WO2006046243A1

WO2006046243A1 - Sv40 for gene therapy of renal disorders

Info

Publication number: WO2006046243A1
Application number: PCT/IL2005/001119
Authority: WO
Inventors: Ariella Oppenheim; Yoram Weiss; Yosef Haviv
Original assignee: Hadasit Medical Research Services and Development Co; Yissum Research Development Co of Hebrew University of Jerusalem
Current assignee: Hadasit Medical Research Services and Development Co; Yissum Research Development Co of Hebrew University of Jerusalem
Priority date: 2004-10-26
Filing date: 2005-10-26
Publication date: 2006-05-04
Anticipated expiration: 2007-04-26
Also published as: IL164850A0

Abstract

The present invention relates to an SV40 renal delivery system for targeting an exogenous nucleic acid molecule to kidney cells. More particularly, the delivery system of the invention comprises as an active ingredient an SV40 infectious particle comprising said nucleic acid molecule packaged therein. This nucleic acid molecule may be a molecule encoding a product involved in a renal pathological disorder or alternatively, a molecule directed against a product involved in a renal pathological disorder.

Description

SV40 FOR GENE THERAPY OF RENAL DISORDERS

Field of the Invention

The present invention relates to gene therapy as a new treatment approach for acute and chronic renal disorders, by using SV40 derived vectors to deliver genes that modulate renal tissue injury and recovery.

Background of the Invention

All publications mentioned throughout this application are fully incorporated herein by reference, including all references cited therein.

SV40 and deriυates as potential gene therapy vectors

SV40 virus is relatively harmless to humans [StrickLer et al., (1998) J. Am. Med. Asso. 279: 292-295] and can be readily made replication incompetent by deleting T antigen [Strayer, D.S., (2000) Curr. Opin. MoI. Ther. 2: 570-578; Arad et al., (2002) Virol. 304: 155-159]. This virus can. infect wide varieties of cells, both dividing and non-dividing [Rund et al., (1998) Hum. Gene Ther. 9: 649-657; Strayer et al., (2000) Gene Ther. 7:886-895] and therefore, offers long term transgene expression [Strayer D. S., (1999) Exp. Opin. Invest. Drugs 8: 2159-2172; Sauter et al., (2000) Gastroenterol. 119: 1348-1357; Strayer et al., (2000) ibid; Strayer et al., (2002) MoI. Ther. 6: 227-237]. Another advantage of SV40 is that it does not elicit immune response [Rondo et al., (1998) Gene Ther. 5: 575-582; Arad et al., (2005) Human Gene Therapy 16: 361-371]. Also viral vectors can be easily prepared at high titer stocks on large-scale basis [Strayer (2000) ibid; Vera et al., (2004) MoI. Ther. 10: 780-791].

Due to all the above-mentioned characteristics, SV40 seems to be an attractive vector for gene therapy. Molecular vehicles resembling viral particles for accomplishing gene therapy can be generated using a variety of technologies. For- example, SV40 vectors may be prepared in tissue culture packaging cell-lines that supply T-antigen with or without the virus capsid proteins [Strayer (2000) ibid; Arad et al., (2002) Virology 340: 155-159.; Vera et al., (2004) DNA Cell. Biol. 23: 271-282; Vera (2004) ibid].

Helper-free SV40 vectors may be constructed by replacing the T-antigen coding-sequence with a transgene. Such vectors are propagated in cell-lines that supply the T-antigen in trans. The T-antigen in the packaging cells activates the replication of the vector DNA and also trans-activates the viral late-proteins genes that would produce the viral capsids. These vectors have a cloning capacity of only 2.2 kb and transduce a variety of cells and tissues [Strayer (2000) ibid; Vera (2004) ibid].

Valuable packaging cell-lines, that eliminate the generation of T-antigen positive replication competent recombinants, were constructed previously by the inventors [Arad (2002), ibid and WO03/025189]. Those cell lines allow the production of SV40 viruses and pseudoviruses that are safe for medical use. In addition, plasmids that contain just the ori and ses elements of SV40, a total of about 250 bp [Oppenheim, A. et al., (1992) J. Virol. 66: 5320-5328], may be packaged into vectors in tissue culture cells, when T-antigen as well as the capsid proteins are supplied in trans [Oppenheim, A., et al., (1986) Proc. Natl. Acad. Sci. USA 83: 6925-6929; and Oppenheim, A., (1992) In: New Perspective on Genetic Markers and Diseases among the Jewish People. B. Bonne-Tamir and A. Adam eds. Oxford University Press Inc., pp . 365-373]. The cloning capacity of such vectors is about 4.5 Kb.

SV40 vectors can be also prepared in vitro, from DNA propagated in bacteria and recombinant capsid proteins [Sandalon et al., (1997) Hum. Gene Ther. 8: 843-849 and WO 97/17456]. These vectors have packaging capacity of about 17 Kb of plasrnid DNA.

In vitro p ackaging is the ideal and safe way to prepare pseudovirions for human tre atment, because all steps of preparation can be well controlled. SV40 pseudovirions can be produced in vitro from purified DNA and cell-free extracts containing: recombinant viral capsid proteins [Sandalon et al., (1997) ibid]. VPl, the SV40 major structural protein expressed as a recombinant protein in insect cells, can be self-assembled in the nucleus and form vmxs-like particles (VLPs), which are morphologically indistinguishable from wild-type SV40 capsids [Sandalon, Z. and Oppenheim, A. (1997) Virol. 237: 414-421]. Such in vitro constructed vectors can be used as vehicles for gene transfer [Sandalon (1997) ibid; KimcM-Sarfaty (2002) ibid; Kimchi-Sarfaty (2003) ibid; and Arad (2005) ibid]. The DNA enclosed within said capsid can be delivered intact to the nucleus of a cell targeted for gene therapy.

SV40-based vectors are efficient in transduction of hematopoietic cell [Kimchi- Sarfaty et al., (2002) Hum. Gene Ther. 13:299-310; Kimchi-Sarfaty et al., (2003) Hum. Gene Ther. 14:167-177], adult human bone marrow cells [Rund (1998) ibid], CD34+ cells derived from human fetal ^"bone marrow [Strayer (2000) ibid], and human T-lymphocytes, [BouHamdan et al., (1999) Gene Ther. 6:660-666] . It also transduces NT2 cells, a human neuronal precursor cell line and mature neurons derived from NT2 precursor cells (neuronally committed human teratocarcinoma cell line) [Cordelier et al., (2003) MoL Ther. 7:801-810].

In vivo and in vitro produced SV40-based vectors are successful for in vivo gene delivery to the liver [Strayer D. S. and Zern, M. A. (1999) Sem. Liv. Dis. 91:71- 81; Arad (2005) ibid]. In liver-targeted gene therapy experiments in mice, the SV40 vector showed a long-term transgene expression and no detectable immune response directed against its proteins was found, even after repeated injections of the vector [Kondo (1998) ibid; Arad (2005) ibid]. SV40-foased vectors were shown to be successful also for in vivo gene delivery and gene expression in a variety of organs, kidneys among them [Strayer, D. S. (1996) J. Biol. Chem. 271: 24741-24746].

Acute renal failure

Acute renal failure (ARF) is a syndrome characterized by an abrupt kidney dysfunction that is reversible if the underlying condition has resolved. ARF is a serious condition that affects as many as 20% of intensive care unit (ICU) patients. The most common causes of acute renal failure in the ICU patient are severe sepsis, ischemia and drug toxicity. The mortality of acute renal failure in septic critically ill patients remains high despite the increasing ability to support vital organs [Wan, L. et al, (2003) Curr. OpL Crit. Care 9: 496-502].

ARF has an incidence of more than 200 patients per million persons per year, and among them, Acute Tubular Necrosis (ATN) patients are approximately 90 patients per million per year. The estimated patient population with acute renal disorders is stated at 100,000 patients per year.

Many causes can trigger pathophysiological mechanisms leading to acute renal failure. ARF syndrome is characterized by a sudden decrease in kidney function and loss of homeostasis. The primary evident marker is the increase of the nitrogenous components concentration in blood. Oliguria (a decreased urine output) is a second marker seen in 50% to 70% of the cases [Berl, T., et al., (1999) Atlas of diseases of the kidneys, Blackwell Publishers, First Edition].

Usually, the cause of ARF is multifactorial. Acute txibular necrosis (ATN) has been reported as the leading cause of acute renal failure among hospitalized patients and patients in IGUs, accounting for 38% and 76% of cases of acute renal failure, respectively. The total incidence of ATN in the general population is 88 cases per 1 million persons, per year. ATN is less frequent (P=O.004) in the very old than in the youngest patients [Berl C1999) ibid]. However, the number of Americans who are 65 years old and older, is continuously increasing and it is expected to double by the year 2030 [End of life ICU utilization may require re-evaluation according to University of Pittsburgh-led study: One of five terminally ill Americans dies in an ICTJ, (2004) University of Pittsburgh News, April 6, 2004].

Chronic renal failure is increased in the elderly population and an acute on chronic event that develops as ATN may result in severe consequences for those patients. It is now established that ATN independently and significantly affects patient survival. In a controlled study of ATN following radiocontrast procedures, it was found that ATN increased the odds rratio for death by 5.5. ATN patients are more likely to have hospital courses complications such as sepsis, respiratory failure, delirium, and bleeding, when Compared to other inpatients [Esson, M. L. et al., (2002) Ann. Inter. Med. 137: 744-752].

There are differences in the causes of ARF and lack of conformity in the use of the term "acute tubular necrosis." Acute tubular necrosis is a pathological diagnosis, and patients with ischemic or toxic insults to their kidneys might be expected to have tubular necrosis, but patients with ARF due to other causes might not [Thadhani, R. et al., (1996) New Engl. J. Med. 334: 1448-1460]. In acute tubular necrosis, if volume replacement does not restore renal function and mrine output remains low (less than 30ml/h), there is no evidence that any treatment improves renal function or accelerates renal recovery [Ashley, C. et al., (2001) Pharm. J. 266: 625-628]. Despite several experimental trials that tried to find a therapy to this entity, studies have not shown that medical therapy alters the course of established ATN [Esson (2002) ibid].

Mortality rates in ARF range from approximately 7% among patients admitted to a hospital with prerenal azotemia to more than 80°/6 among patients with postoperative acute renal failure. Mortality from ATN in. hospitalized and ICU patients is 37.1% and 78.6%, respectively. Despite major advances in dialysis and intensive care treatments, the mortality rate among patients with severe ARF (primarily iscliemic in origin) requiring dialysis have remained fairly constant over the past five decades [Berl (1999) ibid]. This may be explained by two demographic changes: the age of patients continues to rise, and coexisting serious illnesses common among these patients. When ARFoccurs in the setting of multioxgan failure, especially in patients with severe hypotension or the acute respiratory distress syndrome, the mortality rate ranges from 50 to 80 percent [Berl (1999) ibid; Esson (2002) ibid; Thadhani (1996) ibid].

Mortality rate among ARF patients was shown to increase slowly but constantly over a 40-year period (starting in 1951) in a follow-up study, despite the better availability of therapeutic armamentarium (mainly antibiotics and vasoactive drugs), deeper knowledge of dialysis techniques, and wider access to intensive care facilities [Berl (1999) ibid]. It was also shown that most of the episodes of ARF are resolved in the first month and the mean duration of ARF is 14 days. Among ARF casualties, seventy-eight percent of the patients die within the first two weeks after the renal insult. In the same way, 60% of survivors recover renal function by that time. After 30 days, the survival outcome of the ARF episode is conclusive in 90% of thte patients. Unfortunately, patients who eventually lose renal function, need to be included in a chronic dialysis program [Berl (1999) ibid].

ATN arising as a consequence of septic, toxic, or ischemic insult is a potentially reversible process, but frequently patients die from eo-morbid illness and ATN itself before renal recovery is achieved. Pharmacological therapies intending to prevent or alter the course of ATN are unsuccessful. Despite the technological advances in the dialysis treatments, the mortality xate among ATN patients who require dialysis remains between 50% and 80%. Alternative renal replacement therapy does not eliminate the death risk in ATN patients [Esson (2002) ibid]. Sepsis causes 30% to 70% of deaths in patients with ATN; therefore intravenous lines, bladder catheters, and respirators should be avoided. The large fluid volume administered to vasodilated septic patients triggers fluid accumulation in the lung interstitial of these patients and consequently a ventilatory support is required. Prolonged ventilatory support leads to acute respiratory distress syndrome (ARDS), multiorgran failure, and increased mortality [Esson (2002) ibid].

Apoptosis, or programmed cell death, is a normal component of the development and health of multicellular organisms. Cells die in response to a variety of stimuli and during apoptosis they do so in a controlled, regulated fashion. This makes apoptosis distinct from another form of cell death called necrosis in which uncontrolled cell death leads to lysis of cells, inflammatory responses and, potentially, to serious health problems. Apoptosis, by contrast, is a process in which cells play an active role in their own death (which is why apoptosis is often referred to as cell suicide). Upon receiving specific signals instructing the cells to undergo apoptosis a number of distinctive biochemical and morphological changes occur in the cell.

In experimental models of acute ischemic and toxic renal injury it was shown that renal tubular cells die by apoptosis as well as by necrosis and endothelial cells undergo apoptosis in response to a variety of stimuli. [Wan (2003) ibid].

The loss of renal tubular epithelium associated with ARF is attributed to both apoptotic and necrotic cell death, and patient recovery depends, in part, on the protection of renal tubular cells from death. Better understanding of the apoptotic intracellular signals, may allow molecular manipulation at different steps in these pathways that may avoid cellular apoptosis and consequently prevent or reduce renal damaged and improve patient outcome.

Heat shock causes the intracellular expression of a specific group of proteins called lieat shock proteins (HSPs) that have broa_d cytoprotective properties. Induction of HSPs also protected cells and whole organs against nonthermal cytotoxic agents such as oxidants, tumor necrosis factor, and endotoxin. Oxidative stress agents such as nitric oxide (NO) and reactive oxygen intermediates (ROI) are known to be major mediators of cell damages during inflammation.

HSPs can be divided into six subfamilies according to their molecular weights. The HSP 70 family is the most conserved and best-studied family. Human cells contain several HSP 70 family members including highly stress-inducible HSP 70 (also known as HSP 72 or HSP 7Oi), constitixtively expressed heat shock cognate protein HSC 70 (HSP 73), a mitochondrial HSP 75 (mtHSP 75) and Grp78 (BiP) localized in the endoplasmic reticulum..

Under normal conditions, HSP 70 proteins function as ATP-dependant molecular chaperones. Under various stress conditions, the synthesis of stress- inducible HSP 70 can effectively inhibit cellular death processes, apoptosis or necrosis, and thereby increase the survival of cells exposed to a wide range of letlial stimuli. It has been demonstrated that hea.t stress and the subsequent induction of HSP 72 protect renal tubular cells from ischemia-induced apoptosis [Meldrum K. K. et al., (2001) Am. J. Physiol. Regul. Integ. Comp. Physiol. 281: R359-R364].

Necrosis results from severe ATP depletion and leads to rapid and uncoordinated collapse of cellular homeostasis [Wan (2003) ibid]. HSP 72 mediates acquired resistance to ischemic injury in REC-(ATP)-depleted renal epithelial cells [Wang, Y. H. et al., (2002) Cell Stress & Chaperones 7: 137-145].

Although the beneficial effects of inducing HSP 72 in the intact kidney have yet to he demonstrated in vivo, insults that indu.ce HSP 72 are frequently ameliorated when this molecular chaperone is expressed before injury. Pharmacologic ischemia in vitro and renal ischemia in vivo markedly increase the HSP 72 expression. Therefore, HSP 72 may provide a novel therapeutic tool for ameliorating the untoward effects of renal ischemia on organ function [Wang (2002) ibid].

The concomitant upregulation of HSP 72 expression observed in renal cortex of iNOS knockout mice (under basal conditions) and the fact that iNOS knockout mice kidneys are protected against ischemic acute renal failure, support the notion that the protective effect is connected to the compensatory upregulation of HSP 72 [Ling, H. et al., (1999) Am. J. Physiol. 277: F383-F390].

It is the intention of this invention to provide a gene delivery system and method for the treatment of renal diseases. This system may comprise HSP70 or any other factor that might interfere with apoptotic signals, avoid and/or restore renal tissue damage.

It is also the intention of this invention to provide a cost effective treatment, which will significantly reduce hospitalization expenses of ARF patients.

Summary of the Invention

As a first aspect, the invention relates to an SV40 renal delivery system for targeting an exogenous nucleic acid molecule to the kidney in vivo. More particularly, the delivery system of the invention comprises as an active ingredient an SV40 infectious particle comprising said nucleic acid molecule packaged therein. This nucleic acid molecule nxay be a molecule encoding a product involved in a renal pathological disordex or alternatively, a molecule directed against a product involved in a renal patliological disorder.

In one preferred embodiment, the exogenous rmcleic acid molecule comprised "within the delivery system of the invention, or a product of said molecule, is capable of reducing and/or preventing a kidney tissue pathology and thereby prevent said renal pathological disorder. More particularly, such pathologic disorder may be caused by apoptosis, necrosis, fibrosis or inflammation of kidney cells or tissue. In such case, a preferred delivery system according to the invention may comprise a nucleic acid molecule encoding a HSP70 family member protein, preferably, any HSP 70 isoform from prokaryotic or eukaryotic origin. A preferred eukaryotic HSP70 may be a mammalian, preferably human HSP70 family member protein.

In a preferred embodiment, the SV40 renal delivery system of the invention comprises as an active ingredient SV40 infectious particle, which may be any one of an in vitro constructed SV40 pseudo viral particle and an SV40 pseudoviral or viral vector propagated in tissue culture cells that provide in trans large T antigen, required for vector propagation.

In another aspect, the invention relates to a pharmaceutical composition for the treatment of a renal pathological disorder in a mammalian subject in need thereof. The composition of the invention comprises as an effective ingredient the SV40 renal delivery system of the invention, and optionally further comprising pharmaceutically acceptable carrier, diluent, excipient and/or additive. The invention further provides a method for the treatment of a renal pathological disorder.

Still further, the invention relates to the use of tlie renal SV40 delivery system in the preparation of a pharmaceutical composition for the treatment of a renal pathological disorder in a subject in need.

In a further aspect, the invention relates to a method of preparing a therapeutic composition for the treatment of a renal pathological disorder, which method comprises the steps of: (a) providing the renal SV40 delivery system of the invention; and (b) admixing said delivery system with a pharmaceutically acceptable carrier. Brief Description of the Figures

Figure 1A-1D Renal expression of a reporter gene delivered by an SV40 viral vector

The figure shows different rat kidney histological sections expressing the luciferase reporter gene introduced by the vectors of the invention (brown stained areas) and control sections.

Fig. IA shows untransfected rat kidney (control) [xlO magnification].

Fig. IB shows luciferase expression in txansfected rat kidney [xlO magnification].

Fig. 1C shows untransfected rat kidney (control) [x20 magnification]. Fig. ID shows luciferase expression in txansfected rat kidney [x20 magnification] .

Figure 2A-2C Renal expression of a reporter gene delivered by an SV40 viral vector

Mice were injected via the tail vain with 1O⁸ SV40/lαc i.e. SV40 vectors encoding the reporter gene luciferase, prepared, as described in experimental procedures. Mice were sacrificed and kidney tissue sections were stained with anti Luciferase antibody [x 40 magnification].

Fig. 2A shows uninfected mouse kidney (PBS control) [x40 magnification]. Fig. 2B shows Luciferase expression in SV40/luc infected mouse kidney [x40 magnification]. Fig. 2C shows magnification (x4) of the framed region in Fig. 2B.

Detailed Description of the Invention

The invention relates to a new therapy approach to treat renal disorders. This approach is aimed to use organ-targeted gene delivery, which expression generates a therapeutic effect.

Therefore, in. a first aspect, the invention relates to an SV40 renal delivery system for targeting an exogenous nucleic acid molecule to a kidney cell. More particularly, the delivery system of the invention comprises as an active ingredient an SV40 infectious particle comprising said nucleic acid molecule packaged therein. This nucleic acid molecule may be a molecule encoding a product involved in a renal pathological disorder or alternatively, a molecule directed against a product involved in a renal pathological disorder.

As used herein, the term "nucleic acid" refers to polynucleotides such as deoxyribonucleic acid (DNA), and, where appropriate, ribonucleic acid (RNA). The term should also be understood to include, as equivalents, analogues of either RNA or DNA made from nucleotide analogu.es, and, as applicable to the embodiment being described, single-strand.ed and double-stranded polynucleotides. Recombinant nucleic acid sequence is a molecule made of segments, which are naturally not normally linked, in the same manner. Thus, a recombinant nucleic acid molecule comprised within the delivery system of invention may be a continuous nucleic acid molecule having sequences operably linked, typically translated to a single product that exhibits properties derived from the original segments.

Thus, according to one embodiment, the nucleic acid molecule comprised within the SV40 renal delivery system of the invention may be a purified exogenous naked DNA, or a purified exogenous naked DNA encoding a protein or peptide, or a purified naked DNA encoding RNA, wherein said DNA, protein, peptide or RNA, is involved in a renal pathological disorder. Et should be noted that any of these purified exogenous naked DNAs may be comprised within a vector. Such molecule may be a circular or linear exogenous recombinant DNA.

The exogenous nucleic acid molecule of the infectious particles comprised within the delivery system of the invention, may further comprises operably linked control and regulatory elements necessary for the replication and/or the expression of said molecule. It also may include elements necessary for the prevention of the expression of undesired protein or proteins in a mammalian infected cell.

It should be noted that the term "operably linked" is used herein for indicating that a first nucleic acid sequence is operably linked with a second nucleic acid sequence when the first nucleic acid sequence is placed in a functional relationship with the second nucleic acid sequence. For instance, a promoter is operably linked to a coding sequence if the promoter affects the transcription or expression of the coding sequence. Generally, operably linked DNA sequences are contiguous.

Control and regulatory elements mentioned in the invention include promoters, terminators and other expression control elements. Such regulatory elements are described by Goeddel [Goeddel, Gene Expression Technology: Methods in Enzymology 185, Academic Press, San Diego, Calif. (1990)].

The promoter used in the infectious particle comprised within the delivery system of the invention may be a constitutive promoter or regulable, preferably, a promoter which can be induced by a protein or any other element which trans-activates transcription.

In another alternate embodiment, the purified exogenous nucleic acid molecule comprised within the delivery system of the invention, may be an RNA molecule, a purified exogenous RNA encoding an exogenous protein or peptide involved in a renal pathological disorder, or a vector comprising any of these purified exogenous RNAs.

As indicated above, the DNA or RNA encompassed within the delivery system of the invention may be comprised within an expression vector. "Vectors", as used herein, encompass plasmids, viruses, bacteriophage, any DNA fragment capable of integration, and other vehicles, which enable the integration of such DNA fragments into the genome of the host trans fected cell. Expression vectors are typically self-replicating DNA or RNA constructs containing the desired gene or its fragments, and operably linked genetic control elements that are recognized in a suitable host cell and effect expression of the desired genes. These control elements influence the expression of the cloned sequences within a suitable host. Generally, the genetic control elements can include a prokaryotic promoter system or a eukaryotic promoter expression control system. This typically includes a transcriptional promoter, an optional operator to control the onset of transcription, transcription enhancers to elevate the level of RNA expression, a sequence that encode s a suitable ribosome binding site, RNA splice junctions, sequences that "terminate transcription and translation and so forth. Expression vectors "usually contain an origin of replication that allows the vector to replicate independently of the host cell.

A vector may additionally include appropriate restriction sites, antibiotic resistance or other markers for selection of vector containing cells. Plasmids are the most commonly used form of vector but other forms of vectors which serves an equivalent function and which are, or oecome, known in the art are suitable for use herein. See, e.g., Pouwels et al. Cloning Vectors: a Laboratory Manual (1985 and supplements), Elsevier, N. Y- ; and Rodriquez, et al. (eds.) Vectors: a Survey of Molecular Cloning Vectors and their Uses, Buttersworth, Boston, Mass (1988), which are incorporated here in by reference.

Still further, the nucleic acid molecule comprised within the delivery system of the invention may be a purified exogenous antisense RNA, RNAi (interfering RNA), or other nucleic acids that produce RNA interference, such as siRNA (short interfering RNA), or DNA encoding shRNA purified exogenous ribozyme RNA or a purified exogenous RNA or purified exogenous naked DNA, or DNA encoding ribozyme, etc. It should be noted that these RNA based molecules are specifically directed against a gene product involved in a renal pathological disorder and thereby inhibits or prevents the expression of such gene product in mammalian kidney cells.

The term "siRNAs" refers to short interfering RNAs. The term "RNA interference" or "RNAi" refers to the silencing or decreasing of gene expression by siRNAs. It is the process of sequence-sp ecific, post-transcriptional sequence- specific gene silencing in animals and plants, initiated by siRNA that is homologous in its duplex region to the sequence of the silenced gene. The gene may be endogenous or exogenous to the organism, integrated into a chromosome or present in a transfection vector which is not integrated into the genome. The expression of the gene is eittier completely or partially inhibited. RNAi may also inhibit the function of a ta.rget RNA, and said function may be completely or partially inhibited.

The term "ribozyme RNA" or "ribozymes" refers to RNA molecules having an enzymatic activity which is able to repeatedly cleave other separate RNA molecules in a nucleotide base sequence sp ecific manner.

Different basic varieties of naturally-occurring enzymatic RNAs are known presently. Each can catalyze the hydrolysis of RNA phosphodiester bonds in trans (and thus can cleave other RNA molecules) under physiological conditions. In general, enzymatic nucleic acids, such as ribozymes, act by first binding to a target RNA. Such binding occurs through the target binding portion of an enzymatic nucleic acid which is held in close proximity to an enzymatic portion of the molecule that acts to cleave the target RNA. Thus, the enzymatic nucleic acid first recognizes and then binds a target RNA through complementary base-pairing, and once bound to the correct site, acts enzymatically to cut the target RNA. Strategic cleavage of such a target RNA will destroy its ability to direct synthesis of an encoded protein. After an enzymatic nucleic acid has bound and cleaved its RNA target, it is released from that RNA to search for another target and can repeatedly bind and cleave new targets.

The terms "infectious particle", "transduction vehicle or transfection vehicle", "virus, viroid, virion or pseudoviral particle", "SV40-derived particles, virus or pseudovirus" and "viral vectors" are used in a similar manner, and they relate to a viral-like particle useful for transmitting new genetic information to a cell.

In yet another preferred embodiment, the exogenous nucleic acid molecule comprised within the delivery system of the invention, or a product of said molecule, is capable of reducing and/or preventing kidney tissue pathology and thereby prevent said renal pathological disorder.

According to a particular embodiment, the product encoded by the nucleic acid molecule may be any one of an enzyme, a receptor, a structural protein, a regulatory protein, hormone, protein that participates in signal transduction and protein that participates in sorting and trafficking of molecules.

According to a specific embodiment, the renal pathological disorder to be prevented by the delivery system of the invention may be caused by apoptosis, necrosis, fibrosis or inflammation of kidney cells or tissue. In such case, a preferred delivery system according to the invention may comprise a nucleic acid molecule encoding a HSP70 family menxber protein, preferably, any HSP 70 isoform from prokaryotic or eukaryotic origin. A preferred eukaryotic HSP70 may be a mammalian, preferably human HSP70 family member protein. Therefore, according to a particular embodiment, the delivery system of the invention comprises a nucleic acid molecule encoding a human HSP70 family member protein. Preferably, said nucleic acid molecule comprises the nucleotide sequences as denoted in SEQ ID NO: 1 or SEQ ID NO: 2. Apoptosis is a normal cellular process also called programmed cell death. This programmed cell death, follows a specific sequence of events triggered in the course of normal development or as a means of preserving normal function. Cells that die by apoptosis do not usually elicit the inflammatory responses that are associated with necrosis.

In contrast, "necrosis" relates to localized death of cells in an organ or tissue, caused by a variety of chemicals and toxic substances, or by a disease or injury. Necrosis represents the pathological death of part of a tissue due to irreversible damaged.

In a preferred embodiment, the SV40 renal delivery system of the invention may be capable of conferring cytoprotection to a cell infected therewith, wherein said cell was subjected to physical and/or chemical stress. According to another embodiment, such kidney cell may be selected from the group consisting of human kidney cells, renal mesangial cells, epithelial tubular cells, renal endothelial cells, podocytes; and/or infiltrating leukocytes within the glomerulus or the renal interstitium.

In yet another embodiment, the delivery system of the invention may comprise as an active ingredient an infectious particle containing a nucleic acid molecule which encodes a product involved in a renal disorder, or a product which is specifically directed against a renal disorder. Preferably, the renal disorder may be acute and reversible or may be caused by a chronic infliction. According to a preferred embodiment, the renal pathological disorder may be any one of acute renal failure due to acute tubular necrosis, acute glomerulonephritis, acute renal allograft rejection, and chronic renal disorders such as diabetic nephropathy, nephritic syndrome, hypertensive renal disease, polycystic renal disease, chronic glomerulonephritis, chronic rejection of kidney allograft and HIV nephropathy. Thus, according to a particularly preferred embodiment, the SV40 renal delivery system of the invention may be capable of conferring cytoprotection to a cell infected therewith in a mammalian subject suffering from a renal pathological disorder.

More specifically, cytoprotection conferred by the delivery system of the invention may results in the preservation of cell functions and improves renal cell survival and tissue functioning by inhibiting cellular death processes such as apoptosis, necrosis, and tissue fibrosis or infL animation.

The term "cytoprotection" or "cytoprotective" a_s used herein implies a process by which chemical compounds provide protection to cells against harmful agents.

In a preferred embodiment, the SV40 renal ielivery system of the invention comprises as an active ingredient SV40 infectious particle which may be an in vitro constructed SV40 pseudoviral particLe or alternatively, an SV40 pseudoviral or viral vector propagated in tissue culture cells.

According to one embodiment, the SV40 infectious particle may be an in vitro packaged SV40 pseudoviral particle. One preferred example may be an in vitro constructed SV40 pseudovirus comprising at least one pure or partly purified SV40 capsid protein (VPl, VP2 or VP3). Preferably, VPl, and an exogenous nucleic acid molecule as described above.

In one particular embodiment, the in vitro paclsaged SV40 pseudoviral particle may be prepared for example by a method com/prising the steps of: (a) allowing a pure or partly purified SV40 capsid protein or a mixture of at least two SV40 capsid proteins to self-assemble into SV40-like particles; and (b) bringing the SV40-like particles assembled in step (a) into contact with said exogenous nucleic acid molecule to give an in vitro constructed pseudoviral particles. In an alternative embodiment, the SV40 infectious particle comprised within the SV40 renal delivery system of the invention may be an in vivo SV40 pseudoviral or viral vector. Preferably, a vector prepared in vivo in tissue culture.

According to a specific embodiment of such delivery system, the viral vector may be a recombinant T-Ag deleted SV40 viral or pseudoviral vector. Such vector may preferably prepared by a method comprising the steps of: (a) providing a complementation SV40 packaging cell line for in trans complementation of viral T-antigen; (b) infecting or transfecting said cell line with any one of SV40 viral or pseudoviral vector; (c) culturing the infected cells under suitable conditions for permitting the production of said SV40 viral or pseudoviral vector; and (d) harvesting the viruses or pseudoviruses.

As indicated in the Experimental procedures, the viral vectors of the invention may be prepared in vivo in COS-I cells. It should be noted that these vectors may be prepared in vivo using any appropriate complementation cell line.

Complementation cell line refers to a. eukaryotic cell permissive or semi- permissive for SV40 and SV40-derived viral propagation and capable of providing in-trαns the function(s) for which an SV40 or SV40-derived vector is defective. In other words, it is capable of producing the protein or proteins needed for replication and indirectly for encapsidation of said SV40 vectors.

The complementation cell line may be derived either from an immortalized cell line capable of dividing indefinitely, or from a primary line.

A suitable complementation SV40 packaging cell line for in trans complementation of viral T-antigen, is devoid of the capability of generating viable virus carrying the T-antigen gene and has reduced homologous recombination in the packaging process of an SV40 vector. This strategy eliminates the production of viable SV40 viruses during the preparation of SV40 viral or SV40 pseudoviral vector stocks.

A particular example may be a cell line transformed with at least one expression cassette having minimal sequence identity to SV40 sequences comprised within said SV40 vector. This cassette may comprise for example, a nucleic acid encoding SV40 T-antigen, a heterologous promoter, a heterologous termination signal and optionally additional operably linked control elements and/ or selectable markers.

According to a particular embodiment, such complementation packaging cell line may be COT18 (DSM Accession ISTo. ACC2525) or a cell line derived therefrom.

In another aspect, the invention relates to a pharmaceutical composition for the treatment of a renal pathological disorder in a mammalian subject in need thereof. The composition of the invention, comprises as an effective ingredient the SV40 renal delivery system of the invention, and optionally further comprising pharmaceutically acceptable carrier, diluent, excipient and/or additive.

Therapeutic formulations may be administered in any conventional dosage formulation. Formulations typically comprise at least one active ingredient, as defined above, together with one or more acceptable carriers thereof.

Composition dosages may be any that induce a cytoprotective response. It is understood by the skilled artisan that the preferred dosage would be individualized to the patient following good laboratory practice and standard medical practice.

Each carrier should be both pharmaceutically and physiologically acceptable in the sense of being compatible with the other ingredients and not injurious to the patient. While formulations include those suitable for topical, oral, rectal, nasal, preferred formulations are intended for parenteral administration, including intramuscular, intravenous, intratracheal, intradermal and subcutaneous administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any methods known in the art of pharmacy.

The pharmaceutical forms suitable for injection include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases the form must be sterile and must be fluid to the extent that easy syringeability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. The carrier can be solvent or dispersion medium containing, for example, phosphate-buffered saline, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glyol, and the like), suitable mixtures thereof, and vegetable oils. The proper fluidity can be maintained, for example, by the use of a coating, such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.

As used herein "pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, coatings and. the like. The use of such media and agents for pharmaceutical active substances is well known in the art. For any conventional media or agent that is compatible with the active ingredient, use in the therapeutic composition is contemplated.

The therapeutic agent should be delivered in sufficient dose to each target cell. Preferred means of administration is via the blood system (e.g. injection). Administration may, depending on the case, also be done by organ perfusion, catheterization through blood vessels to the target organ, orally, direct injection into an organ or by absorption through the lower gastrointestinal tract. The invention further relates as a third aspect to a method for the treatment of a renal pathological disorder in a mammalian subject in need. The method of the invention comprises the step of administering to said subject the SV40 delivery system or the composition of the invention, in an amount sufficient to induce in said subject a cytoprotective effect.

The term "effective amount" or "sufficient amount" means an amount necessary to achieve a selected result. The "effective treatment amount" is determined by the severity of the disease in conjunction with the therapeutic objectives, the route of administration and the patient's general condition (age, sex, weight and other considerations known to th.e attending physician). For example, an effective amount of the composition of the invention useful for the treatment of said pathology will be an amount sufficient to induce a cytoprotective effect to achieve a selected result and avoid cellular death and tissue dysfunction. For example, an effective amount of the delivery system or the composition of the invention will be conferring cytoprotective effect to renal cells against the treated of a chemical or physical insult or to detrimental effects of an acute or chronic renal disorder, a kidney allograft rejection, or a severe infection, reducing the possibility of these cells to undergo any one of the processes that will result in cell death, such as apoptosis or necrosis.

The term "pathology" or "pathological" used therein the invention relates to the structural and functional deviations from the normal that constitute disease or characterize a particular disease. In a broader definition, pathology concerns with all aspects of disease, with special reference to the essential nature, causes, and development of abnormal conditions, as well as the structural and functional changes that result from the disease processes.

The term "condition" or "disorder" refers to a situation in which there is a disturbance of normal functioning condition and that includes chronic or permanent health defects resulting from disease, injury, or congenital m alfor mations . A "Renal disorder" is a disease or disorder characterized by an abnormality or malfunction of the kidney. One category of renal disorders refers to acute or chronic conditions, deriving from immune or* fibrotic processes; other category refers to renal transplant immune rejections and other renal conditions may result as a consequence of severe bacterial or viral infections or fibrotic processes.

As used herein to describe the present indention, "renal failure" or "renal dysfunction" is defined as a sustained decrease in estimated glomerular filtration rate (GFR). Renal failure may be caused by a physical or chemical insult, or by a pathological infliction resulting in acute or chronic renal failure, which could be prevented or reversed upon treatment.

"Tissue damaged" is a condition that impedes normal tissue function. Tissue damaged could be terminal or reversible. Terminal tissue damaged will result in cellular death and permanent tissue dysfunction. Reversible tissue damaged may result as a consequence of an inflammatory or fibrotic process of the tissue which can result in a temporarily or partial organ dysfunction.

According to a preferred embodiment, the nxethod of the invention is intended for the treatment of a renal disorder which may be any one of acute disorders consisting of: acute renal failure (ARF) due to acute tubular necrosis (ATN), acute glomerulonephritis (AGN), acute tubulointerstitial nephritis, necrotising vasculitis or acute renal allograft rejection.

The term "Acute renal failure" disorders includes, but is not limited to, acute tubular necrosis (ATN), acute glomerulonephritis (AGN), acute tubulointerstitial nephritis, necrotizing vasculitis or an acute renal allograft rejection. In one particular embodiment renal disorder may be chronic, selected from a group consisting of: diabetic nephropathy, nephrotic syndrome, hypertensive renal disease, polycystic renal disease and chronic glomerulonephritis.

In another particular embodiment the renal disorder may be due to acute or chronic renal allograft rejection, or to an HIV nephropathy.

According to a preferred embodiment, the cytoprotective effect caused by the method of the invention results in the preservation of cell functions and improves renal cell survival and tissue functioning by inhibiting cellular death processes such as apoptosis, necrosis, and tissue fibrosis or inflammation.

Treatment" refers to therapeutic treatment. Those in need of treatment are mammal subjects suffering from a renal dysfunction. By "patient" or "subject in need" is meant any mammal for which the gene therapy treatment is desired in order to overcome the renal infliction.

"Mammal" for purposes of treatment refers to any animal classified as a mammal including, human, research animals, domestic and farm animals, and zoo, sports, or pet animals, su.ch as dogs, horses, cats, cows, etc. Preferably, the mammal is human.

The method for the treatment of a renal disorder described in the invention when administered to a mammalian subject in need, exerts a cytoprotective effect which results in the preservation of cell functions and consequently improves renal cell survival and tissue functioning by inhibiting cellular death processes such as apoptosis or necrosis, and tissue fibrosis or inflammation.

The invention further relates to the use of the renal SV40 delivery system of the invention in the preparation of a pharmaceutical composition foar the treatment of a renal pathological disorder in a subject in need. In a further aspect, the invention relates to a method of preparing a therapeutic composition for the treatment of a renal pathological disorder, which method comprises the steps of: (a) providing the renal SV40 delivery system of the invention; and (b) admixing said delivery system with a pharmaceutically acceptable carrier.

As used in the specifications and the appended claims and in accordance with long-standing patent Law practice, the singular forms "a" "an" and "the" generally mean "at least one", "one or more", and other plural references unless the context clearly dictates otherwise. Thus, for example "a cell", "a peptide" and "a nucleic acid" include mixture of cells, one or more peptides and a plurality of nucleic acids of the type described.

Throughout this specification and the claims which follow, unless tlhe context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any otfcier integer or step or group of integers or steps.

The contents of all publications quoted to herein are fully incorporated by reference.

The following examples are representative of techniques employed by the inventors in carrying out aspects of the present invention. It should be appreciated that while these techniques are exemplary of preferred embodiments for the practice of the invention, those of skill in the sjct, in light of the present disclosure, will recognize that numerous modifications can be made without departing from the spirit and intended scope of the invention. Examples

Experimental procedures

Cell cultures and media,

*COS-1 cell-line is a derivative of CVl (ATCC # CCL70) harboring tie SV40 T- antigen gene. COS-I cells were cultured in high glucose Dulbecco's modified Eagle's medium containing glutamine, penicillin, streptomycin, and 10% FBS. These cells were used for in vivo preparation of SV40 viral vectors.

Luciferase activity

Luciferase activity was used as a measure of the titer of functionally transducing vector. It was quantified by infecting COS-I cells with SV/luc vector preparations in a 24-well plate. All infections were performed in triplicate. Forty-eight liours post-infection luciferase activity was analyzed with the Steady-glo™ kit (Promega) and a SPECTRAFluor Plus microplate luminometer (TECAN).

Preparation of viral vectors for in-vivo targeting in mice

Vector harvesting and concentration included lysing the cells in 0.5% dexoycholate and 1% TirtonXIOO, pelleting cellular debris by ceotrifugation and pelleting the virus in the supernatant by ultracentrifugation (80,000 g for 6 hours). The virus pellet was re-suspended in PBS by sonication amd residual detergents were removed by batch treatment with Bio-Beads (BioRa.d).

Immunohistochemical detection of luciferase

Immunohistochemical detection was performed according to [La.von et al., (2000) Nat. Med. 6(5): 573-7]. Briefly, paraffin-embedded sections were pretreated by incubation in 0.01 M citrate buffer and heated in a microwave oven twice for 5 mim. Samples were then incubated for 60 min at room temperature with rabbit polyclonal antibodies against luciferase (1:100 dilution; Cortex Biocliem, San Leandro, CA), washed, and incubated with biotin-conjugated anti-rabbit goat antibody (1:100 dilution; Jackson Immunoresearch, West Grove, PA). The samples were then labeled with peroxidase-conjugated streptavidin and luciferase was detected using 3-amino- 9-ethyl carbazole substrate.

Example 1

Renal gene delivery and expression of a reporter gene

SV40 derived viruses particles which encode for the lucifer/ase gene were prepared on COS-I cells as described in Arad U. et al., (2002) Vixology 304: 155- 159, WO03/025189 and in Experimental procedures.

Anesthetized rats were intravenously injected with 1.25xL0⁸ transducing units/animal of SV40 derived viruses expressing the reporter gene luciferase (Luc). Luciferase expression was evaluated, 24 or 48 hoxirs after viral administration, using a cooled charge -coupled devise (CCCD) camera and by immunohistochemistry, using rabbit polyclonal anti-Luc and biotin conjugated goat anti-rabbit antibodies. As seen in Figure 1, significant SV40 luciferase expression was observed in renal paraffin-embedded sections of rat transfected with the SV40 derived virus carrying the luciferase gene (Figαire IB and ID) while no detectable luciferase expression was observed in control rats (Figure IA and 1C).

As shown in Figure 2, significant luciferase expression was also detected in kidney cells in sections of mice that were injected (iv) with IxIO⁸ transducing units/animal of SV40 derived viruses expressing this reporter gene (Luc).

The SV40 derived viruses particles described in the experiment were able to reach the target organ (kidney) and express the reporter gene carried within the viral particle. Therefore, these particles can successfully be used for gene delivery and gene expression in renal tissue, representing an appealing tool for gene therapy treatment of renal disorders. Example 2

Optimization of SV40 viral particle number transfection

Mice, rats and psammomys obesus are injected with increasing- amount of SV40 viruses in order to calibrate the number of SV40 viral particles needed for optimal renal delivery in the different animal species. Viral detection is performed by immunohistochemistry analysis of kidney sections. Note, this experiment involved SV40 viruses rather than vectors and the antibody reacted with the SV40 capsid protein VPl rather th.an luciferase. Immunohistoch.emistry was otherwise performed as described above.

Example 3

SV40 vectors delivery to ATN animals

ATN was induced chemically in Balb/C mice using the nephrotoxic heavy metal mercury at 4-6 mg/kg. ATN was reproducible as evident by elevated levels of nitrogenous kidney function markers measured 16-18 hours, 48 hours and 5 days after administration. This model serves to examine the utility and feasibility of SV40-based HSP70 gene delivery into animals developing ATN.

Claims

1. An SV40 nrenal delivery system for targeting an exogenous nucleic acid molecule to a kidney cell, comprising as an active ingredient an SV40 infectious particle comprising said nucleic acid molecule packaged therein, wherein said nucleic acid molecule is any one of a molecule encoding a product involved in a renal pathological disorder and a molecule directed against a product involved in a renal pathological disorder.

2. The SV4O renal delivery system according to claim 1, wherein said nucleic acid molecule is selected from the group consisting of:

(a) a purified exogenous naked DNA, or a purified exogenous naked DNA encoding a protein or peptide, or a purified naked DNA encoding RNA, wherein said any one of DNA, a protein, a peptide and RNA, is involved in a renal pathological disorder;

(b) a vector comprising any of the purified exogenous naked DNAs of (a);

(c) a purified exogenous RNA, or a purified exogenous RNA encoding an exogenous protein or peptide involved in a renal pathological disorder ;

(d) a vector comprising any of the purified exogenous RNTAs of (c); and

(e) a purified exogenous antisense RNA, RNAi, siRNA (short interfering RNA), purified exogenous ribozyme RNA or a purified exogenous RNA or purified exogenous naked DNA, wherein said any one of antisense RNA, RNAi, siRNA and ribozyme RNA is directed against a gene product involved in a renal pathological disorder and inhibits or prevents the expression of said gene product in a mammalian kidney cell;

Wherein said exogenous nucleic acid molecule or a product of said molecule, is capable of reducing and/or preventing kidney tissue pathology and thereby prevent said renal pathological disorder.

3. The SV40 renal delivery system according to any of claims 1 to 2, wherein said product encoded by said nucleic acid molecule, is any one of an enzyme, a receptor, a structural protein, a regulatory protein, a hormone. A protein participating in signal transduction and a protein involved in sorting and traffic-king.

4. The SV40 renal delivery system according to any one of claims 1 to 3, wherein said renal pathological disorder is caused by apoptosis, necrosis, fibrosis or inflammation of kidney cells or tissue and wherein said exogenous protein exerts cytoprotective effect on kidney cells.

5. The SV40 renal delivery system according to any of claims 1 to 4, wherein said nucleic acid molecule encodes a HSP70 family member protein, wherein said HSP70 is any HSP 70 isoform.

6. The SV40 renal delivery system according to claim 5, wherein said HSP70 is from prokaryotic or eukaryotic origin.

7. The SV40 renal delivery system according to claim 6, wherein said HSP70 from eukaryotic origin is a mammalian, preferably liuman HSP70 family member protein.

8. The SV40 renal delivery system according to claim 7, wherein said nucleic acid molecule comprises the nucleotide sequences as denoted in SEQ ID NO: 1 or SEQ ID NO: 2.

9. The SV40 renal delivery system according to any of claims 5 to 8, being capable of conferring cytoprotection to a cell infected therewitti, wherein said cell was subjected to physical and/or chemical stress.

10. The SV40 renal delivery system according to any of the claims 1 to 9, wherein said kidney cell is selected from the group consisting of human kidney cells, renal mesangial cells, epithelial tubular cells, renal endothelial cells, podocytes, and/or infiltrating leukocytes within the glomerulus or the renal interstitixxm.

11. The SV40 renal delivery system as defined in any one of claims 1 to 10, wherein said renal disorder is acute and reversible or is caused by a chronic infliction.

12. The SV40 renal delivery system according to claim 11, wherein said renal pathological disorder is any one of acute renal failure due to acute tubular necrosis, acute glomerulonephritis, acute renal allograft rejection, and chronic renal disorders such as diabetic nephropathy, nephritic syndrome, hypertensive renal disease, polycystic renal disease, chronic glomerulonephritis, chronic rejection of kidney allograft and HIV nephropathy.

13. The SV40 renal delivery system according to any one of claims 1 to 12, being capable of conferring cytoprotection to a cell infected therewith in a mammalian subject suffering from a renal pathological disorder.

14. The SV40 renal delivery system according to claim 13, wherein said cytoprotection results in the preservation of cell functions and improves renal cell survival and tissue functioning by inhibiting cellular death processes such as apoptosis, necrosis, and tissue fibrosis or inflammation.

15. The SV40 renal delivery system according to any on.e of claims 1 to 14, wherein said SV40 infectious particle is any one of an in vitro packaged SV40 pseudoviral particle and an SV40 pseudoviral or viral vector.

16. The SV40 renal delivery system according to claim 15, wherein said SV40 infectious particle is an in vitro packaged SV40 pseudoviral particle comprising at least one pure or partly purified SV40 capsid protein, preferably VPl, and said exogenous nucleic acid molecule.

17. The SV40 renal delivery system according to claim 15, wherein said SV40 infectious particle is an SV40 pseudoviral or viral vector.

18. The SV40 renal delivery system according to claim. 17, wherein said viral vector is a recombinant T-Ag deleted SV40 viral or pseudoviral vector prepared by a method, comprising the steps of:

(a) providing a complementation SV40 packaging cell line for in trans complementation of viral T-antigen;

(b) infecting or transfecting said cell line with any one of SV40 viral or pseudoviral vector;

(c) culturing the infected cells under suitable conditions for permitting the production of said SV40 viral or pseudoviral vector; and

(d) harvesting the viruses or pseudoviruses.

19. A pharmaceutical composition for the treatment of a renal pathological disorder in a mammalian subject in need thereof, comprising as an effective ingredient an SV40 renal delivery system as defined in any one of claims 1 to 18, and optionally further comprising pharm.aceutica.lly acceptable carrier, diluent, excipient and/or additive.

20. A method for the treatment of a renal pathological disorder in a mammalian, subject in need, comprising the step of administering to said subject an SV40 delivery system as defined in any of claims 1 to 18 or a composition as defined in claim 19, in an amount sufficient to induce in said subject a cytoprotective effect.

21. The method according to claim 20, wherein said renal disorder is any one of acute disorders consisting of: acute renal failure (ARIF) due to acute tubular necrosis (ATN), acute glomerulonephritis (AGN), acute tubulointerstitial nephritis, necrotising vasculitis or renal allograft rejection.

22. The method according to claim 20, wherein said renal disorder is chronic, selected from a group consisting of: diabetic nephropathy, nephrotic syndrome, hypertensive renal disease, polycystic renal disease and chronic glomerulonephritis.

23. The method according to claim 20, wherein said renal disorder is due to acute or chronic renal allograft rejection, or to an HIV nephropathy.

24. The method according to any of the claims 20 to 23, wherein said cytoprotective effect results in the preservation of cell functions and improves renal cell survival and tissue functioning by inhibiting cellular death processes such as apoptosis, necrosis, and tissue fibrosis or inflammation.

25. Use of a renal SV40 delivery system as defined in any one of claims 1 to 18, in the preparation of a pharmaceutical composition for the treatment of a renal pathological disorder in a subject in need.

26. A method of preparing a therapeutic composition for the treatment of a renal pathological disorder, which method comprises the steps of:

(a) providing a renal SV40 delivery system as defined in any one of claims 1 to 18; and

(b) admixing said delivery system with a pharmaceutically acceptable carrier.