Immune checkpoint inhibitors and BRAF-targeted therapies have redefined the management of malignant melanoma. Overall survival (OS) has increased significantly with immunotherapy, with >60 months reported.1 Stabilization of the survival curve in approximately one-third of patients is further suggestive of durable responses. Likewise, BRAF and MEK inhibitors targeting melanomas harboring BRAF V600E mutations have demonstrated excellent, although short-lived, responses.2–5 Despite these advances, a significant number of patients develop progressive disease, intolerable adverse side-effects, or become resistant to targeted therapies. There remains a need for viable alternative treatments for these patients with treatment-refractory melanoma.
PARP inhibitors (PARPi) have emerged as a promising treatment in cancers with alterations in homologous recombination repair (HRR) genes, one of the DNA damage repair genes (also termed HR-DDR).6 Cancer cells with homologous recombination deficiency (HRD) treated with PARPi are unable to repair double-strand breaks, leading to cell death by synthetic lethality. The prevalence of detected HRR mutations is common in melanoma, with frequencies ranging from 18.1% to upwards of 41%.7,8 In general, patients with metastatic melanoma without selection for PARPi sensitivity have only modest responses to rucaparib, with only 17.4% having a partial response and 17.4% having stable disease (34.8% disease control rate) when combined with chemotherapy.9 However, there are multiple reports of patients with HRR mutations responding to PARPi.10–12 Taken together, PARPi may be a promising treatment in immunotherapy-refractory melanoma, but patient selection will be critical.
PARPi response prediction necessitates an accurate assessment of HRD status. HRD score is not uniformly calculated but rather derived as a composite score from either a single or combination of the genomic findings that reflect genomic instability, genome-wide loss of heterozygosity (GW-LOH),13 telomeric allelic imbalance (TAI),14 and/or large-scale state transitions (LST).15 This composite HRD score derived from all 3 measures (GW-LOH, TAI, and LST) has been shown to correlate with response to PARPi in ovarian cancers.16 Direct gene testing may underappreciate secondary factors leading to genomic instability, such as epigenetic changes. Furthermore, various causes of genomic instability may have differential effects on response to treatment. A small but thorough study in high-grade ovarian cancer suggests that high HRD score through genetic alterations holds greater prognostic significance than high HRD score through epigenetic changes,17 but this finding has not been validated in other cancers. It remains unclear whether direct gene testing for common HRR-associated genes alone is sufficient to assess HRD status and potential treatment response to PARPi in melanoma.
Here, we examine the relationship between PARPi response and HRD scores derived from GW-LOH in 4 patients with metastatic melanoma, expanding on earlier reports, including our 2 prior reports as well as another study showing a profound and durable response in a patient with both HRR mutations and elevated HRD score.12,18,19 We next examined the prevalence of homologous recombination defects in 933 patients with melanoma derived from both GW-LOH and traditional direct gene testing to provide rationale for targeted therapy with a PARPi-based regimen based on HRD-LOH scoring.
Case 1
An 89-year-old female with a history of rheumatoid arthritis on methotrexate initially presented with rectal bleeding and a palpable anal mass. Workup revealed mucosal melanoma, PD-L1 expression of 3% to 5%, and BRAF wild-type tumor cells. Imaging revealed involvement limited to the lymph nodes in the right groin. She underwent partial thickness transanal excision of the tumor and lymph node dissection. Pathology demonstrated a 3.3-cm malignant melanoma with 8 mm of tumor thickness and lymphovascular space invasion extending to the peripheral margin with 3 of the 9 excised lymph nodes positive for metastatic disease.
Initial treatment included 30 Gy of hypofractionated radiotherapy followed by adjuvant immunotherapy with nivolumab, 480 mg every 4 weeks. After 4 cycles, restaging scans and biopsy of a left gluteal nodule revealed disease recurrence. Nivolumab was discontinued after 6 cycles. The recurrent tumor was biopsied with HRD testing (Tempus) revealing elevated HRD-LOH of 43.9% (supplemental eTable 1, available with this article at JNCCN.org).
Given the elevated HRD score, olaparib was started at 200-mg tablets twice daily. Unfortunately, prolonged recovery from an unrelated mechanical fall interrupted outpatient follow-up, with family reporting poor compliance with olaparib during this period. Restaging imaging revealed progression of metastatic disease. After careful discussion with the patient, olaparib was re-trialed under caregiver supervision. She tolerated therapy well with no adverse events. Restaging imaging after 5 months showed nearly complete resolution of previously FDG-avid left inguinal adenopathy (Figure 1A–C). The follow-up liquid biopsy using Signatera (Natera, Inc.) showed clearance of tumor-informed circulating tumor DNA (ctDNA) (Figure 1D). Subsequent patient care was plagued with intermittent drug compliance, including one documented time point that correlated with subsequent increase in ctDNA.
PET/CT showing FDG avidity of metastatic melanoma at (A) recurrence, (B) 4 months after olaparib therapy initiation, and (C) noncompliance on olaparib therapy. Liquid biopsy with (D) MTMs detected in the plasma shown as a function of days on treatment with olaparib, with red arrow indicating documented medication noncompliance.
Abbreviation: MTM, mean tumor molecule.
Citation: Journal of the National Comprehensive Cancer Network 21, 7; 10.6004/jnccn.2022.7102
Case 2
A 61-year-old female was diagnosed with a stage IIC (T4N0M0) malignant melanoma of the scalp that grew quickly over 5 months. She underwent wide local excision followed by adjuvant immunotherapy with pembrolizumab, 200 mg every 3 weeks. Restaging scans 3 months later revealed numerous new hypermetabolic soft tissue nodules and lymph nodes consistent with disease progression. Pembrolizumab monotherapy was discontinued and combination ipilimumab and nivolumab started. Unfortunately, after only 1 cycle she developed grade 2 colitis requiring hospitalization, steroids, and ipilimumab discontinuation. She then developed grade 2 pneumonitis after just 1 cycle of nivolumab monotherapy, forcing discontinuation of immune checkpoint inhibitors.
Next-generation sequencing (NGS) and HRD testing (Tempus) revealed elevated HRD-LOH at 57.7% (supplemental eTable 1), prompting treatment with olaparib, 300-mg tablets twice daily. Liquid biopsy with Signatera testing showed clearance of ctDNA. Restaging scans 1 week later showed mixed response, with complete response at 6 months (Figure 2) and continued response at 10 months.
PET/CT showing FDG avidity of metastatic melanoma at (A) recurrence, (B) 1 week after olaparib therapy, and (C) 6 months after olaparib therapy. Liquid biopsy with (D) MTMs detected in the plasma is shown as a function of days on treatment with olaparib.
Abbreviation: MTM, mean tumor molecule.
Citation: Journal of the National Comprehensive Cancer Network 21, 7; 10.6004/jnccn.2022.7102
Case 3
A 38-year-old male was diagnosed with stage IIIC (pT3a pN2 cM0) melanoma, BRAF wild-type, which was treated with wide local excision and adjuvant ipilimumab.19 Unfortunately, he developed grade 4 colitis after 2 cycles requiring hospitalization, steroids, and immunotherapy discontinuation. After 3.5 years of surveillance an enlarged paratracheal lymph node was found on imaging. Biopsy revealed metastatic melanoma (supplemental eTable 1). Tempus HRD testing revealed GW-LOH of 32.9%. He was started on combination nivolumab (480 mg intravenously every 4 weeks) and olaparib (300-mg tablet twice daily) with interval response after 2 months and clearance of ctDNA. Grade 3 hepatitis prior to cycle 3 prompted steroids and discontinuation of cancer therapy. Repeat CT after 18 months of treatment discontinuation demonstrated no active disease (Figure 3).
CT imaging of metastatic melanoma at (A) recurrence, (B) 2 months after initiating combination nivolumab and olaparib, and (C) 18 months after treatment discontinuation.
Citation: Journal of the National Comprehensive Cancer Network 21, 7; 10.6004/jnccn.2022.7102
Case 4
A 64-year-old male was diagnosed with metastatic melanoma with oligometastatic brain lesion when he presented after several weeks of neurologic deficits. He was initially treated with craniotomy with gross total resection followed by gamma knife surgery. After completion of radiation, he was started on combination ipilimumab and nivolumab, which he tolerated for 3 cycles before developing grade 3 pneumonitis, requiring discontinuation of therapy. He was then found to have recurrent brain metastasis, for which he received gamma knife surgery, as well as liver metastasis. Sequencing showed a germline mutation in MUTYH (p.G396D), and tumor mutations in NRAS (pQ61K, variant allele fraction 40.6%) with tumor mutational burden at 22.6 mutations per megabase (mut/Mb), and an HRD-LOH score of 28% (Tempus). Given the high LOH score, he was started on olaparib, 300-mg tablets twice daily. Restaging scans 2 months after starting therapy showed partial response to therapy in both liver and brain lesions.
HRD Incidence
We next compared the incidence of HRD in 933 patients with melanoma (Table 1). HRD was determined in 3 ways: (1) a previous version of Tempus HRD (referred to here as Tempus HRDv1),20,21 using a GW-LOH threshold of 33% or biallelic BRCA calculated from Tempus xT NGS DNA data22–24; (2) arbitrarily dropping the GW-LOH threshold to 25%; and (3) the presence of a pathogenic or likely pathogenic somatic mutation in 1 of the HRR genes.
Core Clinical Characteristics of Melanoma Cases in Tempus National Database
Records of patients with melanoma were selected from the Tempus database,20,25 which includes longitudinal structured and unstructured deidentified data from geographically diverse oncology practices, and relevant inclusion criteria were applied. Inclusion criteria included a histologic diagnosis of melanoma with available sequencing data.
In this cohort, 8.5% (n=79) of samples harbored HRD positivity derived from Tempus HRDv1, 28% (n=261) by dropping the GW-LOH threshold to 25%, and 9.4% (n=88) by considering pathogenic/likely pathogenic somatic mutations in HRR (supplemental eTable 2). However, only 14.7% (n=13) of samples harboring HRD by somatic mutations in HRR were HRD-positive by Tempus HRDv1. Most HRD calls from both Tempus HRDv1 and GW-LOH ≥25% did not harbor somatic mutations in HRR (84% [66/79] and 91% [238/261], respectively) (supplemental eFigure 1).
Discussion
This case report reviewed 4 patients with recurrent melanoma with a GW-LOH result ≥25%. All demonstrated favorable response to PARPi despite none having an identifiable HRR gene mutation. We further showed that the proportion of patients with HRD-high melanoma was almost 9% according to Tempus HRDv1. Decreasing the GW-LOH threshold to 25% increased the prevalence considerably to 28%, although the sensitivity and positive predictive value of this threshold is unknown.
Our study has several shortcomings, including a lower percentage of patients who harbored an identifiable HRR mutation (9.4%) compared with other studies. Kim et al26 reported 33.5% of patients with HRR alteration in a Foundation Medicine cohort, 41% in a cBioPortal cohort, and 21.4% (18/84 patients) in their institutional cohort. In a pan-cancer analysis, Heeke et al7 reported that 18.1% of patients with melanoma had a detectable HRR mutation. These discordances are possibly related to differences in the HRR mutational panel used to identify these patients, demonstrate the limitations of retrospective methodologies, and highlight the fact that additional prospective studies are needed. Additionally, the first 2 cases in this report feature somatic mutations not usually associated with melanomas, and it is unclear whether they represent atypical cases of melanoma or instead demonstrate passenger mutations detected on the unbiased next-generation sequencing. Furthermore, although we advance the use of GW-LOH as a biomarker for PARPi responsiveness in patients with melanoma in our study, it underscores the sensitivity or specificity of other DNA-based HRD biomarkers, such as TAI and LST. More recently, functional tests analyzing the amount of DNA repair protein accumulation at double-strand DNA breakage sites, including RAD51 and γH2AX,27 have shown relevance in melanoma cell lines,28 suggesting yet another biomarker for HRD assessment. Ultimately, more studies comparing the prevalence of these different HRD biomarkers with correspondence to PARPi response in a disease-specific setting will be critical to determining which HRD assessment would be most appropriate for patients with melanoma.
Our findings, although limited, combined with the lack of association between presence of a known HRR gene and GW-LOH as a marker of HRD, suggest that a traditional HRD gene panel would not identify these patients. Taken together, GW-LOH, even in the absence of detectable mutations in HRR, may be a promising biomarker for response to PARPi in patients with melanoma.
Several factors may contribute to HRD status or response to PARPi in the absence of a mutation in a traditional HRR gene. These include both epigenetic alterations such as DNA methylation29,30 and activity in nontraditional HRR pathways such as TP53 expression signature.31 Targeted gene panels may also underappreciate low(er)-frequency single loci or combinations of mutations in genes of canonical HRR pathways that in concert may affect global HRR status. Likewise, still-unknown pathways may exist that are key for HRR under certain circumstances. This collectively results in a significant undersampling of HRD status when assaying a limited set of mutations directly, favoring an HRD-LOH as a more sensitive approach.
An outstanding issue remains the lack of consensus on GW-LOH approach and cutoffs. This issue itself is multifactorial. First, calculations for HRD status vary with several accepted methods.13–15,17 Second, cutoffs associated with potential response to PARPi therapy vary by cancer type, with cutoffs not well established for certain cancers. Here we built on a reliable, validated commercial assay available through Tempus. Tempus HRDv1 issues specific recommendations for well-established cancer types, including ovarian (≥25%), breast (≥29%), and pancreatic cancer (≥28%), and a basket threshold for all other cancer types at a GW-LOH of ≥33%. This general cutoff is likely conservative in melanoma given that 2 of the patients with HRD scores of 28% and 32.9% exhibited a response to PARPi. Furthermore, the presence of a notable cluster of patients with melanoma between 0.25 and 0.33 (supplemental eFigure 2) suggests this intermediate range HRD score is likely a frequent occurrence in melanoma. Hence, there is a need for further research to establish an LOH threshold specific to melanoma.
There is a growing body of evidence showing that combining PARPi with immunotherapy has the potential for synergistic effect in patients with cancer. PARPi can enhance tumor cell–intrinsic immunity in DNA damage response–deficient tumors by generating cytoplasmic chromatin fragments that activate the cGAS/STING pathway, downstream IFN signaling, and cytokine production.32 Deficiencies in DNA damage response pathways can result in increased tumor mutational burden,33,34 which has been correlated with better outcomes with immunotherapy treatment in melanoma.35 Independent of BRCA mutational status, PARPi can upregulate PD-L1 expression in breast cancer models in vitro by inactivating glycogen synthase kinase 3β, which can regulate expression of PD-L1 and sensitizing tumor cells to T-cell–mediated killing.36 Urothelial carcinomas that have a known mutation in DNA damage response genes are more likely to have a greater response to checkpoint inhibitors,37 suggesting that synthetic lethality with PARPi may generate even more neoantigen formation to further augment response to checkpoint inhibitors. This has led to multiple clinical trials examining the combination of PARPi and immunotherapy in different cancers,38,39 including several clinical trials that are ongoing to evaluate the efficacy of PARPi and immune checkpoint blockade in metastatic melanoma with HRD gene mutations (ClinicalTrials.gov identifiers: NCT04633902 and NCT04187833).
Conclusions
HRD-LOH is a frequent finding in melanoma, and more common than HRD derived from directly assaying canonical HRR mutations. This comparatively sensitive biomarker may identify a significant number of previously unknown patients who may experience a response to PARPi. Further studies are needed to determine the optimal cutoff and paradigm for integration with established immunotherapy protocols.
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