Making a mouse out of a molehill: How precision modeling repurposes drugs for congenital giant nevi

Abstract

Patients with congenital giant nevi (CGN), which can compromise quality of life and progress to melanoma, have limited treatment options. Choi et al. demonstrate that topical application of a pro-inflammatory hapten for alopecia treatment (SADBE) caused nevus regression and prevented melanoma in an Nras mouse CGN model. Their results demonstrate the promise of repurposing drugs through precision modeling.

We all occasionally read a story about someone who was bullied because of large, “brown birthmarks” covering their body that gave them cruel nicknames. These are phenotypically heterogeneous congenital giant nevi (CGN), which can grow more than 40 cm in diameter in adults and occur in one in every 20,000 newborns. CGN originate from a somatic mutation in melanocytes during embryonic development, typically activating the NRAS oncogene, resulting in extended pigmented areas on the neonatal skin. When the mutation occurs in multipotent progenitor cells multiple CGN can appear, and other organs can be affected, such as neurocutaneous melanosis in the central nervous system. These lesions can develop into melanoma, which can be aggressive, resistant to therapy, and often deadly. The risk for melanoma is estimated at 10–15% for giant nevi (>40 cm) and 2.5–8% for large nevi (>20cm) and rises as nevus size increases [1].

Surgical excision is the current standard-of-care for CGN. Unfortunately, multiple, serial excisions may be required with a possibility of infection, scarring, excruciating pain, and discomfort in everyday life. Other approaches are application of epithelial cultured autografts after curettage of CGN and pigment-specific laser treatment. However, re-pigmentation can occur, rendering unclear whether these procedures can prevent melanomagenesis. Importantly, they are not widely available to CGN patients in many low-income regions, reflecting public health disparities. An effective, low-cost, non-invasive treatment is urgently demanded for CGN patients worldwide. Ideally, it should have proven efficacy for the prevention of melanoma.

Sequencing studies have found NRAS hotspot mutations, predominantly either Q61K/R, in 80% - 95% of CGN cases, whereas smaller congenital nevi mostly carry BRAF^V600E mutations [2]. With no other known recurrent mutations to date, it is most likely a single-driver disease. In fact, it has been shown that an Nras mutation alone is sufficient to induce highly pigmented nevi in genetically engineered mouse (GEM) models [3]. Recognizing this connection, Choi and colleagues (2022) designed and generated several GEM models in which expression of Nras^Q61R was inducibly or constitutively targeted to melanocytes [4]. Triggering Nras^Q61R expression in melanocytes either during embryonic development or in neonatal mice resulted in highly pigmented, spreading nevi. Interestingly, like melanosis in the leptomeninges of some CGN patients, ectopic melanocytes were found in the mouse meninges. After proliferating to form nevi, the melanocytes entered the senescence phase. As in human CGN, some nevi escaped senescence in a loss-of-heterozygosity manner and evolved into melanoma, which eventually killed the mice.

To demonstrate the relevance and value of their models to human CGN, the authors attempted to identify low-cost therapies for local treatment. As proof of principle, inhibitors of pathways activated by Nras, including MEK and/or PI3K inhibitors, as well as a c-KIT inhibitor, showed efficacy resulting in depigmentation. Ingeniously, the authors decided to test a pro-inflammatory hapten, squaric acid dibutylester (SADBE), used clinically for topical therapy of alopecia areata, where it can induce vitiligo as a side effect. When SADBE alone was applied topically to the pigmented area, it caused regression of the mouse nevi while effectively preventing melanomagenesis. Interestingly, such an effect was mediated by the influx of innate proinflammatory M1 macrophages, rather than adaptive immune cells (e.g., T or B cells). GSEA pathway analysis demonstrated that TNF-α was upregulated after SADBE treatment, providing a possible mechanism for melanocyte/nevocyte reduction through enhanced macrophage-based cytotoxic activity or apoptosis (Figure 1). The relevance of SADBE was confirmed through preclinical studies of human congenital nevus xenografts, which exhibited macrophage accumulation and partial but significant nevus cell clearance upon treatment.

Figure 1. From human congenital nevi to murine congenital nevus models:

CGN as a single-driver disease was modeled in mice by inducing Nras expression in melanocytes of fetal/neonatal mice. Preclinical studies using the mouse model demonstrated that, SADBE, a hapten eliciting a contact hypersensitivity-like response, resulted in the recruitment of M1 macrophages, which could in turn elevate TNFα levels to cause cytotoxic activity, reducing melanocytes/nevocytes by apoptosis. SADBE treatment alone could also prevent melanomagenesis.

Although further investigations are required for advancement to clinical use, several features of SADBE identified in this study have demonstrated notable potential as an effective CGN treatment. First, it recruits innate immunity against nevi, which can lead to better clearance of mutant melanocytes, thus preventing recurrence and progression to melanoma. Second, topical treatment can be applied repetitively and sequentially for a longer duration until it reaches almost all target cells, sparing patients from expensive and often ineffective surgery/laser procedures. Third, it is a repurposed (off-label) drug whose safety and production were proven in alopecia areata and warts and can be prescribed by clinicians once the new indication is approved. A clinical study will be needed to unravel the effect.

The authors’ strategy of asking clinically relevant questions in precision animal models that could facilitate target validation and drug discovery has proven to be an attractive and productive research approach for solving the clinical puzzle of CGN. The authors precisely modeled CGN by turning on Nras^Q61R in melanocytes during the matched developmental stage (fetal/neonatal), resulting in relevant disease phenotypes and pathology. Accordingly, several studies have shown that CGN patient-derived xenografts respond to MEK inhibitors [5]; in one study a young patient with CGN carrying a MEK-activating fusion gene responded to Trametinib, significantly relieving symptoms [6]. Therefore, mouse models with relevant genotypes, phenotypes, and therapeutic responses are highly desirable for screening and repurposing drugs.

With these promises, several questions concerning the clinical application of SADBE alone or in combination remain. (1) Could CGN driven by other mutations respond to SADBE? (2) As an immunogen and contact sensitizer, could SADBE generate severe side effects? (3) Could long-term use result in resistance? (4) Could SADBE be used to treat cutaneous dysplastic nevi and prevent melanoma, or after removal of early-stage melanoma to prevent recurrence/metastasis? The answers to these questions require further research into the immune environment of the skin. In fact, cutaneous macrophages are known for their critical role in skin homeostasis [7] and unique responses following stimuli [8]. Nevertheless, the promising results of Choi and colleagues [4], together with their insights into skin immunity, are indications that targeted therapies and immunogen therapy may soon enter standard care practices for the management of CGN. They also have important implications for the development of therapeutic procedures to improve patient quality of life and public health disparities.