The heterogeneity of pancreatic ductal adenocarcinoma (PDAC) suggests that successful treatment might rely on simultaneous targeting of multiple genes, which can be achieved by RNA interference-based ...therapeutic strategies. Here we show a potent combination of microRNA and siRNA delivered by an efficient nanocarrier to PDAC tumors. Using proteomic-microRNA profiles and survival data of PDAC patients from TCGA, we found a novel signature for prolonged survival. Accordingly, we used a microRNA-mimic to increase miR-34a together with siRNA to silence PLK1 oncogene. For in vivo dual-targeting of this combination, we developed a biodegradable amphiphilic polyglutamate amine polymeric nanocarrier (APA). APA-miRNA-siRNA polyplexes systemically administered to orthotopically inoculated PDAC-bearing mice showed no toxicity and accumulated at the tumor, resulting in an enhanced antitumor effect due to inhibition of MYC oncogene, a common target of both miR-34a and PLK1. Taken together, our findings warrant this unique combined polyplex's potential as a novel nanotherapeutic for PDAC.
Background: Immune checkpoint inhibitors have transformed clinical oncology. However, their use is limited as response is observed in only ~20–50% of patients. Previously, we demonstrated that ...treating CT26 tumor-bearing mice with ultra-high-concentration gaseous nitric oxide (UNO) followed by tumor resection stimulated antitumor immune responses. Accordingly, UNO may improve tumor response to immune checkpoint inhibitors. Here, we investigated the ability of UNO to improve the efficacy of a programmed cell death protein-1 (PD-1) antibody in vitro and in treating CT26 tumor-bearing mice. Methods: CT26 cells were injected into the flank of Balb/c mice (n = 15–16 per group). On day 6, CT26 cells were injected into the contralateral flank, and anti-mPD-1 injections commenced. Primary tumors were treated with intratumoral UNO on day 8. Tumor volume, response rates, toxicity, and survival were monitored. Results: (1) Short exposure to 25,000–100,000 parts per million (ppm) UNO in vitro resulted in significant upregulation of PD-L1 expression on CT26 cells. (2) UNO treatment in vivo consistently reduced cell viability in CT26 tumors. (3) Treatment reduced regulatory T-cell (Treg) levels in the tumor and increased levels of systemic M1 macrophages. UNO responders had increased CD8+ T-cell tumor infiltration. (4) Nine days after treatment, primary tumor growth was significantly lower in the combination arm vs. anti-mPD-1 alone (p = 0.0005). (5) Complete tumor regression occurred in 8/15 (53%) of mice treated with a combination of 10 min UNO and anti-mPD-1, 100 days post-treatment, compared to 4/16 (25%) of controls treated with anti-mPD-1 alone (p = 0.1489). (6) There was no toxicity associated with UNO treatment. (7) Combination treatment showed a trend toward increased survival 100 days post-treatment compared to anti-mPD-1 alone (p = 0.0653). Conclusion: Combining high-concentration NO and immune checkpoint inhibitors warrants further assessment especially in tumors resistant to checkpoint inhibitor therapy.
Abstract The progress in medical research has led to the understanding that cancer is a large group of heterogeneous diseases, with high variability between and within individuals. This variability ...sprouted the ambitious goal to improve therapeutic outcomes, while minimizing drug adverse effects through stratification of patients by the differences in their disease markers, in a personalized manner, as opposed to the strategy of “one therapy fits all”. Nanotheranostics, composed of nanoparticles (NPs) carrying thera peutic and/or diag nostics probes, have the potential to revolutionize personalized medicine. There are different modalities to combine these two distinct fields into one system for a synergistic outcome. The addition of a nanocarrier to a theranostic system holds great promise. Nanocarriers possess high surface area, enabling sophisticated functionalization with imaging agents, thus gaining enhanced diagnostic ability in real-time. Yet, most of the FDA-approved theranostics approaches are based on small molecules. The theranostic approaches that are reviewed herein are paving the road towards personalized medicine through all stages of patient care: starting from screening and diagnostics, proceeding to treatment and ending with treatment follow-up. Our current review provides a broad background and highlights new insights for the rational design of theranostic nanosystems for desired therapeutic niches, while summoning the hurdles on their way to become first-line diagnostics and therapeutics for cancer patients.
Abstract Polymeric nanocarriers conjugated with low molecular weight drugs are designed in order to improve their efficacy and toxicity profile. This approach is particularly beneficial for ...anticancer drugs, where the polymer–drug conjugates selectively accumulate at the tumor site, due to the enhanced permeability and retention (EPR) effect. The conjugated drug is typically inactive, and upon its pH- or enzymatically-triggered release from the carrier, it regains its therapeutic activity. These settings lack information regarding drug-release time, kinetics and location. Thereby, real-time non-invasive intravital monitoring of drug release is required for theranostics ( thera py and diag nostics ). We present here the design, synthesis and characterization of a theranostic nanomedicine, based on N -(2-hydroxypropyl) methacrylamide (HPMA) copolymer, owing its fluorescence-based monitoring of site-specific drug release to a self-quenched near-infrared fluorescence (NIRF) probe. We designed two HPMA copolymer-based systems that complement to a theranostic nanomedicine. The diagnostic system consists of self-quenched Cy5 (SQ-Cy5) as a reporter probe and the therapeutic system is based on the anticancer agent paclitaxel (PTX). HPMA copolymer–PTX/SQ-Cy5 systems enable site-specific release upon enzymatic degradation in cathepsin B-overexpressing breast cancer cells. The release of the drug occurs concomitantly with the activation of the fluorophore to its Turn-ON state. HPMA copolymer-SQ-Cy5 exhibits preferable body distribution and drug release compared with the free drug and probe when administered to cathepsin B-overexpressing 4T1 murine mammary adenocarcinoma-bearing mice. This approach of co-delivery of two complementary systems serves as a proof-of-concept for real-time deep tissue intravital orthotopic monitoring and may have the potential use in clinical utility as a theranostic nanomedicine.
Complete tumor removal during surgery has a great impact on patient survival. To that end, the surgeon should detect the tumor, remove it and validate that there are no residual cancer cells left ...behind. Residual cells at the incision margin of the tissue removed during surgery are associated with tumor recurrence and poor prognosis for the patient. In order to remove the tumor tissue completely with minimal collateral damage to healthy tissue, there is a need for diagnostic tools that will differentiate between the tumor and its normal surroundings.
We designed, synthesized and characterized three novel polymeric Turn-ON probes that will be activated at the tumor site by cysteine cathepsins that are highly expressed in multiple tumor types. Utilizing orthotopic breast cancer and melanoma models, which spontaneously metastasize to the brain, we studied the kinetics of our polymeric Turn-ON nano-probes.
To date, numerous low molecular weight cathepsin-sensitive substrates have been reported, however, most of them suffer from rapid clearance and reduced signal shortly after administration. Here, we show an improved tumor-to-background ratio upon activation of our Turn-ON probes by cathepsins. The signal obtained from the tumor was stable and delineated the tumor boundaries during the whole surgical procedure, enabling accurate resection.
Our findings show that the control groups of tumor-bearing mice, which underwent either standard surgery under white light only or under the fluorescence guidance of the commercially-available imaging agents ProSense® 680 or 5-aminolevulinic acid (5-ALA), survived for less time and suffered from tumor recurrence earlier than the group that underwent image-guided surgery (IGS) using our Turn-ON probes. Our "smart" polymeric probes can potentially assist surgeons' decision in real-time during surgery regarding the tumor margins needed to be removed, leading to improved patient outcome.
BackgroundImmune checkpoint inhibitors have shown dramatic activity transforming clinical oncology.1 Yet, their activity is limited to a subset of highly sensitive tumors (e.g., melanoma). Even then ...response is observed in only 50% of patients.2Nitric Oxide (NO) is a signaling molecule in multiple diseases, including cancer and has been shown to activate anti-tumor immune responses.3 Previously, we reported that treatment of CT26 tumor-bearing mice with high-concentration NO (UNO) stimulated anti-tumor immune responses leading to the rejection of a secondarily-induced tumor. More specifically, a significant increase of tumor-infiltrating T-cells and blood and spleen B and T-cells were observed 14-21 days post-UNO treatment.In this study we investigated the ability of UNO to improve the efficacy of anti-PD-1 antibody.MethodsDay zero, 5.0 X 106 CT26 cells were injected to the right flank of Balb/c mice (n=15-16 per group). Day six, CT26 cells were injected to the contralateral flank and anti-PD1 injections (5mg/kg mouse, q2d, x 5) commenced.Day eight, tumors (average size 71.9±37.2mm3) were treated intratumorally with UNO (50,000 ppm, 5 or 10 minutes, flow rate ~0.2 liter per minute). Post-treatment tumor volume and survival were monitored thereafter.ResultsComplete regression of the primary tumor occurred in 9/15 (60%) of mice treated with combination 10-minute NO and anti-PD-1, post-treatment day 26. This compared to 4/16 (25%) of controls treated with anti-PD-1 alone (p=0.13) and 0/15 (0%) treated with UNO alone (p=0.0027).Survival was drastically increased in the 10-minute UNO/anti-PD-1 combination arm compared to anti-PD-1 alone (p<0.05), post-treatment day 32.Secondary, contralateral flank tumor take in the anti-PD-1 alone arm was 21.4% but reduced by 38% to 13.3% in the 10-min combination arm and reduced by 67% to 7.14% in the 5-min combination arm, post-treatment day 19.Survival was significantly improved for both the 5- and 10-minute combination arms compared to the 5- and 10-minute UNO controls (p=0.02 and p<0.0001, respectively).ConclusionsCombination of UNO with anti-PD-1 significantly improved outcomes compared with UNO or anti-PD-1 alone. A strong possibility is that high-concentration NO assists the immune system in overcoming anti-PD-1 resistance. Thus, the combination of high-concentration NO and immune checkpoint inhibitors such as anti-PD-1 can be a breakthrough therapy with important clinical implications.ReferencesPardoll DM. The blockade of immune checkpoints in cancer immunotherapy. Nat Rev Cancer 2012; 12(4):252–264. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4856023/Schoenfeld AJ, Hellmann MD. Acquired resistance to immune checkpoint inhibitors. Cancer Cell 2020; 37(4):443–455. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7182070/Khan FH, Dervan E, Bhattacharyya DD, McAuliffe JD, Miranda KM, Glynn SA. The role of nitric oxide in cancer: master regulator or not? Int J Mol Sci 2020; 21(24):9393–9423. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7763974/
Cutaneous melanoma is the most aggressive and deadliest of all skin malignancies. Complete primary tumor removal augmented by advanced imaging tools and effective post-operative treatment is critical ...in the prevention of tumor recurrence and future metastases formation.
To meet this challenge, we designed novel polymeric imaging and therapeutic systems, implemented in a two-step theranostic approach. Both are composed of the biocompatible and biodegradable poly(α,L-glutamic acid) (PGA) nanocarrier that facilitates extravasation-dependent tumor targeting delivery. The first system is a novel, fluorescent, Turn-ON diagnostic probe evaluated for the precise excision of the primary tumor during image-guided surgery (IGS). The fluorescence activation of the probe occurs via PGA degradation by tumor-overexpressed cathepsins that leads to the separation of closely-packed, quenched FRET pair. This results in the emission of a strong fluorescence signal enabling the delineation of the tumor boundaries. Second, therapeutic step is aimed to prevent metastases formation with minimal side effects and maximal efficacy. To that end, a targeted treatment containing a BRAF (Dabrafenib - mDBF)/MEK (Selumetinib - SLM) inhibitors combined on one polymeric platform (PGA-SLM-mDBF) was evaluated for its anti-metastatic, preventive activity in combination with immune checkpoint inhibitors (ICPi) αPD1 and αCTLA4.
IGS in melanoma-bearing mice led to a high tumor-to-background ratio and reduced tumor recurrence in comparison with mice that underwent surgery under white light (23%
33%, respectively). Adjuvant therapy with PGA-SLM-mDBF combined with ICPi, was well-tolerated and resulted in prolonged survival and prevention of peritoneal and brain metastases formation in BRAF-mutated melanoma-bearing mice.
The results reveal the great clinical potential of our PGA-based nanosystems as a tool for holistic melanoma treatment management.
Targeted therapies against cancer can relieve symptoms and induce remission; however, they often present limited duration of disease control, cause side effects, and may induce acquired resistance. ...Therefore, there is great motivation to develop a unique delivery system, targeted to the tumor, in which several active entities can be combined, the therapeutic index can be increased by reducing systemic exposure, and their synergistic activity can be enhanced. To meet these goals, the biocompatible and biodegradable poly(α,l‐glutamic acid) (PGA) is chosen as a nanocarrier that facilitates extravasation‐dependent tumor targeting delivery. The RAS/RAF/MEK/ERK pathway when aberrantly activated in melanoma, can lead to uncontrolled cell proliferation, induced invasion, and reduced apoptosis. Here, two drugs targeting this pathway are selected: a MEK1/2 inhibitor (selumetinib, SLM) and a modified BRAF inhibitor (modified dabrafenib, mDBF) that exhibit synergism in vitro. The combination of PGA conjugated to SLM and mDBF (PGA–SLM–mDBF) is synthesized and characterized. PGA–SLM–mDBF inhibits the proliferation of melanoma cells and decreases their migratory and sprouting abilities without inducing a hemolytic effect. Moreover, it exhibits superior antitumor activity in a mouse model of primary melanoma and prolonged survival at a lower dose than the free drugs.
This study presents a novel polymeric system that simultaneously delivers a combination of drugs that target BRAF/ MEK mutations in melanoma. This system possesses an improved solubility, stability, and selective release by cathepsins in the tumor site. Hence, it exhibits synergistic activity, with superior efficacy and safety compared to each monodrug polymer conjugate and combinations of the free drugs.
In article number 2000028 Ronit Satchi‐Fainaro and co‐workers present a novel polymeric system that simultaneously delivers a combination of drugs that target BRAF/ MEK mutations in melanoma. This ...system possesses an improved solubility, stability, and selective release by cathepsins in the tumor site. Hence, it exhibits synergistic activity, with superior efficacy and safety compared to each monodrug polymer conjugate and combinations of the free drugs.
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