Molecular pathogenesis and prognostic factors in testicular tumor

Aim of this review article was to critically analyse the recently described zytogenetic and molecular markers for testicular germ cell tumors with regard to their clinical utility. The isochromosome i(12p) represents the most common and characteristic cytogenetic finding which already appears in testicular carcinoma in situ. A number of proto-oncogenes (Cyclin D and PTHLH) as well as putative tumor suppressor genes are localized on 12p; however, their role in pathogenesis and prognosis of testicular germ cell tumors has not been defined yet. Clinical characteristics of patients with familial testicular germ cell tumors indicate a genetic background for the development of testicular tumors. Although a number of chromsomal loci encoding potential testicular tumor susceptibility genes have been identified, the genetic basis of testicular cancer pathogenesis is still unknown. With regard to molecular prognostic risk factors most of the reported data on proliferation markers, tumor suppressor genes, proteases and adhesion molecules have to be confirmed in prospective randomized trials prior to their widespread clinical use. Based on the available data on prospective studies percentage of embryonal carcinoma and vascular invasion appear to be the most significant prognosticators. Investigation and identification of those factors determining the aggressive biologic behavior of embryonal carcinoma compared to all other histological components appear to be most promising in research for prognosticators of metastatic disease. In conclusion, the increasing knowledge of molecular genetic events involved in pathogenesis and prognosis of testicular germ cell tumors will not only help to better understand development and progression of testicular cancer, but it also will define new approaches to classification and management of germ cell tumors.     

Effect of the vitreous on the prognosis of full-thickness idiopathic macular hole

Cancer is a genetic disease; tumor cells differ from their normal progenitors by genetic alterations that affect growth-regulatory genes. There exist 2 classes of such cancer genes: the oncogenes, which function as positive growth regulators, and the tumor suppressor genes, which function as negative growth regulators. Oncogenes are widely conserved among diverse forms of life and are active in transmitting growth signals from the cell periphery to the cell nucleus. These signaling functions can be disturbed by many types of genetic change; the result of an altered growth signal is often cancer. Tumor suppressor genes have an attenuating effect on cell growth that is lost as a result of inactivating mutations or deletion of the gene; in DNA virus-transformed cells, it is abrogated by neutralization of the tumor suppressor protein through a viral gene product. Tumor suppressor genes were first recognized in inherited cancers; defects in a tumor suppressor transmitted through the germ line can lead to increased tumor incidence in the offspring. Tumor suppressors also play important roles in nonheritable cancer, however; many tumors in humans show defects in tumor suppressor genes. Most cancers harbor multiple genetic changes in oncogenes as well as tumor suppressor genes. Oncogenes induce aberrant growth through a gain in function; tumor suppressor genes contribute to oncogenesis through a loss of function. Both types of mutation work together to produce cancer; the changes are not constant but increase in number as the tumor develops from benign to more and more malignant. Cancer results from the accumulation of genetic changes. Oncogenes and tumor suppressor genes provide important insights into the regulation of cell growth. This knowledge can now be used to develop gene-specific therapies for cancer.     

Genetic changes in inherited and sporadic ovarian carcinomas by comparative genomic hybridization: extensive similarity except for a difference at chromosome 2q24-q32

Germ-line mutations in the BRCA1 and BRCA2 genes confer a predisposition to breast as well as ovarian carcinoma. Except for loss of the respective wild-type allele, somatic genetic changes needed for the progression of inherited ovarian tumors are unknown. A genome-wide search for such alterations was performed by comparative genomic hybridization analysis on BRCA1 and BRCA2 mutation-positive (n = 20) ovarian carcinoma specimens. Comparison with sporadic ovarian carcinomas (n = 20) revealed extensive genetic similarity between the inherited and sporadic carcinomas with the sole exception of a frequent gain of 2q24-q32 in the inherited group, suggesting the presence of an oncogene at 2q24-q32 operating in the absence of BRCA1 function. The overall similarity of gains and losses by comparative genomic hybridization suggests a common main pathway in tumor progression of both inherited and sporadic ovarian carcinomas.     

Effect of sperm genotype on the segregation of homologs in male mice, heterozygous for an aberrant chromosome 1

The aberrant chromosome 1 with two large homogeneously staining insertions was isolated from wild populations of Mus musculus. Specific features of aberrant chromosome were described elsewhere. These include preferential entry of the chromosome into the oocyte of heterozygous females, increased mortality of homozygotes, decreased fertility of homozygous females. The data obtained indicated that segregation of homologues in heterozygous females depended on which spermatozoon entered the oocyte, prior to the second meiotic division: meiotic drive is powerful when the spermatozoon bore the normal chromosome 1, and homologue segregation got normalized during MII when the spermatozoon bore the chromosome 1 with insertions. Experimental data are adduced and explanations offered for the phenomenon observed.     

Isolation of COM1, a new gene required to complete meiotic double-strand break-induced recombination in Saccharomyces cerevisiae

We have designed a screen to isolate mutants defective during a specific part of meiotic prophase I of the yeast Saccharomyces cerevisiae. Genes required for the repair of meiotic double-strand breaks or for the separation of recombined chromosomes are targets of this mutant hunt. The specificity is achieved by selecting for mutants that produce viable spores when recombination and reductional segregation are prevented by mutations in SPO11 and SPO13 genes, but fail to yield viable spores during a normal Rec+ meiosis. We have identified and characterized a mutation com1-1, which blocks processing of meiotic double-strand breaks and which interferes with synaptonemal complex formation, homologous pairing and, as a consequence, spore viability after induction of meiotic recombination. The COM1/SAE2 gene was cloned by complementation, and the deletion mutant has a phenotype similar to com1-1, com1/sae2 mutants closely resemble the phenotype of rad50S, as assayed by phase-contrast microscopy for spore formation, physical and genetic analysis of recombination, fluorescence in situ hybridization to quantify homologous pairing and immunofluorescence and electron microscopy to determine the capability to synapse axial elements.     

The origin and biology of CIS cells: general discussion

Participants at the 4th Copenhagen Workshop on Carcinoma in situ and Cancer of the Testis, representing cell biologists and tumour biologists, met together to discuss the similarities and differences between primordial germ cells (PGCs) of the embryo, and the carcinoma in situ (CIS) stem cell of human testicular germ cell tumours (GCTs). Much has been discovered about PGCs in the last 10 years and we still do not know the exact nature of CIS cells. Knowledge of PGCs comes mainly from mouse experiments and knowledge of CIS comes from the study of human tumours. A mouse model of human GCT would help to investigate the nature of CIS cells. Grafting mouse male genital ridges into mouse fetal testes results in the development of testicular tissue and the formation of teratomatous tumour components. Amplification of PGCs in culture is possible but this results in their transformation into embryonic germ (EG) cells. CIS cells die by apoptosis if they are isolated, and short-term culture is only possible if the CIS cells are cultured in their normal environment within seminiferous tubules. It may be possible for CIS cells to differentiate in culture although they cannot be maintained in culture as isolated cells. Human CIS cells are likely to be formed as a result of in utero factors rather than agents acting on normal adult testicular germ cells. EG cells stimulate feeder cells by paracrine factors but it is not known if these cells produce autocrine factors.     

Chromatin condensation behaviour of the Y chromosome in the human testis. I. Evidence for decondensation of distal Yq in germ cells prior to puberty with a switch to Sertoli cells in adults

The condensation behaviour of the human Y chromosome in germ cells and Sertoli cells of pre- and post-pubertal testes was followed by fluorescence in situ hybridisation using probes for three different regions of the Y chromosome. Patterns of expansion or contraction of signal are taken to reflect degrees of condensation of the related Y chromatin and hence its potential for genetic activity. For probe pHY2.1, which labels the distal non-fluorescent and fluorescent heterochromatin of the Y chromosome (Yq12), an expanded signal seen in gonocytes of the prepubertal testis is superseded by a condensed signal seen in adult germ cells at all but the zygotene stage of meiotic prophase when meiotic pairing takes place. In contrast, Sertoli cells show a condensed signal pre-pubertally but a greatly expanded signal in the adult testis. A totally condensed pHY2.1 signal is found in a chromosomally normal man with Sertoli-cell-only syndrome. It is hypothesised that control over at least some facets of spermatogenesis may not, in the adult, be autonomous to the germ cells, but rather may emanate from the Sertoli cells. Chromatin expansion at zygotene could, however, be important for pairing and crossing over in the XY bivalent, successful synapsis ensuring survival of spermatocytes into the post-meiotic stages.     

Involvement of serotonin and dopamine in the mechanism of action of novel antidepressant drugs: a review

BACKGROUND: By the time patients are diagnosed with ovarian carcinoma, peritoneal dissemination of the tumor often has occurred. The progressive growth and spread of ovarian carcinoma depend, in part, on the formation of an adequate blood supply. We determined whether the expression of genes that regulate distinct steps in angiogenesis (i.e., the formation of new blood vessels) was associated with the pattern and progressive growth of human ovarian carcinomas implanted in the peritoneal cavity of nude mice. METHODS: Five different human ovarian carcinomas were injected individually into the peritoneal cavity of female NCr-nu/nu nude mice. The expression of basic fibroblast growth factor, vascular endothelial growth factor/vascular permeability factor (VEGF/VPF), interleukin 8 (IL-8), and collagenase type IV (MMP-2 [matrix metalloproteinase-2] and MMP-9) was determined by northern blot analysis, in situ hybridization of messenger RNA, and immunohistochemical analysis. Blood vessel distribution and density, macrophage infiltration pattern, and stromal reaction were determined by immunohistochemical analysis with specific antibodies. RESULTS: Three of the carcinomas produced both solid lesions and ascitic tumors, whereas the remaining two produced only solid lesions. Two of the carcinomas produced rapidly progressive disease, two produced slow disease, and one produced intermediate disease. The formation of ascites was directly associated with expression of VEGF/ VPF, and survival was inversely associated with expression of IL-8. In rapidly growing tumors, the number of blood vessels was high throughout the lesion; in contrast, in slow-growing tumors, most vessels (and infiltrating macrophages) were located at the periphery. CONCLUSIONS: The expression of various genes that regulate angiogenesis in human ovarian carcinomas is associated with the pattern of the disease and its progression. Therefore, targeting specific genes that regulate angiogenesis could offer new approaches to the treatment of ovarian cancer.     

Allelic loss on a chromosome 17 in ductal carcinoma in situ of the breast

Multiple tumor suppressor genes are implicated in the oncogenesis and progression of invasive carcinoma of the breast. To investigate the chronology of genetic changes we studied loss of heterozygosity on chromosome 17 in ductal carcinoma in situ, a preinvasive breast cancer. A microdissection technique was used to separate tumor from normal stromal cells prior to DNA extraction and loss of heterozygosity was assayed mainly using simple sequence repeat polymorphism markers and the polymerase chain reaction. Loss of heterozygosity on 17p was observed in 8 of 28 tumors (29%) when compared with normal control DNA, whereas no loss was seen on 17q, suggesting that at least one locus on 17p is involved early in the development of breast cancer.     

Treatment of hemoglobin Bart''s hydrops with bone marrow transplantation

OBJECTIVE: It is currently proposed to perform tumour enucleation in mono-orchid patients presenting with a small germ cell tumour, in order to preserve physiological endocrine function. However, this conservative surgery must be accompanied by treatment of any carcinoma in situ lesions present in the remaining testicular parenchyma. MATERIAL AND METHODS: The presence of carcinoma in situ was investigated in 35 patients with germ cell tumour of the testis on samples obtained from the presumably healthy pulp of the orchidectomy specimen, adjacent to and away from the tumour. Samples away from the tumour were performed according to principle of surgical testicular biopsy. Histological examination also concerned the tissue situated in contact with the tumour. RESULTS: 25 patients (74.22%) presented intratubular germ cell dysplasia in the testicular parenchyma presumed to be healthy. These lesions were always concomitantly observed on samples performed adjacent to and away from the tumour. It was observed more frequently in seminomas, but was also observed in other histological forms. CONCLUSION: Because of the diffuse nature of carcinoma in situ in germ cell tumours of the testis, pulp samples adjacent to the tumour are sufficient to ensure the diagnosis during conservative surgery. Carcinoma in situ is currently treated by local radiotherapy.     

Dissemination with malignant changes from a pineal tumor through the corpus callosum after total removal

Within the germinal neoplasms of the central nervous system, the embryonal carcinoma is a very rare tumor. The authors report such a lesion arising on the corpus callosum in a child 8 months after a differentiated teratoma was totaly removed from the pineal region. The classification of the germ cell tumors of the central nervous system and their relation with similar tumors of the gonads are discussed. The treatment of the pineal tumors is also discussed.     

Cytological characterization of four meiotic mutants of Arabidopsis isolated from T-DNA-transformed lines

A secondary screen of the Feldmann collection of T-DNA transformed Arabidopsis lines identified several meiotic mutants. We used a spreading technique combined with DAPI staining in a detailed cytogenetic analysis of meiotic chromosome behaviour in four of these mutants, all of which are putatively T-DNA tagged and therefore candidates for molecular and functional analysis of the mutated genes. Two of them are defined as 'synaptic' mutants, showing greatly reduced association of homologous chromosomes at metaphase I: one is asynaptic, showing failure of synapsis during prophase I, whereas the other is desynaptic and is characterized by normal but non-maintained synapsis. Another mutant is defective in meiotic cell cycle control and undergoes a third meiotic division, resembling a second division but without an additional round of chromosome duplication. A further mutant shows meiosis-limited chromosome disruption, resulting in extensive chromosome fragmentation combined with other defects. All four mutants experience very irregular chromosome distribution during the meiotic divisions, resulting in abnormal numbers and/or sizes of microspores, with resulting reduced fertility.     

Induction of centromeric activity in maize by suppressor of meiotic drive 1

The Abnormal chromosome 10 (Ab10) in maize causes normally-quiescent blocks of heterochromatin called knobs to function as meiotic centromeres. Under these circumstances genetic markers associated with knobs exhibit meiotic drive, i.e., they are preferentially transmitted to progeny. Here we describe a mutation called suppressor of meiotic drive (smd1) that partially suppresses meiotic drive, and demonstrate that smd1 causes a quantitative reduction in the mobility of knobs on the meiotic spindle. We conclude that Smd1 encodes a product that is necessary for the activation of ectopic centromeres, and that meiotic drive occurs as a consequence of the resulting change in chromosome movement. As a genetic system, Ab10 offers a new and powerful approach for analyzing centromere/kinetochore function.     

In situ cancer vaccination: an IL-12 defective vector/replication-competent herpes simplex virus combination induces local and systemic antitumor activity

Intratumoral inoculation of replication-competent, attenuated herpes simplex virus (HSV) mutants inhibits tumor growth by direct cytotoxic viral replication and induction of a tumor-specific immune response. To boost the antitumor response, we describe a defective HSV vector encoding IL-12 as an adjuvant to in situ vaccination by the replication-competent HSV helper virus. The defective HSV vector system consists of defective particles containing tandem repeats of the cytokine genes (p40 and p35) in combination with a HSV helper virus. Heterodimeric IL-12 was expressed and secreted after IL-12 defective vector infection of tumor cells. In a syngeneic, bilateral established tumor model with CT26 murine colon carcinoma, unilateral intratumoral inoculation with an IL-12 defective/replication-competent HSV vector combination significantly reduced tumor growth of the inoculated and noninoculated contralateral tumors. This antitumor effect was significantly greater than with a lacZ-defective/replication-competent HSV vector combination, which itself was significantly greater than the mock inoculation. Efficacy is associated with enhancement of tumor-specific CTL activity, including specificity against the CT26 immunodominant MHC class I restricted Ag AH1, and IFN-gamma production. There was no significant tumor growth inhibition after intratumoral inoculation of s.c. CT26 tumors in athymic mice. We conclude that this defective HSV vector system is an effective method for cytokine gene delivery to tumors in situ and IL-12 expression in tumors synergizes the antitumor activity mediated by the replication-competent HSV helper virus.     

Mouse models in tumor suppression

Genetic lesions found in tumors are often targeted to the negative growth regulatory tumor suppressor genes. Much of our understanding of tumor suppressor gene function is derived from experimental manipulations in cultured cells. Recently, however, the generation of mice with germ line tumor suppressor gene mutations through gene targeting in embryonic stem cells has provided another dimension by allowing experimental studies of tumor suppressor function in an organismal context. Novel insights into the role of tumor suppressors in development, differentiation, cell cycle control, and tumor suppression have been obtained from the studies on these 'knockout' mice. In addition, such mice may serve as disease models for humans with inherited cancer predisposition syndromes. Perhaps the greatest advantage of many of the mouse tumor suppressor models is that they facilitate study of the roles of tumor suppressor gene loss in tumor initiation and progression in vivo. Moreover, derivation of primary cells from tumor suppressor-deficient mice has provided an important resource for in vitro studies on the role of targeted genes in cell cycle regulation, DNA damage response, regulation of apoptotic pathways, and preservation of genomic stability. In this review, we discuss some of the mechanistic insights provided by tumor suppressor-deficient mice and their utility as models for human cancer syndromes.     

Systemic effects of cytokines released by gene-transduced tumor cells: marked hyperplasia induced in small bowel by gamma-interferon transfectants through host lymphocytes

Cells transduced with cytokine genes are currently used to enhance the anti-tumor and immunomodulatory effects of these molecules in cancer therapy. The sustained release of cytokine thus obtained can perturb many homeostatic systems of the host. We have previously shown that the murine mammary adenocarcinoma TS/A transfected with the murine gamma-interferon (IFN-gamma) gene stimulates a strong immune response that impairs tumor growth. Mice bearing tiny tumors have serum IFN-gamma levels constantly exceeding 100 IU/ml. Therefore, we asked which systemic effects can be triggered in mice by such transfectants. BALB/c mice bearing tumors produced by clone 16.6000 cells (which release 6,000 IU/ml of IFN-gamma in culture) were compared to normal mice and to mice with tumors produced by parent cells transfected with the neomycin resistance gene (NEO cells, no IFN-gamma release). Histological studies revealed a marked hyperplasia of small bowel in mice bearing 16.6000 tumors; the villi and crypts of these mice were > 1.5 times longer than those of normal mice and of mice bearing NEO tumors. In vivo administration of bromodeoxyuridine evidenced a 2.5-3 times increase in the proliferative score of the intestinal crypts of mice bearing 16.6000 tumors compared to control mice. No intestinal alterations were observed in nude mice bearing 16.6000 tumors. T lymphocytes thus appear to play a causal role in this phenomenon.