Izraženost proteina NOTCH1 i JAGGED1 u monoklonalnoj gamapatiji neodređenog značenja i multiplom mijelomu

Škrtić, Anita (2011) Izraženost proteina NOTCH1 i JAGGED1 u monoklonalnoj gamapatiji neodređenog značenja i multiplom mijelomu. PhD thesis, Sveučilište u Zagrebu.

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Abstract

Multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS) share phenotypic profile and similar genomic aberrations but rarely MGUS progresses into overt MM. Notch signalling is implicated in the pathogenesis of multiple myeloma expressing high level of active Notch proteins NOTCH1 and JAGGED1 in tumor plasma cells. We investigated expression of NOTCH1, JAGGED1, CYCLIN D1, CD20 and PAX5 in bone marrow trephine biopsies of 80 newly diagnosed multiple myeloma and 20 monoclonal gammopathy of undetermined significance patients using immunohistochemical methods. The number of NOTCH1 and JAGGED1 positive tumor cells was counted per 1000 tumor cells and the intensity of staining was assessed semi quantitatively. Immunohistochemical results were compared with the morphological characteristics of tumorous plasma cells in bone marrow, the presence of t(11;14)(q13,q32), t(14;16)(q32;q23), t(4;14)(p16;q23), del13q34 and PAX5(9p13) translocation, the clinical and laboratory data, as well as overall survival in multiple myeloma patients. MGUS and MM showed no significant difference in the pattern of bone marrow infiltration, plasma cell differentiation, immunohistochemical staining of CYCLIN D1, CD20, PAX5 as well as analyzed chromosomal changes in tumorous plasma cells. In contrast, MGUS and MM differ according to type of infiltration of tumorous plasma cells in bone marrow with significant occurrence of diffuse pattern in MM (P<0.001). A similar occurrence of PAX5(9p13) translocation in MGUS and MM plasma cells may indicate its early occurrence in the pathogenesis of plasma cell neoplasia. Multiple myelomas expressed NOTCH1 in 92.31% and JAGGED1 in 92.21% cases with strong staining of NOTCH1 in the majority of cases (59.7%), whereas JAGGED1 was predominately weak (67.6% of cases). In contrast, both markers were negative in all MGUS cases (P<0.001). However, upon progression of disease from MGUS to MM (seen in 4 patients) analysis of the subsequent bone marrow biopsies showed weak expression of both markers in tumorous plasma cells. A significant correlation was found in strong NOTCH1 staining with diffuse type of bone marrow infiltration (P=0.020), an immature morphologic type of plasma cells (P=0.043) and MM subgroup with cytogenetic aberrations in plasma cells (P=0.021). In MM subgroup with cytogenetic aberrations in tumorous plasma cells a statistically significant occurrence of immature morphological type of plasma cells (P=0.024), serum β2 microglobulin (sβ2m) >3,5 mg/L (P=0.029) and serum albumin ≤35 g/L (P=0.044) was found. In this MM subgroup the patients were significantly more frequent in Durie/Salmon (DS) stage III (A or B) (P=0.012). After a median follow-up of 20.3 months, multiple myeloma patients with DS stage III (A or B) had statistically shorter overall survival which confirms DS system as the standard for prognostication in MM (P=0.046). Frequent occurrence of sβ2m >3,5 mg/L and serum albumin ≤35 g/L in MM patients with DS stage III (A or B) was found to be statistically significant. Correlations between serum level sβ2m, albumin and patients survival imply connections to underlying mechanisms. t(14;16)(q32;q23) (P=0.019) and PAX5(9p13) translocation (P=0.048) present prognostic factors with statistically shorter survival in MM patients. In multiple myeloma patients no difference in overall survival between JAGGED1 positive and negative cases were found. In conclusion, our results indicate an importance of chromosomal aberrations in pathogenesis of plasma cell neoplams, their role in the diagnostic and therapeutic algorithms, as well as prognostic value in MM and MGUS. A difference in NOTCH1 and JAGGED1 expression in MGUS and MM may imply possible role of both proteins in pathogenesis of plasma cell neoplasms, and a possible diagnostic value of NOTCH1 and JAGGED1 immunohistochemical evaluation of bone marrow infiltrates for multiple myeloma.

Abstract in Croatian

Multipli mijelom (MM) i monoklonalna gamapatija neodređenog značenja (MGUS) imaju zajedničke fenotipske i citogenetičke karakteristike u tumorskim plazma stanicama, no bolesnici s MGUS-om rijetko razviju kliničku sliku MM-a. Izraženost aktivnih Notch proteina NOTCH1 i JAGGED1 u tumorskim plazma stanicama multiplog mijeloma upućuje na Notch signalni put kao jedan od mogućih čimbenika u patogenezi bolesti. Imunohistokemijski smo analizirali izraženost NOTCH1, JAGGED1, CIKLIN D1, CD20 i PAX5 proteina u tumorskim plazma stanicama u koštanoj srži 80 bolesnika s klinički potvrđenom dijagnozom MM-a i 20 bolesnika s klinički potvrđenom dijagnozom MGUS-a. Broj NOTCH1 i JAGGED1 imunohistokemijski pozitivno obilježenih tumorskih stanica određivan je na ukupnom broju od minimalno 1000 tumorskih plazma stanica te je semikvantitativno određivan intenzitet bojanja. Usporedili smo rezultate imunohistokemijskog bojanja s morfološkim stupnjem diferenciranosti i tipom infiltrata tumorskih plazma stanica u koštanoj srži, prisutnošću t(11;14)(q13,q32), t(14;16)(q32;q23), t(4;14)(p16;q23), del(13q34) i PAX5(9p13) translokacije, kliničkim i laboratorijskim parametrima te s ukupnim preživljenjem bolesnika s MGUS-om i oboljelih od MM-a. MGUS i MM nisu se razlikovali prema morfološkom stupnju diferenciranosti, imunohistokemijskom bojanju CIKLIN D1, CD20, PAX5 proteina, kao ni prema učestalosti citogenetičkih promjena u tumorskim plazma stanicama. Suprotno tome, MGUS i MM razlikovali su se prema tipu infiltrata tumorskog tkiva u koštanoj srži sa statistički značajnom pojavnošću difuznog tipa infiltrata u MM-u (P<0,001). Slična učestalost PAX5(9p13) translokacije u MGUS i MM plazma stanicama najvjerojatnije je rani događaj u patogenezi plazmastaničnih novotvorina. U MM-u NOTCH1 protein bio je pozitivan u 92,31%, a JAGGED1 u 92,21% slučajeva. Jaki intenzitet bojanja NOTCH1 proteina u tumorskim plazma stanicama nađen je u većini slučajeva (59,7%), dok je intenzitet bojanja JAGGED1 proteina bio slab (67,6%). U svih bolesnika s MGUS-om oba proteina bila su negativna u tumorskim plazma stanicama (P<0,001). Međutim, četiri bolesnika s MGUS-om razvila su kliničku sliku MM-a u razdoblju od 3,7 do 29,5 mjeseci (medijan 17,65 mjeseci). Analiza i usporedba prve i druge biopsije koštane srži pokazala je razliku u izraženosti obaju proteina u tumorskim plazma stanicama. U drugoj biopsiji koštane srži tumorske plazma stanice bile su NOTCH1 i JAGGED1 pozitivne, slabog intenziteta bojanja. Značajna učestalost jakog intenziteta bojanja NOTCH1 proteina nađena je u MM-u s difuznim tipom infiltrata koštane srži (P=0,020), morfološkim stupnjem III diferenciranosti plazma stanica (P=0,043) te u podskupini MM-a s citogenetičkim promjenama u tumorskim plazma stanicama (P=0,021). U podskupini MM-a s citogenetičkim promjenama u tumorskim plazma stanicama nađena je značajna pojavnost morfološkog stupnja III diferenciranosti plazma stanica (P=0,024), serumskog β2 mikroglobulin (sβ2m) >3,5 mg/L (P=0,029) i serumskog albumina ≤35 g/L (P=0,044). U ovoj podskupini MM-a bolesnici su bili značajno češće u kliničkom stadiju III (A/B) prema Durie Salmonu (DS) (P=0,012). Bolesnici oboljeli od multiplog mijeloma u DS kliničkom stadiju III (A/B) imali su značajno kraće preživljenje (P=0,046). Rezultati potvrđuju vrijednost DS sustava kao standarda u prognostičkoj podjeli kliničkih stadija u bolesnika oboljelih od MM-a. U bolesnika oboljelih od MM-a u DS kliničkom stadiju III (A/B) nađena je značajna pojavnost sβ2m >3,5 mg/L i serumskog albumina ≤35 g/L, što upućuje na povezanost mehanizama koji sudjeluju u patogenezi i progresiji MM-a. t(14;16)(q32;q23) (P=0,019) i PAX5(9p13) translokacija (P=0,048) prognostički su pokazatelji bolesti sa statistički značajnim kraćim srednjim vremenom preživljenja u bolesnika oboljelih od MM-a. U bolesnika oboljelih od MM-a nije se našla statistički značajna razlika u preživljenju između NOTCH1 i JAGGED1 pozitivnih i negativnih rezultata bojanja u tumorskim plazma stanicama. Naši rezultati upućuju na važnost citogenetičkih promjena u patogenezi plazmastaničnih novotvorina, njihovu ulogu u dijagnostičkim i terapijskim protokolima te na njihovu prognostičku vrijednost u MM-u i MGUS-u. Različita izraženost NOTCH1 i JAGGED1 proteina u MGUS-u i MM-u upućuje na moguću ulogu navedenih proteina u patogenezi plazmastaničnih novotvorina te na moguću dijagnostičku vrijednost analize NOTCH1 i JAGGED1 imunohistokemijskog bojanja infiltrata koštane srži u multiplom mijelomu.

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