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LYMPHATIC DRUG TARGETING AND TRANSPORTATION
@kotla.swetha sagar(M.Pharm)

INTRODUCTION: Lymphatic drug targeting refers to targeting of drugs and therapeutic agents into the lymphatic system for the drug action in the lymphatic system itself or for their transportation in the lymph to specific tissues of interest. Targeting of drugs to lymphatic system is usually attained by utilizing carriers like microspheres, nanoparticles and liposomes. .                                                   The first objective of targeting is exemplified by the targeting of certain anti-cancer agents in some lymphomas and in tumor detection in lymph nodes using radio labeled liposomes and the other objective is exemplifies by the transport of anti-inflammatory agents to the site of inflammation, the per oral uptake and the transport of certain macromolecules by subcutaneous administration. Drug targeting allows a substantial reduction in the amount of drug needs to be administered. It is of interest to point out that Koff et al while studying the effect of immunomodulators encapsulated in liposomes on macrophages, found about 800 times lower dose of modulator encapsulated in lipopsomes in comparison to free modulator. This order of magnitude would reduce the dose of steroids in arthritis treatment to safe levels. Moreover, peroral delivery of macromolecules, if they can be transported via lymphatics into the systemic system would be very advantageous where chronic therapy is indicated. Components of the Lymphatic System The lymphatic system consists of a fluid (lymph), vessels that transport the lymph, and organs that contain lymphoid tissue. Lymph: Lymph is a fluid similar in composition to blood plasma. It is derived from blood plasma as fluids pass through capillary walls at the arterial end. As the interstitial fluid begins to accumulate, it is picked up and removed by tiny lymphatic vessels and returned to the blood. As soon as the interstitial fluid enters the lymph capillaries, it is called lymph. Returning the fluid to the blood prevents edema and helps to maintain normal blood volume and pressure. Lymphatic Vessels Lymphatic vessels, unlike blood vessels, only carry fluid away from the tissues. The smallest lymphatic vessels are the lymph capillaries, which begin in the tissue spaces as blind-ended sacs. Lymph capillaries are found in all regions of the body except the bone marrow, central nervous system, and tissues, such as the epidermis, that lack blood vessels. The wall of the lymph capillary is composed ofendothelium in which the simple squamous cells overlap to form a simple one-way valve. This arrangement permits fluid to enter the capillary but prevents lymph from leaving the vessel. The microscopic lymph capillaries merge to form lymphatic vessels. Small lymphatic vessels join to form larger tributaries, called lymphatic trunks, which drain large regions. Lymphatic trunks merge until the lymph enters the two lymphatic ducts. The right lymphatic duct drains lymph from the upper right quadrant of the body. The thoracic duct drains all the rest. Like veins, the lymphatic tributaries have thin walls and have valves to prevent backflow of blood. There is no pump in the lymphatic system like the heart in the cardiovascular system. The pressure gradients to move lymph through the vessels come from the skeletal muscle action, respiratory movement, and contraction of smooth muscle in vessel walls. Lymphatic Organs Lymphatic organs are characterized by clusters of lymphocytes and other cells, such as macrophages, enmeshed in a framework of short, branching connective tissue fibers. The lymphocytes originate in the red bone marrow with other types of blood cells and are carried in the blood from the bone marrow to the lymphatic organs. When the body is exposed to microorganisms and other foreign substances, the lymphocytes proliferate within the lymphatic organs and are sent in the blood to the site of the invasion. This is part of the immune response that attempts to destroy the invading agent. The lymphatic organs include: •	Lymph Nodes •	Tonsils •	Spleen •	Thymus Lymph Nodes Lymph nodes are small bean-shaped structures that are usually less than 2.5 cm in length. They are widely distributed throughout the body along the lymphatic pathways where they filter the lymph before it is returned to the blood. Lymph nodes are not present in the central nervous system. There are three superficial regions on each side of the body where lymph nodes tend to cluster. These areas are the inguinal nodes in the groin, the axillary nodes in the armpit, and the cervical nodes in the neck. The typical lymph node is surrounded by a connective tissue capsule and divided into compartments called lymph nodules. The lymph nodules are dense masses oflymphocytes and macrophages and are separated by spaces called lymph sinuses. Several afferent lymphatic vessels, which carry lymph into the node, enter the node on the convex side. The lymph moves through the lymph sinuses and enters an efferent lymphatic vessel, which carries the lymph away from the node. Because there are more afferent vessels than efferent vessels, the passage of lymph through the sinuses is slowed down, which allow time for the cleansing process. The efferent vessel leaves the node at an indented region called the hilum.

Tonsils

Tonsils are clusters of lymphatic tissue just under the mucous membranes that line the nose, mouth, and throat (pharynx). There are three groups of tonsils. The pharyngeal tonsils are located near the opening of the nasal cavity into the pharynx. When these tonsils become enlarged they may interfere with breathing and are called adenoids. The palatine tonsils are the ones that are located near the opening of the oral cavity into the pharynx. Lingual tonsils are located on the posterior surface of the tongue, which also places them near the opening of the oral cavity into the pharynx. Lymphocytes and macrophages in the tonsils provide protection against harmful substances and pathogens that may enter the body through the nose or mouth. Spleen The spleen is located in the upper left abdominal cavity, just beneath the diaphragm, and posterior to the stomach. It is similar to a lymph node in shape and structure but it is much larger. The spleen is the largest lymphatic organ in the body. Surrounded by a connective tissue capsule, which extends inward to divide the organ into lobules, the spleen consists of two types of tissue called white pulp and red pulp. The white pulp is lymphatic tissue consisting mainly of lymphocytes around arteries. The red pulp consists of venous sinuses filled with blood and cords of lymphatic cells, such as lymphocytes and macrophages. Blood enters the spleen through the splenic artery, moves through the sinuses where it is filtered, then leaves through the splenic vein. The spleen filters blood in much the way that the lymph nodes filter lymph. Lymphocytes in the spleen react to pathogens in the blood and attempt to destroy them. Macrophages then engulf the resulting debris, the damaged cells, and the other large particles. The spleen, along with the liver, removes old and damage derythrocytes from the circulating blood. Like other lymphatic tissue, it produces lymphocytes, especially in response to invading pathogens. The sinuses in the spleen are a reservoir for blood. In emergencies such as hemorrhage, smooth muscle in the vessel walls and in the capsule of the spleen contracts. This squeezes the blood out of the spleen into the general circulation.

Thymus The thymus is a soft organ with two lobes that is located anterior to the ascending aorta and posterior to the sternum. It is relatively large in infants and children but after puberty it begins to decrease in size so that in older adults it is quite small. The primary function of the thymus is the processing and maturation of special lymphocytes called T-lymphocytes or T-cells. While in the thymus, the lymphocytes do not respond to pathogens and foreign agents. After the lymphocytes have matured, they enter the blood and go to other lymphatic organs where they help provide defense against disease. The thymus also produces a hormone, thymosin, which stimulates the maturation of lymphocytes in other lymphatic organs.

THE PHYSIOLOGY OF LYMPHATIC SYSTEM: The lymphatic system was first recognized by Gasparo Aselli. The lymphatic system is a drainage system, collecting and returning intestinal fluids. As with the blood network the lymph vessels form a network throughout the body, unlike the blood the lymph system is a one-way street draining lymph from the tissue and returning it to the blood. This system is a network of capillaries and tubes called lymphatics. The main components of the lymphatic system are bone marrow, lymph nodes, spleen and the thymus gland. There are five main categories of conduits in the lymphatic system: the capillaries, collecting vessels, lymph nodes, trunks and ducts. Their sizes range from 10 to 2 mm in diameter. Lymphatic capillaries are 10-60 mm in diameter and are comprised of one endothelial cell layer, typically made up of one or two nonfenestrated, highly attenuated cells in cross section. They have a discontinuous or absent basement membrane and with the exception of the initial lymphatics in the bat’s wing, are non-contractile.

Fig. 3:	Effect of particle size on lymphatic transport Figure 3: shows that particles up to 100 nm in diameter are preferentially transported into the lymph capillaries and phagocytised in the lymph nodes. All collecting vessels pass through lymph nodes which are capsular and organized in clusters through-out the lymphatic system. There is hundreds of lymph nodes in the adult human body and vary in size from 1 to 10 mm in diameter. Lymphocytes develop in the thymus gland or in the bone marrow. Lymphocytes in the lymph nodes aid the body in fighting infection by producing antibodies that destroy bacteria and viruses. The main functions of the lymphatic system are fluid and protein balance, immunity and spread of infection, digestion and solute uptake. SOLUTE UPTAKE BY LYMPHATIC SYSTEM The uptake of inert particulate matter (as opposed to viral/bacterial uptake) by the gastrointestinal epithelium and peripheral lymphatic duct are now a widely accepted phenomenon and has prompted a number of biotechnology companies and researchers to focus on this route for the delivery of gastrointestinal (GIT) labile molecules, anti-HIV, anti-cancer and immunosuppressant drug using micro particulate carriers (Hussain et al.,). The endothelial cell junctions of the initial lymphatics are not tight and considered freely permeable to most proteins, the physicochemical properties of the extracellular membrane can affect interstitial solute transport. Since, the solute must travel at least some distance through the interstitium before entering the lymphatics, the interstitial resistance to molecular transport greatly affects the apparent lymphatic uptake rate. Furthermore, significant changes in lymph concentration occur as the fluid passes through various components of the lymphatic system. It becomes concentrated along the contracting lymphangion segments, possibly due to water filtration across the vessel wall. Then, protein concentration decreases during its residence in the lymph nodes from osmotically driven fluid exchange with nodal blood vessels and phagocytosis by white cells. The issue of protein sieving during lymphatic uptake is somewhat controversial. The endothelium of the initial lymphatic poses very little hindrance to solute uptake and when fluid colloid osmotic pressure is directly compared locally between the interstitium and the initial lymphatic vessels, little to no differences are seen. Interstitial protein concentration is inversely related to trans vascular fluid flux and/or blood pressure and lymph protein concentration reflects interstitial protein concentration. The size, shape, charge and lipophilicity of a molecule affect its uptake rate into the lymphatics may actually reflect its interstitial hindrance prior to lymphatic uptake Indeed, interstitial transport cannot be easily decoupled from lymphatic uptake and the path through the extracellular matrix to the lymphatics should be considered when interpreting uptake data for proteins, colloids, drugs, or drug carriers. For the remainder of this discussion, the term lymphatic uptake will refer to the coupled process of interstitial and translymphatic transport, recognizing the importance of the interstitium and its physicochemical properties and architecture in controlling molecular transport. Size is one of the most important determinants of lymphatic uptake and lymph node retention. Molecules that are smaller than 10 nm are preferentially reabsorbed into the blood capillaries while the optimal size for lymphatic uptake is between 10 and 100 nm. The larger the particle or molecule, the more is the selectivity for uptake into the lymphatic system but the slower the uptake. For example, liposomes of 30-60 nm were found to have faster uptake rates than those of 400 nm but the smaller ones also showed higher level in the blood circulation. The upper size limit for lymphatic uptake has not been strictly defined. Particles up to 1 mm have been taken up by the lymphatics following interstitial injection; but above 100 nm, a percentage of injected solute will remain trapped in the interstitial spaces for longer periods of time and thus have lower uptake efficiencies. The composition of the molecule or particle is also important in determining uptake and lymph node retention. Colloids and lipids seem to have high uptake efficiencies. Depending on the size, charge, method of preparation and composition, various molecules such as monoclonal antibodies, peptide drugs and anticancer agents may be encapsulated into liposomes, nanoparticles, dendrimers etc and optimally targeted to lymph nodes. Furthermore, the microparticulate systems can be coated (e.g., with polyethylene glycol) or surface engineered with specific Ligands/chemicals (e.g., Folic acid, Lectin, L-selectin etc.) to improve lymph node retention by avoiding white blood cell phagocytosis; such stealth liposomes are well-characterized and can be designed for a number of specific purposes. Other than intercellular pathways, there is ample evidence for transendothelial pathways for solute and lipid transport in the initial lymphatics. After food uptake, lymphatic endothelial cells may be able to phagocytose particular matter and 7-8 mm trans endothelial channels were seen following milk lipid adsorption. Furthermore, these trans endothelial channels seem to be permanent structures since they are found even after periods of fasting. LYMPHATIC TARGETING AREAS: The lymph as a therapeutic target, number of specific investigations of drug delivery or drug targeting to the lymphatics is much more modest. This may reflect the historical belief that the lymphatics play a minor role in drug absorption (which indeed is the case for the majority of hydrophilic or moderately lipophilic small molecules), however recent results from this and other laboratories suggest that under certain circumstances, the lymphatics may provide the primary route of drug absorption and lead to drug concentrations in the lymph some 5-10,000 times higher than in systemic plasma. Recent advances in drug design and delivery, have also led to the development of an increasing number of (1) highly lipophilic drug molecules which may be substrates for intestinal lymphatic transport, (2) macromolecular biotechnology products which appear to be absorbed into the peripheral lymphatics after SC injection and (3) a range of particulate colloidal systems (micro particles, lipid carriers etc.) which may facilitate the lymphatic transfer of drug molecules with little intrinsic lymph-directing capacity. Recent data that suggests that for some compounds intestinal lymphatic transport may be both the primary route of absorption and responsible for the transport of the majority of the drug dose to the systemic circulation. The lymphatics are the primary conduit for the dissemination of metastases from many solid tumors, play a pivotal role in the generation of immune responses and have been implicated in the pathogenesis of diseases including HIV and metastitial tuberculosis. For lymph resident diseases, lymphatic targeting of therapeutic drugs (e.g., antivirals, cytotoxics or immunemodulators) is therefore expected to provide advantage over conventional approaches that focus on drug delivery via the blood. Pharmacokinetically, promotion of drug transport into the lymph may also reduce hepatic first pass metabolism thereby enhancing oral bioavailability. LYMPHATIC DRUG TARGETING THROUGH THE ORAL ROUTE The possibility of uptake and absorption of nanoparticles and microparticles by the gastrointestinal tract has been a controversial issue although there is now accumulated evidence that it can and does occur (10-15) The stomach, intestines and related organs of the gastrointestinal tract are drained along the lymphatics and through nodes lying in the mesenteries and omenta with the vessels supplying these organs. These nodes are finally drained into the cisterma chili. These are regions in the git, especially in small intestine, called Payer’s Patches and Gut Associated Lymphoid Tissue (GALT) which drain into the lymph vessels. These are useful for the transport of particulates from git into the lymphatic system. Absorbed microspheres then reaches the mesentery via the mesenteric lymph nodes and were transported from the lymphatic circulation into the venous circulation and subsequently into the liver. LYMPHATIC DRUG TARGETING THROUGH THE PARENTERAL ROUTES Various parenteral routes like subcutaneous (sc), intramuscular (im) and intra-peritonial (ip) have been tried for lymphatic targeting. Most compounds of relatively small molecular weight are exclusively absorbed via splenic blood capillaries into the portal vein. In contrast, the lymphatic system can be a major absorption route for compounds impermeable to capillary membranes because of their large molecular size. Many drugs in solution injected into sc or im sites behave as if their absorption were taking place passively by diffusion. For most drugs it has not been determined whether it is through capillaries, lymphatics, or both. Although there are other factors such as area of the drug depot formed after injection, volume of the injection, drug concentration, age, anatomical region and the tissue condition at the site of injection affect the rate of diffusion, the molecular size of the drug or the particle size of the carrier determines whether it will be absorbed via the capillaries or the lymphatics. Molecules with MW>16,000 are absorbed mainly by the lymphatics but compounds with MW<1000 are hardly absorbed at all by the lymphatic vessels. An increasing tendency towards lymphatic absorption was determined for molecules with a MW between 1000 and 16000. Although passive diffusion is an important mechanism for drug absorption the process of endocytosis may also be involved in drug absorption from sc and im sites. Beresford and co-workers showed the most of the absorption of iron polysaccharide complexes injected into the rabbit muscle tissue occurred during the initial 72 hours. The absorption during this time was mediated in part by lymphatic transport of the iron complexes.

APPLICATIONS: CONTROLLED DRUG DELIVERY SYSTEMS: Controlled drug delivery technology offers numerous advantages compared to conventional dosage forms, including improved efficacy, reduced toxicity, and improved patient compliance and convenience. Two types of controlled drug release can be achieved, temporal and distribution control. In temporal control, drug delivery systems aim to deliver the drug over an extended duration or at a specific time during treatment. It is highly beneficial for drugs that are rapidly metabolized and eliminated from the body after administration. In distribution control, drug delivery systems aim to target the release of the drug to the precise site of activity within the body. There are two principle situations in which distribution control can be beneficial. The first is when the natural distribution causes drug molecules to encounter tissues and cause major side effects that prohibit further treatment. The second situation is when the natural distribution of the drug does not allow drug molecules to reach their site of action. Systems using either of these two mechanisms have been developed for clinical therapy. However, integration of these two mechanisms of controlled release in a single device to improve cancer chemotherapy is rarely seen. Delivering a therapeutic agent to the lymphatic system poses a significant challenge in treating cancer. With conventional systemic administration of anticancer agents, effective drug concentrations are not usually reached in the lymphatic system without incurring significant toxicity, presumably because of the ‘blood-lymph barrier’. Such dose limiting toxicities significantly decrease the therapeutic efficacy of anticancer agents. Furthermore, the postoperative systemic chemotherapy to control lymphatic metastasis is often compromised from surgical disruption of the blood supplies to regional lymph nodes.

CANCER THERAPY: Pancreatic cancer is a highly malignant neoplasm of GI system, and radical surgery is its only curative treatmentoption1. Unfortunately, the probability of locoregional recurrence approaches 80% after complete resection, and long-term survival is less than 25% even for patients treated for early stage disease [2-4].

Adjuvant treatment is an integral part of definitive treatment of resectable pancreatic carcinoma; however, the optimal therapeutic modalities in adjuvant setting remain a focus of debate. Radiation therapy is commonly used in adjuvant treatment for pancreatic cancer after radical surgery in the United States. The effect of radiation with concurrent 5-FU based chemotherapy has been suggested in a number of randomized clinical trials [5-7]. In addition, concurrent cheamoradiation therapy has been the mainstay treatment for non metastatic and inoperable pancreatic cancer [8,9]. Radiation fields utilized in these trials encompassed not only subclinical nodal regions but also adjacent normal tissues. Despite such generous coverage, however, locoregional control remains a major mode of recurrence. The underlying reason for such suboptimal outcome is probably due to, at least in part, insufficient radiation dose (i.e., 45-50 Gy in conventional fractionation) to the surgical bed and draining lymph node regions limited by the organs at risk (OARs) adjacent to the pancreas and lymphnodal regions such as liver, small intestine, stomach, spinal cord, and kidneys. The prevailing utilization of intensity-modulated radiation therapy (IMRT) in cancer treatment including upper GI malignancies enabled dose differentiation between target volumes and adjacent normal tissues and organs thereby improved therapeutic ratio. Results from recently published dosimetry studies have suggested the advantage of IMRT in the treatment of tumors of upper abdomen including pancreatic, gastric, and billiary cancers as compared to 3-dimentional conformal radiation therapy (3D-CRT) [10-13]. Proper defining of high-risk regions especially the lymph nodal regions (i.e., CTV-N) forms an imperative basis for dose escalation using IMRT. However, selection and delineation of nodal regions in both adjuvant IMRT after pancreaticoduodenectomy and in definitive setting have never been addressed. The aim of this analysis is to address the selection of high-risk subclinical lymph nodal regions in conformal radiation therapy for resectable pancreatic cancer, by reviewing and summarizing the probability of lymph node metastases in resectable pancreatic cancer patients treated with radical surgery with lymph node dissection and pathological investigation of the resected regional node.

TREATMENT OF INFECTION: A number of human diseases have been linked to abnormal or defective lymphatic vessels. While the theory of anti-angiogenesis therapy has been extensively studied, the concept of targeting lymphangiogenesis to gain a therapeutic advantage in human disease is only a recent development. Advances in our understanding of the molecular signaling pathways that control lymphatic vessel formation therefore provide an opportunity to explore the value of inhibiting these processes. A good example of this is cancer biology, where the spread of tumor cells appears highly dependent on the vessels of the lymphatic system and the protein factors which drive their growth and differentiation. As a consequence, therapeutic options which target these cellular pathways may provide a means to prevent growth or metastasis from the primary tumor. Therapeutics may be either anti-lymphatic (targeting functions of the existing vessels) and/or anti-lymphangiogenic (targeting the generation of new lymphatic vessels). An understanding of the key signaling components and cellular processes that are critical for lymphatic vessel function and growth is essential to enable the rational design of effective inhibitors. One family of molecules, the protein tyrosine kinases, are known to be key drivers of angiogenesis and studies have shown they also play a pivotal role in lymphatic biology/lymphangiogenesis. In this review we explore the potential for this family of molecules to be used as targets for anti-lymphatic/anti-lymphangiogenesis and the ways in which we can gain insight into how these family members might contribute to key signaling pathways within the lymphatic endothelium.

CONCLUSION: Lymphatic drug targeting and transportation can be an important phenomenon for the absorption and delivery of drugs. While lymphatic targeting and transportation may be aimed at specifically in the case of carriers for peptides for avoiding hepatic first pass metabolism, for tumor detection and its treatment, it may also be normal route of absorption even when not aimed at some conventional drugs with high lipophilicity may be incorporated into chylomicrons inside the intestinal epithelial cells and be absorbed into systemic circulation via the lymphatic system. The use of carrier systems such as microspheres and nanoparticles to target specific areas of git such as Payer’s Patches have been tried so that peroral delivery systems can be developed for therapeutic molecules, which are used for chronic treatment in conditions like diabetes, rheumatic disorders and others. Oral immunization is another incentive for developing peroral formulations that target the lymphatics. Parenteral routes such as sc, im and intra peritoneal offer an alternative route for lymphatic targeting. Lymphatic drug delivery is in its infancy, but localized treatments of the lymphatics will decrease systemic toxicities associated with cytotoxic chemotherapy and reduce recurrence owing to residual local disease. By increasing our understanding of lymphatic transport and uptake and the role of lymphatics in cancer spread, we can design new therapeutics that may one day supplement or even replace radiotherapy for local disease control. In summary, we believe a future direction in cancer therapy will involve combining emerging nanocarrier technologies with locoregional therapy to the lymphatics. This combination has the potential to both reduce non-specific organ toxicities and increase the chemotherapeutic dose to the most likely sites of locoregional cancer metastasis. In addition, advanced lymphatic imaging tools will improve cancer staging and reduce the need for destructive nodal dissections.

REFERENCES: 1. Alpar, H.O., Field, W.N., Hyde, R., Lewis, D.A.:J. Pharm. Pharmacol.41,194 (1989) 2. Jani, P., Halbert, C.W., Langridge, J., Florence, A.T.:Ibid. 42,821 (1990) 3. Kochiro,H, Anthony, H.C.:J.Pharm. Sci. 74, 915(1985) 4. Rayman, B.E., Gillman, M.B.: Liposomes-Further considerations of their possible role as carriers of therapeutic agents ( In) Gregoriadis. G., Senior, J., Trout, A. (Eds)  Targeting of Drugs, Plenum Press, London 1981 5. Supersaxo, A., Mein, W., Gallati, H. Steffen, H.:Pharm. Res. 5, 472(1988) 6.Koff, W.C., Fidler, I.J., Schwalter, S.D., Chakrabarty, M.K., Hamper, B., Ceccorulli, L., Kleineiman, E.S.:   Science   224, 1007 (1984) 7. Best, C.H., Taylor, N.B.: The Living Body 4th edn. pp52 Champans Hall Ltd., London 1952 8. Charles, M.C.: Gray’s Anatomy  29th edn. 731 Leaf and Febeger, Philadelpia,1973 9. White A., Handles, P.,Smith, E.C.: Principles of Biochemistry 3rd edn. Pp 431 Mc Graw Hill/Kogakusha Co.Ltd. London/Tokyo1964 10. Thompson, A.R., Payne, J.M., Sansom, F.B., Gamer, R.J., Miles, B.J,: Nature  186, 586 (1960) 11. Volkheimer, G.: Ann.N.Y. Acad.Sci. 246, 164 (1975) 12. Aprahanian, M., Michel, C., Humber, W., Devissaguet,J.P., Dange, C.: Biol. Cell 61,69 (1987) 13. Leferve, M.E., Vanderhoff, J.W., Laisue,  J.A., Joel,D.D.: Experimentia34,120 (1978) 14. McClugage,  S,G.,  Low,  F.N., Zimmy, M.L.: Gastroenterology 91,  1128 (1986) 15. Tilney, N.L.: J. Ant, 109, 369 (1971)