Colorectal cancer (CRC) is the third most frequently diagnosed cancer, with 694,000 deaths of 1.4 million prevalent cases each year worldwide according to GLOBOSCAN 2020. Unfortunately, the numbers of new cases seem to be rising and it is believed that by the year 2030 the global burden of colorectal cancer (CRC) is expected to increase by 60% to more than 2.2 million new cases and 1.1 million deaths [1, 2].
CRC can be either sporadic or hereditary. The majority of CRC cases are sporadic of which more than 80% arise from somatic mutations in the Adenomatous Polyposis Coli (APC) and additional 13% are attributed to deficiencies of the DNA Mismatch Repair (MMR) genes .
One of the most prevalent hereditary cancer prone syndromes is Lynch Syndrome (LS) also known as Hereditary Non-Polyposis Colon Cancer (HNPCC). The molecular basis of LS is deficiency in the MMR system .
The direct consequence of impaired MMR activity is microsatellite instability (MSI) – alteration in the length of tandem repeats within microsatellite regions . Current laboratory assays for MSI and MMR include either a polymerase chain reaction (PCR) or an immunohistochemical (IHC) panel which demonstrates absence of 1 of 4 MMR enzymes MLH1, PMS2, MSH2,MSH6 .
It is estimated that of all new cases of colorectal cancer, 3% are attributable to LS .
The LS is characterized by predisposition to certain types of cancers, of which CRC and endometrial cancer (EC) are the most common. Among patients who have the LS, CRC tend to occur at younger age compared with patients with sporadic CRC (45 to 60 vs. 69 years of age) .
In the colon, LS associated cancers usually manifest as right sided tumors with a propensity for synchronous and metachronous CRC. Histologically, these tumors can present with poorly differentiated histologic features which may include mucinous features or a medullary growth pattern. These tumors tend to be infiltrated by lymphocytes which can be found between the cancerous glands [8–10].
Colorectal cancers, either sporadic or hereditary, which harbor a deficiency in DNA MMR mechanism, are diagnosed at early stages, have lower metastatic potential have better prognosis and higher disease free survival (DFS) when compared to proficient MMR tumors (pMMR) ]11[. Furthermore, the disease free survival of Stage III patients receiving the FOLFOX (fluorouracil, leucovorin, and oxaliplatin) therapy protocol was significantly longer in patients with deficient MMR (dMMR) CRC ]12[.
As mentioned above, MMR deficient CRC is characterized by increased density of tumor-infiltrating lymphocytes. This property makes these cancers possible candidates for immunotherapy with checkpoint inhibitors. Indeed, the anti–programmed death 1 (PD-1) antibodies pembrolizumab and nivolumab have been evaluated in patients with dMMR metastatic colorectal cancer in whom previous treatment with cytotoxic agents had failed. Pembrolizumab monotherapy and nivolumab monotherapy resulted in objective response rates that ranged from 31 to 52% (median follow-up time, 12 to 12.5 months) that were durable; similar responses were achieved in patients with LS associated CRC compared with non-LS CRC patients [13–15].
Therefore, identifying patients with deficient MMR CRC (either sporadic or hereditary) has a prognostic value and may be beneficial in tailoring a more suitable therapy for their disease.
Recent guidelines supported by multiple organizations including the Collage of American Pathologists (CAP) and the National Comprehensive Cancer Network (NCCN) have recommended the universal testing of all newly diagnosed CRC cases for deficient MMR or MSI in order to identify LS associated CRC cases [16, 17]. However, most CRC cases are not evaluated for MMR status because of the high costs associated with universal testing .
Tissue Micro Array (TMA) is an automatic system which enables allocating a few dozen tissue samples from their original standard paraffin blocks to a single TMA paraffin block, and subsequently to section and stain those samples simultaneously. In other words, as each standard paraffin block represent an individual patient, one TMA paraffin block represents many patients. Therefore, this method can be used with the potential benefit of significant reduction in cost and time 
Understanding the tumoral MMR status is of utmost importance not only for identifying patients with the LS or other MMR related familial CRC cases (and their family members), it is also important for accurate prognostication and for tailoring the best treatment to the patient. Indeed, the NCCN and the CAP recommend this test for every new case of CRC. With regard to testing MMR status in every new case of CRC, Israel is no different from other countries and the guidelines are not followed due to the high costs of the test.
The primary objective of this study is to demonstrate the accuracy of using TMA core evaluation compared to whole slide (i.e., ground truth). For this aim, a comparison between the core and the slide MMR status will be performed. As there are four proteins results per core, it was determined that the core had pathological finding if at least one of proteins was not stained. In case all of the proteins were stained, the core was classified as normal.
The secondary objective of this study is to evaluate and validate the automatic digital image analysis QuantCenter software by 3DHistech LTD. in differentiating pathological and normal cores. MMR proteins are stained by IHC and the software, which identify the nucleus, produces an automatic H-score for the nuclear staining of MMR protein. However, little is known about this score and it is very challenging to interpret its results. Also, the score is continuous, and no cutoff values were proposed. Thus, it is impossible to utilize this score in practice. Therefore, this study will propose a new staining score and examine both scores (H-score and the new staining score) for cutoff values to determine a pathological result as defined above.