Table 1.

Overview of recent studies that used exosomes and viral vectors for CRISPR/Cas delivery in cancer cells.

Study	Vector	Type of cancer	Approach	Outcome
A New Tool for CRISPR-Cas13a-Based Cancer Gene Therapy [55]	AAVs	HCC	Decoy minimal promoter (DMP)-controlled CRISPR-Cas13a system used to knock down several oncogenes (TERT, EZH2, and RelA)	Significant inhibition of HCC cell growth
CRISPR-Cas9 disruption of PD-1 enhances activity of universal EGFRvIII CAR T cells in a preclinical model of human glioblastoma [56]	AAV6	Glioblastoma (GBM)	Allogeneic EGFRvIII CAR T-cell deficient in PD-1, TCR and B2M generated by CRISPR-Cas9	Enhanced antitumor efficacy in preclinical models of GBM.
Targeting HPV16 DNA using CRISPR/Cas inhibits anal cancer growth in vivo [57]	AAVs	Anal cancer	Cleaving the HPV16 E6 or E7 genes in primary human anal cancer cells utilizing CRISPR/Cas9	Significant and selective tumor suppression
CRISPR/Cas9-mediated cervical cancer treatment targeting human papillomavirus E6 [58]	AAVs	Cervical cancer	Causing multiple mutations in targeted HPV E6 gene in cervical cancer cells by CRISPR/Cas9	Increased expression of tumor suppressors, tumor growth inhibition, and increased apoptosis in the cancer cells
Disruption of PD-1 Enhanced the Anti-tumor Activity of Chimeric Antigen Receptor T Cells Against Hepatocellular Carcinoma [59]	Lentivirus	Hepatocellular Carcinoma (HCC)	The PD-1 disruption in the second-generation GPC3-targeted CAR T cells by CRISPR/Cas9.	Promoted Anti-tumor activity of the CAR T cells against HCC
CRISPR knock out of programmed cell death protein 1 enhances anti-tumor activity of cytotoxic T lymphocytes [60]	Lentivirus	Multiple Myeloma (MM)	Impairing PD-1/PD-L1 pathway in CTLs with CRISPR-Cas9 system.	CTLs repressed MM tumor growth and prolonged survival.
Safety and feasibility of CRISPR-edited T cells in patients with refractory non-small-cell lung cancer [61]	Lentivirus	Refractory non-small-cell lung cancer (NSCLC)	CRISPR-Cas9-mediated knock down of PD-1 of T cells in patients with NSCLC	Results showcased the clinical safety and feasibility of CRISPR-Cas9 gene-edited T cells in NSCLC patients.
CRISPR-engineered T cells in patients with refractory cancer [62]	Lentivirus	Refractory cancer	NY-ESO-1 TCR-expressing engineered cells deprived of PD-1 (PDCD1), TCRα (TRAC), and TCRβ (TRBC)	Enhanced anti-tumor immunity and feasibility multiplex CRISPR-Cas9 editing at clinical scale
Activation of concurrent apoptosis and necroptosis by SMAC mimetics for the treatment of refractory and relapsed ALL [63]	Lentivirus	Refractory acute lymphoblastic leukemia (ALL)	Disruption of receptor-interacting protein kinase 1 (RIP1) by using CRISPR and pharmacologic interference (SMAC mimetics).	Hampering cancer cells evading apoptosis, and activating concurrent apoptosis and necroptosis
Genome-wide CRISPR screen identifies HNRNPL as a prostate cancer dependency regulating RNA splicing [64]	Lentivirus	Prostate cancer	CRISPR/Cas9-based gene knockout of essential spliceosome and RNA binding protein (RBP) genes.	A RBP gene aclled HNRNPL is capable of increasing prostate cancer growth.
Genome-wide CRISPR/Cas9 library screen identifies PCMT1 as a critical driver of ovarian cancer metastasis [65]	Lentivirus	Ovarian cancer	PCMT1 (protein-L-isoaspartate (D-aspartate) O-methyltransferase) knockdown by CRISPR/Cas9	PCMT1 increases in vivo metastasis formation and its serum levels may serve as a potential metastatic marker.
Genome-wide CRISPR screen reveals SGOL1 as a druggable target of sorafenib-treated hepatocellular carcinoma [66]	Lentivirus	HCC	In combination with NGS, the genome-wide CRISPR screen was used to determine loss-of-function mutations bestowing sorafenib resistance upon HCC cells.	SGOL1 found to be a druggable target that its inhibition may reduce drug resistance against sorafenib treatment.
Genome-Wide CRISPR-Cas9 Screen Identifies MicroRNAs That Regulate Myeloid Leukemia Cell Growth [67]	Lentivirus	Acute Myeloid Leukemia (AML)	miRNA loss-of-function screening was exerted by CRISPR-Cas9 technology.	Disruption of miR-150 (targeting p53) and miR-155 are therapeutic targets in AML.
Genome-wide CRISPR screen identifies LGALS2 as an oxidative stress-responsive gene with an inhibitory function on colon tumor growth [68]	Lentivirus	Colon cancer	The CRISPR-based screening alongside NGS evaluated the genetic factors involved in the regulation of oxidative stress.	It is reported that Glycan-binding protein Galectin 2 (Gal2) overexpression reduces the human colon tumor growth.
Genome-wide CRISPR-Cas9 screen identified KLF11 as a druggable suppressor for sarcoma cancer stem cells [69]	Lentivirus	Osteosarcoma	The genome-wide CRISPR screening of cancer stem cells (CSCs) of Osteosarcoma identified the regulator of osteosarcoma.	Results showed that Low KLF11 correlates with osteosarcoma’s poor prognosis and inadequate chemotherapy response.
Genome-Scale CRISPR-Cas9 Transcriptional Activation Screening in Metformin Resistance Related Gene of Prostate Cancer [70]	Lentivirus	Prostate cancer	CRISPR-based screening of metformin resistance in prostate cancer to find genes involved in metformin insensitivity.	Activation of ECE1, ABCA12, BPY2, EEF1A1, RAD9A, and NIPSNAP1 associated with in vitro resistance to metformin.
Genome-wide CRISPR-Cas9 knockout library screening identified PTPMT1 in cardiolipin synthesis is crucial to survival in hypoxia in liver cancer [71]	Lentivirus	HCC	Genome-wide CRISPR-Cas9 screening showcased therapeutic factors responsible for hypoxic survival in HCC.	Knockout of PTPMT1 provokes ROS and apoptosis in hypoxic HCC cells.
Identifying novel therapeutic targets in gastric cancer using genome-wide CRISPR-Cas9 screening [72]	Lentivirus	Gastric cancer	The genome-scale CRISPR-Cas9 knock-out library of gastric cancer cells	Among 184 novel genes involved in gastric cancer, methyltransferase 1 (METTL1) inhibition was the most validated approach for cancer-targeted therapy.
In vivo CRISPR/Cas9 targeting of fusion oncogenes for selective elimination of cancer cells [73]	Adenovirus	PDX (patient-derived xenograft) cancer models	CRISPR/Cas9-mediated targeting of two introns of the translocated genes, results in	Disruption of the fusion oncogene in cancer cells, followed by a selective and efficient activity for cancer cell elimination.
Pancreatic cancer modeling using retrograde viral vector delivery and in vivo CRISPR/Cas9-mediated somatic genome editing [74]	Lentivirus and Adenovirus	Pancreatic cancer	CRISPR/Cas9-mediated genomic manipulation of pancreatic cancer cells to develop transgenic mouse lines, allowing titratable initiation of pancreatic tumors	This method paves the way for the investigation of molecular alterations, driving each step of pancreatic cancer development.
Cancer-derived exosomes as a delivery platform of CRISPR/Cas9 confer cancer cell tropism-dependent targeting [75]	Exosome	Ovarian cancer	Tumor-derived exosomes loaded with cas9 and PARP-1 sgRNA expression plasmids via electroporation	CRISPR/Cas9-induced inhibition of PARP-1 resulted in ovarian cancer cell apoptosis and increased sensitivity to chemotherapeutic agent (cisplatin).
Exosome-mediated delivery of CRISPR/Cas9 for targeting of oncogenic KrasG12D in pancreatic cancer [76]	Exosome	Pancreatic cancer	Exosomes loaded with CRISPR/Cas9 capable of targeting the mutant KrasG12D oncogenic	Suppressed proliferation and hampered tumor growth in syngeneic subcutaneous and orthotopic models of pancreatic cancer.
Tropism-facilitated delivery of CRISPR/Cas9 system with chimeric antigen receptor-extracellular vesicles against B-cell malignancies [77]	EVs	B cell malignancies	The CRISPR/Cas9 system aiming at the MYC oncogene, was loaded into selective EVs having anti-CD19-CAR on their surface.	The induced CRISPR/Cas9-mediated loss-of-function mutations of the MYC gene in CD19 + cells exhibited the significant potential of this approach.
Efficient RNA drug delivery using red blood cell extracellular vesicles [78]	RBC extracellular vesicles (RBCEVs)	AML M5	Electroporation of HA-tagged Cas9 mRNA and gRNA of human mir-125b-2 into RBCEVs, and used them to treat MOLM13 cells.	Exosomes successfully transfected both human cells and xenograft mice, with no notable cytotoxicity.
Exosome–Liposome Hybrid Nanoparticles Deliver CRISPR/Cas9 System in MSCs [79]	Exosome	Mesenchymal stem cells (MSCs)	Hybrid exosomes-liposomes capable of carrying large cargoes such as CRISPR/Cas9 System	Hybrid exosomes efficiently delivered CRISPR/dCas9 to inhibit the expression of mRunx2 and hCTNNB1 in MSCs
In vitro and in vivo RNA inhibition by CD9-HuR functionalized exosomes encapsulated with miRNA or CRISPR/dCas9 [80]	Exosome	Recipient cells	Since Hur is an RNA binding protein, CD9-HuR exosomes could efficiently encapsulate the miR-155 or CRISPR/dCas9	Increased RNA cargo loading into engineered exosomes
Activation of Necroptosis by Engineered Self Tumor-Derived Exosomes Loaded with CRISPR/Cas9 [55]	Exosome	GBM, Thyroid cancer, lung adenocarcinoma	Engineered exosomes for TNFR activation and impairment of IAP 1/2 and Caspase 8 expression loaded with CRISPR/CAS9	Activation of TNFR and subsequently inactivation of IAP 1/2 and Caspase results in blocked cell survival and Necroptosis activation.