WO2001046405A2 - Modulation of pax-2 for controlled apoptosis or survival of cells - Google Patents
Modulation of pax-2 for controlled apoptosis or survival of cells Download PDFInfo
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- WO2001046405A2 WO2001046405A2 PCT/CA2000/001545 CA0001545W WO0146405A2 WO 2001046405 A2 WO2001046405 A2 WO 2001046405A2 CA 0001545 W CA0001545 W CA 0001545W WO 0146405 A2 WO0146405 A2 WO 0146405A2
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Definitions
- the invention relates to modulators of PAX-2 gene for controlled therapeutical apoptosis of cells or therapeutical survival of cells,
- PAX2 is a transcription factor critically required during the development of the nervous and excretory systems, including the mid/hindbrain, spinal cord, eye, ear and urogenital tract (Dressier, G.R. et al. (1990) Developmen t, 109, 787-795; Eccles, .R. et al. (1992) Cell Growth & D ⁇ ff. , 3, 279-289; Dahl, E. et al. (1997) BioEssays, 19, 755-764).
- PAX2 encodes a conserved 128 a ino acid paired box DNA binding domain in the N- terminal portion of the molecule (Dahl, E. et al . (1997) BioEssays, 19, 755-764). While PAX2 is a transcriptional regulator, there are no proven target genes regulated by PAX2, and little is yet known of the exact role of PAX2 during development of the nervous or excretory systems . Eight of the nine Pax genes cause phenotypic abnormalities when mutated in humans or mice, and in four of these (including abnormalities caused by PAX2 mutations) developmental abnormalities are observed in the heterozygotes, revealing haploinsufficiency (Dahl, E. et al.
- the ureteric bud During embryonic life the ureteric bud emerges from the nephric duct, growing outward into and arborizing within the undifferentiated mesenchyme. Signals from each branch of the ureteric bud induce adjacent mesenchymal cells to transform into proximal tubules and glomeruli of individual nephrons which ultrafilter fluid from blood. Each nephron fuses with its parent branch of the ureteric bud, providing an outlet to the bladder. Thus the size and functional capacity of each kidney is ultimately determined by the complexity of ureteric bud branching and the number of individual nephrons which have been induced by the time the pool of metanephric stem cells has been consumed and nephrogenesis is terminated.
- PAX2 is expressed throughout the branching ureteric bud, Wolffian and Mullerian ducts. PAX2 is subsequently expressed in each focus of induced nephrogenic mesenchyme and its derivatives, the early epithelial structures of the emerging nephron (Dressier, G.R. et al. (1990) Development, 109, 787-795; Eccles, M.R. et al. (1992) Cell Growth & Diff. , 3, 279-289). The importance of PAX2 to nephrogenesis is evident in mice with targeted disruption or spontaneous mutations of the Pax2 gene (Dahl, E. et al.
- mice lack kidneys, ureters, vas deferens, epidydimis, seminal vesicles, uterus, oviducts and vagina and have developmental defects of the eyes, ears and CNS; these defects are lethal in the perinatal period (Eccles, M.R. (1998) Pedia tr . Nephrol . , 12, 712-720).
- INeu carrying a Pax2 mutation (Koseki, C. et al. (1992) J. Cell Biol . , 119, 1327-1333) were analyzed. This mutation is identical to the G619 insertion mutation in some humans.
- the fetal kidney size of heterozygous mutants was reduced to 60% of that of wild-type littermates, closely resembling renal hypoplasia in humans.
- Heterozygous INeu mice showed reduced branching and increased apoptotic cell death in fetal kidney collecting ducts, but the increased apoptosis was not associated with random stochastic inactivation of Pax2 expression in the mutant kidneys, and Pax2 was biallelically expressed during kidney development.
- One aim of the present invention is to provide modulators of PAX-2 gene for controlled therapeutical apoptosis of cells or therapeutical survival of cells.
- a method of treating cancer and cystic kidney disease in a patient comprising the step of administering to said patient a therapeutically effective amount of an agent which selectively prevents the function of PAX-2 .
- the agent prevents the expression of PAX-2 , such as an antisense of PAX-2 .
- the agent may inhibit the activity of PAX-2 , such as a ribozyme.
- Cancer treated is selected from the group consisting of prostate, ovarian, bladder and kidney cancers .
- a method of rescuing cells from apotosis in a patient comprising the step of administering to said patient a therapeutically effective amount of an agent which selectively stimulate the function of PAX-2 .
- the agent may activate the expression of PAX-2 .
- a method to enhance resistance of normal tissues to apoptotic cell death induced by chemotherapy or radiation therapy in a patient comprising the step of administering to said patient a therapeutically effective amount of an agent which selectively stimulate the function of PAX-2 .
- the agent may activate the expression of PAX-2 .
- a genetic construct comprising a nucleic acid encoding a molecule capable of preventing or stimulating the function of PAX-2 .
- a method of diagnosing prostate, ovarian, bladder and/or kidney cancer, renal hypoplasia, renal failure, renal-coloboma and cystic kidney diseases in a patient comprising the steps of a) obtaining a sample containing nucleic acid or protein from prostate, ovary, bladder and/or kidney cells or tissue of said patient; and b) determining whether the sample contains a PAX-2 mutation or a level of PAX-2 nucleic acid or protein associated with inappropriate postnatal PAX-2 overexpression, thereby being indicative of neoplast c phenotype or disease phenotype associated with abnormal PAX-2 function.
- Fig. 1 illustrates the pedigree of the Brazilian family with renal-coloboma syndrome
- Fig. 2 illustrates the detection of PAX2 mutations in the Brazilian family, and in a sporadic Japanese patient
- Fig. 3 illustrates a graph of kidney size in 10 patients with PAX2 mutations
- Fig. 4 illustrates the morphology of E15 INeu mutant and wild type kidneys
- Fig. 5 illustrates the analysis of nephrogenesis in E15 INeu mutant and wild type kidneys
- Fig. 6. illustrates the analysis of apoptosis
- Fig. 7 illustrates graphs showing the number of nick-end-labeled cells per kidney area in fetal and postnatal kidneys from Pax21Neu/+ mutant and wildtype mice.
- Fig. 8 illustrates biallelic expression of Pax2 in fetal kidney epithelial cells
- PAX2 mutations cause renal-coloboma syndrome (RCS) , a rare multi-system developmental abnormality involving optic nerve colobomas and renal abnormalities. End-stage renal failure is common in RCS, but the mechanism of how PAX2 mutations lead to renal failure is unknown.
- RCS renal-coloboma syndrome
- PAX2 is a member of a family of developmental genes containing a highly conserved "paired box" DNA-binding domain, and encodes a transcription factor expressed primarily during fetal development in the CNS, eye, ear and urogenital tract. Presently, the role of PAX2 during kidney development is poorly understood.
- kidney anomalies were analyzed in patients with RCS, including a large Brazilian kindred in whom a new PAX2 mutation was identified.
- renal hypoplasia was the most common congenital renal abnormality.
- Genomic DNA from each individual was extracted from peripheral blood (collected with informed consent) using a DNA extraction kit (Promega) . Fragments spanning exons 1-12 of PAX2 were amplified from genomic DNA using PCR primers in the introns flanking the exons.
- PCR products were labeled by incorporation of 32p_ c ⁇ c ⁇ p in reactions containing 100 ng DNA, 62.5 ⁇ M dNTPs, 1 ⁇ Ci [ ⁇ - 32 P] dCTP (3000 Ci/mmol), 20 pmol of each primer, reaction buffer (50mM KCl, lOmM Tris pH 8.3, 1.0-3.0 mM MgCl2) and Pwo (Boehringer) or AmpliTaq Gold (Perkin Elmer) DNA polymerase, and electrophoresed in non-denaturing 6% or 12% polyacrylamide gels as described previously (Sanyanusin, P. et al. (1995) Na ture Genet . , 9, 358- 364) to reveal single-strand conformational variants. Subcloning and DNA sequencing
- PCR products were directly sequenced using ⁇ 33p_ ATP radiolabeled kinased primers and a cycle sequencing kit (GIBCO-BRL) .
- GEBCO-BRL cycle sequencing kit
- mutant and normal alleles of PAX2 were amplified using Pwo DNA polymerase (Boehringer Manheim) , and subcloned into EcoRV digested pBluescript II (Stratagene) . Positive clones were identified by blue/white selection, and plasmid DNA was isolated using Wizard mini-preps kits (Promega). Sequencing reactions were carried out on plasmid DNA as described previously (Sanyanusin, P. et al . (1995) Na ture Genet . , 9, 358-364) .
- Wild type and INeu mouse colonies were bred and maintained at Neuherberg, Germany and at University of Otago, New Zealand.
- Timed matings of male mice heterozygous for the Pax2 lneu mutant allele were carried out with female C57BL/6 mice, and the embryos were genotyped. Genotyping was performed as described (Koseki, C. et al. (1992) J. Cell Biol . , 119, 1327- 1333) , or by PCR amplification of genomic DNA extracted from tail slices by using primers 2F and 2R (see Fig. 8) .
- PCR amplification reactions were as previously described (Sanyanusin, P. et al . (1995) Na ture Genet . , 9, 358-364). The presence of a 167 bp fragment in addition to a 166 bp fragment when the PCR products were electrophoresed on a 6% polyacrylamide gel indicated the presence of the mutant Pax2 allele. Fetal kidneys (E15-E18) were dissected from heterozygous Pax2 1Neu /+ mutant or wild-type mice.
- the expression of the mutant Pax2 1N u allele was demonstrated in fetal kidney RNA from Pax2 Weu /+ mutant mice by RT-PCR amplification of total kidney RNA using primers IF and 3R, spanning exons 1-3 of Pax2. Nested PCR amplification of the RT-PCR product from exons 1-3 was carried out using primers 2F and 2R. PCR products were electrophoresed on 6% polycrylamide gels. Primer sequences were based on the murine Pax2 sequence (Dressier, G.R. et al. (1990) Development, 109, 787- 795) and were as follows;
- Heterozygous PAX2 mutant E15 embryos and their wild type littermates were incubated in freshly prepared 4% paraformaldehyde in PBS for 16-18 hours and stored at 4° in 70% ethanol prior to embedding in paraffin.
- Serial 5 ⁇ m sagital sections of both kidneys from each mutant and wild type embryo were prepared on Superfrost/Plus slides (Fisher) . Sections were stained with hematoxylin-eosin and analyzed by bright field microscopy: the number of glomeruli and early epithelial structures (renal condensates, comma- and S- shaped bodies) in the nephrogenic layer was counted in serial cross-sections from both kidneys of INeu and wild type embryos.
- PAX2 antibody Polyclonal anti-mouse PAX2 antibody was purchased from Zymed, California. Mouse sections were deparaffinized and incubated with primary antibody (10 ug/ml) for 30 min. at room temperature in PBS supplemented with 20mM glycine and 1% BSA. PAX2 antibody was detected with Vecstatin ABC Rabbit IgG kit (Vector Laboratories) as described by the manufacturer, followed by incubation with DAB substrate (Sigma) . Sections were washed in PBS, counterstained with hematoxylin, dehydrated and mounted with Permount (Fisher) . Wild type and mutant tissues were stained simultaneously; each analysis was repeated 3 times. Detection of apoptosis (TUNEL staining)
- the TUNEL assay was performed on paraffin- embedded tissue sections as described (Gavrielli, Y. et al. (1992) J. Cell Biol . , 119, 493-501) with modifications.
- the paraffin sections were dewaxed, rehydrated, partially digested with proteinase K (15 ⁇ g/ml; Boehringer Mannheim) in Tris-Cl (10 mM, pH 7.6) at room temperature (RT) for 15 minutes.
- TdT terminal deoxynucleotidyl transferase
- the sections were then washed 3 times in PBS, incubated with extravidin peroxidase (1:250 dilution in PBS, SIGMA) for 60 minutes at RT, rinsed 3 times in PBS, stained with diaminobenzidine (Boehringer Mannheim) for 5 minutes at RT, and counterstained with 1% methyl green.
- renal features were compared in a total of 29 patients with PAX2 mutations, including 10 new patients from 2 families. The 10 new patients are described in the following.
- a three generation Brazilian kindred (pedigree shown in Fig. 1) was evaluated for optic nerve and kidneys defects consistent with diagnosis of renal-coloboma syndrome.
- Individuals in three generations of the pedigree (I, II and III) are shown in Fig. 1. Filled symbols represent affected individuals. Open symbols represent unaffected individuals.
- PCR-SSCP revealed a variant pattern in exon 7 that was present in 5 of the affected members of the family, but was not present in 18 unaffected members (Fig. 2A) .
- PCR-SSCP analysis of PAX2 exon 7 showed the SSCP variant band in the affected individuals, but not in the unaffected individual (arrow; Fig. 2A) .
- the undigested band in the uncut unaffected, and affected lanes and in the Cac ⁇ l digested affected lanes is 234 bp, which is digested to products of 137 and 97 bp by Ca c ⁇ l .
- Unaffected family members contained only sequences that digested with Cac ⁇ l . Additionally, a Japanese patient was examined who had sporadic occurrence of optic nerve colobomas, renal anomalies and bilateral cryptorchidism (see Table 1, patient X2003 for phenotypic details) . Table 1
- each exon of the PAX2 gene was amplified by PCR to identify a PAX2 mutation.
- a variant SSCP pattern was observed for exon 2, and sequencing of this PCR product revealed a guanosine nucleotide insertion at position 619 (Fig. 2D).
- the mutant and normal PAX2 sequence in exon 2 of the Japanese patient are shown in Fig. 2D.
- the mutation (arrowhead) is a G insertion resulting in an additional G (underlined) in a homonucleotide tract of 7 G's.
- IV-3 1 46 + - + - +
- IV-6 1 40 + - + - +
- IV-7 1 35 + - + - -
- renal hypoplasia small kidneys
- Renal failure, histological abnormalities and proteinuria are features commonly seen in association with other disease processes, and are not specific for renal-coloboma syndrome. These abnormalities were probably secondarily acquired since birth.
- renal hypoplasia although seen in association with a number of congenital conditions, is caused by failure of embryonic growth of the kidney, and would therefore be consistent with an abnormality arising primarily from the PAX2 mutation.
- kidneys were found in 17/29 patients (59%), and in 10 of these patients the sizes of the kidneys were measured by ultrasound and graphed as a percentage of the mean normal size of kidneys for individuals at that age (Fig. 3) .
- kidney lengths are shown as a percentage of the mean normal kidney length for a person of the same age and sex; black bar, left kidney; grey bar, right kidney.
- the patients in order were 1, 3, TRN, 579, III-8, IV-2, IV-3, IV-6, IV-7, V-2 in Table 2.
- Renal hypoplasia was observed even in the youngest patients in this study, consistent with it being a congenital abnormality associated with renal- coloboma syndrome.
- Homozygous Pax2 mice have renal agenesis, bilateral optic nerve colobomas, abnormalities of ear development and missing midbrain-hindbrain regions. These animals die within 24 hours of birth.
- heterozygous INeu mice like their human counterparts, are viable and have no gross midbrain-hindbrain phenotype. Their main characteristics are abnormalities of optic nerve development and hypoplastic kidneys (u) .
- Fetal mice carrying a heterozygous Pax2 mutation were
- the nephrogenic zones of fetal (E15-E16) INeu kidneys were thin and contained fewer nephrons in comparison to those of normal littermates.
- the number of early epithelial structures derived from induced metanephric mesenchyme (vesicles, comma- and S-shaped bodies) was reduced to 30-40% of normal (Fig. 5b).
- the number of mature glomeruli was even more sharply reduced (20% of the wild type) .
- early tubular structures and glomeruli in Pax2 mutant kidneys appeared to be of normal size and morphology (Fig. 4) .
- Renal hypoplasia in Pax2 mice is associated with enhanced apoptosis of the ureteric epithelium
- INeu heterozygous Pax2 mutant mice was associated with increased apoptosis.
- An increase in TUNEL-positive staining was observed in the kidneys of embryonic day
- INeu mutant Pax2 /+ was equivalent to that of wildtype mice from embryonic day E15 through postnatal day 6 (Fig. 6C, 6D) .
- PCNA staining was unchanged in E15 INeu embryos compared with their wild type littermates.
- INeu cells m Pax2 /+ mutant (white bars) and wildtype mice (black bars) for (Fig. 7A) over the whole kidney, (Fig. 7B) specifically in collecting ducts, or (Fig. 7C) specifically in renal cortex.
- INeu renal cortex in E15-E16 Pax2 /+ mutant fetal kidneys (Fig. 7C) , which mostly corresponded to ureteric buds. Increased apoptosis was not observed in glomeruli, or proximal or distal tubules at either E15-E16 or later in development. The rate of apoptosis in fetal INeu kidneys was not determined prior to E15 because successfully mated females did not have vaginal plugs, and excessive numbers of randomly pregnant female mice would have been required to obtain the Figures for apoptotic rates.
- Pax2 mutant kidneys contain transcripts of both PAX2 alleles and express a reduced level of PAX2 protein
- PAX gene PAX5
- PAX2 PAX2 expression in developing kidney is also monoallelic.
- heterozygous mutant kidney would be comprised of a mixture of normal and null mutant cells.
- Apoptotic cell death might occur in those cells expressing the mutant allele, and viable collecting duct cells could consist of those randomly expressing normal amounts of PAX2 protein from the wild type allele. Alternatively, if PAX2 is expressed from both
- nephrons of heterozygous Pax2 kidneys should express PAX2 protein with perhaps a slight reduction in intensity.
- Pax2 allele and the locations of the primers (IF, 2F, 2R, 3R) used for PCR and RT-PCR. Reverse transcribed RNA was amplified using primers IF and 3R (DNA amplification was precluded by introns 1 and 2). Primers 2F and 2R were used to amplify genomic DNA.
- Immunohistochemical staining for Pax2 protein was observed in all identifiable nuclei of ureteric bud and collecting duct cells of heterozygous INeu fetal kidneys (Fig. 8C) , but the intensity of Pax2 staining was uniformly less than that in fetal kidneys of wild-type littermates analyzed simultaneously.
- Immunohistochemistry of E15 heterozygous mutant INeu mouse fetal kidney, and Fig. 8C D E15 wild type mouse fetal kidney using anti-Pax2 primary antibody. Nuclei stained brown in the sections correspond to cells of the collecting duct and ureteric bud expressing the Pax2 protein.
- RT-PCR-SSCP was used to discriminate between the two PAX2 alleles (Fig. 8f, lanes 1-3) .
- Biallelic transcription of PAX2 in fetal kidney (Fig. 8F, lane 7) was not unexpected since monoallelic transcription of PAX2 in one cell (if present) would be statistically matched by monoallelic transcription of the opposite allele in another cell.
- Lanes 4-6 heterozygous Wilms tumors. Lane 7, heterozygous human fetal kidney (12 weeks gestation) . Lanes 8, 9, renal carcinoma cell line (A704) before cell cloning (lane 8), and after cell cloning (lane 9) .
- Wilms tumors are predominantly comprised of clonal blastemal and epithelial tissues, derived originally from embryonic kidney. Biallelic PAX2 transcription was observed in mRNA isolated from a series of 3 Wilms tumor tissues (Fig. 8f, lanes 4-6) . In addition, biallelic (albeit slightly unbalanced) PAX2 transcription was observed in clonally expanded renal carcinoma cells (Fig. 8F, lanes 8, 9) . The small imbalance of PAX2 allelic expression observed in the cells may be due to alterations in the number of copies of each PAX2 allele in the cells, and was an indication that the RT-PCR-SSCP technique was sensitive enough for this purpose. Discussion
- renal phenotype associated with renal-coloboma syndrome we have further characterized the renal phenotype associated with renal-coloboma syndrome and have identified elevated levels of apoptosis in fetal kidneys of individuals carrying a PAX-2 mutation. End- stage renal failure is commonly associated with this syndrome, and one goal of this research was to determine how PAX2 mutations could cause renal failure. This was addressed firstly by characterizing the renal phenotype in renal-coloboma syndrome patients. Renal hypoplasia was the most common congenital abnormality in a series of 29 patients, including 10 new patients reported here.
- PAX2 protein lacking the transactivation domain is unlikely to be fully functional, as studies have shown that this region in the closely related PAX5 protein is required for transcriptional activation of target genes .
- a Japanese boy we identified a G619 insertion mutation of PAX2 exon 2.
- the G619 insertion mutation has previously been documented in other patients (Favor, J. et al . (1996) Proc . Na tl . Acad. Sci . USA, 93, 1380-1387; Gavrielli, Y. et al . (1992) J. Cell Biol . , 119, 493-501).
- mice harbor a mutation that is identical to a mutation in approximately one third of families with renal-coloboma syndrome. These mice transmit a single base pair insertion amid a string of seven guanidine residues (positions 613-619) in the second exon of Pax2 , which produces a frameshift and presumably a null allele (Koseki, C. et al. (1992) J.
- Pax2 mice have optic nerve and kidney abnormalities, similar to humans with renal-coloboma syndrome. Although the homozygous mutants die within 24 hours of birth and are phenotypically similar to "knockout" mice generated by targeted homologous recombination, heterozygotes survive and reach adulthood. The heterozygous mutant mice therefore afford a useful animal model in which to analyze fetal kidney defects associated with Pax2 mutations .
- Pax2 is first detected along the nephrogenic cord at the sites from which ureteric buds will emerge (Dressier, G.R. et al . (1990) Development, 109, 787-795) .
- Intense Pax2 expression persists in cells of the ureteric buds as they invade the metanephric blastema to each side and begin to undergo dichotomous branching (Dressier, G.R. et al. (1990) Development, 109, 787-795, Eccles, M.R. et al. (1992) Cell Growth & Diff. , 3, 279-289).
- INeu heterozygous Pax2 mutant mice Cell division, as assessed by PCNA immunohistochemistry and BrdU uptake, was normal. However, apoptosis in the medullary segments of the collecting duct (as assessed by TUNEL assay) was strikingly increased at the time when Pax2 is maximally expressed in kidney development, and in cells which normally express Pax2 (Dressier, G.R. et al. (1990) Developmen t, 109, 787-795). Indeed, the amount and localization of the apoptosis correlated temporally and spatially with the known expression patterns of PAX2 (Dressier, G.R. et al. (1990) Development, 109, 787-795, Eccles, M.R.
- Pax8 was able to compensate for heterozygous Pax2 mutations in the mesenchymally-derived structures of kidneys from Pax2 mutant mice, and that this ultimately rescued these cells from apoptosis.
- Our data do not determine whether PAX2 directly regulates genes in apoptotic pathways during kidney development.
- PAX2, PAX5 and PAX8 have all been reported to inhibit p53 transcription, which can promote apoptosis.
- transgenic mice overexpressing wild type p53 have altered differentiation of the ureteric bud, and have small kidneys.
- Pax5 knockout mice the B cells fail to differentiate and undergo apoptotic cell death at an early stage.
- large-scale apoptosis of photoreceptor precursors occurs in the eye discs of
- Drosophila with the eyeless mutation ( ey ) (homologous to the mammalian Pax ⁇ gene) .
- ey eyeless mutation
- these authors also found no difference in BrdU uptake studies in wild type and mutant eye discs. As in our studies, this suggests that the mutation affects cell survival rather than proliferation.
- Pax3 mutant mice were shown to have enhanced apoptosis in the somatic mesoderm during embryonic development, and treatment of rhabdomyosarcoma cells lines with PAX3 antisense oligonucleotides has been shown to result in apoptosis.
- our studies were carried out on heterozygous mutant animals.
- PAX2 haploinsufficiency on the renal phenotype are similar to the developmental effects of haploinsufficiency for other PAX genes.
- PAX3 mutations cause an autosomal dominant defect in epidermal pigmentation
- PAX6 mutations cause autosomal dominant aniridia
- PAX ⁇ mutations cause autosomal dominant hypothyroidism. In each case, inactivation of one allele is sufficient to interfere with normal organ development.
- haploinsufficiency others have hypothesized that monoallelic Pax gene expression may be related to the semi-dominant effects of Pax gene mutations.
- Pax2 mutant kidneys immunostaining of Pax2 was detected in virtually every cell in the ureteric buds.
- these observations suggest that Pax2 is biallelically transcribed in wild- type fetal kidney, and that ureter and collecting duct abnormalities produced by Pax2 haploinsufficiency probably result from sensitivity to reduced gene dosage in each cell.
- Pax2 is monallelically expressed at embryonic stages earlier than E15, or in stem cells which were not amenable to study by immunohistochemistry in E15 kidney.
- the mu ⁇ ne Pax2 mutation adds much to our understanding of the human disease.
- renal hypoplasia is the most common congenital renal anomaly in humans and mice with PAX2 mutations, and that in mice this phenotype is associated with significantly enhanced apoptosis.
- the Pax2 gene was found to be biallelically expressed in mutant mice, therefore it seems unlikely that the enhanced apoptosis was due to random stochastic inactivation of Pax2 expression in cells mono-allelically expressing Pax2.
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CA002394589A CA2394589A1 (en) | 1999-12-22 | 2000-12-21 | Modulation of pax-2 for controlled apoptosis or survival of cells |
AU21390/01A AU2139001A (en) | 1999-12-22 | 2000-12-21 | Modulation of PAX-2 for controlled apoptosis or survival of cells |
EP00984747A EP1240321A2 (en) | 1999-12-22 | 2000-12-21 | MODULATION OF i PAX /i -2 FOR CONTROLLED APOPTOSIS OR SURVIVAL OF CELLS |
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EP1934331A2 (en) * | 2005-10-14 | 2008-06-25 | MUSC Foundation For Research Development | Targeting pax2 for the induction of defb1-mediated tumor immunity and cancer therapy |
US8080534B2 (en) | 2005-10-14 | 2011-12-20 | Phigenix, Inc | Targeting PAX2 for the treatment of breast cancer |
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2000
- 2000-12-21 WO PCT/CA2000/001545 patent/WO2001046405A2/en not_active Application Discontinuation
- 2000-12-21 AU AU21390/01A patent/AU2139001A/en not_active Abandoned
- 2000-12-21 EP EP00984747A patent/EP1240321A2/en not_active Withdrawn
- 2000-12-21 CA CA002394589A patent/CA2394589A1/en not_active Abandoned
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Non-Patent Citations (8)
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EP1934331A2 (en) * | 2005-10-14 | 2008-06-25 | MUSC Foundation For Research Development | Targeting pax2 for the induction of defb1-mediated tumor immunity and cancer therapy |
EP1934331A4 (en) * | 2005-10-14 | 2009-01-21 | Musc Found For Res Dev | Targeting pax2 for the induction of defb1-mediated tumor immunity and cancer therapy |
US7964577B2 (en) | 2005-10-14 | 2011-06-21 | Donald Carlton D | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
EP2392647A1 (en) * | 2005-10-14 | 2011-12-07 | MUSC Foundation For Research Development | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
EP2392646A1 (en) * | 2005-10-14 | 2011-12-07 | MUSC Foundation For Research Development | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
EP2392645A1 (en) * | 2005-10-14 | 2011-12-07 | MUSC Foundation For Research Development | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
EP2395076A1 (en) * | 2005-10-14 | 2011-12-14 | MUSC Foundation For Research Development | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
US8080534B2 (en) | 2005-10-14 | 2011-12-20 | Phigenix, Inc | Targeting PAX2 for the treatment of breast cancer |
EP2402435A3 (en) * | 2005-10-14 | 2012-03-28 | MUSC Foundation For Research Development | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
US8318692B2 (en) | 2005-10-14 | 2012-11-27 | Donald Carlton D | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
US8394780B2 (en) | 2005-10-14 | 2013-03-12 | Phigenix, Inc. | Targeting PAX2 for the treatment of breast cancer |
US8431546B2 (en) | 2005-10-14 | 2013-04-30 | Phigenix, Inc. | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
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US8653021B2 (en) | 2005-10-14 | 2014-02-18 | Phigenix, Inc. | Targeting PAX2 for the treatment of breast cancer |
US8735365B2 (en) | 2005-10-14 | 2014-05-27 | Phigenix, Inc. | Targeting PAX2 for the induction of DEFB1-mediated tumor immunity and cancer therapy |
US11033628B1 (en) | 2005-10-14 | 2021-06-15 | Phigenix, Inc. | Targeting PAX2 for the treatment of breast cancer |
Also Published As
Publication number | Publication date |
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CA2394589A1 (en) | 2001-06-28 |
EP1240321A2 (en) | 2002-09-18 |
WO2001046405A3 (en) | 2002-03-07 |
AU2139001A (en) | 2001-07-03 |
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