TUMOR TARGETED DELIVERY

1. TUMOR
TARGETING
METHODS

2. WHAT IS A TUMOR?
A tumor is a mass of tissue that's formed by an
accumulation of abnormal cells. Normally, the
cells in our body age, die, and are replaced by
new cells. With cancer and other tumors,
something disrupts this cycle. Tumor cells grow,
even though the body does not need them, and
unlike normal old cells, they don't die. As this
process goes on, the tumor continues to grow as
more and more cells are added to the mass.

3. DIFFERENCE BETWEEN BENIGN AND
MALIGNANT TUMOR

4. TDDS FOR TUMOR THERAPY
• ACTIVELY TARGETING DDS
• PASSIVELY TARGETING DDS
• INVERSE TARGETING
• DUAL TARGETING
• DOUBLE TARGETING

5. ACTIVELY TARGETING DDS
Active targeting, sometimes called ligand-mediated targeting, is achieved by attaching specific ligands/antibodies to the
nanocarriers. This approach is theoretically based on the paired ligand–receptor or antigen– antibody recognition on the
tumor cell surfaces and can improve the uptake of TDDSs in tumors and enhance the therapeutic efficacy of the anticancer
agents.
It could also significantly reduce the undesired side effects, since the TDDS is designed to bind specifically to a receptor
overexpressed on tumors (including tumor cells and tumor vasculature) but not expressed by normal cells. Moreover, the
smart TDDS could effectively deliver antitumor drugs to the cytoplasm in response to various endogenous stimuli (pH,
redox, hypoxia, etc.) and exogenous stimuli (light, ultrasound, magnetic, etc.)
Considering these above advantages, the concept of active targeting has attracted increasing attention and is extensively
integrated into DDS, and these actively targeting DDSs have shown averagely better performance than those non-targeted
DDSs, such as increased cytotoxicity to tumor cells and the reduction of side effects. In the following section, we introduce
the most widely utilized active targeting ligands in typical advanced active targeting based DDSs.

6. TARGETING APPROACHES/MECHANISMS
FOR TUMOR THERAPY
1. TARGETING THE CYTOMEMBRANE
2. LYSOSOMAL TARGETING
3. TARGETING THE ENDOPLASMIC RETICULUM
4. TARGETING THE MITOCHONDRIA
5. TARGETING THE CELL NUCLEUS
6. TARGETING THE TUMOR MICROENVIRONMENT

7. The cell membrane is a biological protective barrier that surrounds the cell surface and is mainly composed of membrane lipids and
proteins.
An intact cell membrane could maintain the relative stability of the microenvironment within cells and thus sustain the normal life
activity. It also has critical roles in many biological processes such as cell recognition, signal transduction as well as the exchange of
substances and energy.
The composition and structure of the cell membrane changes significantly during canceration of cells. Studies show that some
characteristics of tumor cells, such as invasion and migration, could also be associated to the abnormal changes found on the cell
membrane based on this, the specific antigens or receptors overexpressed on the tumor cytomembrane could be used as specific
target sites for targeted tumor therapy. In a typical targeted drug delivery process, the nanocarriers are functionalized with the
corresponding ligand or antibody to target over-expressed receptors or antigens on the cell surface. After nanocarriers are leaked
from the vessel, they can accumulate in the tumor tissue and then be internalized through specific recognition effect between
receptor and ligand (or antigen and antibody). This targeted strategy not only achieves the targeted delivery of drugs but also greatly
increases the endocytosis amount.
.
TARGETING THE CYTOMEMBRANE

8. For example, an elevated concentration of sialic
acid was found due to the unusual sialyl behaviour
on the surface of tumor cells. Benzene boric acid,
which has a strong affinity with sialic acid, has
been used to target tumor cells
The asialoglycoprotein receptor (ASGPR) is one
type of overexpressed receptor located on the
surface of the tumor cell membrane that could
specially bind to several sugar ligands (e.g.,
Galactose, lactobionic acid) with high affinity;
however, its expression on normal cells/tissues is
almost negligible. Thus it could be used as a
promising active site for DDS conjugated with
targeting ligands

9. Lysosomes are an important organelle in cells and are essentially monolayered cystic particles. Lysosomes contain large amounts of
hydrolase, within which more than 60 kinds of hydrolytic enzymes have been found so far, such as proteolytic enzymes, nucleic acid
hydrolase, lipase, glycosyl hydrolase, etc.
Its main function is to decompose intracellular macromolecules and proteins. When foreign substances enter the lysosome, they are
decomposed by the hydrolase within. Should the membrane integrity of the lysosome be damaged its abundant hydrolases would be
released into the cell, leading to cell autolysis.
Studies have indicated that cationic substances and external stimuli (such as surfactants and heat, etc.) can lead to increased permeability
of the lysosomal membrane, as a result of which the enzymes are released into the cytoplasm, causing cell death. Therefore, therapies
targeting the lysosome shave drawn considerable interest in tumor treatment. Compared with other targeting strategies, the specific
targeting of lysosomes is relatively simple to achieve because most of the nanoparticles will enter the lysosome after cellular uptake.
Specifically, after the surface ligand modification, the nanomaterials are endocytosed into primary endosomes through the interaction
between the ligand and receptor on the cell membrane surface, and eventually fuse with the lysosome. At this time, the destabilizing
agents loaded in nanoparticles, such as magnetic nanoheaters, can readily initiate the permeabilization of a variety of hydrolytic enzymes
through the lysosomal membrane, resulting in the effective cell apoptosis
LYSOSOMAL TARGETING

11. The endoplasmic reticulum (ER) is a closed-mesh piping system comprising
primarily of internal membranes. It is responsible for the synthesis, folding and
assembly of nascent peptide chains.
A broad range of diseases is the direct result of ER-misfolded protein. The
endoplasmic reticulum can also maintain homeostasis of intracellular calcium
ions, which are important intracellular signalling substances stored in the ER. ER
will release calcium ions and initiate caspase-8-related program apoptosis when
subjected to external stimuli.
Typical ER stresses, such as protein misfolding and environmental stimulation
(incubation with anticancer drug), could induce certain biological response of ER
and then lead to cell death through the initial programmed apoptosis pathway.
This applies to many cancer cells.
Therefore, the endoplasmic reticulum is also a potential target for tumor
therapy. Ligand-mediated endoplasmic reticulum localization is a common
targeting approach for tumor therapy that relies on the conjugation of ER-
targeting signal peptides.
TARGETING THE ENDOPLASMIC RETICULUM

12. PERK – PROTEIN KINASE
ENDOPLASMIC RETICULUM
CHOP – TRANSCRIPTION
FACTOR
Luminescent platinum(II) complexes bearing bis(NHC)
ligands selectively target ER and induce ER stress, eventually
leading to apoptotic cell death.

13. The mitochondria is the main site of aerobic respiration in cells. In addition to the role of ‘‘power house,’’ mitochondria also
play critical roles in many regulatory mechanisms concerning cell growth and metabolism, apoptosis, calcium homeostasis,
free radical generation, lipid metabolism and so on.
Mitochondria can control programmed cell death via regulating the membrane potential. However, in most tumor cells,
mitochondria are in a constant dysfunctional state, such as poor membrane permeability and abnormal release of
apoptotic signals. Mitochondria are potential targets for tumor therapy.
Mitochondria have a highly negative membrane potential that is derived from its function in the cell. Mitochondria provide
ATP energy to support various cellular activities. They are constantly pumping out protons (H+, na+ , etc.) From the intima.
Moreover, the proliferation ability of cancer cells is higher than that of normal cells, so cancer cells need more energy
supply to meet the growth of cells. Consequently, the membrane potential of mitochondria in tumor cells is more negative
than that of normal cells. Therefore, mitochondria-targeted therapy could be developed based on this characteristic.
A lipophilic tpp could efficiently bind to mitochondria due to the higher mitochondrial membrane potential. Jung and co-
workers constructed mitochondria-targeting iron oxide NPs (IO NP) through coating with TPP. Compared to IO NP, TPP-IO
NP was gathered more in the mitochondria because TPP has a higher affinity with the mitochondria.
TARGETING THE MITOCHONDRIA

14. The cell nucleus is the control centre of a cell and plays an important role in cell metabolism, growth and differentiation. It is also
the main storage site of genetic materials. More importantly, the action sites of most therapeutic anticancer drugs, such as DNA
intercalators and topoisomerase inhibitors, are in the nucleus.
Thus, directly targeting drug delivery to the nucleus can effectively increase the therapeutic effect due to the bypassing of drug
efflux pumps, which makes nuclei-targeting an important delivery strategy for tumor therapy. From a structural perspective, the
nuclear membrane is constituted by two layers of membrane and decorated by nuclear pore complex (NPC) on the membrane
surface. Nanoparticles with a diameter of less than 9 nm could gain entry to the nuclear area via NPC. However, such small
nanoparticles would be easily removed from blood circulation before they could even reach the tumor site. Large nanoparticles can
be delivered to the nucleus through NPC facilitated by the nuclear localization signal (NLS). The NLS is a signal sequence located in
the c-terminal of nucleoplasmins. It usually contains 4–8 amino acids and is positively charged. NLS complexes with NPC and forms a
hydrophilic channel on the nuclear membrane. Subsequently, the cargo that previously bound to NLS can enter the nuclear area in
an energy-dependent manner. In addition, there are adequate amounts of glucocorticoid receptors on the nucleus; therefore, small
molecular glucocorticoids such asdexamethasone, triamcinolone acetonide, betamethasone, etc. Could be used as nucleus-
targeting molecules.
More interestingly, when the nanoparticles are transported to the nucleus mediated by glucocorticoid, the NPC channel could be
expanded up to 60 nm, so that the nanoparticles may enter thenucleus more easily
TARGETING THE CELL NUCLEUS

15. The tumor microenvironment is closely related to the pathological state of tumors. It mainly consists of the
surrounding blood vessels, fibroblasts, lymphocytes, immune cells and the extracellular matrix (ECM).
It is widely involved in tumor occurrence, growth, invasion and metastasis. Thus, the tumor microenvironment
sensitivity is another extensively studied area in the field of tumor-targeted DDS, taking advantage of the
numerous differences in tumor microenvironments to the surrounding normal tissues, including pH value,
vascular abnormalities and ECM composition.
Meanwhile, many distinctive characteristics have been discovered in cancerous ECM, including slightly acidic
environment and low oxygen concentration, which differ greatly from the normal tissue ECM and could facilitate
the development of novel targeted TDDSs. Few studies reported that optimized zwitterionic nanoparticles rapidly
aggregated in cancerous ECM in response to the slight pH change from 7.4 to 6.5, which could be utilized for
tumor-targeted therapy.
TARGETING THE TUMOR MICROENVIRONMENT

16. Schematic representation of the actively targeting DDS. Targeting ligands grafted on
the surface of nanocarriers bind to receptors overexpressed by tumor cells and
facilitate internalization via receptor mediated endocytosis. Target receptor/sites
mainly exist on the tumor (1) cytomembrane, (2) endo/lysosome, (3) endoplasmic
reticulum, (4) mitochondria and (5) nucleus. Intracellular drug release is triggered

17. PASSIVELY TARGETING DDS
Most solid tumors possess unique pathophysiological characteristics that are not observed in normal tissues
or organs, such as extensive angiogenesis, defective vascular architecture, impaired lymphatic
drainage/recovery system, and greatly increased production of a number of permeability mediators.
Therefore, the passive targeting of solid tumor largely relies on the enhanced permeation and retention (EPR)
effect that has been universally observed in solid tumors. EPR was first discovered by Matsumara and Maeda
in 1986. In general, rapid vascularization, through which the external nutrients and oxygen are provided, is
essential for rapid tumor growth. Many growth factors, including VEGF, are involved in cancer angiogenesis.
However, the newly formed tumor vessels are usually abnormal in form and architecture, characterized by
poorly aligned endothelial cells, lack of a smooth muscle layer, impaired functional receptors for angiotensin
II and limited lymphatic drainage. The defective blood vessels allow the extravasation of large molecules and
nanoparticles and their retention in the tumor tissues. Therefore, the EPR effect also provides a great
opportunity for more selective targeting of lipid- or polymer-conjugated anticancer drugs/nanocarriers for
tumor tissue/cells

18. PASSIVELY TARGETING DDS AND THE INFLUENCE OF NANOPARTICLES
FOR EPR. NANOPARTICLES PASSIVELY EXTRAVASATE THOUGH THE
LEAKY VASCULATURE AND ACCUMULATE IN TUMORS DUE TO THE
DAMAGED LYMPHATIC DRAINAGE.

19. PHYSIOLOGICAL FACTORS AFFECTING THE EPR EFFECT : EPR effect in tumors has been
frequently exploited for the delivery of anticancer drugs. However, it is also significantly affected by
the state and types of tumors on account of tumor heterogeneity. In addition, the structure and pore
dimensions of tumor vessels would also vary significantly within the same tumor or between different
tumor types. Nanomaterial factors improving the EPR effect size and shape.
(1) tumor growth environment, for example, the nature of ECM the vascular bed and surrounding
stroma,
(2) several vascular factors and special inhibitors existing in body the body, which could lower blood
pressure
(3) co-medications and their impact on stroma and blood pressure
NANOMATERIAL FACTORS IMPROVING THE EPR EFFECT :
1 Size and shape : Due to the high interstitial fluid pressure and the poor lymphatic drainage in the
tumor microenvironment, the EPR effect would be profoundly affected by the size of
nanocarriers/macromolecular drugs. Specifically, the long-circulating nanocarriers with suitable size
(around 100 nm) are prone to accumulation in the tumor. The sphere-shaped nanoparticles of sub-
100 nm usually show relatively superior uptake efficiency compared to other shapes, such as rods,
cylinders, cubes and so on
2 Charge : The surface charges of the nanocarriers can significantly influence phagocytosis and
their circulation in blood as well. It was reported that negatively charged or neutral nanoparticles
have random effect on the blood clearance of NPS, but positively charged nanoparticles are generally

20. Surface wettability : in addition to particle size, shape and charge status, surface
hydrophobicity is another important factor concerning EPR - dependent drug delivery.
in general, hydrophobic nanoparticles are more likely coated by special proteins such
as immunoglobulin and plasma proteins when they enter the blood circulation and
then get cleared by res, which would lower the efficiency of drug delivery for tumor
therapy. To address this challenge, a commonly used approach was to coat
nanoparticles with highly biocompatible amphiphilic molecules; for example,
polyethylene glycol PEG and its analogue were conjugated to the surface of fabricated
nanocarriers to maximize the EPR effect as well as to minimize the RES clearance.
Advanced drug delivery system based on passive targeting : inspired by the
discussion above, many nanoparticles with optimal size, shape and surface
properties have been utilized to fabricate various advanced ddss, which were
commonly conjugated with multiple functional motifs to cooperatively regulate
the multi-stimuli-responsive mechanisms on TDDSs and enhance the EPR effect,
thus to enhance the accumulation of nanocarriers in tumor sites and facilitate the
localized cargo release. In this part, we mainly focus on those advanced TDDSs

21. INVERSE TARGETING
It is a result of the avoidance of passive uptake of colloidal carriers by the RES.
It can be achieved by suppressing the function of RES by pre- junction of a large amount of
blank colloidal carriers or macromolecules like dextran sulphate.
Other strategies include modification and defined manipulation of the size, surface charge,
composition, surface rigidity & hydrophilicity characteristics of carriers for desirable
biofate.

22. DUAL TARGETING
In this targeting approach, carrier molecule, itself have
their own therapeutic activity and thus increase the
therapeutic effect of drug.
A carrier molecule having its own antiviral activity can be
loaded with antiviral drug and for the synergistic effect of
drug conjugate.

23. DOUBLE TARGETING
SPATIAL CONTROL - Targeting drugs to specific organs, tissues, cells or even sub
cellular compartment.
TEMPORAL CONTROL - Controlling the rate of drug delivery to target site.