Description:

CRISPR-CAS9 mediated gene editing for targeted therapy of pancreatic cancer
Field of the Invention
[0001] The present disclosure relates to CRISPR-Cas9 mediated genetic engineering, more particularly, to targeted therapy of pancreatic cancer using selective KRAS disruption.
Background
[0002] The background description includes information that may be useful in understanding the present invention. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication specifically or implicitly referenced is prior art.
[0003] Pancreatic cancer remains one of the most aggressive malignancies, with five-year survival rates persistently below ten percent despite advancements in chemotherapy, radiation, and surgical interventions. Conventional therapeutic regimens often demonstrate limited efficacy due to extensive desmoplastic stroma, poor vascularization, and intrinsic resistance mechanisms within pancreatic tumor cells. Chemotherapy agents such as gemcitabine, fluorouracil, and nab-paclitaxel exhibit modest survival benefits, yet systemic toxicity and rapid tumor relapse severely constrain clinical outcomes. Radiotherapy is hindered by surrounding healthy tissue radiosensitivity, while surgical resection is feasible only in a minority of patients presenting with localized disease.
[0004] Genetic analysis of pancreatic adenocarcinoma has revealed a consistent prevalence of KRAS mutations, particularly the G12D and G12V variants, which act as principal oncogenic drivers through constitutive activation of downstream signaling pathways including MAPK and PI3K-AKT cascades. Existing targeted therapies directed toward KRAS downstream effectors, such as MEK inhibitors or PI3K antagonists, have demonstrated limited clinical utility owing to pathway redundancy and rapid compensatory adaptation. Immunotherapy, while transformative in melanoma and lung cancer, has shown only modest success in pancreatic cancer due to the profoundly immunosuppressive tumor microenvironment.
[0005] The absence of clinically validated methods for directly targeting KRAS mutations has long been recognized as a central therapeutic gap. Small-molecule inhibitors selectively binding KRAS-G12C have entered clinical evaluation, yet these molecules fail to address non-G12C mutations prevalent in pancreatic tumors. Accordingly, there exists a pressing demand for therapeutic modalities that can directly disrupt oncogenic KRAS sequences at the genomic level, thereby halting tumorigenesis at its source. CRISPR-Cas9 gene editing technology offers unprecedented precision in genomic manipulation, yet challenges remain in terms of delivery, off-target editing, and controlled repair pathway modulation. The disclosed system addresses these shortcomings by integrating high-fidelity Cas9 variants, chemically stabilized guide RNAs, optimized lipid nanoparticle carriers, and context-specific therapeutic protocols tailored for pancreatic cancer treatment.
[0006]
Summary
[0007] The following presents a simplified summary of various aspects of this disclosure in order to provide a basic understanding of such aspects. This summary is not an extensive overview of all contemplated aspects, and is intended to neither identify key or critical elements nor delineate the scope of such aspects. Its purpose is to present some concepts of this disclosure in a simplified form as a prelude to the more detailed description that is presented later.
[0008] The following paragraphs provide additional support for the claims of the subject application.
[0009] The disclosure pertains to a CRISPR-Cas9 mediated gene editing system for targeted therapy of pancreatic cancer is provided. The system includes a guide RNA construct designed to recognize mutant KRAS sequences frequently encountered in pancreatic adenocarcinoma cells. A Cas9 endonuclease, operably linked with a nuclear localization signal, is configured to achieve cleavage of the targeted locus. A lipid nanoparticle formulation, composed of ionizable cationic lipids, cholesterol, helper phospholipids, and polyethylene glycol conjugates, facilitates effective delivery across pancreatic tumor barriers. The system is further configured with therapeutic administration protocols including systemic intravenous injection, endoscopic ultrasound-guided intratumoral injection, and adaptive dosing cycles tailored for tumor regression monitoring.
[00010] In one embodiment, the system integrates a homology-directed repair template comprising wild-type KRAS sequence flanked by homology arms to restore normal gene function in edited pancreatic cells. In another embodiment, co-administration of small-molecule repair pathway modulators biases repair outcomes toward corrective replacement rather than error-prone non-homologous end joining. In yet another embodiment, the delivery vehicle incorporates fluorescent tags to enable pharmacokinetic monitoring and real-time biodistribution tracking. The therapeutic method includes cycles of administration, genetic disruption, monitoring of tumor regression biomarkers, and adaptive guide RNA redesign in response to mutational shifts. The disclosed system further supports combination with immune checkpoint inhibitors to enhance immune activation and amplify anti-tumor effects. Through integration of selective targeting, efficient delivery, and controlled editing outcomes, the system provides a comprehensive therapeutic framework capable of directly addressing oncogenic KRAS mutations in pancreatic cancer.
Brief Description of the Drawings
[00011] The features and advantages of the present disclosure would be more clearly understood from the following description taken in conjunction with the accompanying drawings in which:
[00012] FIG. 1 illustrates a system architecture diagram showing the interconnection of core modules within the CRISPR-Cas9 mediated gene editing therapeutic system, including guide RNA design unit, Cas9 nuclease assembly, nanoparticle delivery vehicle, administration protocol interface, and therapeutic monitoring system, in accordance with the embodiments of the present disclosure.
[00013] FIG. 2 illustrates a method flow diagram showing sequential therapeutic steps beginning with guide RNA design, Cas9 complex formation, nanoparticle encapsulation, systemic or intratumoral delivery, genomic disruption of mutant KRAS sequence, and monitoring of tumor regression outcomes., in accordance with the embodiments of the present disclosure.
[00014] FIG. 3 illustrates a data flow diagram showing exchange of therapeutic information across modules including donor DNA template integration, repair pathway modulation, pharmacokinetic tracking signals, adaptive guide RNA redesign, and clinical biomarker data integration., in accordance with the embodiments of the present disclosure.
Detailed Description
[00015] In the following detailed description of the invention, reference is made to the accompanying drawings that form a part hereof, and in which is shown, by way of illustration, specific embodiments in which the invention may be practiced. In the drawings, like numerals describe substantially similar components throughout the several views. These embodiments are described in sufficient detail to claim those skilled in the art to practice the invention. Other embodiments may be utilized and structural, logical, and electrical changes may be made without departing from the scope of the present invention. The following detailed description is, therefore, not to be taken in a limiting sense, and the scope of the present invention is defined only by the appended claims and equivalents thereof.
[00016] The use of the terms “a” and “an” and “the” and “at least one” and similar referents in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. The use of the term “at least one” followed by a list of one or more items (for example, “at least one of A and B”) is to be construed to mean one item selected from the listed items (A or B) or any combination of two or more of the listed items (A and B), unless otherwise indicated herein or clearly contradicted by context. The terms “comprising,” “having,” “including,” and “containing” are to be construed as open-ended terms (i.e., meaning “including, but not limited to,”) unless otherwise noted. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., “such as”) provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.
[00017] Pursuant to the "Detailed Description" section herein, whenever an element is explicitly associated with a specific numeral for the first time, such association shall be deemed consistent and applicable throughout the entirety of the "Detailed Description" section, unless otherwise expressly stated or contradicted by the context.
[00018] The disclosed CRISPR-Cas9 mediated gene editing system for targeted therapy of pancreatic cancer shall now be described in expanded operational detail.
[00019] The system operates by selecting a guide RNA construct specifically designed to recognize the KRAS gene sequence harboring oncogenic mutations. Said guide RNA is synthesized through a process comprising sequential assembly of a crRNA and tracrRNA, chemically stabilized with modifications such as 2′-O-methyl ribose substitutions and phosphorothioate linkages, thereby extending half-life in serum and improving accumulation within pancreatic tumor cells. The guide RNA construct is hybridized with a Cas9 endonuclease, forming a ribonucleoprotein complex that is subsequently encapsulated within a lipid nanoparticle delivery vehicle. Said nanoparticle comprises ionizable cationic lipids configured to promote nucleic acid condensation, cholesterol providing membrane rigidity, helper phospholipids facilitating bilayer fusion, and polyethylene glycol conjugates enabling steric stabilization and prolonged circulation time in systemic administration.
[00020] Upon systemic delivery, such as intravenous infusion, the nanoparticles traverse the circulatory system, accumulate in the tumor microenvironment via enhanced permeability and retention effect, and undergo endocytosis by pancreatic tumor cells. Following cellular uptake, endosomal escape is mediated through protonation of ionizable lipids, leading to disruption of endosomal membranes and release of the Cas9-guide RNA ribonucleoprotein complex into the cytoplasm. Nuclear localization signals appended to the Cas9 protein facilitate active transport through the nuclear pore complex, thereby enabling access to genomic DNA. Target recognition occurs when the guide RNA sequence hybridizes with the complementary KRAS mutant locus, initiating cleavage through the catalytic domains of the Cas9 nuclease. Double-strand breaks at the KRAS locus disrupt oncogenic signaling, leading to apoptosis or senescence of tumor cells.
[00021] In one embodiment, a donor DNA template comprising wild-type KRAS sequence flanked by 800-base pair homology arms is co-delivered with the Cas9-guide RNA complex. Homology-directed repair is thereby stimulated, resulting in replacement of the mutant sequence with physiologically functional wild-type sequence. Such correction halts constitutive KRAS signaling, restores normal cell signaling homeostasis, and potentially reprograms malignant pancreatic cells toward normal physiology. In a second embodiment, editing outcomes are biased toward corrective replacement by administering small-molecule inhibitors such as SCR7 targeting DNA ligase IV, thereby suppressing non-homologous end joining and promoting homology-directed repair. This embodiment enhances precise correction efficiency, thereby reducing indel formation and maintaining genomic integrity. In a third embodiment, the lipid nanoparticle vehicle is modified with fluorescent lipids such as DiR or DiD dyes, enabling real-time imaging and pharmacokinetic tracking of therapeutic biodistribution in vivo. Such visualization permits dose optimization, patient-specific tailoring, and enhanced safety monitoring.
[00022] The therapeutic administration protocol incorporates flexibility in delivery routes. In one operational flow, intravenous administration delivers nanoparticles to systemic circulation, exploiting tumor permeability for passive accumulation. In another flow, endoscopic ultrasound-guided intratumoral injection introduces nanoparticles directly into the tumor mass, achieving higher localized concentration while minimizing systemic toxicity. In yet another flow, intra-arterial infusion through celiac artery branches is employed to directly perfuse the pancreatic region, providing selective vascular targeting. Each protocol yields distinct technical benefits: systemic infusion permits repeated administration cycles, intratumoral injection ensures focused delivery, and intra-arterial infusion balances penetration with minimized systemic exposure.

assessment of tumor biomarkers such as carbohydrate antigen 19-9, imaging modalities including positron emission tomography and magnetic resonance imaging, and biopsy evaluation of KRAS allele disruption efficiency. Adaptive treatment cycles are then configured wherein guide RNA constructs are redesigned in response to mutational evolution of pancreatic tumors, such as emergence of resistance-associated substitutions. Continuous cycles of administration, monitoring, adaptation, and re-administration establish a dynamic therapeutic framework capable of maintaining efficacy against evolving tumor genotypes.
[00024] Alternative embodiments further expand the technical architecture. In one alternative embodiment, Cas9 is replaced by a high-fidelity SpCas9 variant containing engineered amino acid substitutions that reduce tolerance for mismatched sequences, thereby lowering off-target editing frequency. In another alternative embodiment, SaCas9, derived from Staphylococcus aureus, is utilized due to its smaller molecular size, enabling encapsulation within adeno-associated viral vectors for sustained delivery. In a third alternative embodiment, Cas12a (Cpf1) is incorporated as an alternative nuclease, providing staggered cuts with distinct repair pathway engagement and enabling multiplexed editing of multiple KRAS mutations simultaneously.
[00025] The integration with immunotherapy provides further benefit. Following genetic correction of KRAS, residual tumor cells are sensitized to immune clearance. Combination with checkpoint inhibitors such as anti-PD-1 or anti-CTLA-4 antibodies stimulates immune activation, overcoming pancreatic tumor immunosuppression. This dual therapeutic mechanism results in synergistic tumor regression, combining direct genomic disruption with immune system engagement.
[00026] Thus, the disclosed CRISPR-Cas9 mediated gene editing system presents a comprehensive therapeutic approach wherein operational flows are expanded into multiple embodiments, delivery vehicles are optimized for pancreatic microenvironment, editing outcomes are directed toward precise correction, and adaptive treatment cycles sustain long-term efficacy. Through expanded embodiments, technical benefits include selective KRAS disruption, restored signaling homeostasis, enhanced delivery, minimized off-target effects, real-time monitoring, and synergistic immune activation. Each embodiment reiterates data processing flows in distinct operational contexts, thereby illustrating versatility and robustness of the disclosed therapeutic system for targeted treatment of pancreatic cancer.
[00027] Figure 1 depicts the system architecture of the CRISPR-Cas9 mediated gene editing therapeutic platform designed for targeted therapy of pancreatic cancer. The figure includes a guide RNA design unit configured to generate highly specific sequences directed against mutant KRAS alleles, a Cas9 nuclease assembly configured for ribonucleoprotein formation, and a nanoparticle delivery vehicle incorporating lipid components optimized for tumor penetration. The architecture further incorporates an administration protocol interface managing systemic and intratumoral delivery routes, as well as a therapeutic monitoring system configured to integrate imaging, biomarker, and sequencing data for adaptive feedback. The overall architecture functions as an integrated therapeutic pipeline. The guide RNA design unit provides precision targeting by computationally screening KRAS variants. The Cas9 nuclease assembly ensures high-fidelity cleavage at the selected locus. The nanoparticle delivery vehicle facilitates transport of ribonucleoprotein complexes into pancreatic tumors with enhanced endosomal escape efficiency. The administration protocol interface governs the clinical deployment of therapeutic doses through either intravenous infusion or direct tumor injection. The therapeutic monitoring system assesses outcomes by measuring regression biomarkers, imaging tumor response, and evaluating allele correction efficiency. The system collectively provides improved selectivity, reduced toxicity, and enhanced therapeutic durability. Figure 2 depicts a method flow diagram for the therapeutic operation of the disclosed system. The diagram begins with design of a guide RNA targeting oncogenic KRAS variants, followed by formation of a Cas9-guide RNA ribonucleoprotein complex. This complex is encapsulated within a lipid nanoparticle delivery vehicle, subsequently prepared for administration. The therapeutic administration step may involve systemic intravenous infusion or localized intratumoral injection. Following delivery, the nanoparticle facilitates endosomal escape, releasing the ribonucleoprotein complex into the cytoplasm and subsequently into the nucleus. Genomic disruption occurs at the KRAS locus, optionally followed by homology-directed repair using a donor DNA template. Tumor regression is then monitored using biomarker assays and imaging modalities. The described operational flow provides a sequential therapeutic pathway from design through outcome assessment. Each step contributes to therapeutic efficacy by ensuring accurate targeting, efficient delivery, genomic disruption, and adaptive monitoring. The flow diagram highlights the interdependence of design, delivery, editing, and feedback mechanisms in achieving targeted therapeutic outcomes for pancreatic cancer.
[00028] Figure 3 depicts a data flow diagram for therapeutic information exchange within the CRISPR-Cas9 mediated system. The donor DNA template integrates with the Cas9-guide RNA ribonucleoprotein complex to promote homology-directed repair. Repair pathway modulation data, generated by co-administered small-molecule inhibitors, is integrated with the editing outcome analysis module. Pharmacokinetic tracking signals derived from fluorescently labeled nanoparticles are transmitted to a biodistribution monitoring module, which feeds into an adaptive therapy controller. Clinical biomarker data, including tumor regression markers, is continuously captured and integrated with the adaptive guide RNA redesign module. The data exchange culminates in an updated therapeutic configuration for subsequent treatment cycles. The described flow illustrates feedback-driven adaptation. Integration of donor DNA repair, pathway modulation, pharmacokinetics, and biomarker monitoring ensures that therapeutic strategies remain responsive to dynamic tumor evolution. The data flow architecture supports iterative refinement of therapeutic sequences, thereby improving long-term outcomes in pancreatic cancer treatment through controlled, data-driven editing interventions.
[00029] Operations in accordance with a variety of aspects of the disclosure is described above would not have to be performed in the precise order described. Rather, various steps can be handled in reverse order or simultaneously or not at all.
[00030] While several implementations have been described and illustrated herein, a variety of other means and/or structures for performing the function and/or obtaining the results and/or one or more of the advantages described herein may be utilized, and each of such variations and/or modifications is deemed to be within the scope of the implementations described herein. More generally, all parameters, dimensions, materials, and configurations described herein are meant to be exemplary and that the actual parameters, dimensions, materials, and/or configurations will depend upon the specific application or applications for which the teachings is/are used. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific implementations described herein. It is, therefore, to be understood that the foregoing implementations are presented by way of example only and that, within the scope of the appended claims and equivalents thereto, implementations may be practiced otherwise than as specifically described and claimed. Implementations of the present disclosure are directed to each individual feature, system, article, material, kit, and/or method described herein. In addition, any combination of two or more such features, systems, articles, materials, kits, and/or methods, if such features, systems, articles, materials, kits, and/or methods are not mutually inconsistent, is included within the scope of the present disclosure.
 

Claims
I/We Claim:
1. A CRISPR-Cas9 mediated gene editing system configured for targeted therapy of pancreatic cancer, comprising: a guide RNA construct specifically designed to recognize a mutant KRAS gene sequence prevalent in pancreatic adenocarcinoma cells; a Cas9 endonuclease operably linked to a nuclear localization signal to promote entry into a tumor cell nucleus; a delivery vehicle comprising a lipid nanoparticle formulation optimized for pancreatic tumor microenvironmental penetration; and a therapeutic administration protocol including systemic intravenous injection and tumor-specific accumulation, wherein cleavage of the mutant KRAS locus results in disruption of oncogenic signaling pathways and induction of apoptotic cascades in pancreatic tumor cells.
2. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the guide RNA construct comprises a dual-guide configuration incorporating a tracrRNA and crRNA chemically stabilized with 2′-O-methyl modifications, thereby enhancing stability in serum and improving editing efficiency in pancreatic tumor tissues.
3. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the Cas9 endonuclease is selected from a high-fidelity SpCas9 variant or SaCas9 variant engineered to minimize off-target cleavage, thereby conferring enhanced therapeutic selectivity within a heterogeneous pancreatic tumor cell population.
4. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the lipid nanoparticle formulation comprises ionizable cationic lipids, cholesterol, helper phospholipids, and polyethylene glycol-lipid conjugates blended in a molar ratio optimized for endosomal escape and cytoplasmic release of the Cas9-guide RNA ribonucleoprotein complex.
5. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the therapeutic administration protocol further comprises image-guided infusion through endoscopic ultrasound-guided fine-needle injection into a pancreatic tumor site, thereby achieving higher local concentration and reduced systemic toxicity.
6. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein a pharmacokinetic monitoring module is integrated with a nanoparticle tracking fluorescence tag, thereby enabling real-time biodistribution assessment and therapeutic dose adjustment in pancreatic cancer patients.
7. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the cleavage of the mutant KRAS locus is followed by homology-directed repair using a supplied donor DNA template comprising a wild-type KRAS sequence flanked by homology arms, thereby restoring physiological function in edited pancreatic cells.
8. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein editing efficiency is further enhanced by co-administration of small-molecule inhibitors targeting DNA repair pathways that compete with homology-directed repair, thereby biasing the repair mechanism toward corrective replacement of the KRAS sequence.
9. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the therapeutic administration protocol is configured in multiple dosing cycles with sequential monitoring of tumor regression markers, thereby enabling adaptive modification of guide RNA sequences in response to observed mutational evolution of pancreatic cancer cells.
10. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the therapeutic outcome is augmented through integration with an immunomodulatory checkpoint inhibitor therapy, thereby inducing synergistic anti-tumor activity through simultaneous genetic correction and immune activation in pancreatic cancer treatment.

CRISPR-CAS9 mediated gene editing for targeted therapy of pancreatic cancer
Abstract
A CRISPR-Cas9 mediated gene editing system for targeted therapy of pancreatic cancer is disclosed. The system comprises a guide RNA construct configured to recognize mutant KRAS sequences, a Cas9 endonuclease operably linked with a nuclear localization signal, and a lipid nanoparticle formulation optimized for pancreatic tumor microenvironmental delivery. Therapeutic administration includes systemic and intratumoral injection strategies. The editing process disrupts oncogenic KRAS signaling and optionally restores wild-type sequence through homology-directed repair using a donor template. Additional embodiments include co-administration of DNA repair modulators, incorporation of fluorescent tracking tags, and integration with immune checkpoint inhibitors. The disclosed system enables direct genomic disruption of oncogenic KRAS variants, thereby achieving selective therapeutic efficacy in pancreatic adenocarcinoma with reduced systemic toxicity and adaptive treatment flexibility.
Fig. 1

, Claims:Claims
I/We Claim:
1. A CRISPR-Cas9 mediated gene editing system configured for targeted therapy of pancreatic cancer, comprising: a guide RNA construct specifically designed to recognize a mutant KRAS gene sequence prevalent in pancreatic adenocarcinoma cells; a Cas9 endonuclease operably linked to a nuclear localization signal to promote entry into a tumor cell nucleus; a delivery vehicle comprising a lipid nanoparticle formulation optimized for pancreatic tumor microenvironmental penetration; and a therapeutic administration protocol including systemic intravenous injection and tumor-specific accumulation, wherein cleavage of the mutant KRAS locus results in disruption of oncogenic signaling pathways and induction of apoptotic cascades in pancreatic tumor cells.
2. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the guide RNA construct comprises a dual-guide configuration incorporating a tracrRNA and crRNA chemically stabilized with 2′-O-methyl modifications, thereby enhancing stability in serum and improving editing efficiency in pancreatic tumor tissues.
3. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the Cas9 endonuclease is selected from a high-fidelity SpCas9 variant or SaCas9 variant engineered to minimize off-target cleavage, thereby conferring enhanced therapeutic selectivity within a heterogeneous pancreatic tumor cell population.
4. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the lipid nanoparticle formulation comprises ionizable cationic lipids, cholesterol, helper phospholipids, and polyethylene glycol-lipid conjugates blended in a molar ratio optimized for endosomal escape and cytoplasmic release of the Cas9-guide RNA ribonucleoprotein complex.
5. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the therapeutic administration protocol further comprises image-guided infusion through endoscopic ultrasound-guided fine-needle injection into a pancreatic tumor site, thereby achieving higher local concentration and reduced systemic toxicity.
6. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein a pharmacokinetic monitoring module is integrated with a nanoparticle tracking fluorescence tag, thereby enabling real-time biodistribution assessment and therapeutic dose adjustment in pancreatic cancer patients.
7. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the cleavage of the mutant KRAS locus is followed by homology-directed repair using a supplied donor DNA template comprising a wild-type KRAS sequence flanked by homology arms, thereby restoring physiological function in edited pancreatic cells.
8. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein editing efficiency is further enhanced by co-administration of small-molecule inhibitors targeting DNA repair pathways that compete with homology-directed repair, thereby biasing the repair mechanism toward corrective replacement of the KRAS sequence.
9. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the therapeutic administration protocol is configured in multiple dosing cycles with sequential monitoring of tumor regression markers, thereby enabling adaptive modification of guide RNA sequences in response to observed mutational evolution of pancreatic cancer cells.
10. The CRISPR-Cas9 mediated gene editing system of claim 1, wherein the therapeutic outcome is augmented through integration with an immunomodulatory checkpoint inhibitor therapy, thereby inducing synergistic anti-tumor activity through simultaneous genetic correction and immune activation in pancreatic cancer treatment.