Bortezomib appears to overcome the poor prognosis conferred by chromosome 13 deletion in phase 2 and 3 trials.

In multiple myeloma, deletion of chromosome 13 (del(13)) is associated with poor prognosis regardless of treatment. This study analyzed the impact of del(13) status on response and survival following treatment with either bortezomib or high-dose dexamethasone in patients in the SUMMIT and APEX trials. Additionally, matched-pairs subset analyses were conducted of patients with and without del(13), balanced for age and International Staging System parameters. In both SUMMIT and APEX, prognosis appeared to be poorer in bortezomib-treated patients with del(13) compared with patients with no del(13) by metaphase cytogenetics. In the SUMMIT and APEX matched-pairs analysis, response and survival appeared comparable in bortezomib-treated patients with or without del(13) by metaphase cytogenetics. However, patients with del(13) receiving dexamethasone in APEX appeared to have markedly decreased survival compared with those without del(13) by metaphase cytogenetics. These matched-pairs analyses suggest that bortezomib may overcome some of the poor impact of del(13) as an independent prognostic factor. However, sample sizes were very small; these findings require confirmation from further studies.

Cytogenetic analysis of melanocytes from premalignant nevi and melanomas.

We karyotypically analyzed cultured melanocytes from a variety of lesions, including congenital and dysplastic nevi, primary melanoma, and metastatic melanoma. The cells derived from congenital nevi had normal karyotypes, as did 22 of the 26 cultures derived from dysplastic nevi. The karyotypes of melanocytes from primary and metastatic melanomas were all abnormal. The only chromosome change in common between the nevi with abnormal karyotypes and the melanomas was the loss of one copy of chromosome 9 (two of four nevi and four of 11 melanomas, including three from the same patient) or the loss of the short arm of chromosome 9, especially of region 9pter-p22 (three of 11 melanomas). We suggest that deletion of a gene or genes on 9p, possibly interferon genes, is an initial step in the malignant transformation of melanocytes.

Cytogenetic evidence of the multistep origin of head and neck squamous cell carcinomas.

BACKGROUND: Head and neck squamous cell carcinomas are associated with tobacco and alcohol use; therefore, the incidence of this type of tumor is expected to rise in the future as a result of the increasing numbers of female and adolescent smokers. Previous reports of cytogenetic analysis of this type of tumor have implicated a number of chromosomal regions in recurring changes, but no clear pattern of characteristic changes has emerged. PURPOSE: We have undertaken cytogenetic analysis of 10 cell lines which were established from squamous cell carcinomas of the head and neck, to determine the possible sites of additional tumor suppressor genes and oncogenes that may contribute to malignant transformation. METHODS: Metaphases were harvested from cultures of cells in the exponential growth phase, following exposure to Colcemid (demecolcine) at a final concentration of 30 ng/mL for 5 hours. Air-dried slides were G-banded using trypsin and Giemsa. Fifteen metaphases were photographed and fully karyotyped. RESULTS: We observed that several chromosomal regions were lost at high frequency, including 18q (10 of 10 lines), 10p (eight of 10 lines), 3p (six of 10 lines), 8p (seven of 10 lines), and the short arms of the acrocentric chromosomes (seven of 10 lines). Nine of 10 lines had additional copies of 7p. We also noted clustering of breakpoints in a number of chromosome bands, including 1p22, 10q11.2, 11q13, and the short arms of the acrocentric chromosomes. CONCLUSION: The observation of loss of multiple chromosomal regions in a significant number of lines analyzed is consistent with the theory that tumorigenesis occurs as the result of the accumulation of a number of genetic alterations, as proposed for colorectal carcinoma. The high frequency with which these changes are seen suggests that genes located in these regions have a role in the etiology of this type of tumor.

Chromosome changes characterizing in vitro response to radiation in human squamous cell carcinoma lines.

We have analyzed the karyotypes of a series of 16 cell lines derived from human squamous cell carcinomas of the head and neck. When the cell lines were grouped according to their in vitro response to radiation, it was observed that some recurrent chromosomal changes occurred with differing frequencies between groups. Radiation resistance was associated with clusters of breakpoints in bands 1p22, 3p21, and 8p11.2 and deletion of distal 14q, while relative radiation sensitivity was associated with a high frequency of breakpoints in 11q13 and duplication of distal 14q. The regions identified point to the possible locations of genes involved in the response to radiation and could provide a series of markers with which to predict response to radiation therapy.

The role for NES1 serine protease as a novel tumor suppressor.

Previously (Liu et al, Cancer Res., 56: 3371-3379, 1996), we isolated a novel serine protease-like gene--Normal Epithelial Cell Specific-1 (NES1)--that is expressed in normal mammary epithelial cells but is down-regulated in most breast cancer cell lines. Here, we demonstrate that stable expression of NES1 in the NES1-negative MDA-MB-231 breast cancer cell line suppressed the oncogenicity as revealed by inhibition of the anchorage-independent growth and tumor formation in nude mice. Fluorescence in situ hybridization localized the NES1 gene to chromosome 19q13.3, a region that contains genes for related proteases (including the prostate-specific antigen) and is rearranged in human cancers. Similar to breast cancer cell lines, prostate cancer cell lines also lacked NES1 mRNA and protein expression. Together, these results strongly suggest a tumor-suppressor role for NES1 in breast and prostate cancer.

Molecular analysis of deletions of the short arm of chromosome 9 in human gliomas.

Previous studies have suggested that structural abnormalities involving the short arm of chromosome 9 are frequently associated with gliomas. The alpha-, beta-, and omega-interferon (IFNA, IFNB1, and IFNW, respectively) and the methylthioadenosine phosphorylase (MTAP) genes have been mapped to the short arm of chromosome 9, band p22. Homozygous deletions of these genes have been reported in many leukemia- and glioma-derived cell lines. In this report, we present a detailed analysis of partial and complete homozygous or hemizygous deletions of DNA sequences on 9p in human cell lines and primary tumor samples of glioma patients. Ten of 15 (67%) glioma-derived cell lines had hemizygous or homozygous deletion of IFN genes or rearrangement of sequences around these genes, while 13 of 35 (37%) primary glioma tumor samples had hemizygous (8 tumors) or homozygous (5 tumors) deletion of the IFN genes. The shortest region of overlap of these deletions maps in the interval between the centromeric end of the IFN gene cluster and the MTAP gene. In the cell lines and primary tumors examined, these gross genomic alterations were seen only in association with high grade or recurrent gliomas. Our observations confirm that loss of DNA sequences on 9p, particularly the IFN genes, occurs at a significant frequency in gliomas, and may represent an important step in the progression of these tumors. These results are consistent with a model of tumorigenesis in which the development or progression of cancer involves the loss or inactivation of a gene or several genes that normally act to suppress tumorigenesis. One such gene may be located on 9p; this gene may be closely linked to the IFN genes. Nevertheless, loss of the IFN genes, when it occurs, may play an additional role in the progression of these tumors.

Prolonged N-myc protein half-life in a neuroblastoma cell line lacking N-myc amplification.

Genomic amplification of the oncogene N-myc is associated with rapid tumor progression and poor prognosis in patients with neuroblastoma (NB). However, 40% of NBs which lack N-myc amplification are also clinically aggressive. Factors other than N-myc copy number must therefore play a role in determining tumor progression in these NBs. We have established an unusual human NB cell line (NBL-S) from the primary tumor of a patient with rapidly progressive disease which lacks N-myc amplification. The doubling time in vitro (48 h) and the time from injection of 2 x 10(7) cells to detectable tumors in nude mice (46 days) in similar to NB cell lines with amplified N-myc. However, karyotype analysis reveals no evidence of double minutes (DMs), homogeneously staining regions (HSRs), or chromosome 1p deletions, features commonly seen in NB cell lines. The cells have the cell surface phenotype typical of N-myc amplified NB (HLA-A,B,C negative and HSAN 1.2 positive), and similar to other NB cell lines, N-myc RNA and protein are expressed. Interestingly, the half-life of the N-myc protein in NBL-S is prolonged (approximately 100 min) compared to the short N-myc protein half-life previously described in N-myc amplified NB cell lines (approximately 30 min). Because N-myc protein is thought to have a regulatory role, prolongation of the half-life of this protein may be an important factor in the regulation of growth in NBs which lack N-myc amplification and rapidly progress.

Cytogenetic and immunohistochemical analysis of an adult anaplastic neuroblastoma.

Neuroblastomas in children are common tumors and are characterized by a number of recurrent cytogenetic and molecular changes. Adult neuroblastomas are rare, and their relationship to pediatric neuroblastomas is not clear. We report an anaplastic neuroblastoma presenting in a 28-year-old man. Histopathologic identification of the tumor as a neuroblastoma was problematic, and the initial diagnosis was poorly differentiated sarcoma. Tumor cells expressed immunoreactivity for tyrosine hydroxylase in addition to generic neuroendocrine markers, consistent with catecholamine-synthesizing ability. They also extended long, branching neurites in vitro. The tumor was positive for immunoreactive trkA. The karyotype after 6 days in culture was found to be 42,XY with multiple chromosomal abnormalities. The only abnormality shared with pediatric neuroblastomas was a rearrangement of chromosome 17q. Double minute chromosomes or homogeneously staining regions associated with N-myc amplification were not present. To our knowledge, this is the first reported karyotype of an adult neuroblastoma. The cytogenetic findings, together with expression of trkA, suggest that the tumor was more closely related to the favorable prognosis neuroblastomas of infancy than to the poor prognosis tumors that occur in older children, despite its unfavorable histology.

Physical findings in 21q22 deletion suggest critical region for 21q- phenotype in q22.

Multiple abnormalities were observed in a newborn infant with a deletion in the long arm of chromosome 21, from band 22q22.1-->qter. The phenotype of this infant was similar to that previously described in infants with deletions spanning the long arm of chromosome 21, from the centromere to 21q22 [Rethoré et al., 1972, Exp Cell Res 70:455-456, 1973, Ann Genet (Paris) 16:271-275]. However, as a phenotypically normal child with normal intelligence and with deletion of 21q11.1-21q21.3 has also been identified [Korenberg et al., 1991, Hum Genet 87:112-118], this case suggests that the critical region of deletion for the 21q- phenotype lies distal to 21q21, within 21q22.1-22.2.

SC phocomelia syndrome, premature centromere separation, and congenital cranial nerve paralysis in two sisters, one with malignant melanoma.

Two middle age sisters had most manifestations of the SC phocomelia syndrome including postnatal growth retardation, symmetric limb deficiencies with radial aplasia and absent thumbs, facial anomalies with microcephaly, microphthalmia, hypoplastic nasal alae, and borderline to mild mental retardation. Unusual findings included congenital paralysis of some cranial nerves in both patients and malignant melanoma in the proposita. Cultured lymphocytes from both patients, and skin fibroblasts, Epstein Barr virus-transformed lymphocytes, and tumor cells from the proposita demonstrated premature separation of centromeric heterochromatin (PCS) of many chromosomes, a finding noted previously in the SC phocomelia syndrome and the similar but more severe Roberts syndrome. Extensive overlap of the phenotypes of the sisters and 15 other patients with either syndrome and PCS confirms that these are either allelic conditions or the same disease--designated Roberts-SC phocomelia syndrome. The role of PCS in the syndrome(s) remains uncertain since some patients with the characteristic clinical phenotypes are reported to lack it.

Symmetrical chromosome rearrangements in cell lines established from human radiation-induced sarcomas.

We report the establishment of cell lines from six radiation-induced tumors, three osteosarcomas, and three soft tissue sarcomas. Cytogenetic analysis of the lines showed mostly balanced rearrangements in the osteosarcomas; the soft tissue sarcomas had more unstable karyotypes. The rearrangements observed were unlike those previously reported for spontaneously occurring tumors of the same histological type and were more typical of the long-term changes observed after radiation exposure.

The involvement of 6p in melanoma.

Karyotype analysis of first passage cells from a melanoma, occurring in a patient with a giant congenital nevus, revealed monosomy of chromosomes #6 and #11, and trisomy of chromosomes #8 and #22. Five marker chromosomes were present, including two ring chromosomes. The origin of three of the marker chromosomes was determined, and all were found to contain the region 6p21.1-6p23. The ring chromosomes were also thought to contain this region, to which the major histocompatibility locus has been mapped. The significance of these findings is discussed in the light of recent reports of chromosome studies of melanoma cells.

Cytogenetic and molecular genetic studies of a patient with atypical lymphoid hyperplasia.

We have karyotyped cells from a lymph node of a patient with atypical lymphoid hyperplasia. Among other clonal chromosomal abnormalities, a t(2;19) translocation was observed with breakpoints at 2p11.2 and 19q13. The genes for transforming growth factor alpha and beta have been mapped to 2p11-p13 and 19q13, respectively, but Southern blot analysis did not reveal any alteration in the structure of these genes. Similarly, the kappa immunoglobulin gene, which maps to 2p11-p12 was not rearranged. In addition, Southern blot analysis using immunoglobulin and T-cell receptor genes as probes, did not demonstrate any clonality of either B or T cells. We propose that this patient represents an early, polyclonal stage of atypical hyperplasia. The chromosome changes observed may have been one of the etiologic factors causing this disorder.

Dominant transmission of imperforate hymen.

OBJECTIVE: Imperforate hymen is an uncommon anomaly of the reproductive tract, occurring in approximately 0.1% of newborn females. The familial occurrence of imperforate hymen in a child, her mother, and her mother's monozygotic twin is reported. DESIGN: Case report. SETTING: Academic medical center. PATIENT(S): Three affected family members. MAIN OUTCOME MEASURE(S): Karyotype and pedigree analysis. RESULT(S): The proband, presenting with peritonitis, was evaluated at age 12 for imperforate hymen because this condition was diagnosed in her mother at age 14. At age 14, the mother's monozygotic twin was asymptomatic except for primary amenorrhea and was also demonstrated to have imperforate hymen. No other reproductive system abnormalities were known to be present in the remaining family members. Chromosomal structural analysis confirmed that the mother of the proband had no chromosomal abnormalities. CONCLUSION(S): The occurrence of imperforate hymen in two consecutive generations of a family is consistent with a dominant mode of transmission, either sex-linked or autosomal. Previously reported examples of siblings with imperforate hymen suggested a recessive mode of inheritance. Taken together, these cases suggest that imperforate hymen can be caused by mutations in several genes. This case highlights the importance of evaluating all family members of affected patients. Familial examples of other developmental anomalies of the female reproductive tract also suggest a multifactorial genetic etiology.

The interpretation of electromyographic responses to electrical stimulation of the motor cortex in diseases of the upper motor neurone.

The complexities of interpreting results of electrical stimulation of the motor cortex in pathological states are discussed and illustrated by reference to results from a variety of patients with diseases affecting the upper motor neurone (multiple sclerosis, cervical spondylosis and myelopathy, motor neurone disease, hemiparesis due to cerebral infarction, and hereditary spastic paraplegia). The abnormalities of the electromyographic (EMG) responses after anodal cortical stimulation consisted of delay in the latency to onset, dispersion or reduction in response size or even absence of EMG responses. These changes were not confined to any specific condition or pathology. Previous work has suggested that the sequence of events that follow anodal cortical stimulation involves repetitive excitatory inputs to spinal motoneurones and transmission across at least one central synapse. Accordingly, delayed latencies may not exclusively indicate slowing of motor conduction, while the absence of any response may not indicate complete failure of conduction in corticomotoneurone pathways.

Cytogenetic analysis in melanoma and nevi.

We have presented data from the cytogenetic analysis of melanocytes derived from four dysplastic nevi and 14 melanomas. We have confirmed previous results from melanomas, including the loss of chromosome 6q and duplication of 1q and 7p. We did not observe an excess of rearrangements involving chromosomes 2 and 3, but we found frequent deletion of chromosomes 9p, 10p, 10q, and 11q. We observed the loss of chromosome 9 in 2 of 4 dysplastic nevi, and combining this observation with that from the melanomas suggests that deletion or inactivation of a gene on 9p may be a primary event in melanocyte transformation. Other tumor suppressor genes are likely to be involved in the transformation process, and these are most likely located on 10p, 10q, 11q, and 6q. We observed many rearrangement involving chromosome band 1p13 and suggest that activation of a gene in that band may also contribute to the transformation process.

An in vitro study of the effects of acetaldehyde on Intoxilyzer 4011AS-A results.

A series of practical experiments were conducted to determine whether physiologic concentrations of acetaldehyde could produce an apparent ethanol concentration in breath alcohol analyses using the Intoxilyzer 4011AS-A. Aqueous ethanol:acetaldehyde standards were prepared and analyzed following the same protocol used for testing individuals arrested for driving while intoxicated. The average response of the Intoxilyzer to concentrations of acetaldehyde vapor are reported. Results clearly demonstrate that physiologic breath concentrations of acetaldehyde will not significantly affect the results of a breath alcohol test when an Intoxilyzer 4011AS-A is used for the analysis.

Determination of volume of distribution for ethanol in male and female subjects.

Ten nonalcoholic subjects gave written informed consent. Six men (aged 25-43) and four women (aged 25-35) were hydrostatically weighed to determine their percentage of body fat and lean weight. Each subject fasted for at least 10 h and then received an oral dose of alcohol (0.9 g per kilogram of lead body weight) calculated to yield a peak alcohol concentration of 0.100 g/210-L breath. Breath alcohol measurements were conducted at 20-min intervals until each subject's alcohol concentration returned to 0.000 g/210-L breath. All alcohol analyses were conducted on the Intoxilyzer 5000 and reported as g/210-L breath. Female subjects on average reached a lower peak alcohol concentration (mean, 0.086; range, 0.074-0.091 g/210 L) than male subjects (mean, 0.096; range, 0.093-0.101 g/210 L). Females demonstrated a higher average rate of elimination (mean, 0.017; range, 0.014-0.021 g/210 L) than males (mean, 0.015; range, 0.013-0.017 g/210 L). Female subjects on average had a higher percentage of body fat (mean, 26.0; range, 16.7-36.8%) than males (mean, 18.0; range, 10.2-25.3%). The average volume of distribution (Vd), as calculated from percentage of body fat, for the women (mean, 0.63; range, 0.54-0.71) was less than for the men (mean, 0.69; range, 0.63-0.76). The average Vd as calculated from linear regression of the alcohol concentration curve, for the women (mean 0.64, range, 0.56-0.71) was also less than for the men (mean, 0.72; range, 0.67-0.77). The data from this limited study indicate that hydrostatic weighing is an acceptable way of determining Vd for both men and women.

Cytogenetics in head and neck cancer.

Cytogenetic analysis of cell lines has identified a number of chromosomal regions that are likely to contain the genes important in malignant transformation in HNSCC. Further work needs to be done comparing results from cells grown in culture with tumors from which they are derived to determine the applicability of cytogenetic analysis. Preliminary results suggest that some of the chromosome changes observed may correlate with a cell's response to radiation and as such could provide a tool for evaluating a patient's response to planned therapy.

The response of the Intoxilyzer 4011AS-A to a number of possible interfering substances.

Five Intoxilyzer 4011AS-As were tested for their response to eleven chemicals and one mixture of chemicals. The air/water partition ratios were also determined for these eleven chemicals and one mixture. The chemicals tested and their approximate partition ratios were the following: acetaldehyde (190:1), acetone (341:1), acetonitrile (578:1), isoprene (1:1), isopropanol (1671:1), methanol (3229:1), methylene chloride (11:1), methyl ethyl ketone (229:1), toluene (5.5:1), 1,1,1-trichloroethane (14:1), trichloroethylene (20:1), and a 50:50 mixture of 1,1,1-trichloroethane and trichloroethylene (14:1). Of the eleven chemicals and one mixture studied during this experiment, only three, isopropanol, toluene, and methyl ethyl ketone, could reasonably interfere with the test, and then only under unusual circumstances--those circumstances being a slight additive effect to a breath ethanol concentration near the level required for prosecution. Any substantial additive effect from these three substances would illuminate the interference light which invalidates the test. The mean illumination point of the interference light was 0.0286 g/210 L for methyl ethyl ketone, 0.0294 for toluene, and between 0.0116 and 0.0292 for the apparent alcohol concentration for isopropanol, depending on the amount of isopropanol metabolized to acetone. Even with these unusual circumstances considered, the Intoxilyzer 4011AS-A must be viewed as an effective way of determining the ethanol concentration in human breath for evidential purposes.