Patent Application: US-29344002-A

Abstract:
a method to aid in identifying a familial or sporadic pattern of risk in at least one individual for developing cancer of a mucosal epithelial tissue , the method comprising screening said at least one individual for heterozygosity or homozygosity for a mutation in a gene coding for a poly - ig receptor or a poly - ig - like receptor capable of mediating inhibition of cancer cell growth by an immunoglobulin inhibitor . a method of treating an individual so identified includes enhancing the amount of immunoglobulin inhibitor contacting a mucosal epithelial tissue of said individual , and , especially in individuals homozygous for the defective receptor , may also include prophylactic surgery . other methods include implementation of a risk reduction or prevention program in individuals identified as being at risk .

Description:
recent discoveries disclosed in co - owned , co - pending u . s . patent application ser . nos . 09 / 852 , 547 and 09 / 852 , 958 , and in international patent application nos . pct / us01 / 15171 and pct / us01 / 15183 ( also identified as items 21 and 22 in the references , below ), each of which is hereby incorporated herein by reference , are expected to lead to resolution of the problem of how to identify genes that are key to the 60 to 70 % of breast cancer cases that are today termed “ sporadic .” the isolation of new “ serum factor ( s )” that regulate estrogen responsive breast cancer cell growth in culture is described in the preceding applications . the purification yielded dimeric / polymeric immunoglobulin a ( iga ) and pentameric immunoglobulin m ( igm ) as the active regulators . these immunoglobulins (“ immunoglobulin inhibitors ”) arrested estrogen target tumor cell growth completely at low nanomolar concentrations , and their inhibitory effects were entirely reversible by picomolar concentrations of estrogens . that disclosure revealed a previously unknown function for the secretory immune system . in the above - identified patent applications , a major role for tgfβ in breast growth regulation is also identified : it is a cytokine that controls iga / igm immunocytes . breast cancer growth is best defined as negative paracrine control by secretory immunoglobulins ( immunoglobulin inhibitors ) and positive direct control by estrogens . in conjunction with this work , the longstanding problem of the regulation of estrogen dependent cell growth in culture under serum - free defined medium conditions was solved . these results have great physiological relevance . iga and igm are secreted by b immunocytes located in the lamina propria of estrogen target tissues including breast . they are more than 90 % of the immunoglobulins secreted into breast milk . the positioning of the immunocytes in the tissue adjacent to the epithelial cells and the secretion of the immunoglobulins is hormone regulated . it was found that the secretory immune system products immunoglobulins a and m ( iga and igm ) inhibit the growth of estrogen - sensitive ( early ) breast cancer by suppressing cell replication . this is the first time a connection was established between the secretory immune system and early estrogen receptor positive ( er + ) breast cancer growth . in addition , when breast cancer becomes most malignant ( i . e . er − hormone - insensitive stages ), control by immune system iga and igm is lost . evidence points to the poly - ig receptor or a very similar poly - ig - like fc receptor as the mediator of the inhibitory effects of iga and igm . this receptor may be the long sought after gene that explains many of the “ sporadic ” breast cancers . here , the term “ sporadic ” refers to breast cancer originating from unknown genetic origins . the poly - ig receptor gene is located at locus d1s58 on chromosome 1 ( 23 ) which has been proven to be a “ hot spot ” for allelic imbalances in more than 70 % of breast cancers ( 24 ). a hot spot is a chromosomal loci or gene that is frequently altered in breast cancer specimens . this site has 46 % loss of heterozygosity ( loh ) in breast cancer specimens ( 24 ) and 30 % incidence of allelic imbalance ( al ) ( 24 ). this research , and that described in the above - identified u . s . and pct patent applications ( 21 , 22 ), support the conclusion that this gene will have notably broader significance in “ sporadic ” breast cancer etiology than brca1 ( located on chromosome 17 ) or brca2 ( located on chromosome 13 ). mutations in genes that are critical to cell growth or are known to be predisposing for breast cancer have been described . such mutations can either cause activation of oncogenes to promote cell replication or cause inactivation of suppressors to release cells to growth without control . in studies related to the present disclosure and described in the above - identified u . s . and pct patent applications , the poly - ig ( fc ) receptor or a similar poly - ig - like ( fc ) receptor has been identified as a tumor suppressor . binding the ligands iga or igm results in growth arrest in er + breast cancer cells . when this receptor is absent , cells replicate without immune control . the gene identified in this invention has not previously been recognized to have growth regulating properties . as this study developed the application of this new receptor gene function indicated an important concern for both germ line and somatic mutations . although the majority of cancers are thought to arise by somatic mutation , the fact that “ familial aggregation ” of breast cancers exists is strong evidence in favor of germ line analysis . the line of thought is to study both types of mutations in the poly - ig - like receptor . germ line mutations will be sought with blood cells or mucosal cells ( i . e . mucosal scrapings ) obtained from women . somatic mutations are revealed via examination of breast cancer specimens and cells aspirated from breast ducts or in breast fluid samples . key models of germ line breast cancer predisposing genes are readily available from sources known to investigators in this field . these include tp53 , atm , pten , mlh1 and the msh2 genes . if the full expression of these predisposing genes is to be realized , other mutations must contribute . the poly - ig - like receptor is found in every steroid and thyroid hormone responsive cell line examined to date . when the receptor is lost , cells achieve full autonomy . this indicates that the receptor gene may be a key contributor to the development of breast cancer as well as other cancers arising from mucosal tissues . genetic analysis of the poly - ig ( fc ) receptor or poly - ig - like ( fc ) receptor to determine breast cancer susceptibility in heterozygotes and homozygotes individuals seeking to determine breast cancer risk for the reasons cited above will be screened for predisposing genetic damage / mutations in the gene for the poly - ig ( fc ) receptor or poly - ig - like ( fc ) receptor . this will be done using lymphocytes and technology suited to rapid but accurate screening ( e . g . pyrosequencing and rapid pcr methodology ). this analysis will be preformed similarly to those used to identify heterozygotes and homozygotes for the atm mutation and for hereditary nonpolyposis colorectal cancer ( hncc ). studies done with control volunteers will be carried out to determine natural innocuous mutations in the gene and to determine if selected populations have different mutations and hence may be at greater risk . it is expected that one damaged gene will confer greater risk than controls because inactivation of the other functional gene will eliminate immune negative growth regulation by iga and igm . suitable techniques that will be used for this genetic analysis are well known in the art . such an analysis has not been recognized previously as useful in determining risk for breast cancer in populations before development of the disease . individuals showing homozygous mutations are considered at highest risk and will be counseled to decide on surgical prophylactic measures or on the use of tamoxifen or other anti - estrogen as preventative measures . thus , genetic screening can be used to not only to define potential risk , but to assistant in initiating preventative life saving actions , as discussed in more detail in the following example . screening for loss of heterozygosity ( loh ) or allelic imbalance ( ai ) in the poly - ig ( fe ) receptor or poly - ig - like ( fe ) receptor using blood cells , mucosal scrapings , breast fluid derived cells and other body and tissue samples and fluids , the presence of loh and al in the poly - ig ( fc ) receptor or a poly - ig - like ( fc ) receptor gene will be determined by methods commonly applied and well known . preferably the d1s58 locus of chromosome 1 will be a primary point of focus . this analysis can begin at very young ages ( i . e . nine or ten years old ) or can be initiated at age 25 when breast cancer rates are still very low . analysis of breast fluid cells can be continued with women showing loh or al at early ages . this information is used along with methods such as mammography to monitor women at high risk . early genetic analysis , as described in this example and in examples 1 , 3 and 4 , is especially valuable for identifying women at risk so that appropriate steps for risk reduction or prevention can be taken . if germline mutations are found or if somatic mutations are found , the issue then becomes “ what to do ?”. preventative and therapeutic compositions and methods are described in co - pending u . s . patent application ser . no . 09 / 852 , 547 and in international patent application no . pct / us01 / 15171 ( also identified as item 21 in the references , below ), or in co - pending u . s . patent application ser . no . ______ ( atty . dkt . no . 1944 - 01201 ) entitled “ breast cancer eradication program .” additional preventative and risk reduction methods and compositions are described in co - pending u . s . patent application ser . no . ______ ( atty . dkt . no . 1944 - 01301 ) entitled “ anti - estrogen and immune modulator combinations for treating breast cancer .” the disclosures of these co - owned patent applications are hereby incorporated herein by reference . double screening for poly - ig ( fc ) receptor or poly - ig - like ( fc ) receptor mutations and mutations in other breast cancer predisposing genes because the development of cancer most likely depends on more than one mutation ( 50 ), and may involve several cell types ( 50 ), it is useful to screen for mutations in genes that will lead to damage in other genes . both the lynch syndrome ii genes and the cowden &# 39 ; s disease gene are candidates for double screening along with the poly - ig ( fc ) receptor or a poly - ig - like ( fe ) receptor . for example , changes in the effectiveness of the lynch syndrome genes can lead to a gradual accumulation of mutations . if mutations are also present in the poly - ig - like receptor , risk can be expected to be substantially higher than controls . heterozygotes for the lynch syndrome genes have not been previously analyzed to determine if they possess an increased risk of breast cancer . likewise , heterozygotes for the cowen &# 39 ; s mutation have not been examined before to determine breast cancer susceptibility . these results will then be compared to results of screens for the poly - ig ( fe ) receptor or poly - ig - like ( fe ) receptor to identify those individuals at greatest risk . suitable risk reduction or preventative measures can then be implemented , as discussed in the preceding example . screening for heterozygous atm mutations and comparison to mutations in the poly - ig ( fc ) receptor or poly - ig - like receptor the atm mutation , associated with the disorder ataxia telangiectasia , is not found in breast cancers . nonetheless , it is clear that even heterozyogotes are at substantial risk for breast cancer . in light of the present disclosure of the importance of breast cancer cell growth control by the secretory immunoglobulins , it is of particular significance that the at disorder is known to be accompanied by chromosome fragility ( i . e . lack of repair after ionizing radiation ) and a marked deficiency in iga . because the atm mutation is so widely distributed in the population , it is important to initiate a screening for this mutation as a first line defense against breast cancer . identification of meaningful mutations will then permit decisions by women to elect preventative measures . pending results indicating a potential problem , these same individuals will then be offered another screening for the poly - ig ( fc ) receptor or a poly - ig - like ( fc ) receptor as a further indicator of risk status . if significant mutations are found in one or both receptor genes , prophylactic or preventative measures can be initiated , as discussed above in example 2 . screening of breast cancer specimens for mutations in growth regulating genes specimens from breast cancer patients will be screened for alterations in the poly - ig ( fe ) receptor or a poly - ig - like receptor to assist in therapy decisions and to identify individuals at greatest risk and requiring more intense intervention . this approach identifies somatic mutations . conventional screening techniques will also be used to identify heterozygous genes including the lynch syndrome gene and the cowden &# 39 ; s disease gene as well as alterations in tp53 . the molecular fingerprinting of tumors is expected to increase the effectiveness of treatment programs by allowing each to be adapted to the individual patient . future use of molecular methods is expected to provide a genetic profile of a patient &# 39 ; s primary tumor as well as to provide information relevant to family members concerning their potential risks . evaluation of breast fluid derived cells for molecular changes in genes cells will be obtained from breast fluids by any of a number of well known methods , or , alternatively , by newer methods that are known in the art and have been described in the literature for direct aspiration of the breast milk ducts . commercial milk pumps are available for this application . premalignant changes in the poly - ig ( fc ) receptor or a poly - ig - like ( fc ) receptor gene will be evaluated as will be changes in the other breast cancer predisposing genes discussed above . this technology is expected to identify somatic mutations at the site of breast cancer development . identification of sets of changes consistent with long - term development of the disease will permit immediate intervention to eradicate the altered cells or arrest the mutation process . after parturition , cells will be harvested from expressed breast milk and evaluated for somatic mutations . this analysis can be readily carried out during routine postpartum doctor &# 39 ; s office visits . the cells will be collected directly on to filters ( supplied ) and dna screening conducted , as previously described . this method permits direct assessment of the genetic status of a subset of reproductive age women without disruption of daily routines . fluid aspiration can also be done during routine mammography examinations . nipple pumps , which are available from well known commercial suppliers , can be used to withdraw only the few milliliters of fluid required . once a person or a group of persons have been identified as being at risk for breast cancer using one or more of the foregoing procedures , a program of prevention or risk reduction can be implemented , as described in , for example , co - pending u . s . patent application ser . no . ______ ( atty . dkt . no . 1944 - 01201 ) entitled “ breast cancer eradication program ,” and u . s . patent application ser . no . ______ ( atty . dkt . no . 1944 - 01301 ) entitled “ anti - estrogen and immune modulator combinations for treating breast cancer .” the disclosures of these co - owned patent applications are hereby incorporated herein by reference . such program can include ( a ) enhancing the amount of an immunoglobulin inhibitor (“ immune modulator ”) of cancer cell growth that contacts the breast ductal tissue of said individuals ; and / or administering an immunoglobulin inhibitor mimicking compound ; administering an anti - estrogenic compound ; ( b ) administering an aromatase inhibitor ; ( c ) enhancing the number of b immunocytes producing iga or igm in breast tissue ; and / or ( d ) immunizing individuals at risk of developing breast cancer against microorganisms known to or suspected of causing breast cancer . some immunoglobulin inhibitor mimicking compounds that may be used include : tamoxifen and mer - 25 and chemically substituted or modified derivatives thereof . to reduce possible side effects of mer - 25 , or its derivative compound , in some cases it may also be desirable to co - administer progesterone or another hormone . some immune enhancers that may be used include : levimisole , imiquimod , picibanil , and dhea . some useful anti - estrogens include : tamoxifen , toremifene , ici 16384 , ici 182780 , em - 800 , ru 58688 and em - 139 . 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( 50 ) beckmann m w , niederacher d , schnurch h - g et al ( 1997 ) mulitstep carcinogenesis of breast cancer and tumor heterogeneity . j mol med ( review ) 75 : 429 - 439 . while the preferred embodiments of the invention have been shown and described , modifications thereof can be made by one skilled in the art without departing from the spirit and teachings of the invention . the embodiments described herein are exemplary only , and are not intended to be limiting . many variations and modifications of the invention disclosed herein are possible and are within the scope of the invention . the disclosures of all patents , patent applications and publications cited herein are hereby incorporated herein by reference . the discussion of certain references in the description of related art , above , is not an admission that they are prior art to the present invention , especially any references that may have a publication date after the priority date of this application .