Optimización y análisis crítico del estudio de la monotorización de enfermedad mínima residual mediante ASO RQ:PCR en pacientes con mieloma múltiple. Comparación con la citometría de flujo
[EN] In patients with multiple myeloma ( MM) , the quality of response to treatment has been shown to have prognostic value , so that today the complete response (CR ) represents a therapeutic target in all MM schemes . However, we know that the current response criteria are suboptimal , since they are based on low sensitivity techniques such as immunofixation and morphology. Several studies indicate that immunophenotypic and molecular techniques are more sensitive and allow assessing minimal residual disease (MRD ) in bone marrow (BM ) , so that a better quality of response would be associated with longer survival. In the molecular field , strategies based on the use of specific primers and probes to quantify a patient and the tumor mass (ASO RQ- PCR ) has shown clinical value in patients with MM . Since the applicability of the same low, we consider first objective of this thesis improve its applicability , the search for new molecular markers and / or the use of samples enriched in plasma cells by CD138 + selection . Of molecular markers used in MM , VDJH somatic mutations frequently occurs and is detected only DJH in 60 % of patients . Alternatively, the Kappa deleting element rearrangements ( Kde ) are detected in all lymphoproliferative lambda B and 1/3 of the kappa and somatic mutations have not compromise the efficiency of primers and probes . Therefore, as in acute lymphoblastic leukemia , may be successfully used as markers for EMR in MM . This rearrangement studied in 96 patients with MM and found that is detected in 50 % of them , which can be easily sequenced and no somatic mutations. Then analyze its value as an additional marker in MM and saw EMR increases the applicability of these studies in 9% of cases overall and in 20 % of cases of MM lambda group that would be a significant advance . Then investigated using samples enriched MO plasma cells by immunomagnetic selection CD138 + as a material for obtaining molecular markers of EMR by PCR in MM . For this, of the 25 samples analyzed, a part was subjected to CD138 + immunomagnetic selection and the rest , not processed , was used as control. In both sample groups try to identify three molecular targets : VDJH , DJH and Kde . Seen that the use of selected samples increased from 60% to 96 % the proportion of cases with at least one molecular marker is available at the time of diagnosis . Its use , therefore, increase the applicability of such studies. The second objective of the thesis, we propose identifying problems responsible for failures in each of the steps in the molecular study of EMR following the recommendations of the European Group Euro - MRD , standardized but not in MM . Also, take advantage of this prospective study to compare the results of molecular techniques against immunophenotypic , in order to determine which of the two is ideal for monitoring in MM EMR . ASO saw RQ -PCR enabled monitoring of the EMR only in 42 % of patients due to pre- analytical problems responsible for a 28 % loss and post- analytical , 30% of them. In a series of 103 evaluable cases , we found that the quantification of MRD in patients with MM by ASO RQ -PCR allows monitoring the effectiveness of treatment and demonstrated to have prognostic value , stratifying patients into groups with different risk of progression. When we compare both techniques , we found that while the ASO RQ -PCR has a slightly higher sensitivity, the applicability of the CMF ( > 90 % ) is significantly higher. Therefore, the CMF is to be considered the technique of choice for monitoring EMR in MM .
Tesis y disertaciones académicas, Universidad de Salamanca (España), Tesis Doctoral, Academic dissertations, Hematología, Hematopatología, Hematology, Hematopathology
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