The CRISPR/Cas9 system is a programmable genome editing tool that uses guide RNA (gRNA) to direct Cas9 nuclease to target specific DNA sequences and induce double-strand breaks (DSBS) at this point. This break can be repaired through non-homologous end joining (NHEJ) or homologous directed repair (HDR) mechanisms, thereby achieving gene knockout, correction or insertion.

The CRISPR/Cas9 system enables the precise targeting and removal of resistance pathways in gynecological diseases that disrupt the E6 and E7 oncogenes, promoting apoptosis, restoring p53 and Rb function, and enhancing sensitivity to radiotherapy and cisplatin. In ovarian cancer, drug resistance mechanisms involve ABCB1 overexpression and BRCA1/2 dysfunction, both of which challenge genomic repair precision. The efflux of doxorubicin and paclitaxel is reduced by eliminating ABCB1 through CRISPR/Cas9 editing, thereby improving their therapeutic potency (18, 19).

Moreover, repairing BRCA1 or TP53 mutations via CRISPR in HR-deficient malignancies re-establishes sensitivity to Pt compounds and PARP inhibitors. These studies highlight the dual role of gene editing in both suppressing disease progression and restoring chemotherapeutic efficacy. Innovative strategies that integrate RISPR with pharmacological agents are being adopted to enhance the effectiveness of killing tumor cells (20).

Gene delivery is a key bottleneck in gene therapy, which requires vectors that can efficiently transport genetic material to target cells while minimizing immune activation and off-target effects. Vectors are mainly classified into viral vectors (such as AAV and lentiviruses) and non-viral vectors (such as lipid nanoparticles and polymers), each having different advantages in terms of transduction efficiency, payload capacity and safety.

Effective gene therapy relies strongly on the development of benign and efficient delivery vehicles. Viral vectors, particularly adeno-associated viruses (AAVs), remain the most validated platforms, offering sustained expression of therapeutic genes in both malignant and benign reproductive tissues. AAV-mediated delivery of shRNAs targeting HPV-E7 has resulted in durable tumor regression and enhanced chemo-radio-sensitivity in cervical cancer models (21).

In endometriosis, AAVs are employed to deliver anti-inflammatory cytokines, such as IL-10, which modulates local immune responses and potentially enhances sensitivity to hormonal treatments. Targeted gene delivery also holds potential in fibroid management, where gene silencing of TGF-β receptors may improve responsiveness to selective progesterone receptor modulators (SPRMs) (22).

Non-viral delivery systems provide greater customization for therapeutic applications and exhibit lower immunogenic potential compared to viral vectors (23). These vectors are functionalized with targeting ligands to enable selective delivery of therapeutic payloads to endometrial and ovarian tissues. The development of dual-function nanoparticles has shown the delivery of siRNAs targeting resistance pathways and chemotherapeutics, as exhibited by the co-delivery of paclitaxel and BCL2 siRNA to treat multidrug-resistant ovarian cancer (24).

Oncogenes and tumor suppressor genes are the two basic categories of cancer drivers. Oncogenes (such as PI3K and AKT) acquire functions through mutations or overexpression, promoting cell survival and proliferation. However, tumor suppressor genes (such as TP53 and BRCA1) lose their function, leading to uncontrolled cell growth. Targeting these genes through gene silencing or alternative strategies provides a reasonable path to correct the molecular imbalance behind gynecological tumors and benign proliferative diseases.

Gynecologic cancers exhibit aberrant activation of oncogenic drivers and frequently harbor inactivating mutations in tumor suppressor genes, which contribute to disease growth and therapeutic resistance. Gene therapy can reinstate therapeutic responsiveness and potentiate the effects of co-administered treatments by directly targeting abnormal genetic pathways (25).

Therapeutic restoration of wild-type p53 has improved the responsiveness to radiotherapy and cisplatin in both cervical and endometrial cancers (26). Similarly, silencing of PI3K/Akt/mTOR via siRNA or shRNA has enhanced doxorubicin-induced apoptosis in ovarian cancer models and reversed pro-survival signaling. In hormone-resistant endometriosis, suppression of SF-1, COX-2, and IL-6 via RNA interference (RNAi) has been shown to reduce lesion volume and improve sensitivity to progesterone therapy (27).

CAR-T cells and T cell receptor (TCR)-modified T cells are at the forefront of immunotherapy for solid tumors. In gynecologic cancers, particularly ovarian and cervical, drug resistance frequently arises from antigenic drift, immune escape, or a suppressive tumor microenvironment (TME), which limits the effectiveness of Pt-based drugs, taxanes, and immune checkpoint inhibitors (29).

CAR-T therapies are designed to bypass these resistance mechanisms by recognizing surface antigens independent of MHC presentation. In ovarian cancer, CAR-T cells targeting MUC16 (CA125), mesothelin, and folate receptor-α have demonstrated potent activity in preclinical models and early-phase clinical trials (30). Importantly, these therapies may eradicate tumor clones resistant to cisplatin or bevacizumab, thus complementing standard regimens. Dual-targeting CARs, also known as “logic-gated” CARs (e.g., SynNotch systems), are being tested to mitigate off-target toxicity and address tumor heterogeneity.

In HPV-associated cervical cancer, TCR-T cells that recognize viral antigens (e.g., HPV E6/E7) offer a potential route to eliminate tumor cells that are unresponsive to radiotherapy and chemotherapy. These antigen-specific T cells can target intracellular proteins processed on MHC class I, thereby extending the therapeutic reach beyond surface markers and providing a strategy against tumors with defective DNA damage responses or altered p53 (31).

NK cells are emerging as effective agents targeting drug-resistant gynecologic tumors, providing innate cytotoxicity prior to sensitization. These cells retain their cytotoxic function against tumor cells that downregulate MHC expression, a common mechanism of immune escape in chemoresistant tumors. Unlike T cells, they are not restricted by MHC molecules (32).

CAR-modified NK cells have been designed to specifically target ovarian cancer-associated antigens, enhancing their targeted cytotoxicity against tumor cells. The use of CAR-NK cells in patients with Pt-resistant ovarian cancers has shown promising results. These include tumor clearance and reduced cytokine-related toxicity, resulting in their safe and effective role. For patients who have relapsed and not responded to drugs such as paclitaxel or bevacizumab, these CAR-NK cell platforms are beneficial, as tumor antigen profiles remain stable, allowing for sustained targeting of the disease (33). Furthermore, NK cells can exert synergistic effects; for instance, beneficial efficacy in endometrial and HER2-positive uterine tumors can be improved by integrating NK cells with anti-HER2 or anti-PD-L1 antibodies.

Due to their immunomodulatory, regenerative, and anti-fibrotic characteristics, MSCs present promising alternatives for treating benign uterine disorders where conventional therapies are ineffective. Circumstances such as uterine synechiae, Asherman’s syndrome, and thin endometrium often arise from the repeated intrauterine procedures or unresponsive endometritis. These cells have demonstrated the potential to restore endometrial thickness and promote angiogenesis through paracrine signaling and direct tissue integration, making them a valuable tool for uterine regeneration (34). This regenerative effect may reverse endometrial damage caused by the prolonged use of GnRH analogues, presenting a fertility-preserving therapeutic alternative for the patients.

Beyond structural repair, MSCs are being used as carriers of therapeutic genes or microRNAs. Engineered MSCs expressing IL-10 or VEGF can attenuate fibrosis and improve hormonal responsiveness in endometriosis. Additionally, exosome-derived MSC therapies are emerging as cell-free options for delivering regulatory RNAs that target inflammation, estrogen metabolism, or fibrotic remodeling, thereby enhancing responsiveness to progesterone therapy or aromatase inhibitors (35).

Safety concerns related to tumorigenicity, immune rejection, or ectopic differentiation are being closely monitored; however, early-phase clinical trials have demonstrated good tolerability. Scaffold-based delivery systems, autologous sourcing, and controlled gene loading are being developed to improve localization and therapeutic outcomes. Figure 1 presents the main therapeutic mechanisms underlying gene and cell therapies in gynecological disorders, including CRISPR/Cas9-enabled gene editing, RNA interference, immune cell cytotoxicity, and MSC-based regeneration (see Table 1).

Synthetic biology is enabling the creation of “intelligent” therapeutic platforms, such as logic-gated CAR-T cells, inducible promoters, and tumor-selective gene circuits that can dynamically respond to cues of drug resistance. For instance, SynNotch-based CAR systems can restrict cytotoxic activity to dual-antigen environments, thereby minimizing off-target toxicity and addressing antigen heterogeneity, a common cause of CAR-T treatment failure in ovarian cancer (36).

Additionally, synthetic promoters responsive to hypoxia, inflammatory cytokines, or estrogen receptor activation are being developed to regulate therapeutic gene expression in drug-resistant microenvironments. These designs enable the combinatorial use of hormone therapies or checkpoint inhibitors, thereby optimizing the spatiotemporal control of therapeutic payloads. Self-amplifying RNA systems and base editors are also under exploration for non-disruptive correction of resistance mutations (BRCA1 reversion, TP53 dysfunction) (37).

TME presents both physical and molecular barriers to therapy, particularly in cases of chemoresistant ovarian and endometrial tumors. Dense stroma, hypoxia, and high levels of TGF-β contribute to immune exclusion, suppressing the activity of engineered T or NK cells. These factors also drive epithelial-mesenchymal transition (EMT), a known contributor to platinum resistance and metastasis (38).

To overcome these difficulties, strategies such as co-administration of collagenase, nanoparticle-based TGF-β inhibitors, and armored CAR-T cells capable of secreting IL-12 or DNase I are under investigation, which improve drug penetration and enhance immune access to persistent tumor islets. Gene therapies targeting matrix metalloproteinases (MMPs), angiogenesis factors, or immune checkpoints within the TME are also being pursued as adjuncts to standard chemotherapeutic regimens. Such modulation may reinstate chemosensitivity and prolong responses to agents like paclitaxel or bevacizumab (39).

Despite the therapeutic promise, gene and cell therapies for gynecological applications face complex regulatory landscapes. Safety concerns include insertional mutagenesis, off-target gene edits, and immunogenicity of engineered cells. For reproductive-age women, reproductive toxicity, placental transmission, and germline editing remain under strict scrutiny, especially when delivery is intrauterine or intraperitoneal (40).

Manufacturing and scalability present additional bottlenecks. Autologous CAR-T cell production is time-consuming and resource-intensive, limiting accessibility in low-resource settings where gynecologic cancers are most prevalent. To address this issue, allogeneic “off-the-shelf” CAR and NK cell platforms are being developed in conjunction with point-of-care processing units (41).

Clinical trial pipelines also lag behind hematologic indications. Many gynecological trials are early-phase or exploratory, lacking large-scale randomized data. Multimodal trial designs, integrating gene or cell therapies with conventional drugs, are necessary to clarify sequencing, synergy, and the reversal of resistance (42). Figure 2 displays an overview of translational challenges and emerging novelties in synthetic biology, TME modulation, and regulatory solutions in the clinical regulatory frameworks supporting the implementation of gene- and cell-based interventions in gynecology. Additionally, a summary of gene- and cell-based therapeutic trials for gynecologic disorders is presented (see Table 2).