Metastasis suppressor genes: from gene identification to protein function and regulation

In the past decade, findings from various disciplines of research have stimulated a reevaluation of fundamental concepts of the biology of metastasis. The convergence of two avenues of research has largely been responsible for this shift. First, clinical and experimental studies of specific steps of the metastatic cascade have shown that cancer cells often disseminate early in the natural history of disease and can persist at secondary sites for extended periods of time. These findings suggest that disseminated cells remain subject to growth regulation at distant sites as "dormant" single cells or microscopic metastases consisting of small numbers of cells. Second, complementary functional, biochemical, and signal transduction studies have identified a specific class of proteins that suppress the formation of overt metastases. These proteins are encoded by metastasis suppressor genes, which are operationally defined as genes that suppress in vivo metastasis without inhibiting primary tumor growth when expressed ectopically in metastatic cell lines. While metastasis suppressor proteins may affect many steps in metastatic development, recent evidence specifically implicates several of these proteins in the regulation of growth of disseminated cells at secondary sites. This review describes the evolving understanding of rate-limiting steps of metastatic growth, and the role of metastasis suppressor proteins in the regulation of these processes. We will give an overview of the studies of metastasis suppressor protein function, which have shifted our attention toward mechanisms of growth control at the secondary site (i.e., "metastatic colonization"). Emphasis is placed upon the complimentary research in the fields of metastasis and signal transduction that has identified signaling pathways controlling metastatic colonization. We also discuss the regulation of metastasis suppressor proteins and the potential biological and biochemical mechanisms responsible for their organ-type specificity. Finally, the implication of these emerging concepts on the development of therapeutic strategies will be presented.     

Keynote address: mechanisms of cellular resistance to cytotoxic drugs and X-irradiation.

Lack of response to X-irradiation and cytotoxic chemotherapy is the major cause of treatment failure in patients with cancer. However, "resistance" to these modalities may be considered a normal cellular response. The ability to cure patients with particular tumor types may be related to hypersensitivity to these modalities caused by loss or abnormality of certain normal cellular constituents such as enzymes involved in DNA repair. It is likely that the initial chemo or radiosensitivity of a tumor will broadly reflect the intrinsic resistance of the tissue type from which the tumor arose. There are many cellular biochemical mechanisms responsible for this relative resistance to drugs and radiation. Many of these mechanisms, although present in normal tissues, may be inducible and can result in enhanced resistance to DNA damaging agents. Although certain resistance mechanisms would appear to be specific for drug resistance or for radiation resistance, there are other resistance mechanisms that potentially affect both modalities. In particular, the study of DNA repair genes in mammalian cells may give us greater insight into common mechanisms of resistance to these modalities.     

The secretion of pancreatic juice in response to stimulation of the vagus nerves in the pig

1. The secretory response to stimulation of the vagus nerves has been examined in the pig and compared with that in the dog under similar experimental conditions.2. In the pig, stimulation of the vagus nerves caused a profuse flow of pancreatic juice with a high content of bicarbonate, in addition to a secretion of digestive enzymes; atropine suppressed the secretion of enzymes but failed to diminish the flow of bicarbonate-rich juice.3. Intra-arterial injections of acetylcholine closely imitated the effects of stimulation of the nerves, with the difference that both the flow of juice and the secretion of enzymes were abolished together by atropine.4. Stimulation of the nerves and injections of acetylcholine were effective after resection of the stomach and intestine: these stimuli can therefore act directly on the pancreas, independently of the release of gastro-intestinal hormones.5. In the dog, the pancreas differed from that of the pig in that stimulation of the vagus nerves and injections of acetylcholine acted predominantly on the secretion of enzymes and caused only a sparse flow of juice. Atropine annulled all these effects together.6. The results show that the vagus nerves can exert a much wider control of the secretion of pancreatic juice in the pig than in the dog. Possible mechanisms for this action are discussed.     

DNA helicase deficiencies associated with cancer predisposition and premature ageing disorders

Deficiency in a helicase of the RecQ family is found in at least three human genetic disorders associated with cancer predisposition and/or premature ageing. The RecQ helicases encoded by the BLM, WRN and RECQ4 genes are defective in Bloom's, Werner's and Rothmund-Thomson syndromes, respectively. Cells derived from individuals with these disorders in each case show inherent genomic instability. Recent studies have demonstrated direct interactions between these RecQ helicases and human nuclear proteins required for several aspects of chromosome maintenance, including p53, BRCA1, topoisomerase III, replication protein A and DNA polymerase delta. Here, we review this network of protein interactions, and the clues that they present regarding the potential roles of RecQ family members in DNA repair, replication and/or recombination pathways.     

Parental administration of chemical agents: a cause of apparent life-threatening events

Nine infants with apparent life-threatening events that occurred as a result of poisoning by a caretaker are described. These episodes were characterized by apnea plus some combination of color change, choking or gagging, and abnormal muscle tone. Five of the infants responded to vigorous stimulation, and four required cardiopulmonary resuscitation. Most poisonings (seven infants) were detected by a urine drug screen. Medications detected included acetaminophen, amphetamine, benzodiazepines (two infants), cocaine, codeine, meperidine (two infants), Methadone, phenobarbital, and phenothiazines (three infants). Four infants received two or more drugs. Most perpetrators of the poisonings were mothers (seven) and five of the parents admitted administering the various agents. Reasons for the poisonings included an apparent attempt to harm an infant, the need to sedate a fussy infant, or a gross misunderstanding of the potential risk of various agents to infants. Because no history of drug administration was elicited at the time of hospital admission, six infants might have been discharged with a diagnosis of apnea of infancy had not an attempt been made to investigate the possibility of poisoning. These cases suggest that poisoning by a caretaker should be added to the differential diagnosis of any infant brought to medical attention because of an apparent life-threatening event and that urine drug screening should be considered in the evaluation.     

Increased error-prone non homologous DNA end-joining--a proposed mechanism of chromosomal instability in Bloom's syndrome

BS is an inherited cancer predisposition disorder caused by inactivation of the RecQ family helicase, BLM. One of the defining features of cells from BS individuals is chromosomal instability, characterized by elevated sister chromatid exchanges (SCEs), as well as chromosomal breaks, deletions, and rearrangements. Although the basis for chromosomal instability is poorly understood, there is evidence that chromosomal abnormalities can arise through an alteration in the efficiency or fidelity of DNA double strand break (DSB) repair. Here, we show that BS cells demonstrate aberrant DSB repair mediated by the non-homologous end-joining (NHEJ) pathway for DNA repair, one of the two main pathways for the repair of DSBs in mammalian cells. Through a comparison of BS cell lines, and a derivative in which the BS phenotype has been reverted by expression of the BLM cDNA, we show that BS cells display aberrant end-joining of DSBs. Importantly, DNA end-joining in BS cells is highly error-prone and frequently results in DNA ligation at distant sites of microhomology, creating large DNA deletions. This aberrant repair is dependent upon the presence of the Ku70/86 heterodimer, a key component in the NHEJ pathway. We propose that aberrant NHEJ is a candidate mechanism for the generation of chromosomal instability in BS.     

Mitogen-activated protein kinase kinase 4 (MKK4) acts as a metastasis suppressor gene in human ovarian carcinoma

Despite improvements in chemotherapy and the recognition that aggressive surgical cytoreduction is beneficial, the majority of patients diagnosed with ovarian cancer will die as a result of metastatic disease. The molecular changes associated with acquisition of metastatic ability in ovarian cancer are poorly understood. We hypothesize that metastasis suppressor gene inactivation or down-regulation plays a role in ovarian cancer progression. Mitogen-activated protein kinase kinase 4 (MKK4), a member of the stress-activated protein kinase signaling cascade, has been identified recently as a metastasis-suppressor gene. An immunohistochemical approach was taken to test the possibility that MKK4 dysregulation occurs during the development of clinical ovarian cancer metastases. MKK4 expression was evaluated in normal and metastatic ovarian tissues. Normal ovarian epithelial cells showed high intensity staining for MKK4, whereas metastatic tissues showed a statistically significant decrease in expression. These results support a role for MKK4 dysregulation in the development of clinical disease. A functional approach was taken to test the ability of MKK4 to suppress metastatic colonization, the process whereby disseminated cancer cells lodge and grow at a secondary site in vivo. The SKOV3ip.1 human ovarian cancer cell line was chosen for these studies because it lacks endogenous MKK4 expression but retains both upstream and downstream components of the signaling cascade of MKK4. Ectopic expression of MKK4 in these cells, when injected into female SCID mice, suppressed the number of overt metastatic implants by nearly 90%. Furthermore, MKK4 expression increased the life span of the animals by 70%. Taken together, these data support a role for MKK4 in the suppression of metastatic colonization in ovarian cancer.