Cancer Initiation: Mesothelin emerges as a new biomarker for early cancer detection

Tuesday, December 31, 2013

Mesothelin emerges as a new biomarker for early cancer detection

 This note is intended to serve as a scientific background to a remarkable story, the story as told by The Scientist  [1] of Jack Andraka,  "The Cancer-Test Kid". While TS covers one end of this remarkable story, there are actually quite a few ways to look at it.  The first is, how is it that a relative youngster  who just earned his way into the education system beat an entire industry of "translational medicine" research to complete his remarkable find?  Were there things that "grown-ups" could have done to at least facilitate what seems to be a single handed story?  Obviously, the answer to these questions is "complicated". In addition to the usual convoluted emergence of understanding from medical science experiments, the diagnostic story of the mesothelin bio-marker overlaps "causation" and therapeutics in many remarkable and non-traditional ways, at least in terms of our traditional approach to medical research which strongly parallels the foundations of the "central dogma".

Research History

The story  of mesothelin appears to begin in the laboratories of the NIH's national cancer institute in Bethesda Maryland.[2] The initial discovery that led to the isolation and characterization of mesothelin occured in 1992, with an experiment that consisted of inoculating mice with ovarian cancer cells (OVCAR-3), and isolating an antibody that had unique interaction with the ovarian cancer cell surface. This antibody was designated K1. The antibody was found to also be reactive with human mesothelium, the external lining of many organs including the peritoneum. Hence, the future name of the K1 associated  biomarker was an indication of where it would normally be found in human tissues. The significance of this antibody was recognized immediately as this quote from the abstact shows:

The K1 antibody may be useful as a targeting agent for therapy and in the diagnosis of ovarian carcinoma, as well as some other human cancers. ( [2] 1992 )
Even with the significance immediately recognized, it would not be until 1996 when the gene identified by the K1 antibody would be isolated and cloned by that same team[3]. The gene and protein product were the officially named mesothelin, again, for its normal location in mesothelium derived tissues.. Its function remained unknown.
  The next step was to produce a "mono-clonal" antibody which involved immunizing mice with mesothelin produced from their previously cloned gene.  Apparently, the antibody K1 was not what would be called a "mono-clonal" antibody, that is one which arose from a single gene sequence to target a single epitope, or "feature" on a biomarker. Such mono-clonal antibodies are the result of the construction of a hybridoma or immortalized cell line which produces the antibody. The hybridoma was sucessfully produced by the Chang lab in 1997 [4], after what ( from the abstract ) seems to have been technical difficulties.
Several attempts to produce anti-mesothelin hybridomas from spleen cells of mice immunized with recombinant mesothelin were unsuccessful. This report describes the isolation of anti-mesothelin scFvs from a phage display library made from the mRNA of the same spleens.[4]

The story then continues in 1999 in the laboratories of the Pacific Northwest Laboratory and the Fred Hutchinson Cancer center with a publication by Scholler et. al. [5] . What Scholler did was gather cells from ovarian cancer patients and immunize mice with them. The immune systems of the mice then generated antibodies that were specific to biomarkers that would be found on ovarian cancer cells. Among these was a mouse antibody ( mAb ) designated OV569. This antibody ( OV569 ) was found to bind selectively to ovarian cancer cells, as well as certain other cancer cells, while showing no interaction with normal tissue. In addition, a binding target of OV569 was found to be circulating in the blood serum of patients with ovarian cancer, but not in those that are free from cancer. The value of the "mesothelin" type target as a marker for cancer was immediately recognized by the group that published the initial findings of a circulating form of mesothelin, and was noted as follows:
 Thus, there is a need for improvement, either in the form of a more specific and/or sensitive assay or an assay that uses a different marker and can be used to complement CA125 toward the goal to improve patient survival by improving diagnosis. [5]
The current biomarker for ovarian cancer in CA-125, or cancer antigen 125, otherwise known as MUC16 ,  a mucin or carbohydrate rich protein found highly expressed on surface cells of the female reproductive tract. Of note here, the laboratory at Johns Hopkins and Jack Andraka himself were interested in biomarkers for pancreatic cancer, not ovarian cancer. It is not clear that the Scholler team immediately recognized the importance of a non-mucoid biomarker as a means of early diagnosis of other types of cancer.
     The importance of finding a circulating and specific biomarker cannot be overstated. For example, in the current top video under the search "cancer biomarker", Raj Krishnan relates that a current Nobel Prize in medicine holder regards this as a top priority in cancer research, and one itself worthy of a Nobel Prize. Once again, the magnitude of the Jack Andraka story cannot be overstated.

    But, it seems that MUC16, the current and traditional biomarker for ovarian cancer,  remains part of the mesothelin story. MUC16 is not expressed in normal pancreatic ducts, but becomes upregulated and is expressed at the advancing edges of pancreatic cancer[6].  It seems that MUC16 and mesothelin actually bind to each other, and after doing so up regulat Matrix Metaloprotein 7 ( MMP-7)  [6]
     In less aggressive cancer, extra cellular matrix (ECM)  provides a barrier to invasion by the cancer. Extra cellular matrix is a web of protein and polysaccarides that exists outside the cell wall of tissues. It is called "matrix" because it consists of a web like structure, made largely of collagen for structural support and several types of glyco-proteins for signaling to cells.  Cells that "remodel" ECM do so by cleaving it with the aid of "Matrix Metalo-proteins" or MMPs. These MMPs are enzymes that use a particular metallic ion as a "knife" to enigmatically cut through ECM. One that is of relevance to pancreatic cancer is MMP-7. It has been shown, that in addition to cutting through extra-cellular matrix proteins, MMP-7 disrupts tight junctions, or cell to cell bindings in pancreatic epithelium. [7].    As such, cells of the invasive growth not only cut through ECM tissue, but cells dissociate and become free to metastasize through the blood stream.
   Consequently, when MMP-7 expression is blocked, the tissue invasion by  the cancer is reduced [6].

  As mentioned, mesothelin was found to be circulating in the blood stream of patients as early as 1999[5].  As such, mesothelin should be considered among the "gold standard" of potential biomarkers, because, it is very specific for a few types of cancer, it circulates in the blood stream, and is itself a promoter of the invasive qualities of the particular cancer. As such, it should have ( in my opinion ) been advanced more quickly than the article in The Scientist implies.

 Cell surface biomarkers are the foundation of targeted therapy. The first was the combination of human epidermal growth factor receptor 2, (HER-2) and herceptin a humanized monoclonal antibody to HER-2 used for treatment of breast cancer and other HER-2 positive cancer.[10] Based on the model of HER-2 and herceptin, mesothelin provides a new target for immune system targeting of cancer by manufacture of mesothelin specific monoclonal antibodies, or else the use of antibodies to deliver cyto-toxic loads targeted at specific cancer cells.[11][12]

 The nature of cancer research is that it is a competitive endeavor, were proposals are reviewed and compared, and rated at the level of each grant, or round of research. The goal is to provide the maximum rational basis of fairness between competing medical centers and universities. Unfortunately, when looking back over published track record of a particular line of research, such as that of mesothelin, the system gives the appearance of having a hap-hazard quality. The NIH must find a way to provide a fair system of grant awards, while maintaining a focused record of progress on important projects. In other words, there should not be a large number  of Jack Andracka type stories out there, and the National Cancer Institute of the NIH is the organization that should make sure this is not a regular occurrence.


[1] Dan Cossins, The Cancer-Test Kid. The Scientist, Dec. ( 2013) [Full Text]

[2]  Chang K, Pastan I, Willingham MC. Isolation and characterization of a monoclonal antibody, K1, reactive with ovarian cancers and normal mesothelium Int J Cancer. 1992 Feb 1;50(3):373-81. [Abstract]

[3]Chang K, Pastan I Molecular cloning of mesothelin, a differentiation antigen present on mesothelium, mesotheliomas, and ovarian cancers.. Proc Natl Acad Sci U S A. 1996 Jan 9;93(1):136-40.  [PubMed Central]

[4] Chowdhury PS, Chang K, Pastan I Isolation of anti-mesothelin antibodies from a phage display library.  Mol Immunol. 1997 Jan;34(1):9-20. [Abstract]

[5] Nathalie Scholler*, Ning Fu*†, Yi Yang*, Zhengmao Ye*,Gary E. Goodman*‡, Karl Erik Hellström*, and Ingegerd Hellström*§ Soluble member(s) of the mesothelin/megakaryocyte potentiating factor family are detectable in sera from patients with ovarian carcinoma
PNAS September 28, 1999 vol. 96 no. 20 11531-11536 [Full Text]

[6] Shih-Hsun Chen, Wei-Chien Hung, Pu Wang, Colin Paul & Konstantinos Konstantopoulos
Mesothelin Binding to CA125/MUC16 Promotes Pancreatic Cancer Cell Motility and Invasion via MMP-7 Activation Scientific Reports 3,Article number: 1870 doi:10.1038/srep01870 [Full Text]

[7]  Tan X, Egami H, Abe M, Nozawa F, Hirota M, Ogawa M.  Involvement of MMP-7 in invasion of pancreatic cancer cells through activation of the EGFR mediated MEK-ERK signal transduction pathway.   J Clin Pathol. 2005 Dec;58(12):1242-8. [PubMed Central Full Text]

[8] Argani P, Iacobuzio-Donahue C, Ryu B, Rosty C, Goggins M, Wilentz RE, Murugesan SR, Leach SD, Jaffee E, Yeo CJ, Cameron JL, Kern SE, Hruban RH. Mesothelin is overexpressed in the vast majority of ductal adenocarcinomas of the pancreas: identification of a new pancreatic cancer marker by serial analysis of gene expression (SAGE). Clin Cancer Res. 2001 Dec;7(12):3862-8. [PubMed Abstract and Full Text]

[9] Wang Y, Wang L, Li D, Wang HB, Chen QF. Mesothelin promotes invasion and metastasis in breast cancer cells. J Int Med Res. 2012;40(6):2109-16.[Abstract]

[10] Verma S, Joy AA, Rayson D, McLeod D, Brezden-Masley C, Boileau JF, Gelmon KA.HER story: the next chapter in HER-2-directed therapy for advanced breast cancer.
Oncologist. 2013;18(11):1153-66. [PubMed]

[11] Hassan R, Bera T, Pastan I. Mesothelin: a new target for immunotherapy. Clin Cancer Res. 2004 Jun 15;10(12 Pt 1):3937-42. [PubMed]

[12] Haridas D, Chakraborty S, Ponnusamy MP, Lakshmanan I, Rachagani S, Cruz E, Kumar S, Das S, Lele SM, Anderson JM, Wittel UA, Hollingsworth MA, Batra SK. Pathobiological implications of MUC16 expression in pancreatic cancer.  PLoS One. 2011;6(10):e26839.[Full Text]

[12] Tang Z, Qian M, Ho M. The role of mesothelin in tumor progression and targeted therapy. Anticancer Agents Med Chem. 2013 Feb;13(2):276-80. [PubMed Central]

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