Amal M. El-Naggar

The canonical transsulfuration (TSS) pathway enzymes cystathionine β-synthase (CBS) and cystathionine γ-lyase (CTH) are traditionally recognized for their roles in the sequential conversion of homocysteine to cysteine and in endogenous hydrogen sulfide (H₂S) production. Increasing evidence, however, suggests that these enzymes may also exhibit non-canonical (“moonlighting”) functions that extend beyond metabolic regulation. In this review, we evaluate the hypothesis that CTH may participate in translational regulation, particularly in the control of hypoxia-inducible factor-1α (HIF-1α) expression in clear cell ovarian carcinoma (CCOC). We first highlight limitations of the prevailing H₂S- and cysteine-centric view of the TSS pathway, which may not fully explain emerging context-dependent functions of CTH in cancer biology. Current evidence suggests that CTH enhances HIF-1α protein expression through mechanisms independent of transcription, protein stability, or H₂S production, implicating a potential role in translational regulation, although direct mechanistic evidence remains limited. To critically evaluate this emerging hypothesis, we categorize evidence according to its level of experimental support, ranging from direct experimental evidence to indirect mechanistic observations and computational predictions. Within this framework, we examine three non-mutually exclusive models: (1) regulation through PI3K/AKT/mTOR-dependent translational signaling; (2) modulation of translational control through interaction with translation-associated proteins and RNA-binding proteins (RBPs) involved in HIF1A mRNA regulation; and (3) the more speculative possibility of direct interaction between CTH and HIF1A mRNA. Collectively, these observations support a model in which CTH contributes to selective translational regulation beyond its canonical metabolic functions, potentially linking sulfur metabolism to stress-adaptive gene expression in cancer.

Clear cell ovarian carcinoma (CCOC) is a rare, aggressive ovarian cancer subtype, comprising approximately 12% of epithelial ovarian carcinoma (EOC) cases

DICER1 is an RNase III enzyme essential for miRNA biogenesis through cleaving precursor-miRNA hairpins. Germline loss-of-function DICER1 mutations underline the development of DICER1 syndrome, a rare genetic disorder that predisposes children to cancer development in organs such as lung, gynecologic tract, kidney, and brain. Unlike classical tumor suppressors, the somatic “second hit” in DICER1 syndrome–associated cancers does not fully inactivate DICER1 but impairs its RNase IIIb activity only, suggesting a noncanonical two-hit hypothesis. Here, we developed a genetically engineered conditional compound heterozygous Dicer1 mutant mouse strain that fully recapitulates the biallelic DICER1 mutations in DICER1 syndrome–associated human cancers. Crossing this tool strain with tissue-specific Cre strains that activate Dicer1 mutations in gynecologic tract cells at two distinct developmental stages revealed that embryonic biallelic Dicer1 mutations caused infertility in females by disrupting oviduct and endometrium development and ultimately drove cancer development. These multicystic tubal and intrauterine tumors histologically resembled a subset of DICER1 syndrome–associated human cancers. Molecular analysis uncovered accumulation of additional oncogenic events (e.g., aberrant p53 expression, Kras mutation, and Myc activation) in murine Dicer1 mutant tumors and validated miRNA biogenesis defects in 5P miRNA strand production, of which, loss of let-7 family miRNAs was identified as a putative key player in transcriptomic rewiring and tumor development. Thus, this DICER1 syndrome–associated cancer model recapitulates the biology of human cancer and provides a unique tool for future investigation and therapeutic development.Significance:Generation of a Dicer1 mutant mouse model establishes the oncogenicity of missense mutations in the DICER1 RNase IIIb domain and provides a faithful model of DICER1 syndrome–associated cancer for further investigation.

Clear cell ovarian cancer (CCOC) is the second most common ovarian cancer subtype, accounting for 5%–11% of ovarian cancers in North America. Late‐stage CCOC is associated with a worse prognosis compared to other ovarian cancer histotypes, a challenge that has seen limited progress in recent decades. CCOC typically originates within the toxic microenvironment of endometriotic ovarian cysts and is characterized by its intrinsic chemoresistance, a strong hypoxic signature, and abundant expression of cystathionine gamma‐lyase (CTH). CTH is a key enzyme in the transsulfuration pathway and serves as a marker of ciliated cells derived from the Müllerian tract. CTH plays a pivotal role in de novo cysteine synthesis, which is essential for glutathione (GSH) production and redox homeostasis. Using an array of molecular tools and cancer models, including in vivo studies, we demonstrated that CTH expression was induced under various stress conditions, such as exposure to endometriotic cyst content and hypoxia. This induction enables cell survival and creates a differentiation state manifested by CCOC that potentiates tumor progression and metastasis. In addition to regulating redox homeostasis, CTH enhances hypoxia inducible factor 1‐alpha (HIF1α) expression, independently of hydrogen sulfide (H₂S) production. Re‐expression of HIF1α in CTH KO cells fully restored metastatic capacity in in vivo models. Co‐expression of CTH and HIF1α proteins was also observed in human CCOC samples. Importantly, targeting CTH in CCOC significantly reduced its metastatic potential in in vivo models and enhanced sensitivity to chemotherapy. These findings underscore that CTH is both a defining feature of CCOC and a promising therapeutic target, not only for CCOC patients but also for those with other CTH‐expressing cancers. © 2025 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Supplementary Tables from Bi-allelic Dicer1 mutations in the gynecologic tract of mice drive lineage-specific development of DICER1 syndrome-associated cancer

Supplementary Tables from Bi-allelic Dicer1 mutations in the gynecologic tract of mice drive lineage-specific development of DICER1 syndrome-associated cancer

DICER1 is an RNase III enzyme essential for microRNA (miRNA) biogenesis through cleaving pre-miRNA hairpins. Germline loss-of-function DICER1 mutations underlie the development of DICER1 syndrome, a rare genetic disorder that predisposes children to cancer development in organs such as lung, gynecologic tract, kidney, and brain. Unlike classical tumor suppressors, the somatic “second hit” in DICER1 syndrome-associated cancers does not fully inactivate DICER1 but impairs its RNase IIIb activity only, suggesting a noncanonical two-hit hypothesis. Here, we developed a genetically engineered conditional compound heterozygous Dicer1 mutant mouse strain that fully recapitulates the bi-allelic DICER1 mutations in DICER1 syndrome-associated human cancers. Crossing this tool strain with tissue-specific Cre strains that activate Dicer1 mutations in gynecologic tract cells at two distinct developmental stages revealed that embryonic bi-allelic Dicer1 mutations caused infertility in females by disrupting oviduct and endometrium development and ultimately drove cancer development. These multicystic tubal and intra-uterine tumors histologically resembled a subset of DICER1 syndrome-associated human cancers. Molecular analysis uncovered accumulation of additional oncogenic events (e.g. aberrant p53 expression, Kras mutation, and Myc activation) in murine Dicer1 mutant tumors and validated miRNA biogenesis defects in 5P miRNA strand production, of which loss of let-7 family miRNAs was identified as a putative key player in transcriptomic rewiring and tumor development. Thus, this DICER1 syndrome-associated cancer model recapitulates the biology of human cancer and provides a unique tool for future investigation and therapeutic development.