Hypoxia is a critical factor affecting tissue homeostasis that dramatically alters the tumor microenvironment (TME) through genetic, metabolic, and structural changes, promoting tumor survival and proliferation. Hypoxia-inducible factor-1α (HIF-1α) plays a central role in this process by regulating hundreds of genes involved in the processes of tumorigenesis, angiogenesis, metabolic reprogramming, and immune evasion. This review provides a comprehensive examination of the role of HIF-1α in hypoxia and how hypoxia weakens intercellular junctions—including gap junctions, adherens junctions, tight junctions, and desmosomes. The disruption of gap junctions decreases intercellular communication, which alters signal transduction cascades and tumor suppressive properties. Adherens junctions are comprised of proteins that characterize the tissues and link cells to the actin cytoskeleton, whereby their disruption promotes the epithelial-to-mesenchymal transition (EMT). Under hypoxic conditions, the tight junction proteins are dysregulated, altering paracellular transport and cell polarity. In addition, desmosomes provide linkage to intermediate filaments, and hypoxia compromises tissue integrity. Collectively, the influence of hypoxia on cellular junctions promotes tumorigenesis through reducing cell communication, cytoskeletal interactions, and altering signaling pathways. Activation of matrix metalloproteinases (MMPs) further degrades the extracellular matrix and enhances tumor invasion and metastasis. This process also involves hypoxia-induced angiogenesis, regulated by HIF-1α. A comprehensive understanding of the mechanisms of hypoxia-driven tumor adaptation is essential for developing effective therapeutic strategies. Furthermore, this review examines current treatments aimed at targeting HIF-1α and explores future directions to enhance treatment efficacy and improve patient outcomes.

Although melanoma accounts for only 5.3% of skin cancer, it results in >75% of skin-cancer-related deaths. To avoid disfiguring surgeries on the head and neck associated with surgical excision, there is a clear unmet need for other strategies to selectively remove cutaneous melanoma lesions. Mohs surgery is the current treatment for cutaneous melanoma lesions and squamous and basal cell carcinoma. While Mohs surgery is an effective way to remove melanomas in situ, normal tissue is also excised to achieve histologically negative margins. This paper describes a novel combination therapy of nonthermal plasma (NTP) which emits a multitude of reactive oxygen species (ROS) and the injection of a pharmaceutical agent. We have shown that the effects of NTP are augmented by the DNA-damaging prodrug, tirapazamine (TPZ), which becomes a free radical only in conditions of hypoxemia, which is often enhanced in the tumor microenvironment. In this study, we demonstrate the efficacy of the combination therapy through experiments with B16-F10 and 1205 Lu metastatic melanoma cells both in vitro and in vivo. We also show the safety parameters of the therapy with no significant effects of the therapy when applied to porcine skin. We show the need for the intratumor delivery of TPZ in combination with the surface treatment of NTP and present a model of a medical device to deliver this combination therapy. The importance of functional gap junctions is indicated as a mechanism to promote the therapeutic effect. Collectively, the data support a novel therapeutic combination to treat melanoma and the development of a medical device to deliver the treatment in situ.

Hypoxia is a critical factor driving cellular adaptation, reshaping the tumor microenvironment (TME) through genetic, metabolic, and structural changes that promote tumor survival and proliferation. Hypoxia-inducible factor-1α (HIF-1a) plays a central role in this process by regulating angiogenesis, metabolic reprogramming, and immune evasion through numerous pathways, ultimately enabling tumor cells to survive, invade, and develop therapy resistance. This article examines how hypoxia weakens intercellular junctions—including gap junctions, adherens junctions, tight junctions, and desmosomes—thereby disrupting intercellular contacts and communication to promote invasion. Additionally, it explores the activation of matrix metalloproteinases (MMPs) under hypoxia, which further degrades the extracellular matrix and enhances the invasion. Hypoxia-induced metabolic and genetic changes induce the epithelial-to-mesenchymal transition (EMT), facilitating tumor cell invasion and metastasis. This process also involves hypoxia-induced angiogenesis, regulated by HIF-1a. A comprehensive understanding of the mechanisms of hypoxia-driven tumor adaptation is essential for developing effective therapeutic strategies. This article explores the molecular pathways that regulate angiogenesis, metabolic reprogramming, cell migration and immune evasion, illustrating how changes in the tumor microenvironment ultimately lead to metastasis. Given the role of hypoxia in treatment resistance, targeting HIF-1α and its downstream effectors could potentially enhance treatment efficacy and improve patient outcomes.

Pancreatic adenocarcinoma remains a highly lethal malignancy with limited effective treatment options, largely due to late-stage detection and rapid progression to metastatic disease. Therapeutic strategies capable of targeting both pre-metastatic and metastatic tumors are critically needed. In this study, we evaluated a combination therapy consisting of non-thermal plasma (NTP), a generator of reactive oxygen and nitrogen species, and the hypoxia-activated prodrug tirapazamine (TPZ). We further investigated whether sulforaphane (SF), a bioactive phytochemical, could further enhance therapeutic efficacy. NTP and TPZ produced strong cytotoxic effects as single agents and demonstrated additive to synergistic activity when combined, reducing viability by 87% in pre-metastatic BxPC-3 cells and achieving near-complete elimination of metastatic AsPC-1 cells. The addition of sulforaphane (10 µM) further enhanced cytotoxicity across all treatment conditions, with Bliss independence analysis indicating additive to synergistic interactions depending on cell line and treatment combination. Sulforaphane-mediated enhancement occurred without restoration of connexin 43 expression or coordinated reversal of epithelial-to-mesenchymal transition markers, and treatments did not induce N-cadherin upregulation or suggest acquisition of invasive characteristics. Together, these findings support NTP + TPZ as a potent combinatorial strategy for pancreatic adenocarcinoma and identify sulforaphane as an effective adjunct that enhances cytotoxic efficacy through mechanisms that remain to be fully elucidated.

Molecular docking was used to investigate the interactions between tirapazamine (a bioreductive anticancer drug) and bovine serum albumin (BSA), with emphasis on binding site localization and geometry. Docking simulations were performed using SwissDock with the AutoDock Vina 1.2.0 scoring engine. Warfarin (a canonical Sudlow Site I ligand) was docked under the same conditions as a reference to provide a relative comparison of binding orientation within Subdomain IIA. Docking results indicate that tirapazamine occupies a peripheral entrance-region position within Subdomain IIA, with limited polar interactions and substantial solvent exposure. In contrast, warfarin adopts a more enclosed binding orientation consistent with canonical Site I burial. UV–visible measurements across three independent titration experiments yielded apparent association constants of 2.30 ± 1.05 × 10⁴ M⁻¹ (mean ± SD), consistent with a weak interaction regime and supporting the docking results. Geometric analysis further shows that tirapazamine maintains greater minimum ligand–residue distances from core Site I residues than warfarin, consistent with reduced burial. This interpretation is further supported by the physicochemical profile of tirapazamine, which is characterized by low lipophilicity and high polarity, properties that are incompatible with deep binding in hydrophobic pockets (as seen with warfarin). Short-timescale molecular dynamics simulations (2 ns, triplicate) confirmed the stability of the docked poses, with tirapazamine exhibiting markedly lower ligand RMSD variability (0.247 ± 0.002 Å) than warfarin (0.578 ± 0.163 Å), consistent with a stable peripheral binding geometry. Together, these findings support a model in which tirapazamine associates weakly and peripherally with serum albumin, consistent with non-canonical Site I binding.