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Cheng et al. Cancer Drug Resist. 2025;8:46 Page 3 of 28

functional role remains largely unclear . The β subunit includes HIF-1β, which is constitutively expressed in
[18]
all tissues, and HIF-2β, whose expression is limited to organs such as the kidney and brain and whose role is
not yet fully explored [17,19] .

The stability of the α subunits is tightly regulated by oxygen availability. Under normoxic conditions, the α
subunits are rapidly degraded, whereas under hypoxia, they are stabilized [20-22] . In normoxia, prolyl
hydroxylase domain (PHD) enzymes hydroxylate conserved proline residues (Pro402 and Pro564) of the α
subunits, using oxygen and α-ketoglutarate as substrates [20,23-25] . During hydroxylation, one oxygen atom is
incorporated into the prolyl residue, while the other oxidizes α-ketoglutarate, producing succinate and
carbon dioxide . The hydroxylated α subunits are then ubiquitinated at lysine residues by the von
[17]
Hippel-Lindau protein, a component of the tumor suppressor E3 ubiquitin ligase complex. This complex,
composed of Cullin-2, Elongin B, Elongin C, RING box protein 1, and an E2 ubiquitin-protein ligase, targets
the α subunits for degradation by the 26S proteasome [26-28] . In contrast, hypoxia inhibits PHD activity,
allowing HIF-α to accumulate and translocate into the nucleus. There, HIF-α dimerizes with HIF-β and binds
to hypoxia-response elements [5′-(A/G)CGTG-3′] in promoter regions, activating transcription of genes that
promote hypoxia adaptation .
[29]
HIF plays a central role in regulating metabolism and cellular adaptation to oxygen deprivation. Under
hypoxia, it induces the expression of a broad range of genes encoding proteins involved in metabolic
reprogramming, angiogenesis, proliferation, apoptosis, glucose and iron transport, genomic instability,
invasion and metastasis, growth factor signaling, and resistance to chemotherapy and radiotherapy.
Approximately 100 HIF-dependent genes have been identified to date [13,30] . Hypoxia also reduces drug
efficacy by limiting penetration across hypoxic gradients . Clinically, intratumoral hypoxia is a negative
[13]
prognostic factor and a major determinant of treatment failure, poor overall survival, and increased
mortality [31,32] .

Moreover, hypoxia contributes to an immunosuppressive TME and plays a significant role in resistance to
cancer immunotherapy. It promotes polarization of macrophages toward the M2 phenotype
(tumor-associated macrophages, TAMs), induces T cell exhaustion, facilitates immune evasion, and
enhances tumor angiogenesis, all of which support immunotherapy resistance [33-36] . Consequently, targeting
key hypoxia-associated pathways represents a promising strategy to overcome immunotherapy resistance.
Nanotechnologies designed to inhibit HIF-1α signaling as a means of reversing immunotherapy resistance
are discussed in the following section.

MECHANISM OF HYPOXIA-REGULATED RESISTANCE TO CANCER IMMUNOTHERAPY

Hypoxia-induced macrophage polarization and the role of TAMs in resistance to immunotherapy
Mechanisms of hypoxia in macrophage polarization
Tissue-resident macrophages are primarily derived from erythro-myeloid progenitors in the yolk sac or fetal
liver, and partly from bone marrow progenitors. These progenitors differentiate into various types of
macrophages . Macrophages are generally classified into two subtypes: classically activated macrophages
[37]
(M1) and alternatively activated macrophages (M2) [38,39] . Interleukin-12 (IL-12), interferon-γ, bacterial
lipopolysaccharide, tumor necrosis factor (TNF) and Toll-like receptor agonists induce macrophage
polarization toward the M1 phenotype . In contrast, IL, IL-5, IL-10, IL-13, colony-stimulating factor 1
[40]
(CSF-1), transforming growth factor-β1, and prostaglandin E2 promote polarization toward the M2
phenotype . Similarly, TAMs can be divided into the proinflammatory, tumor-suppressive M1 type and the
[40]
anti-inflammatory, tumor-promoting M2-like type .
[38]

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