Ehud J. Margalioth

Copper and zinc levels in normal and malignant tissues.

The copper and zinc levels in 53 malignant and 47 normal human tissue samples were measured. In the malignant tissues, the mean copper concentration was 46% higher (P < 0.001) than in the normal ones. Analysis of the individual organs showed this increment to be statistically significant in malignancies of the large bowel, stomach, urinary bladder and female reproductive organs, while in cancer of the breast, kidney and testis, the increase in copper level was not significant. The mean zinc concentration in the malignant tissues was not significantly different from that in the corresponding normal tissue specimens (‐11%; P < 0.2). In breast cancer, however, tissue zinc levels were increased by 72% (P < 0.01), and decreased markedly in carcinoma of the kidney (‐73%). A hypothesis for the possible mechanism involving elevated tissue copper levels in biological damage (previously caused by free radicals) which may be responsible for the malignant process, is presented and discussed.

PGD on a recombinant allele: crossover between the TSC2 gene and 'linked' markers impairs accurate diagnosis.

Accounting for possible recombinations in developing an accurate preimplantation genetic diagnosis (PGD) protocol based on familial haplotypes.

Real-time reverse linkage using polar body analysis for preimplantation genetic diagnosis in female carriers of de novo mutations

BACKGROUNDnSingle cell diagnosis for preimplantation genetic diagnosis (PGD) requires simultaneous analysis of multiple linked polymorphic markers in addition to mutation analysis in order to reduce misdiagnosis. This type of analysis requires building family haplotypes spanning at least two generations. We present three childless couples in whom the female was a de novo mutation carrier in the Duchenne Muscular Dystrophy (DMD), incontinentia pigmenti (IKBKG) or Neurofibromatosis type 2 (NF2) genes, precluding linkage prior to the PGD cycle. We constructed haplotypes based on linked polymorphic markers in these families and performed concurrent diagnosis enabling embryo transfer from the first PGD cycle.nnnMETHODSnInformative markers flanking the DMD, IKBKG and NF2 genes were used to construct non-linked haplotypes. Polar bodies 1 (PB1) and 2 (PB2) were biopsied and analyzed to determine allelic association between the mutation and markers in multiplex PCR reactions.nnnRESULTSnFor each family, the first PGD cycle allowed the establishment of linked haplotypes based on homozygous PB1 and PB2 analysis; however, no embryos were available for transfer. Subsequent cycles, when performed, confirmed this linkage. A mutation-free child was born to the family affected with DMD and an ongoing pregnancy (32 weeks) was achieved with the carrier of the IKBKG deletion.nnnCONCLUSIONSnPB analysis for reverse linkage in real-time coupled with the PGD cycle is a powerful tool for diagnosis and linkage between markers and de novo mutations for maternal autosomal dominant or X-linked disorders. Simultaneous amplification of multiple informative markers in conjunction with the mutation allows the building of familial haplotypes and accurate PGD analysis.

Serum copper level in ovarian carcinoma.

Serum copper levels (SCL) were determined before any diagnostic procedure was performed or treatment given in 40 women admitted for the investigation of a pelvic mass and, later, in those patients with ovarian carcinoma after chemotherapy and before a second‐look operation was performed. Patients with ovarian carcinoma were found to have significantly higher SCL than patients with benign ovarian lesions. A SCL of 150 μg/dl clearly separated patients with a pelvic mass on the basis of ovarian carcinoma and those with benign noninflammatory pelvic lesions. The same SCL of 150 μg/dl separated patients with ovarian carcinoma that responded to chemotherapy and those with residual disease. It is suggested that SCL be included as a member of the screening panel of biologic tumor markers in general and in ovarian carcinoma in particular.

Treatment of oligoasthenospermia with human chorionic gonadotropin: hormonal profiles and results

Forty-seven oligoasthenospermic patients were treated with human chorionic gonadotropin, 5000 IU twice weekly for 12 to 15 weeks. Only three patients impregnated their wives following treatment. Sperm concentration subsequent to treatment decreased in 26 patients, increasing in only 8 cases. Sperm motility improved in 36% of the patients and deteriorated in 30%. During treatment, mean testosterone levels rose by 72%, mean estradiol levels rose by 140%, and mean follicle-stimulating hormone levels decreased by 80%. Mean prolactin levels remained unaltered. The disappointing results with human chorionic gonadotropin treatment fail to make it a recommendable therapy for patients with idiopathic oligoasthenospermia.

Preimplantation genetic diagnosis (PGD) for nonsyndromic deafness by polar body and blastomere biopsy

PurposeDevelopment of an efficient and reliable PGD protocol for nonsyndromic deafness, by polar body (PB) and blastomere PGD.MethodsThe GJB2/GJB6 mutations along with 12 polymorphic markers were used in PGD analysis of blastomeres or polar bodies in 14 couples for 35 cycles. Marker informativity, diagnosis rates, Allele Drop Out (ADO) rates and PB1 heterozygosity rates were assessed.ResultsSix cycles were performed by PB biopsy, 27 by blastomere and two combined cycles, resulting in delivery of three unaffected children and five ongoing pregnancies. Diagnosis rates for PB and blastomeres were similar. Only 17% PB1s were heterozygote. ADO rates of 19% were observed in both groups.ConclusionsWe have developed a single cell multiplex PGD protocol for nonsyndromic deafness with a high efficiency of diagnosis. Most PB1 are homozygous, and similar ADO rates were observed; therefore, blastomere biopsy appears to be the method of choice for this autosomal recessive disease.

Ovarian stimulation for oocyte cryopreservation for prevention of age-related fertility loss: one in five is a low responder

Abstract Oocyte cryopreservation for age-related fertility loss is gaining interest considering the tendency to postpone motherhood in many societies. Little is currently known about the actual efficiency of this approach. We aimed to explore ovarian response of presumably fertile women undergoing in vitro fertilization for this indication. A total of 105 women underwent 151 stimulation cycles at mean age 37.7u2009±u20092.4. None had known infertility. Mean daily starting FSH dose was 371u2009±u2009110 (225–600). Mean number of mature oocytes cryopreserved at the first completed cycle was 9.7u2009±u20097.5 (0–43). However, 21% of started cycles were either cancelled before egg retrieval or resulted in 0–3 mature oocytes retrieved. Therefore, women considering oocyte cryopreservation for prevention of age-related fertility decline should be encouraged to perform this procedure at younger age than, preferably before 35.

Attenuation of ovarian response by low-dose ketoconazole during superovulation in patients with polycystic ovary syndrome.

OBJECTIVEnTo investigate the clinical efficacy of mild inhibition of ovarian steroidogenesis by very low-dose ketoconazole during induction of ovulation in patients with polycystic ovary syndrome (PCOS).nnnDESIGNnProspective, randomized, cross-controlled study in consecutive cycles.nnnSETTINGnLarge tertiary care center.nnnPATIENT(S)nEighteen patients with PCOS undergoing hMG superovulation with or without ketoconazole.nnnINTERVENTION(S)nA fixed hMG dosage was initiated on cycle days 5-9 in both of the study cycles. Further hMG adjustment was done according to serum E2 levels and follicular measurements. Ketoconazole was administered in one of the cycles by two protocols.nnnMAIN OUTCOME MEASURE(S)nSerum E2 and P levels, lead follicles, pregnancy rate, and development of ovarian hyperstimulation syndrome.nnnRESULT(S)nAlthough higher daily hMG doses were needed in cycles with ketoconazole compared with cycles without the drug, the peak E2 levels were substantially lower in the ketoconazole cycles. Although the number of lead follicles did not differ between treatments, the addition of ketoconazole significantly reduced the number of hyperstimulated cycles. Consequently, the cancellation rate dropped dramatically, thus yielding a higher pregnancy rate per patient in the ketoconazole protocols.nnnCONCLUSION(S)nUse of a very low dose of ketoconazole during ovulation induction effectively attenuates ovarian steroidogenesis in patients with PCOS. This effect may serve as an adjunct to better control the ovarian response in women who are prone to hyperstimulated cycles.

Preimplantation genetic diagnosis (PGD) for a treatable disorder: Gaucher disease type 1 as a model

BACKGROUNDnPreimplantation genetic diagnosis (PGD) is a technique that enables identification of unaffected embryos prior to in vitro fertilization (IVF) transfer in couples at risk for a Mendelian disorder. Most cases involve severe genetic diseases with neurological features and/or major malformations. We present two couples in which PGD was performed for prevention of type 1 Gaucher disease, a non-neuronopathic, non-lethal disorder.nnnMATERIALS AND METHODSnWe developed a multiplex fluorescent PCR protocol, simultaneously amplifying the familial mutations and eight closely spaced, highly polymorphic informative microsatellite markers surrounding the gene, to be used for PGD analysis.nnnRESULTSnCouple #1 mother was homozygous for the N370S mutation and the father carried the 84GG mutation; their first daughter receives specific Gaucher therapy. One PGD cycle resulted in seven embryos of which four had the paternal wild type allele; two were transferred resulting in a healthy baby boy born at term. Couple #2, each a carrier (N370S and R359Q), whose first-born child had died (age 5years) of Gaucher disease, underwent 7 PGD cycles. Only one cycle resulted in a clinical pregnancy but a miscarriage was followed at 10weeks.nnnCONCLUSIONSnPGD is an effective and accurate method for preventing Gaucher disease type I in carrier couples. Since this disease is treatable, special ethical considerations and careful selection of couples should be performed.

An Immunochemical Assay for Natural IgM Antibodies with an Affinity to Galactose and Whose Titer is Reduced in the Sera of Cancer Patients

Normal sera contain an IgM fraction which binds to free galactosyl groups of agarose. This fraction causes complement-dependent lysis of Ehrlich ascites tumor cells (EATC). Cancer sera show a much reduced titer of this lytic activity. In this paper we describe an immunochemical assay for these galactophilins (GALF). Agarose columns were immunoabsorbent and bound proteins were detected by labeled antibody to human IgM. All 157 normal sera contained GALF. The serum GALF concentrations of normal sera were linearly correlated to the serum cytolytic titers. However, EATC absorbed only 15% of normal serum GALF. The titers in normal females are higher than in males. The titers in 147 nonmalignant chronic disease patients were lower than the normals only in women over 40 years of age. The sera of 138 cancer patients, taken before any therapy, were significantly lower than the corresponding normal or chronic disease controls. Lymphoproliferative neoplasms were associated with the lowest titers while melanoma sera showed normal titers. It is likely that GALF is bound to tumor associated antigens with free galactosyl groups. These are on the cell surface and/or are shed into the circulation. GALF titration may prove useful in following treatment efficacy in individual cancer patients.