Raju Chaganti, PhD

The main goal of this laboratory is the analysis of the genetic basis of deregulation of cellular proliferation and differentiation associated with malignant transformation. Several tumor types are investigated, notably non-Hodgkin's lymphoma (NHL) and adult male germ cell tumors (GCTs). A wide range of laboratory techniques are utilized in these studies, which include conventional and molecular cytogenetics, comparative genomic hybridization (CGH), multicolor spectral karyotyping (SKY), molecular cloning, and structure-function analysis of genes. The laboratory emphasizes the investigation of the relevance of newly discovered genetic markers to tumor cell classification and clinical behavior. In order to pursue these latter studies with state-of-the-art approaches, we have incorporated high-throughput microarray technology into our methodology.

NHL arises in normal B and T cells during their lineage differentiation, which is signaled by productive intragenic rearrangements of the immunoglobulin (IG) and T-cell receptor (TCR) genes. Occasional errors in the rearrangement process result in chromosomal translocations, which juxtapose the IG and TCR genes with novel genes from the break junction site of the reciprocal chromosome. The juxtaposed gene is placed under the control of the IG or TCR gene and becomes activated, leading to deregulation of normal differentiation of the lymphoid cell and initiation of proliferation. We have discovered several new translocations in B-cell lymphomas that involve the IG gene sites and isolated the deregulated genes. In these translocations, as a general rule, the coding region of the deregulated gene is not disrupted, while the upstream regulatory regions are replaced by those of the IG genes, leading to abnormal expression of the normal gene product. Among the genes identified by us in these translocations, transcription factors predominate (including NFKB2, PAX5, MUM1/IRF4, and BCL6). Other genes that have been isolated play a role in tumor progression (MUC1), while the function of several novel genes is still to be fully characterized (BCL8, BCL11). Studies of the newly discovered transcription factors are providing new insights into normal development of B cells and the mechanisms underlying their transformation.

Through CGH studies, we have discovered that gene amplification of chromosomal regions is a common phenomenon in NHL. Using a candidate gene approach, we identified REL (mapped at 2p14 to 15) to be frequently amplified and overexpressed in aggressive lymphomas. The consequence of this REL overexpression to the NFKB/REL-mediated anti-apoptotic response of tumor cells is under study. Because CGH identified large genomic segments (~10MB) and candidate gene approaches tend to be slow and labor-intensive, we have introduced high-throughput screening of tumors with chromosomal amplification for simultaneous analysis of gene amplification (chip-CGH) and gene expression using microarrays.

Adult human male germ cell tumors (GCTs) retain totipotentiality for differentiation -- a characteristic of their precursor cells, the germ cells. They present a fascinating biological paradox: to initiate a totipotential tumor, a germ cell committed to a highly defined differentiation path must overcome a restriction on proliferation and initiate differentiation of embryonal and somatic lineages. It must accomplish this without the reciprocal contribution from fertilization. Therefore, an understanding of the mechanisms of GCT development has considerable relevance for the understanding of normal germ-cell development as well as regulation of embryonal differentiation pathways in humans.

An additional feature of male GCTs is their exquisite sensitivity to cisplatin-based chemotherapy, with a small proportion of tumors manifesting lethal therapy resistance. GCTs are thus an ideal system to study pathways that underlie chemotherapy sensitivity and resistance of relevance to these tumors as well as to cancer per se. Our studies of GCTs address 3 questions:

• What is the mechanism of germ cell transformation?
• What is the genetic basis of chemotherapy sensitivity and resistance?
• How are lineage decisions initiated in these pluripotent transformed cells?

For a detailed description of our studies of these 3 aspects, please see the descriptions of the individual projects.