire reversion mutations that restore HRR function, making PARPi ineffective. Hence, there’s a need for dynamic functional assays to complement static genomic assessments.

Basic Test on HRD: What’s Included?

• Germline and/or somatic BRCA1/2 mutation analysis
• Genomic instability scoring using LOH, TAI, and LST
• Optional analysis of additional HRR genes such as PALB2, RAD51, ATM, CHEK2, etc.

Depending on the test, samples can be derived from tumour tissue or a liquid biopsy. Technologies like Next-Generation Sequencing (NGS), Single Nucleotide Polymorphism (SNP) arrays, or even Optical Genome Mapping (OGM) are used for assessment.

Functional HRD Testing and Emerging Technologies

Emerging approaches like the RAD51 foci formation assay provide real-time evaluation of DNA repair competency. It directly measures HRR activity in tumour cells. If RAD51 foci are absent at DNA break sites, the tumour is likely HRD-positive. However, these tests face practical challenges due to technical variability, lack of standardisation and sample limitations.

Additionally, shallow whole-genome sequencing (sWGS) and transcriptomic approaches are being explored for dynamic HRD assessment. Algorithms such as HRDetect and CHORD use mutational signatures to predict HRD status with high accuracy. These models analyse patterns like COSMIC Signature 3 (SBS3) and others associated with BRCA dysfunction.

Clinical Utility and Decision-Making

HRD status informs multiple clinical decisions:

• Selection of patients for PARP inhibitor therapy
• Prediction of response to platinum-based chemotherapy
• Enrollment into HRD-enriched clinical trials

In ovarian cancer, HRD testing guides maintenance therapy with drugs like olaparib, niraparib, and rucaparib. In breast, pancreatic, and prostate cancers, the utility is expanding as new evidence emerges.

Challenges in HRD Testing

• Genetic variability within a tumour can lead to inconsistent or misleading HRD test results.
• Low tumour cell content in samples can reduce test sensitivity.
• Formalin-fixed, paraffin-embedded tissue samples may introduce sequencing errors.
• Over time, tumours can change, making earlier HRD results less reflective of the current status.
• Static genomic scars may produce false positives, while clonal heterogeneity can lead to false negatives.
• Different GIS (Genomic Instability Score) cutoffs across cancer types like breast and ovarian cancer complicate interpretation.
• Combining BRCA status, GIS, and mutational signatures leads to more accurate and context-specific diagnosis.
• Correctly distinguishing HRD-positive from HRP (HRD-proficient) tumours is crucial for selecting effective therapies.
• Assessing both the causes (mutations) and consequences (genomic scars) of HRD is needed.

Conclusion

As research advances, integrating AI-driven analytics and real-time data from functional assays may redefine how HRD status is monitored throughout treatment. Dynamic tracking can reveal shifts in tumour behaviour. Patients and clinicians should also stay informed about clinical trials exploring HRD-targeted therapies beyond PARP inhibitors. Accessing up-to-date resources and consulting with genomics specialists can ensure that treatment remains aligned with the evolving genetic profile of the tumour across different stages of care.

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