<p>B cell lymphoma (BCL) comprises a biologically and clinically heterogeneous group of tumors deriving from different stages of B cell development. The immunoglobulin (Ig) variable heavy chain (V_H) gene rearrangement is unique for each BCL and can be used to reveal cellular origin, to study signs of antigen selection and to quantify tumor cell load.</p><p>The normal counterpart of mantle cell lymphoma (MCL) has been postulated to be a naïve B cell and in hairy cell leukemia (HCL) it is considered to be a post-germinal centre B cell. We analyzed the V_H gene rearrangements in 110 MCLs and 32 HCLs by PCR amplification and sequencing. Most MCLs (84%) displayed V_H genes lacking somatic hypermutation (SHM), thus correlating to a naïve cell origin, whereas a subgroup (16%) showed SHM, implying derivation from a more differentiated B cell. In HCL, a majority of cases (84%) displayed SHM with signs of intraclonal heterogeneity and 16% had unmutated V_H genes, thus questioning the cell of origin in HCL. Biased usage of particular V_H genes was detected in both HCL (V_H3-30) and MCL (V_H3-21 and V_H4-34), which indicates that antigen selection may be involved in lymphoma development. Furthermore, V_H3-21⁺ MCLs showed a highly restricted V_λ3-19 gene use and they also had a superior outcome compared to other MCLs.</p><p>Rearrangement analysis of 67 V_H3-21⁺ chronic lymphocytic leukemia (CLL) cases from three different countries verified, regardless of geographical origin, the short and highly homologous complementarity determining region 3s and the strikingly biased usage of the V_λ2-14 gene (75%), as previously reported in CLL. This further supports that antigen selection by a common antigenic epitope may have occurred in V_H3-21⁺ CLLs. </p><p>In an autologous transplantation study of 30 multiple myeloma patients, we quantified the tumor content in the autografts before and after stem cell selection using clone-specific PCR. We conclude that stem cell selection reduced the number of clonal cells linearly, but purging could not totally eliminate the tumor cells from the graft, thus increasing the risk of a relapse.</p><p>Altogether, our data allowed us to define new BCL subsets and to gain insights into the potential role of antigen selection in BCL development as well as the monitoring of tumor cell load using Ig gene rearrangements analysis. </p>

B cell lymphoma (BCL) comprises a biologically and clinically heterogeneous group of tumors deriving from different stages of B cell development. The immunoglobulin (Ig) variable heavy chain (VH) gene rearrangement is unique for each BCL and can be used to reveal cellular origin, to study signs of antigen selection and to quantify tumor cell load. The normal counterpart of mantle cell lymphoma (MCL) has been postulated to be a naïve B cell and in hairy cell leukemia (HCL) it is considered to be a post-germinal centre B cell. We analyzed the VH gene rearrangements in 110 MCLs and 32 HCLs by PCR amplification and sequencing. Most MCLs (84%) displayed VH genes lacking somatic hypermutation (SHM), thus correlating to a naïve cell origin, whereas a subgroup (16%) showed SHM, implying derivation from a more differentiated B cell. In HCL, a majority of cases (84%) displayed SHM with signs of intraclonal heterogeneity and 16% had unmutated VH genes, thus questioning the cell of origin in HCL. Biased usage of particular VH genes was detected in both HCL (VH3-30) and MCL (VH3-21 and VH4-34), which indicates that antigen selection may be involved in lymphoma development. Furthermore, VH3-21+ MCLs showed a highly restricted Vλ3-19 gene use and they also had a superior outcome compared to other MCLs. Rearrangement analysis of 67 VH3-21+ chronic lymphocytic leukemia (CLL) cases from three different countries verified, regardless of geographical origin, the short and highly homologous complementarity determining region 3s and the strikingly biased usage of the Vλ2-14 gene (75%), as previously reported in CLL. This further supports that antigen selection by a common antigenic epitope may have occurred in VH3-21+ CLLs. In an autologous transplantation study of 30 multiple myeloma patients, we quantified the tumor content in the autografts before and after stem cell selection using clone-specific PCR. We conclude that stem cell selection reduced the number of clonal cells linearly, but purging could not totally eliminate the tumor cells from the graft, thus increasing the risk of a relapse. Altogether, our data allowed us to define new BCL subsets and to gain insights into the potential role of antigen selection in BCL development as well as the monitoring of tumor cell load using Ig gene rearrangements analysis.

This study demonstrates the use of an adenovirus vector system to study the effect of a DNA repair gene on untransformed human fibroblasts. The bacteriophage T4 pyrimidine dimer DNA glycosylase (den V) gene has been inserted into the E3 region of human adenovirus type 5. The resulting recombinant virus Ad Den V was determined to be producing correctly initiated RNA from the RSV 3' LTR promoter used in the den V expression cartridge inserted into the virus. The effect of the den V gene product on human fibroblasts 'liras examined by assaying for the percent host cell reactivation (%HCR) of Vag production for UV irradiated Ad Den V in comparison to that for a control virus. It was shown that the %HCR was significantly greater for Ad Den V as compared to the control virus in xeroderma pigmentosum (XP) cells. UV survival of adenovirus in XP cells exhibited a two component nature. Introduction of the den V gene into XP group A cells increased the D0 value of the first component of the viral survival curve to a level similar to that of XPC cells, which showed no change in this component irrespective of the presence of the den V gene. It has been suggested that the den V gene is able to partially complement the deficiency in some XP cells because of its small size, allowing it to gain access to the DNA damage site where as the cellular repair enzyme complex can not. Since XPC cells are proficient in their alteration of DNA secondary structure prior to DNA excision repair, these results are consistant with the hypothesis that the first component of UV viral survival curves reflects the pathway involved in accessing the damaged sites. The manuscript of a paper has been included as an appendix. The work theorizes on the origin of mammalian immune system diversity and bacteriophage lambda, and their possible relationship to prokaryotic DNA repair genes. / Thesis / Master of Science (MS)