PMC0
10.2147%2FIJN.S47129
introduction
These macromolecules have a different biodistribution profile, a prolonged plasma half-life, and reduced metabolism of the encapsulated drug.>>1<< One of the most commonly cited reasons, however, for the use of nanomedicine is the ability of macromolecular compounds to exploit the enhanced permeability and retention (EPR) effect to specifically target tumor (Figure 1) and inflamed
the most commonly cited reasons, however, for the use of nanomedicine is the ability of macromolecular compounds to exploit the enhanced permeability and retention (EPR) effect to specifically target tumor (Figure 1) and inflamed tissues.>>1<<
factors. The imbalance in expression and activation of proangiogenic and antiangiogenic molecules results in a discontinuous endothelial layer, where fenestrations between the endothelial cells may range from 300 nm to 4,700 nm in size.>>2<< In addition to the formation of this porous vasculature, impaired lymphatic drainage may also occur in tumor tissue due to dysfunctional lymphangiogenesis and compression of the lymphatic vessels by growing numbers of cancer cells.
As a result of this insufficient lymphatic drainage, constructs present in the interstitial fluid of tumors are retained for longer than in normal tissues with functional lymphatics.>>3<<
Exploitation of the EPR effect via use of nanoconstructs has been shown to consistently increase the fraction of the injected drug dose that reaches the tumor tissue.>>7<< Furthermore, the absence of large fenestrations in nonpathological tissues prevents the extravasation and deposition of macromolecular constructs in healthy tissue and thereby reduces the level of systemic side effects.
of macromolecular constructs in healthy tissue and thereby reduces the level of systemic side effects. The two notable exceptions to this rule are the liver and spleen, because both of these organs commonly have large fenestrations.>>7<< As a result of reduced nonspecific extravasation and circulating free drug, adverse effects are greatly reduced with the use of nanomedicine.
drug, adverse effects are greatly reduced with the use of nanomedicine. This effect was clearly demonstrated with a reduction in doxorubicin-induced cardiotoxicity following treatment with Doxil®, a liposomal formulation of doxorubicin.>>8<< However, an increased incidence of palmar–plantar erythrodysesthesia has been observed to occur as a consequence of use of this construct.
8 However, an increased incidence of palmar–plantar erythrodysesthesia has been observed to occur as a consequence of use of this construct.>>9<<
challenges to the epr effect
Interstitial fluid pressure (IFP) in tumor tissue is a significant barrier that prevents the penetration of therapeutic agents.>>10<< The elevation of IFP that is commonly seen in tumor tissue has been found to occur as a result of numerous factors.
and impedes the movement of extravasated nanoconstructs. Thirdly, compression of lymphatic vessels as a result of the pressure of the growing tumor results in a reduction of interstitial fluid drainage and thereby a net gain in IFP.>>11<< Finally, IFP is also increased as a response to contraction of the interstitial space, an effect mediated by stromal fibroblasts.
of the growing tumor results in a reduction of interstitial fluid drainage and thereby a net gain in IFP.11 Finally, IFP is also increased as a response to contraction of the interstitial space, an effect mediated by stromal fibroblasts.>>12<< All of these factors play a key role in elevating the IFP of tumor tissue and consequently reducing the penetration of therapeutic macromolecules.
Treatment with DC101 40 mg/kg increased the penetration of TRITC-BSA (molecular weight 71,000 Da)>>13<< from 7.26 μm to 11.23 μm from the blood vessel one hour post administration.
(molecular weight 71,000 Da)13 from 7.26 μm to 11.23 μm from the blood vessel one hour post administration. This enhanced infiltration was suggested to be the consequence of treatment with DC101 lowering IFP by approximately 45%.>>14<< Repeated administration of a VEGF monoclonal antibody (six doses of 100 μg per mouse) was reported to decrease IFP in tumors by up to 74% in a mouse model of colorectal adenocarcinoma.
14 Repeated administration of a VEGF monoclonal antibody (six doses of 100 μg per mouse) was reported to decrease IFP in tumors by up to 74% in a mouse model of colorectal adenocarcinoma.>>15<< However, given that no investigation into the correlation between IFP and the integrity of the vasculature has been conducted, other potential mechanisms, such as tumor cell death or a reduction in microvessel density, may account for the
Promisingly, a 73% reduction in IFP was observed in four patients with rectal carcinoma treated with bevacizumab, a humanized monoclonal antibody targeting VEGF-A,>>16<< indicating that an efficient reduction in IFP is achievable in the clinic.
normalization of blood vessels and thereby reduced IFP is also expected to significantly reduce extravasation, because vascular normalization also reduces permeability of the vasculature to macromolecules such as albumin by up to 51%.>>14<< Furthermore, the temporal window for efficacy of these drugs is narrow, as little as 6 days, and may show a great deal of heterogeneity depending on the patient and tumor being treated.
14 Furthermore, the temporal window for efficacy of these drugs is narrow, as little as 6 days, and may show a great deal of heterogeneity depending on the patient and tumor being treated.>>17<<
This is due to the observation that it is through functional lymphatics that tumor cells metastasize, so normalization of these vessels may increase the risk of metastasis.>>18<<
Simulation of the platelet-derived growth factor receptor (PDGFR) has been shown to increase IFP via contraction of stromal cells, the integrins of which interact with the extracellular matrix.>>19<<–21 Administration of the PDGFR-β inhibitor, STI-571, was shown to decrease IFP in colon carcinoma by 32%20 and in thyroid carcinoma by 58%,22 while imatinib, a PDGFR inhibitor, reduced IFP in a mouse model of non-small-cell lung cancer by
19–21 Administration of the PDGFR-β inhibitor, STI-571, was shown to decrease IFP in colon carcinoma by 32%>>20<< and in thyroid carcinoma by 58%,22 while imatinib, a PDGFR inhibitor, reduced IFP in a mouse model of non-small-cell lung cancer by 28%.
19–21 Administration of the PDGFR-β inhibitor, STI-571, was shown to decrease IFP in colon carcinoma by 32%20 and in thyroid carcinoma by 58%,>>22<< while imatinib, a PDGFR inhibitor, reduced IFP in a mouse model of non-small-cell lung cancer by 28%.
of the PDGFR-β inhibitor, STI-571, was shown to decrease IFP in colon carcinoma by 32%20 and in thyroid carcinoma by 58%,22 while imatinib, a PDGFR inhibitor, reduced IFP in a mouse model of non-small-cell lung cancer by 28%.>>21<< Inhibition of PDGFR activity, while appearing to be less effective than vascular normalization in terms of lowering IFP, does not result in reduced vascular fenestrations, thereby preserving extravasation of macromolecules.
21 Inhibition of PDGFR activity, while appearing to be less effective than vascular normalization in terms of lowering IFP, does not result in reduced vascular fenestrations, thereby preserving extravasation of macromolecules.>>22<<
Furthermore, variations in IFP measured in human tumors range from −1 mmHg to 94 mmHg, making it unreasonable to generalize the role of elevated IFP to all types of human cancer.>>23<<
efflux small molecules as a protective mechanism. In certain brain tumors, such as glioblastoma multiforme, the blood–brain barrier does show a degree of disruption, allowing macromolecules to penetrate into the central nervous system.>>24<<,25 However, infiltration of macromolecules into this tissue is impeded by the inability of large molecular weight constructs to diffuse easily through the interstitial space.
small molecules as a protective mechanism. In certain brain tumors, such as glioblastoma multiforme, the blood–brain barrier does show a degree of disruption, allowing macromolecules to penetrate into the central nervous system.24,>>25<< However, infiltration of macromolecules into this tissue is impeded by the inability of large molecular weight constructs to diffuse easily through the interstitial space.
Furthermore, glioblastoma multiforme cells are highly invasive, and the presence of even a small number of remaining cells makes post-surgical recurrence essentially universal within this patient population.>>26<< The residual cells that remain in the brain are supported by the existing vasculature and due to the lack of neoangiogenic vessels in the spreading tumor lesion, the blood–brain barrier still prevents chemotherapeutics from reaching the
These factors have traditionally made treatment of central nervous system tumors difficult and resulted in the application of systemic antibodies, macromolecular drug carriers, and cytolytic viruses being negatively impacted.>>27<< For these reasons, convection-enhanced delivery (CED), which involves the utilization of a positive pressure gradient applied during an infusion, has been investigated as a possible mechanism to overcome these challenges.
The use of this procedure is currently being investigated for the treatment of central nervous system pathologies such as glioblastoma multiforme and Parkinson’s disease.>>27<<
The use of this procedure is currently being investigated for the treatment of central nervous system pathologies such as glioblastoma multiforme and Parkinson’s disease.27,>>28<<
Clear evidence of CED-mediated improvement has been demonstrated, with one study showing a 115-fold increase in volume of distribution of 50 nm lipid nanocapsules when compared with a classical injection of the same nanoparticles.>>29<<
administration of liposomal irinotecan or topotecan into the central nervous system of Sprague-Dawley rats via CED has been seen to cause a 65.6-fold and 15-fold increase, respectively, in half-life relative to that of the free drug.>>30<<,31 Similarly, liposomal and micellar doxorubicin administered via CED showed a 4.6-fold and 3.9-fold increase in the volume of distribution when compared with the free drug.
administration of liposomal irinotecan or topotecan into the central nervous system of Sprague-Dawley rats via CED has been seen to cause a 65.6-fold and 15-fold increase, respectively, in half-life relative to that of the free drug.30,>>31<< Similarly, liposomal and micellar doxorubicin administered via CED showed a 4.6-fold and 3.9-fold increase in the volume of distribution when compared with the free drug.
30,31 Similarly, liposomal and micellar doxorubicin administered via CED showed a 4.6-fold and 3.9-fold increase in the volume of distribution when compared with the free drug.>>32<<
Transport through tumor tissue is also highly complex, with macromolecules showing differential distribution depending on the homogeneity of the tumor tissue itself.>>33<< In addition to this, factors such as preinfusion sealing time, rate of infusion, size of the delivery cannula, and concentration and leakage of the infusate into the subarachnoid space or surgical cavity must also be considered when
factors such as preinfusion sealing time, rate of infusion, size of the delivery cannula, and concentration and leakage of the infusate into the subarachnoid space or surgical cavity must also be considered when applying this technique.>>34<<
such as preinfusion sealing time, rate of infusion, size of the delivery cannula, and concentration and leakage of the infusate into the subarachnoid space or surgical cavity must also be considered when applying this technique.34,>>35<<
varies considerably depending on the tumor being treated. For example, the microvessel density in hepatocellular carcinoma and colorectal carcinoma may be up to 127.2 vessels per field and 145 vessels per field, respectively.>>7<< Conversely, squamous cell carcinoma and stage I–II non-small-cell lung cancer may have a microvessel density of only 14.8 vessels per field and 23.6 vessels per field, respectively.
per field and 145 vessels per field, respectively.7 Conversely, squamous cell carcinoma and stage I–II non-small-cell lung cancer may have a microvessel density of only 14.8 vessels per field and 23.6 vessels per field, respectively.>>7<< In tumor tissues where the microvessel density is low, the capacity to deliver nanomedicine effectively is significantly reduced, given that the distance that the macromolecule will have to diffuse through is significantly increased.
The liver and spleen have large endothelial fenestrations (~100 nm and 5 μm, respectively),>>36<<,37 so macromolecules will extravasate into these tissues. However, the extent to which these macromolecules accumulate within the liver and spleen is determined by several parameters.
The liver and spleen have large endothelial fenestrations (~100 nm and 5 μm, respectively),36,>>37<< so macromolecules will extravasate into these tissues. However, the extent to which these macromolecules accumulate within the liver and spleen is determined by several parameters.
Greater accumulation of macromolecules within the liver is sometimes observed in preclinical studies despite the larger fenestration size in the spleen,>>7<< a distribution pattern likely to be a consequence of the high blood flow through the liver relative to the spleen (1.8 mL versus 0.09 mL per minute in mice, 13.8 mL versus 0.63 mL per minute in rats, and 1,450 mL versus 77 mL per minute
pattern likely to be a consequence of the high blood flow through the liver relative to the spleen (1.8 mL versus 0.09 mL per minute in mice, 13.8 mL versus 0.63 mL per minute in rats, and 1,450 mL versus 77 mL per minute in humans).>>38<< Previous research has shown that the degree of accumulation of macromolecules within these organs may be further influenced by the nature of the construct.
Modulation of construct parameters such as size, shape, and charge>>7<<,39 may therefore alter the accumulation of these macromolecules in various organs and prevent the development of unwanted effects.
Modulation of construct parameters such as size, shape, and charge7,>>39<< may therefore alter the accumulation of these macromolecules in various organs and prevent the development of unwanted effects.
For example, cylindrical silicone particles have been shown to accumulate in the liver with a quantity two-fold that of spherical and quasihemispherical particles and five-fold that of discoidal particles.>>39<< Nevertheless, it has been demonstrated that nanomedicines can accumulate and be retained in hepatic tumors at a concentration higher than that of the surrounding healthy liver,3 an effect which can be attributed to differential lymphatic
39 Nevertheless, it has been demonstrated that nanomedicines can accumulate and be retained in hepatic tumors at a concentration higher than that of the surrounding healthy liver,>>3<< an effect which can be attributed to differential lymphatic drainage in healthy and cancerous tissues.
improvement of the epr effect at the tissue level
an external magnetic field. This study showed an 11.5-fold greater accumulation of iron oxide nanoparticles in tumor tissue when the animals were concomitantly subjected to a localized magnetic field (0.4 T) relative to control animals.>>40<< Furthermore, in animals exposed to magnetic targeting, accumulation of iron oxide nanoparticles in tumor tissues was 9.5-fold higher than accumulation within the contralateral brain,40 indicating significant tissue specificity.
40 Furthermore, in animals exposed to magnetic targeting, accumulation of iron oxide nanoparticles in tumor tissues was 9.5-fold higher than accumulation within the contralateral brain,>>40<< indicating significant tissue specificity.
This may result in the applied magnetic strength exceeding the 8 T considered safe by the US Food and Drug Administration.>>41<<
cellular uptake of therapeutics, although not indisputable, is thought to be due to membrane disruption. This disturbance results in increased permeability to macromolecular agents and the drugs they release in response to ultrasound.>>42<< Ultrasound serves as a noninvasive, highly manipulable method to disrupt cell membranes and enhance capillary permeability.
In these studies, the uptake of micelles loaded with doxorubicin was significantly improved following application of ultrasound (30 seconds with 1 mHz)>>42<<,43 and was sustained for at least 8 hours.
In these studies, the uptake of micelles loaded with doxorubicin was significantly improved following application of ultrasound (30 seconds with 1 mHz)42,>>43<< and was sustained for at least 8 hours.
In these studies, the uptake of micelles loaded with doxorubicin was significantly improved following application of ultrasound (30 seconds with 1 mHz)42,43 and was sustained for at least 8 hours.>>42<< However, it must be noted that there has been an indication that areas not subject to direct sonication also show evidence of an increased concentration of doxorubicin, indicating nonspecific effects of ultrasound that could be
be noted that there has been an indication that areas not subject to direct sonication also show evidence of an increased concentration of doxorubicin, indicating nonspecific effects of ultrasound that could be potentially detrimental.>>42<< In addition to these findings, the extended use of ultrasound has the potential to adversely affect the integrity of the nanomedicine formulation.
J6456 lymphoma lesions showed that 120 seconds of low frequency ultrasound application induced release of 70% of the encapsulated cisplatin, representing a 23.3-fold increase compared with the release in the absence of ultrasound.>>44<<
is achieved by a process known as cavitation. This technique uses microbubbles that collapse under ultrasonic pressure to generate mechanical stress that can then act to transiently increase the permeability of the blood–brain barrier.>>45<<,46 This method is already being examined in order to treat pathologies such as glioblastoma multiforme and Alzheimer’s disease.
achieved by a process known as cavitation. This technique uses microbubbles that collapse under ultrasonic pressure to generate mechanical stress that can then act to transiently increase the permeability of the blood–brain barrier.45,>>46<< This method is already being examined in order to treat pathologies such as glioblastoma multiforme and Alzheimer’s disease.
mechanical stress that can then act to transiently increase the permeability of the blood–brain barrier.45,46 This method is already being examined in order to treat pathologies such as glioblastoma multiforme and Alzheimer’s disease.>>47<<–49 Utilization of ultrasound in animals larger than mice may, however, be problematic due to the effect of cranial bone attenuation and scattering.
a porcine model of ultrasound-induced blood–brain barrier disruption, ie, 1 mHz for 30 minutes, the increase in Evans blue dye extravasation in ultrasound-treated portions of the brain increased by ~30% relative to nontreated portions.>>50<< Unfortunately, the groups were highly heterogeneous and the effect was abolished 120 minutes following sonication.
50 Unfortunately, the groups were highly heterogeneous and the effect was abolished 120 minutes following sonication.>>50<<
Examination of this feature has shown the involvement of several factors, including a lack of a functional smooth muscle layer around the blood vessels,>>51<<,52 a significantly reduced density of angiotensin II (AT-II) receptors53–55 and bradykinin receptors,56 and an increased concentration of nitric oxide,57 presumably as a consequence of increased levels of inducible nitric oxide
Examination of this feature has shown the involvement of several factors, including a lack of a functional smooth muscle layer around the blood vessels,51,>>52<< a significantly reduced density of angiotensin II (AT-II) receptors53–55 and bradykinin receptors,56 and an increased concentration of nitric oxide,57 presumably as a consequence of increased levels of inducible nitric oxide synthase58,59
Examination of this feature has shown the involvement of several factors, including a lack of a functional smooth muscle layer around the blood vessels,51,52 a significantly reduced density of angiotensin II (AT-II) receptors>>53<<–55 and bradykinin receptors,56 and an increased concentration of nitric oxide,57 presumably as a consequence of increased levels of inducible nitric oxide synthase58,59 within tumors.
feature has shown the involvement of several factors, including a lack of a functional smooth muscle layer around the blood vessels,51,52 a significantly reduced density of angiotensin II (AT-II) receptors53–55 and bradykinin receptors,>>56<< and an increased concentration of nitric oxide,57 presumably as a consequence of increased levels of inducible nitric oxide synthase58,59 within tumors.
including a lack of a functional smooth muscle layer around the blood vessels,51,52 a significantly reduced density of angiotensin II (AT-II) receptors53–55 and bradykinin receptors,56 and an increased concentration of nitric oxide,>>57<< presumably as a consequence of increased levels of inducible nitric oxide synthase58,59 within tumors.
a significantly reduced density of angiotensin II (AT-II) receptors53–55 and bradykinin receptors,56 and an increased concentration of nitric oxide,57 presumably as a consequence of increased levels of inducible nitric oxide synthase>>58<<,59 within tumors.
a significantly reduced density of angiotensin II (AT-II) receptors53–55 and bradykinin receptors,56 and an increased concentration of nitric oxide,57 presumably as a consequence of increased levels of inducible nitric oxide synthase58,>>59<< within tumors.
Conversely, tumor vasculature has fewer AT-II receptors, a defective smooth muscle layer, and a higher concentration of nitric oxide.>>60<< These parameters mean that, while healthy tissues may respond appropriately to AT-II and maintain the same blood flow, the blood flow through tumor vasculature will increase as the vessel lumen enlarges in response to increased blood
The use of AT-II has been demonstrated to be highly efficacious at enhancing the selective delivery of nanomedicine to tumor tissue while increasing the tumor blood flow by 2–6-fold relative to normal tissues.>>61<< The increase in blood flow will cause a greater amount of drug to pass through the tumor vasculature, which in turn will also augment the EPR effect.
Enhancement of the EPR effect has been demonstrated by administration of the >>51<<Cr-labeled nanomedicine known as styrene-co-maleic-acid polymer conjugated to neocarzinostatin (SMANCS) following AT-II-induced hypertension.
known as styrene-co-maleic-acid polymer conjugated to neocarzinostatin (SMANCS) following AT-II-induced hypertension. This treatment resulted in a 1.3–3-fold increase in tumor accumulation of SMANCS, depending on blood pressure.>>62<< Remarkably, use of AT-II also showed the potential to decrease the quantity of SMANCS in healthy tissues because constriction of healthy blood vessels decreases the size of the endothelial fenestrations.
62 Remarkably, use of AT-II also showed the potential to decrease the quantity of SMANCS in healthy tissues because constriction of healthy blood vessels decreases the size of the endothelial fenestrations.>>62<< The degree of selectivity for tumor tissue achieved using this process is remarkable and constitutes the first example of this type of manipulation, thereby providing a platform for advancement of this technique and the development of
been found to react with membranous matrix metalloproteinase 2, which is specifically expressed in a wide variety of tumor tissues, including glioblastoma multiforme, pilocytic astrocytoma, small cell lung carcinoma, and Ewing’s sarcoma.>>63<< Chlorotoxin targeting produces a significant increase in tumor localization of nanoparticles in mouse models of glioblastoma multiforme64,65 and specific localization of radioactive iodine conjugates in humans with glioblastoma multiforme
63 Chlorotoxin targeting produces a significant increase in tumor localization of nanoparticles in mouse models of glioblastoma multiforme>>64<<,65 and specific localization of radioactive iodine conjugates in humans with glioblastoma multiforme lesions in Phase I/II trials.
63 Chlorotoxin targeting produces a significant increase in tumor localization of nanoparticles in mouse models of glioblastoma multiforme64,>>65<< and specific localization of radioactive iodine conjugates in humans with glioblastoma multiforme lesions in Phase I/II trials.
a significant increase in tumor localization of nanoparticles in mouse models of glioblastoma multiforme64,65 and specific localization of radioactive iodine conjugates in humans with glioblastoma multiforme lesions in Phase I/II trials.>>66<<,67 This method of targeting shows immense promise; however, no extensive Phase III clinical trials involving chlorotoxin-conjugated nanomedicine have been carried out in humans to date.
increase in tumor localization of nanoparticles in mouse models of glioblastoma multiforme64,65 and specific localization of radioactive iodine conjugates in humans with glioblastoma multiforme lesions in Phase I/II trials.66,>>67<< This method of targeting shows immense promise; however, no extensive Phase III clinical trials involving chlorotoxin-conjugated nanomedicine have been carried out in humans to date.
Secreted phospholipase A2 shows increased expression in a range of tumor types (including esophageal adenocarcinoma, prostate cancer, and hepatocellular carcinoma)>>68<<–70 and inflammatory conditions such as psoriasis and rheumatoid arthritis.
phospholipase A2 shows increased expression in a range of tumor types (including esophageal adenocarcinoma, prostate cancer, and hepatocellular carcinoma)68–70 and inflammatory conditions such as psoriasis and rheumatoid arthritis.>>71<<,72 Liposomes sensitive to degradation by secreted phospholipase A2 have been synthesized with the purpose of producing a long circulating nanomedicine that can be selectively degraded at pathological sites.
phospholipase A2 shows increased expression in a range of tumor types (including esophageal adenocarcinoma, prostate cancer, and hepatocellular carcinoma)68–70 and inflammatory conditions such as psoriasis and rheumatoid arthritis.71,>>72<< Liposomes sensitive to degradation by secreted phospholipase A2 have been synthesized with the purpose of producing a long circulating nanomedicine that can be selectively degraded at pathological sites.
and rheumatoid arthritis.71,72 Liposomes sensitive to degradation by secreted phospholipase A2 have been synthesized with the purpose of producing a long circulating nanomedicine that can be selectively degraded at pathological sites.>>73<<,74 Curiously, the degree of liposome degradation by secreted phospholipase A2 increases with the quantity of poly(ethylene glycol) (PEG)-lipid conjugates incorporated into the liposome, in a charge-dependent manner.
and rheumatoid arthritis.71,72 Liposomes sensitive to degradation by secreted phospholipase A2 have been synthesized with the purpose of producing a long circulating nanomedicine that can be selectively degraded at pathological sites.73,>>74<< Curiously, the degree of liposome degradation by secreted phospholipase A2 increases with the quantity of poly(ethylene glycol) (PEG)-lipid conjugates incorporated into the liposome, in a charge-dependent manner.
pathological sites.73,74 Curiously, the degree of liposome degradation by secreted phospholipase A2 increases with the quantity of poly(ethylene glycol) (PEG)-lipid conjugates incorporated into the liposome, in a charge-dependent manner.>>73<<,74 Unfortunately, to the authors’ knowledge, studies to validate the efficacy of this system in vivo have not been published.
sites.73,74 Curiously, the degree of liposome degradation by secreted phospholipase A2 increases with the quantity of poly(ethylene glycol) (PEG)-lipid conjugates incorporated into the liposome, in a charge-dependent manner.73,>>74<< Unfortunately, to the authors’ knowledge, studies to validate the efficacy of this system in vivo have not been published.
ph targeting
to be acidic (as low as pH 5.7). Rapidly growing tumor cells accumulate lactic acid as a result of their high rate of glucose metabolism, insufficient blood supply, poor lymphatic drainage, and reduced rates of oxidative phosphorylation.>>75<< Thus, the acidic pH of the tumor microenvironment can be harnessed for pH-dependent drug delivery (Figure 2C).
drug delivery (Figure 2C). The role of the acidic environment in increased uptake of nanoparticles was first shown by Maeda et al who observed that SMANCS, under acidic conditions, showed a ten-fold increase in tumor cell binding.>>76<< Paclitaxel-loaded acrylate polymers with a pH-sensitive 2,4,6-trimethoxybenzaladehyde linker have also been developed and shown to be stable at neutral pH but cleaved at acidic pH, leaving the hydroxyl groups of the polymer exposed.
paclitaxel were shown to inhibit rapid growth of Lewis lung carcinoma tumors compared with non-pH-sensitive nanoparticles, which were associated with a nonsignificant reduction in tumor size in a 14-day study performed in C57B1/6 mice.>>77<<
These hydrazone-linked hyaluronic acid hydrogels release 60% of their linked immunoglobulin G in 8 hours at pH 5, while 60% release is achieved at 400 hours at pH 7.4.>>78<< A major challenge to utilizing low pH for targeting tumor tissues is the lack of an efficient blood supply for delivery of the nanoconstructs to the hypovascular tumor center.
Rapid endosomal acidification occurs during the endocytosis process due to ATPase-mediated proton influx. This activity results in a drop in pH levels to <6 in early endosomes, sorting endosomes, and multivesicular bodies.>>79<< Subsequently, pH-sensitive liposomes have been developed to release their content in response to acidic pH.
The first liposomes developed containing the pH-sensitive lipid palmitoyl homocysteine showed release of a fluorescent marker in a manner that was inversely proportional to the pH of the serum.>>80<< Furthermore, pH-sensitive liposomes, generated by surface modification of egg yolk phosphatidylcholine liposomes with MGlu-Dex, have been shown to be taken up efficiently by dendritic cells to deliver entrapped ovalbumin molecules into
Subcutaneous administration of MGlu-Dex-modified liposomes loaded with ovalbumin to mice induced antigen-specific humoral and cellular immunity more effectively than the unmodified liposomes loaded with ovalbumin.>>81<<
are often coated with stealth PEG layers, which are stable in the circulation but released in endosomes. PEG layer shedding helps in nanoparticle endosomal escape by reducing the steric and electrostatic hindrance from the PEG layer.>>82<< These PEG layers are grafted onto the nanoparticles with pH-sensitive linkers favoring drug release.
PEG layer shedding helps in nanoparticle endosomal escape by reducing the steric and electrostatic hindrance from the PEG layer.82 These PEG layers are grafted onto the nanoparticles with pH-sensitive linkers favoring drug release.>>83<< This endosomal acidification is used as a trigger for endosomal escape by a mechanism hypothesized to occur via the “proton sponge” effect.
disruption activity induced by the “proton sponge” mechanism. This activity causes disruption of the compartment membrane and release of doxorubicin into the cytosol, thus producing much higher in vitro and in vivo anticancer activity.>>85<< Lysosomal escape is also an important step towards achieving nuclear targeting.
Since nuclear targeting of layered double hydroxide nanorods is rapid and requires an active process, it has been suggested that the active transport occurs via a microtubule-mediated trafficking process.>>86<<
into amidoamine monomers in the reductive endosomal environment and therefore allow release of siRNA into the cytoplasm. Subsequently, PEI polymeric nanoparticles showed two-fold higher suppression of VEGF than linear PEI formulations.>>87<< However, the cationic nature of polyamine nanoparticles leads to increased affinity towards the negatively charged cell membrane, so these nanoparticles tend to have toxicity issues, such as activation of apoptotic pathways.
However, the cationic nature of polyamine nanoparticles leads to increased affinity towards the negatively charged cell membrane, so these nanoparticles tend to have toxicity issues, such as activation of apoptotic pathways.>>88<<
The parietal cells of the stomach lining secrete acids and digestive enzymes such as pepsin that are essential for denaturation of ingested proteins, so the stomach has a pH as low as 1.2.>>89<< The small intestine contains bile salts and other basic enzymes such as pancreatin, resulting in a pH of around 6.5 in this section of the gastrointestinal tract.
of ingested proteins, so the stomach has a pH as low as 1.2.89 The small intestine contains bile salts and other basic enzymes such as pancreatin, resulting in a pH of around 6.5 in this section of the gastrointestinal tract.>>89<< It is also important to note that pH varies throughout the gastrointestinal tract in the fasted and fed state.
89 It is also important to note that pH varies throughout the gastrointestinal tract in the fasted and fed state.>>90<<
At pH 7.4, the copolymer is in the swollen state and releases 90% of its loaded insulin while the remaining 10% is released at pH 1.2 in the complexed state.>>91<< Acrylic-based polymers such as PMAA retain their complexed state in the stomach due to protonation of carboxyl groups, while in the intestine these nanoparticles swell due to the increased pH, causing carboxyl ionization and breakage of
such as PMAA retain their complexed state in the stomach due to protonation of carboxyl groups, while in the intestine these nanoparticles swell due to the increased pH, causing carboxyl ionization and breakage of hydrogen bonds.>>92<<
in the duodenum, while polymethacrylate (PMAA-PMA, commercially available as Eudragit S100), dissolves at pH >7 and is therefore more suitable for ileal drug release or to increase the bioavailability of therapeutics such as cyclosporin.>>93<<
but, when in contact with H. pylori infection along the gastric epithelium (pH 7.4), deprotonation of chitosan occurs, which weakens the electrostatic interactions and leads to collapse of the nanoparticles and release of heparin.>>95<<
exploiting inflammatory mediators
cells lining the capillaries (Figure 3). This pathological response is induced by expression of histamine, bradykinin, leukotrienes, and serotonin in the inflamed tissue and results in increased intraendothelial gaps (Figure 3).>>96<<–99 These fenestrations allow extravasation of nanoparticles specifically into the inflamed tissue, as has been previously demonstrated in the case of polystyrene nanoparticles in the rodent trinitrobenzenesulfonic acid model of colitis.
These fenestrations allow extravasation of nanoparticles specifically into the inflamed tissue, as has been previously demonstrated in the case of polystyrene nanoparticles in the rodent trinitrobenzenesulfonic acid model of colitis.>>100<< Furthermore, in inflammatory conditions induced by infection, the pathogen itself may secrete factors that increase the permeability of blood vessels.
nanoparticles in the rodent trinitrobenzenesulfonic acid model of colitis.100 Furthermore, in inflammatory conditions induced by infection, the pathogen itself may secrete factors that increase the permeability of blood vessels.>>101<<,102 The inflammation also causes expression of inflammatory biomarkers, such as reactive oxygen species, that allow the active and regulated release of drug encapsulated in nanomedicines that are sensitive to these biomarkers (Table 1).
nanoparticles in the rodent trinitrobenzenesulfonic acid model of colitis.100 Furthermore, in inflammatory conditions induced by infection, the pathogen itself may secrete factors that increase the permeability of blood vessels.101,>>102<< The inflammation also causes expression of inflammatory biomarkers, such as reactive oxygen species, that allow the active and regulated release of drug encapsulated in nanomedicines that are sensitive to these biomarkers (Table 1).
Oral administration of thioketal nanoparticles loaded with TNF-α siRNA showed diminished levels of TNF-α messenger RNA in the colon in a murine model of ulcerative colitis.>>103<<
These cell adhesion molecules bind to the sialyl Lewis X (SLX) tetrasaccharide which is present on the plasma membrane of polymorphonuclear leukocytes>>104<< and is therefore considered to be a promising target for drug delivery at sites of inflammation.
molecules bind to the sialyl Lewis X (SLX) tetrasaccharide which is present on the plasma membrane of polymorphonuclear leukocytes104 and is therefore considered to be a promising target for drug delivery at sites of inflammation.>>105<< This observation has led to the development of intravenously administered SLX-conjugated liposomes encapsulating dexamethasone, which resulted in a two-fold higher accumulation of dexamethasone in the inflamed eye of a murine experimental
encapsulating dexamethasone, which resulted in a two-fold higher accumulation of dexamethasone in the inflamed eye of a murine experimental autoimmune uveoretinitis model when compared with intravenous injection of free dexamethasone.>>106<< Nontargeted liposomes loaded with dexamethasone resulted in no detectable level of dexamethasone in the inflamed eye using this model.
autoimmune uveoretinitis model when compared with intravenous injection of free dexamethasone.106 Nontargeted liposomes loaded with dexamethasone resulted in no detectable level of dexamethasone in the inflamed eye using this model.>>106<< SLX conjugated to liposomes encapsulating the dyes CY5 or CY3 showed accumulation in inflamed tissue on in vivo fluorescent imaging in a collagen-antibody induced arthritis model.
dexamethasone in the inflamed eye using this model.106 SLX conjugated to liposomes encapsulating the dyes CY5 or CY3 showed accumulation in inflamed tissue on in vivo fluorescent imaging in a collagen-antibody induced arthritis model.>>107<< Histopathological sections showed fluorescence in the hyperplastic synovium of periarticular soft tissue and pannus invasion with inflammatory cells in the collagen-antibody induced arthritis mouse model.
On administration of targeted SLX-liposome with Cy3, accumulation of the fluorescence signal was observed in the synovial tissue after 24 hours, while no accumulation was seen in control mice administered untargeted liposomes.>>107<<
exploiting unique cell characteristics at the tumor level
There is a wide range of receptor targeting mechanisms being utilized today, including transferrin receptor-mediated, low density lipoprotein receptor-mediated, and folic acid receptor (FR)-mediated endocytosis.>>108<<
Folate has a high affinity for the FR, which is often overexpressed on human tumor cells and taken up by receptor-mediated endocytosis following binding to the receptor.>>111<<,112 In this way, the interaction between folate and the FR can be used for tumor-specific drug delivery.
Folate has a high affinity for the FR, which is often overexpressed on human tumor cells and taken up by receptor-mediated endocytosis following binding to the receptor.111,>>112<< In this way, the interaction between folate and the FR can be used for tumor-specific drug delivery.
111,112 In this way, the interaction between folate and the FR can be used for tumor-specific drug delivery.>>113<< There are two different glycosyl phosphatidylinositol-anchored isoforms of the FR, ie, FR-α, which is found on about 40% of human epithelial cancers, and FR-β, which is expressed on activated macrophages in association with chronic
glycosyl phosphatidylinositol-anchored isoforms of the FR, ie, FR-α, which is found on about 40% of human epithelial cancers, and FR-β, which is expressed on activated macrophages in association with chronic inflammatory diseases.>>114<<,115 Overexpression of FRs has been observed in a variety of tumor types, including ovarian and endometrial cancers, where FR overexpression occurs in 90% of cases.
glycosyl phosphatidylinositol-anchored isoforms of the FR, ie, FR-α, which is found on about 40% of human epithelial cancers, and FR-β, which is expressed on activated macrophages in association with chronic inflammatory diseases.114,>>115<< Overexpression of FRs has been observed in a variety of tumor types, including ovarian and endometrial cancers, where FR overexpression occurs in 90% of cases.
114,115 Overexpression of FRs has been observed in a variety of tumor types, including ovarian and endometrial cancers, where FR overexpression occurs in 90% of cases.>>111<<
Encapsulation of drugs in folate-targeted nanomedicine has been pursued in a variety of nanoconstructs micelles and liposomes.>>110<<,116 In vivo, folate targeting of liposomes has shown the ability to increase the quantity of nanomedicine associated with J6456 lymphoma cells by 17-fold relative to nontargeted liposomes.
Encapsulation of drugs in folate-targeted nanomedicine has been pursued in a variety of nanoconstructs micelles and liposomes.110,>>116<< In vivo, folate targeting of liposomes has shown the ability to increase the quantity of nanomedicine associated with J6456 lymphoma cells by 17-fold relative to nontargeted liposomes.
of nanoconstructs micelles and liposomes.110,116 In vivo, folate targeting of liposomes has shown the ability to increase the quantity of nanomedicine associated with J6456 lymphoma cells by 17-fold relative to nontargeted liposomes.>>117<< It must be noted that activated macrophages also express the FR, so it is possible that the therapeutic activity of the liposomes is partially due to targeted elimination of FR-expressing tumor-associated macrophages.
117 It must be noted that activated macrophages also express the FR, so it is possible that the therapeutic activity of the liposomes is partially due to targeted elimination of FR-expressing tumor-associated macrophages.>>118<<
For example, methotrexate is utilized because of its ability to inhibit dihydrofolate reductase, an enzyme that converts dihydrofolate to tetrahydrofolate, before it is converted to the folate cofactor methylene tetrahydrofolate.>>119<< Therefore, folic acid targeting (such as in PAMAM dendritic polymers conjugated with methotrexate) has been used to increase the concentration of free methotrexate in sensitive tumor cells.
This approach allows reduction of methotrexate toxicity and improvement of methotrexate efficacy relative to the nontargeted construct.>>120<<
As a consequence, antibodies must be engineered to have a high enough affinity so that the antibody is still specific and able to effectively target the nanomedicine while not having an affinity high enough to prevent diffusion.>>121<<
Firstly, the use of antibodies may increase the quantity of the nanomedicine that reaches the pathological tissue; for example, an antibody to CD22 increases the quantity of doxorubicin in non-Hodgkin lymphoma-bearing mice by 80%.>>122<< The second method by which the efficacy of antibody-targeted nanomedicines may be enhanced does not involve increased delivery of the construct to gross tissue but rather increased delivery to specific cell types within the tissue (Figure
indicate that internalization of the liposomes specifically by cells targeted by the EGF receptor antibody was the determining factor for the increased efficacy of the construct, as opposed to increased accumulation at the tumor site.>>123<< This same phenomena was also described by Kirpotin et al who reported that nontargeted and HER-2 targeted immunoliposomes showed comparable accumulation in tumor xenografts, but that only HER-2 targeted immunoliposomes localized primarily
comparable accumulation in tumor xenografts, but that only HER-2 targeted immunoliposomes localized primarily in tumor cells, while nontargeted immunoliposomes showed localization within the stroma and tumor-associated macrophages.>>124<<
combinational approaches
iron oxide nanoparticles (EGFRvIIIAb-IONPs). Mice with orthotopic glioblastoma multiforme lesions treated using CED with EGFRvIIIAb-IONPs had a 15% and 10.5% longer survival time than mice treated with IONs or EGFRvIIIAb, respectively.>>125<< Furthermore, boronated dendrimers conjugated to EGF were developed to target EGFR-expressing glioblastoma multiforme.
Unfortunately, there was no control performed in this study to compare boronated dendrimers without EGF.>>126<<
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