Uveal melanoma (UM) represents a prevalent form of intraocular malignant tumor in adults, accounting for approximately 5 % of all melanomas [1]. As the second most common subtype of melanoma, UM originates from melanocytes in three ocular tissues, namely the choroid (90 %), ciliary body (6 %), and iris (4 %) [2]. Typically affecting individuals at a median age of approximately 62 years, UM predominantly manifests unilaterally. Despite few treatments such as radiotherapy and laser therapy for primary UM, nearly half of the patients ultimately experience hematogenous dissemination leading to metastasis, primarily to the liver [3], [4]. Regrettably, the prognosis is bleak after metastasis, with a median survival rate of less than one year. Unfortunately, current therapeutic approaches in treating cutaneous melanoma have not yielded desirable outcomes in metastatic UM [1]. Consequently, there is an urgent need to develop novel therapeutic strategies to combat this disease.

According to recent studies, GNAQ and GNA11 mutations have been revealed as the primary drivers of UM [5], [6], [7], [8]. Approximately 85 % of UM patients carry either GNAQ or GNA11 mutations [1], while the remaining cases without GNAQ/11 mutations typically involve other gene mutations that activate the same signaling pathways as GNAQ/11. GNAQ and GNA11 encode Gαq and Gα11, which are G proteins that exhibit 90 % amino acid sequence homology. G proteins are heterotrimeric complexes composed of Gα (39‐52 kDa), Gβ (∼37 kDa), and Gγ (6‐9 kDa) subunits [9]. Functioning as molecular switches in GPCR signaling pathways, G proteins transmit upstream GPCR signals into the cells, modulating various essential physiological processes by regulating different second messengers. G proteins undergoes an “activation-inactivation” cycle across membranes in mediating signaling transduction. In the inactive state, the Gα subunit binds to GDP and forms stable heterotrimer with the Gβγ dimer. The activation of the GPCR induces conformational changes that facilitate GDP-GTP exchange in the nucleotide-binding pocket of the Gα subunit. Consequently, Gα-GTP and Gβγ dissociate and independently initiate downstream signaling pathways to regulate diverse physiological processes. Through its intrinsic GTP hydrolytic activity, Gα proteins gradually hydrolyze GTP to GDP, terminate the signaling and enable Gα-GDP to re-associate with Gβγ and the GPCR, thus complete a signal transmission cycle [10], [11]. However, activating mutations at positions Q209 (85 %) and R183 (5 %) are prevalent in GNAQ/GNA11, resulting in constitutive activation of Gαq/11 proteins due to impaired GTP hydrolysis [12]. These Gαq/11 proteins mutations lead to aberrant activation of downstream signaling pathways, including MAPK and PI3K/AKT/mTOR pathways, and play pivotal roles in UM development [13]. To date, most therapeutic interventions for UM were focused on inhibiting the individual downstream signaling pathways of mutant Gαq/11 proteins, but did not yield significant improvements in progression-free survival for UM patients [14], [15], [16]. This might be attributed to the fact that mutated Gαq/11 proteins typically activate multiple independent downstream signaling pathways, making it challenging to achieve the desired therapeutic effects by inhibiting a single pathway. Consequently, directly targeting the mutated Gαq/11 proteins represents a potential and promising therapeutic strategy for combating UM.

To date, no drugs targeting Gαq/11 proteins have been approved by the FDA. Only very few inhibitors, including YM-254890, FR900359, BIM-46174, and BIM-46187 (Fig. 1), have been reported to possess inhibitory activity against Gαq/11 proteins and demonstrate effectiveness against UM cells [17], [18], [19], [20]. Nevertheless, various limitations associated with these compounds hindered their further development. YM-254890 and FR900359, being natural products with intricate macrocyclic structures, present significant challenges in synthesis despite the successful establishment of their total synthetic routes by our group in 2016 [21], [22], [23], [24]. The limitations imposed by their challenging synthesis on a large scale restrict their pharmaceutical development. BIM-46174 and BIM-46187, small molecular pan inhibitors of Gαq/11 proteins with imidazo[1,2-α]pyrazine scaffold, exhibit high toxicity and instability. Consequently, the above compounds are primarily used as pharmacological tools to explore the complexity and diversity of GPCR pharmacology, as well as its downstream G proteins-mediated signaling pathways [25]. In recent years, our group have made significant progress in developing several novel small molecule Gαq/11 proteins inhibitors with imidazo[1,2-α]pyrazine skeleton, including GQ127, GQ262, and GQ352, optimized from the BIM-46174 scaffold. These compounds demonstrate moderate inhibitory activity against Gαq/11 proteins in vitro and exhibit robust anti-UM efficacy in vivo, indicating the potential of directly targeting Gαq/11 proteins as a viable strategy for combating UM [26], [27], [28]. Therefore, the development of small molecule inhibitors for Gαq/11 proteins holds great significance and prospect in providing novel treatments for UM.

In this study, we successfully designed and identified a potent small molecule inhibitor targeting Gαq/11 proteins, designated as F33, which possesses quinazoline scaffold. The discovery of F33 was achieved through a systematic screening of the existing compound libraries, followed by subsequent structural modifications. Comprehensive bio-activity evaluations demonstrated that F33 exhibits remarkable inhibitory activity against Gαq/11 proteins and displays significant anti-UM efficacy in vitro. Additionally, we conducted a series of target verification experiments to validate the target of F33, revealing that F33 directly binds to Gαq/11 proteins, and exerts the anti-UM efficacy through inhibiting Gαq/11 proteins.