| Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimens -> Monitor Keywords |
|
|
  Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimensUSPTO Application #: 20060088871Title: Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimens Abstract: Provided are materials, methods, platforms, and kits for diagnosing, prognosing, and/or determining the biological aggressiveness of a tumor (malignant and non-malignant) based in part on the temporal profile of genomic deletion expansion of the tumor. Also provided are methods of determining marker panels for different diseases and/or tissues and markers identified by these methods. (end of abstract) Agent: Drinker Biddle & Reath (dc) - Washington, DC, US Inventors: Sydney David Finkelstein, Patricia Swalsky USPTO Applicaton #: 20060088871 - Class: 435006000 (USPTO) Related Patent Categories: Chemistry: Molecular Biology And Microbiology, Measuring Or Testing Process Involving Enzymes Or Micro-organisms; Composition Or Test Strip Therefore; Processes Of Forming Such Composition Or Test Strip, Involving Nucleic Acid The Patent Description & Claims data below is from USPTO Patent Application 20060088871. Brief Patent Description - Full Patent Description - Patent Application Claims
REFERENCE TO RELATED APPLICATIONS [0001] This application claims benefit of U.S. Provisional Application Nos. 60/620,926 filed Oct. 29, 2004; 60/631,240 filed Nov. 29, 2004; 60/644,568 filed Jan. 19, 2005; 60/679,969 filed May 12, 2005; and 60/679,968 filed May 12, 2005, all of which are herein incorporated by reference in their entirety for all purposes. FIELD OF THE INVENTION [0002] The application relates to methods, materials, kits and devices for achieving a diagnosis, prognosis, and/or determination of the biological aggressiveness of a tumor based on the temporal profile (time course) of genomic deletion expansion. The materials, methods, kits and devices disclosed herein can also be used for pre-cancerous growths and diagnosis and prognosis of non-malignant tissues and cells. BACKGROUND [0003] Cancer is characterized by the stepwise accumulation of genetic damage. Such genetic damage includes mutations and aberrant expression of oncogenes and tumor suppressor genes that are causally related to phenotypic expression of malignant characteristics such as rapid uncontrolled growth, local invasion, and metastasis (Weir et al., 2004 Cancer Cell. 6(5): 433-8; and Osborne et al., 2004 Oncologist 9(4): 361-77). As individual mutations are acquired by affected cells, clonal expansion of biologically more aggressive tumor cells ensues (Braakhuis et al., 2005 Semin. Cancer Biol. 15(2): 113-20; Breivik et al., 2005 Semin Cancer Biol. 15(1): 51-60). Thus, cancer cells with increasing mutational change come to replace precursor cells with fewer mutations, which is the driving force for clonal expansion. The process is dynamic one occurring across the entire tumor, with the creation of distinct tumor cell clones in topographic regions that undergo neoplastic progression at different rates. To a large extent, the process is irreversible, with the most biologically aggressive cellular elements of a particular cancer overgrowing the relatively less aggressive cellular components. The majority of human cancers are eventually dominated by clonal expanding malignant cells possessing specific, detectable mutational alterations directly responsible for tumor growth and biological behavior. [0004] One of the most common cancer related genetic alterations is genomic deletion (Bishop A J, et al., 2003 Exp. Mol. Pathol. 74(2): 94-105; Popescu, 2003 Cancer Lett. 192(1): 1-17). This form of damage is commonly associated with tumor suppressor gene inactivation which follows a two-step process (Baker et al., 2003 Genes Chromosomes Cancer. 38(4): 329 and Knudson, 2001 Lancet Oncol. 2(10): 642-5). First, one normal copy of the tumor suppressor is inactivated, typically by means of point mutational damage. The second step is genomic deletion of the remaining normal copy, often with adjacent non-coding DNA being included in the deletional region (Baker et al., 2003 and Knudson, 2001). [0005] A primary objective in the management of cancer is predicting the intrinsic biological aggressiveness of an individual neoplasm, so that it may be treated in an appropriate manner. Cancers that are intrinsically indolent should be treated in a conservative manner to avoid the unnecessary morbidity associated with the side effects of therapy. In contrast, cancers that can be predicted to behave in a more aggressive fashion justifiably demand a more aggressive therapy to avoid under the under-treatment of the patient. Thus, it is essential to effectively prognosticate the biological aggressiveness of each cancer on an individual patient basis. [0006] The prediction of cancer aggressiveness, according to conventional teaching, involves searching for specific microscopic features such as high mitotic rate, cellular anaplasia, nuclear pleomorphism, etc. Conventional teaching does not require that the prediction of cancer aggressive require criteria other than those encompassed by microscopic observation. At the extremes (i.e. near normal cellular appearance with no evidence of high mitotic rate, cellular anaplasia, nuclear pleomorphism versus extensive high mitotic rate, cellular anaplasia, nuclear pleomorphism), these criteria are generally reliable. However, the majority of clinical cases fall in between the polar ends of this spectrum with some but not all microscopic features of biological aggressiveness present. Under these circumstances, the prediction of tumor biological aggressiveness cannot be carried out microscopically with certainty. The use of additional technique to derive correlative mutational information can serve as a powerful means to add discriminating information leading to greater certainty. [0007] Unfortunately, in the majority of cases, such clear-cut discrimination is not possible, and there is significant error upon the part of the diagnosing physician in predicting cancer aggressiveness based on microscopic features alone. In fact, patients with the identical histologic type of cancer, including cellular microscopic features, have proven over time not to behave in an equivalent manner. One patient may die after a short interval from rapid tumor progression, while another patient shows a long disease-free survival. The mortality for equivalent stages of human cancer is ex pressed as a probability, although it is recognized that there will be variation across the spectrum of patient outcomes. This renders individualized cancer patient care less effective. [0008] Progress over the last decade has provided abundant information regarding the molecular changes in cancer (Ross et al., 2002 Hematology 7(4): 239-52 and Tomlinson et al., 2002 Genes Chromosomes Cancer 34(4): 349-53). Cancer aggressiveness can be more effectively but not completely predicted by parameters such as the cumulative amount of mutational damage (Ross et al., 2002 and Tomlinson et al., 2002) or the level of altered gene expression (Hughes, 2004 J. Surg. Oncol. 85(1): 28-35; Risques et al., 2003 Cancer Res. 63(21): 7206-14; and Kwon et al., 2004 Dis. Colon Rectum. 47(2): 141-52). In some cases, specific gene involvement has proven capable of characterizing differential cancer biological aggressiveness (Risques et al., 2003 Histol. Histopathol. 18(4): 1289-99). While great strides have been taken, there is much to be learned concerning the control mechanisms for tumor behavior. [0009] The realities of clinical practice demand that cancer diagnosis be accomplished on minute samples of tumor or fluid aspirate. Moreover, all cell or tissue specimens removed from a patient's body must be thoroughly evaluated by microscopic pathology evaluation to ensure that adequate amounts of lesion material have been obtained, and in order to derive conventional histopathologic diagnosis upon which to base a standard diagnosis. Measures that interfere with this basic approach and fail to complement routine pathology practice run the risk of significant error, with serious medical and legal consequences. Rather, molecular analysis must proceed in series after conventional microscopic examination on the very same samples in order to effectively integrate mutational analysis into pathology practice. As a result, the molecular analysis must also be effective on small-sized, fixative-treated and/or stained cells as well as on cells or DNA material containing in biological fluid samples. The most valuable diagnostic methods are those that meet these operational criteria. [0010] Advances such as the sequencing of the human genome have made it possible to design molecular testing strategies to assist in the detection of cancer, precancer and related conditions (Kreiner et al., 2005 Am. J. Health Syst. Pharm. 62(3): 296-305). Among the information generated by the Human Genome Project is the position of polymorphic markers in the form of minisatellites, microsatellites and single nucleotide polymorphisms (SNPs) that can be used to track the presence of both copies of markers and genes that compose the human genome (Beroud et al., 2005 Hum. Mutat. 26(3): 184-91). While the existence of these markers are known, their value and use in the context of searching for and characterizing human mutation is still unknown. It is the desire of many to use these markers to evaluate pathologic specimens. However, their utilization remains unclear. [0011] When endeavoring to use markers to explore cancer formation and progression, the issue arises as to how to formulate a panel of markers that will serve this function best and provide the most information (Berger et al., 2003 Diagn. Mol. Pathol. 12(4):187-92). Conventional teaching on the subject is minimal and repeats the basic tenet that one should choose a panel of markers that are as close as possible to those genes, which research has already shown to be involved in the molecular pathogenesis of a particular form of cancer (e.g., Yoshino et al., 2005 Respir. Med. 99(3): 308-12). Current convention instructs that the best markers are dinucleotide microsatellites and SNPs, because there are many more of these markers scattered across the human genome and thus one can get closer or even within specific genes of interest. Beyond this recommendation to favor the use of dinucleotide microsatellites and SNPs, there are no further recommendations and one is left to determine how best to then utilize these markers and perform and broad marker panel analysis SUMMARY [0012] Herein are provided methods, compositions, kits, and devices for assessing dynamic genomic deletional expansion that can be determined using small amounts of tissue, fixative-treated tissue, cellular specimens, and/or fluid specimens that directly and effectively prognosticate cancer biological aggressiveness and combinations thereof. This observation has heretofore not been described, yet its application in everyday clinical practice can be highly effective and easy to perform. The result is a new modality with which to characterize a tumor in a patient on an individual basis. Although the patient is preferably human, such diagnosis can also be performed on other animals for veterinary diagnosis. It may be expected to have profound consequences for future diagnosis of cancer and pre-cancer states and, by consequence, therapeutic management. [0013] The materials, methods, and kits provided herein can be utilized to diagnose and characterize the biological aggressiveness of a tissue or cells, such as a neoplastic tissue, thereby improving patient diagnosis and treatment of the patient with the tissue abnormality. [0014] Provided herein is a method of determining tumor aggressiveness in a patient comprising the steps of: (a) amplifying DNA from a microdissection section of the biological sample; (b) analyzing two or more genes for the presence of a nucleotide deletion wherein a deletion is the acquisition of genetic mutation; (c) analyzing each gene with an array of markers to determine the extent of nucleotide deletion; (d) determining the order of acquisition of each nucleotide deletion in the patient; and (e) collating the data from steps (a) to (d) to determine tumor aggressiveness of the patient. In one aspect, the reagents used to amplify DNA from the biological sample of the patient, when admixed for amplification, comprises about 1 to about 15 mM magnesium chloride (other valency ions such as manganese can be substituted in the form of MnCl.sub.2), more preferably from about 6.0 to about 10.0 mM (and every 0.1 mM value in between), and most preferably about 8.0 mM magnesium chloride. The solution preferably also comprises when admixed about 5 to about 20 g/100 mL sucrose (and any 0.1 g/100 mL value there between), more preferably the solution contains about 12.0 g/100 mL sucrose. [0015] The biological sample can be pretreated with a proteinase from about 2 hours to overnight. Preferably, the proteinase treatment occurs at about 37.degree. C. and halted by boiling the sample for about 5 minutes. Suitable proteinases include but are not limited to proteinase K, pronase, subtilisin, thermolysin, papain, or a combination of proteinases. Proteinases are present in an amount of about 0.5% to about 2.0% final volume of lysis buffer (and every 0.1% value there between), and preferably about 1.0%. The lysis buffer further includes a nonionic detergent. Nonionic detergents can include but are not limited to nonidet P40, Tween (e.g., Tween 20), Triton X, or Nikkol. The nonionic detergents are present in the amount of about 0.5% to about 2.0% (and any 0.1% value in between). [0016] The biological sample may be a cell-free fluid sample, a blood sample, a cytology sample, a urine sample, a tissue swab, or resected tissue. The cell-free fluid sample contains non-nuclear DNA. The resected tissue may be frozen, fresh, stained, or fixative-treated, or a combination thereof. More than one type of biological sample can be utilized for each patient. For example, a fluid sample and a resected tissue sample can be analyzed and the data from each combined. Additionally, biological samples can be derived from more than one point of origin in a patient. For example, a biological sample can be obtained from a putative primary tumor as well as a biological sample from a putative second unrelated tumor or from a putative metastatic lesion, or combination thereof. Thus, for example, in Wilm's tumor, wherein nodules may each be a primary, samples of each nodule may be analyzed alone or the data combined. [0017] Step (c) of the method may be repeated to obtain replicate data. The method may further comprise the step of identifying a treatment plan to treat the tumor of said patient which best treats the tumor based on the determination of tumor aggression. The biological sample may be microdissected tissue, and the steps of (b) and/or (c) may be performed on DNA obtained from two or more microdissected sections of the tissue sample. Step (c) may be repeated to obtain replicate data. [0018] The patient may also have a fluid sample, and the fluid sample may undergo analysis comprising: (a) amplifying DNA from the fluid sample of the patient; (b) analyzing two or more genes for the presence of nucleotide deletion from the amplified DNA; (c) analyzing each nucleotide deletion with an array of markers to determine the extent of nucleotide deletion; (d) determining the order of acquisition of each nucleotide deletion in the patient; and (e) validating the collated data with the data from the fluid sample. Markers can be used to determine whether the mutation was environmentally induced, such as by exposure to trichloroethylene or other chemical, a germ line mutation, or a spontaneous mutation. [0019] Also provided is a kit for determining tumor aggressiveness comprising (i) a device for amplifying DNA; and (ii) sets of cancer specific markers for assessing nucleotide deletion and extent of nucleotide deletion. The markers can include markers for environmentally-induced mutations, spontaneous mutations, and/or germ line mutations. The kit may also provide for reagents for amplifying DNA as discussed above. The device for amplifying and determining genomic deletion may also comprise a data storage component for storing patient information regarding sex, age, weight, medical history, family medical history, prior cancer history and genetic analysis of a prior cancer, and genomic deletion acquisition data. This information may further be stored in a relational database, wherein factors are weighted in order to best determine treatment strategy. [0020] Another aspect provided is a method of creating a panel of molecular markers for detecting a condition in a patient comprising the steps of: (a) determining gene targets for detection of a mutation to include in a molecular marker analysis for a marker panel; (b) delineating genomic regions for each gene target; (c) identifying a 4 to 1500 nucleotide repeat microsatellite and/or a minisatellite in the genomic region that will constitute the marker panel; (d) identifying at least two 4 to 1500 nucleotide microsatellites and/or minisatellites positioned a certain distance from each other and performing genomic deletional expansion; (e) determining the amount of DNA in the biological sample; (f) determining the quality of DNA in the biological sample by quantitative PCR; (g) determining the quality of DNA in the biological sample by means on competitive template PCR (CT-PCR); (h) determining the amplifiability of DNA for each 4 to 1500 repeat microsatellite and/or minisatellite; (i) defining a normal range of allele variation thereby defining allelic imbalance comprising: [0021] (i) defining different normal ranges for each allele for two or more quantities of DNA; and [0022] (ii) defining different normal ranges for each allele with two or more qualities of DNA; and (j) defining minimum thresholds for significant allelic imbalance thereby obtaining an indication of mutation change in a significant percentage of evaluated cells comprising: [0023] (i) defining minimum thresholds for significant allelic imbalance for different amounts of DNA; and [0024] (ii) defining minimum thresholds for significant allelic imbalance for different qualities of DNA; and (k) calculating a percentage of mutated cells based upon ratios of a tested sample using a calculated normal for each 4 to 1500 microsatellite and/or minisatellite, thereby creating a panel of molecular markers for detecting a condition in a patient. Preferably, the microsatellite is 4 nucleotides to 1500 nucleotides, more preferably 4 to 200, and more preferably 4 to 20 nucleotides (any integer value between 4 and 1500). The markers can be used to preferably diagnose a neoplasia, a hyperplasia, or a benign growth as well as determine if the mutation is germ line (inherited), spontaneous, or due to an environmental factor. Continue reading... Full patent description for Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimens Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimens patent application. Patent Applications in related categories: 20080113379 - Method for the detection of cytosine methylations in immobilized dna samples - A method is described for the analysis of cytosine methylation patterns in genomic DNA samples. In the first method step, the genomic DNA is isolated from cells or other accompanying materials and bound essentially irreversibly to a surface. Then the DNA bound to the surface is treated, preferably with a ... 20080113342 - Plant genome sequence and uses thereof - The present invention is in the field of plant biochemistry and genetics. More specifically the invention relates to nucleic acid sequences from plant cells, in particular, genomic DNA sequences from Arabidopsis thaliana plants. The invention encompasses nucleic acid molecules present in non-coding regions as well as nucleic acid molecules that ... 20080113380 - Sensitizer-labeled analyte detection - The invention provides methods for detecting an analyte in a sample including the steps of: (a) exciting a sensitizer label on an analyte; (b) permitting energy from the excited sensitizer label to be transferred to and excite an acceptor molecule, whereby the sensitizer label returns to an unexcited state; (c) ... 20080113373 - Sirna targeting amyloid beta (a4) precursor protein (app) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113370 - Sirna targeting apolipoprotein b (apob) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113371 - Sirna targeting beta secretase (bace) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113369 - Sirna targeting diacylglycerol o-acyltransferase homolog 2 (dgat2) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113374 - Sirna targeting fructose-1,6-bisphosphatase 1 (fbp1) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113372 - Sirna targeting glucagon receptor (gcgr) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113378 - Sirna targeting interleukin-1 receptor-associated kinase 4 (irak4) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113376 - Sirna targeting myeloid differentiation primary response gene (88) (myd88) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113377 - Sirna targeting proto-oncogene met - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... 20080113375 - Sirna targeting superoxide dismutase 1 (sod1) - Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed ... ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. Start now! - Receive info on patent apps like Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimens or other areas of interest. ### Previous Patent Application: Dual labeled fluorescent probes Next Patent Application: Expression system for stem-loop rna molecule having rnai effect Industry Class: Chemistry: molecular biology and microbiology ### FreshPatents.com Support Thank you for viewing the Dynamic genomic deletion expansion and formulation of molecular marker panels for integrated molecular pathology diagnosis and characterization of tissue, cellular fluid, and pure fluid specimens patent info. IP-related news and info Results in 8.49228 seconds Other interesting Feshpatents.com categories: Software: Finance , AI , Databases , Development , Document , Navigation , Error |
||
