IntroductionTheAmerican Cancer society looks at the number of cases of Cancer in North Americaevery year and compiles the data to provide statistics for the coming year. Atthe beginning of 2017, the three types of cancer expected to be most commonlydiagnosed in women was breast, lung and bronchus, and colorectal.
Breast cancerwas expected to account for 30% of all new cancer diagnoses in women. In menthe cancers to be most commonly diagnosed were prostate, lung and bronchus, andcolon and rectum with prostate expected to account for 19% of new cases. (Siegelet al.
, 2017). In Ireland, Cancer is the second biggest killer withover 9,000 deaths a year. (Irish Cancer Society, 2017). Focusing on Breast,Colon and Prostate Cancer as they are the leading killers and most studied, wesee that an increased amount of knowledge could lead to better therapies. Studieshave shown PDZ_LIM proteins are highly associated with regulating certaincancers. PDLIM2 is a protein coding gene also known as Mystique which islocated on the 8p21.
2 chromosome which encodes the PDZ_LIM protein. Only oneprotein isoform, Mystique 2 was found in cell lines. This protein hasnumerous amounts of cellular functionsincluding cell migration and adhesion, epithelial mesenchymal transition (EMT)and cell polarization. (Loughranet al., 2005) Main BodyPDLIM2and Breast CancerIthas been shown that NF-?B (nuclear factor kappa-light-chain-enhancer ofactivated B cells) plays a major role in Breast Cancer and that PDLIM2 ismajorly repressed in breast cancer cells. It was therefore hypothesised andproven that PDLIM2 may be involved in breast cancer pathogenesis.
NF- ?B is a proinflammatorysignalling pathway which is due to the nuclear factors high expression incytokines, adhesion molecules and chemokines which are all proinflammatorygenes (Lawrence,2009). NF-?B has other functions also including cellproliferation and expressing I?B?, the activated I?B? then enters the nucleusand transports the active NF-?B back to the cytoplasm. P65 is a member of NF-?B and was shown to play a role in breastcancer development and progression allowing it to resist therapeutic treatmentsof cancer. PDLIM2 was found to target P65 specifically for polyubiquitination-mediatedproteasomal degradation which in turn terminates the activation of NF-?B. Ithas been shown that the C-terminal Lim domain is responsible for promoting theubiquitination of P65 and that the N-terminal PDZ domain is responsible fortransporting the P65 into intranuclear compartments for the proteasomaldegradation. The activation of NF-?B isn’t wellunderstood but it’s thought that PDLIM2 plays a role.
It was found that whenPDLIM2 was expressed, NF-?B activation was inhibited as well as malignant tumorcells tumor formation in vivo and anchorage-independent growth in vitro. PDLIM2mutants which lost the NF-?B inhibiting ability also lost the ability tosuppress breast cancer. The protein level of PDLIM2 was tested in many breastcancer cell lines including the two most used model human breast cancer celllines, MCF-7 and MDA-MB-231. It was proven that the breast cancer cell linesexpress much less PDLIM2 irrespective of ER expression in comparison with thenontumorigenic breast cells, HMT3522 and MCF10A, (Fig 1A) (Quet al., 2010a). A study also showed that the PDLIM2 repression wasactually at the mRNA level in colon cancer (Yanet al., 2009). Qu et al.
also discovered a PDLIM2 decrease at themRNA level in breast cancer cell lines compared to normal cells, (Fig 1B). Asmentioned above, PDLIM2 expression inhibited NF-?B activation therefore thedata suggested that the repression of PDLIM2 may then take part in the breastcancer cell NF-?B activation. A luciferase gene reporter assay was carried out andshowed that PDLIM2 which when transiently expressed provided a dose dependantsuppression of activated NF-?B.
A permanent expression of PDLIM2 resulted ininhibition of NF-?B activation and inhibited Bcl-2 and cyclin D1 expression,these are NF-?B targeted genes in which are associated with tumors and found inthe cell lines used to carry out the assay. As PDLIM2 inhibits NF-?B activationby transporting p65 for proteasomal degradation, the impact that PDLIM2 has onthe expression of p65 was examined. MDA-MB-231 cells were used to carry this examinationout. PDLIM2 overexpression caused the p65 expression to decrease in the solublenuclear area but caused an increase of p65 in the insoluble nuclear area.
MG132, a cell permeable proteasome inhibitorwas used to treat the cells resulting in a build up of p65 at the insolublefraction and a slight p65 recovery in the soluble fraction. The MG132 caused adecrease in PDLIM2 in the nuclear soluble fraction but and increase in theinsoluble. This showed that PDLIM2 cannot be released until the proteasomal degradationof p65 is carried out.
These results further the suggestion of the repression ofPDLIM2 being part of the activating mechanism of NF-?B in human breast cancercell lines. CONTINUE FROM: * PDLIM1in Breast Cancer PDLIM2in Colon CancerTherisk of colon cancer is increased with chronic inflammatory disorders like inflammatorybowel diseases including ulcerative colitis and Crohn’s Disease. These resultin constant recurring inflammation elevating the risk of colorectal cancercausing it to be the second most common source of cancer related deaths (Bursteinand Fearon, 2008). The activation of the NF-?b transcription factoralso plays a major role in colon tumorigenesis and colon inflammation.
PDLIM2 aterminator of the activation of NF-?b and was proven to be repressed in humancolorectal cancer cell lines in comparison to a non-cancer cell line (Figure1A), methylation is involved in the repression of the PDLIM2 protein. Theactivity of NF-?b is closely regulated under physiological conditions (Xiao,2007). PDLIM2 inhibits NF-?b activation as mentioned abovewhich leads to tumour supressing activity, demonstrated through the decreasedexpression of PDLIM2 in colon cancer cells. How PDLIM2 is repressed wassuggested by linking DNA methylation regulation and PDLIM2 expression. A DNAmethyltransferase inhibitor (5-aza-dC) replenished the PDLIM2 expression incolon cancer cell lines suggesting that DNA methylation is responsible forPDLIM2 repression (Fig.
1B). More data from a bisulfite genomic DNA sequencing furtheredthe belief that methylation is directly responsible for the expression ofPDLIM2 (Quet al., 2010b). 5-aza-dC is toxic to cancer cells and is non-toxicto normal cells which makes it a hopeful therapy for colon cancer (Garcia-Manero,2008). Qu et al. then looked at PDLIM2 re-expressionand its relationship with NF-?b activation in colon cancer cells.
Luciferase genereporter assays were carried out to research how the restoration of PDLIM2effects NF-?b activation in these colon cancer cells. The re-expression ofPDLIM2 caused a dose-dependent suppression of the NF-?b activation in a numberof colon cancer cell lines, this was linked with a decreased expression of p65.Further studies showed that similarly to the p65 in breast cancer cell lines, PDLIM2is responsible for the shuttling of p65 for proteasomal degradation in thenuclear matrix. The data again implies that the repression of PDLIM2 is acontributing mechanistic factor of NF-?b activation but this time, also in coloncancer cells. To look further at PDLIM2 re-expression, stable expression ofPDLIM2 was done in HCT116 and DLD1whcih are colorectal cancer cell lines. Thesoft agar colony formation activity was measured compares to a control. Thecell lines which expressed PDLIM2 formed smaller and less colonies in the softagar. This showed that in colon cancer cells, tumorigenicity is supressed byPDLIM2 in vitro.
To establish that the same would happen in vivo, a control andcolon cancer cells expressing PDLIM2 were injected subcutaneously into differingflanks of SCID mice. As you can see in Figure3 below, the control and PDLIM2 both developed tumors at the site of theinjection but the tumors at the site including PDLIM2 were noticeably smallerwhich indicates that PDLIM2 does suppress colon cancer tumorigenicity in vivo. PDLIM2in Human Castration-Resistant Prostate CancerProstatecancer is the most common occurring cancer in males, it accounts for nearly 1in 5 new diagnoses in males. The rate of cases is dropping due to an increasein prostate-specific antigen (PSA) testing (Siegelet al., 2017).
Contrary to above where the ability of PDLIM2 tosuppress tumors was mentioned, in this case, a prostate cancer cell line(DU145) showed a high level of expression of PDLIM2. It’s clear that PDLIM2 hasa multifunctional role amongst various cancers. Discussion