Gain of CCND1 May Occur Too Infrequently in Cutaneous Melanoma, and Too Late in Melanomagenesis, to Be Diagnostically Useful: Genomic Analysis of 88 Cases.

ABSTRACT: Genomic analysis is an important tool in the diagnosis of histologically ambiguous melanocytic neoplasms. Melanomas, in contrast to nevi, are characterized by the presence of multiple copy number alterations. One such alteration is gain of the proto-oncogene CCND1 at 11q13. In melanoma, gain of CCND1 has been reported in approximately one-fifth of cases. Exact frequencies of CCND1 gain vary by melanoma subtype, ranging from 15.8% for lentigo maligna to 25.1% for acral melanoma. We present a cohort of 72 cutaneous melanomas from 2017-2022 in which only 6 (8.3%) showed evidence of CCND1 gain by chromosomal microarray. This CCND1 upregulation frequency falls well below those previously published and is significantly lower than estimated in the literature ( P < 0.05). In addition, all 6 melanomas with CCND1 gain had copy number alterations at other loci (most commonly CDKN2A loss, followed by RREB1 gain), and 5 were either thick or metastatic lesions. This suggests that CCND1 gene amplification may be a later event in melanomagenesis, long after a lesion would be borderline or equivocal by histology. Data from fluorescence in situ hybridization, performed on 16 additional cutaneous melanomas, further corroborate our findings. CCND1 gain may not be a common alteration in melanoma and likely occurs too late in melanomagenesis to be diagnostically useful. We present the largest chromosomal microarray analysis of CCND1 upregulation frequencies in cutaneous melanoma, conjecture 3 hypotheses to explain our novel observation, and discuss implications for the inclusion or exclusion of CCND1 probes in future melanoma gene panels.

A Novel Method to Detect Copy Number Variation in Melanoma: Droplet Digital PCR for Quantitation of the CDKN2A Gene, a Proof-of-Concept Study.

ABSTRACT: A definitive diagnosis of nevus or melanoma is not always possible for histologically ambiguous melanocytic neoplasms. In such cases, ancillary molecular testing can support a diagnosis of melanoma if certain chromosomal aberrations are detected. Current technologies for copy number variation (CNV) detection include chromosomal microarray analysis (CMA) and fluorescence in situ hybridization. Although CMA and fluorescence in situ hybridization are effective, their utilization can be limited by cost, turnaround time, and inaccessibility outside of large reference laboratories. Droplet digital polymerase chain reaction (ddPCR) is a rapid, automated, and relatively inexpensive technology for CNV detection. We investigated the ability of ddPCR to quantify CNV in cyclin-dependent kinase inhibitor 2A ( CDKN2A ), the most commonly deleted tumor suppressor gene in melanoma. CMA data were used as the gold standard. We analyzed 57 skin samples from 52 patients diagnosed with benign nevi, borderline lesions, primary melanomas, and metastatic melanomas. In a training cohort comprising 29 randomly selected samples, receiver operator characteristic curve analysis revealed an optimal ddPCR cutoff value of 1.73 for calling CDKN2A loss. In a validation cohort comprising the remaining 28 samples, ddPCR detected CDKN2A loss with a sensitivity and specificity of 94% and 90%, respectively. Significantly, ddPCR could also identify whether CDKN2A losses were monoallelic or biallelic. These pilot data suggest that ddPCR can detect CDKN2A deletions in melanocytic tumors with accuracy comparable with CMA. With further validation, ddPCR could provide an additional CNV assay to aid in the diagnosis of challenging melanocytic neoplasms.

Molecular genetic profiling reveals novel association between FLT3 mutation and survival in glioma.

INTRODUCTION: Recent molecular characterization of gliomas has uncovered somatic gene variation and DNA methylation changes that are associated with etiology, prognosis, and therapeutic response. Here we describe genomic profiling of gliomas assessed for associations between genetic mutations and patient outcomes, including overall survival (OS) and recurrence-free survival (RFS). METHODS: Mutations in a 50-gene cancer panel, 1p19q co-deletion, and MGMT promoter methylation (MGMT methylation) status were obtained from tumor tissue of 293 glioma patients. Multivariable regression models for overall survival (OS) and recurrence-free survival (RFS) were constructed for MGMT methylation, 1p19q co-deletion, and gene mutations controlling for age, treatment status, and WHO grade. RESULTS: Mutational profiles of gliomas significantly differed based on WHO Grade, such as high prevalence of BRAF V600E, IDH1, and PTEN mutations in WHO Grade I, II/III, and IV tumors, respectively. In multivariate regression analysis, MGMT methylation and IDH1 mutations were significantly associated with improved OS (HR = 0.44, p = 0.0004 and HR = 0.21, p = 0.007, respectively), while FLT3 and TP53 mutations were significantly associated with poorer OS (HR = 19.46, p < 0.0001 and HR = 1.67, p = 0.014, respectively). MGMT methylation and IDH1 mutations were the only significant alterations associated with improved RFS in the model (HR = 0.42, p < 0.0001 and HR = 0.37, p = 0.002, respectively). These factors were then included in a combined model, which significantly exceeded the predictive value of the base model alone (age, surgery, radiation, chemo, grade) (likelihood ratio test OS p = 1.64 × 10-8 and RFS p = 3.80 × 10-7). CONCLUSIONS: This study highlights the genomic landscape of gliomas in a single-institution cohort and identifies a novel association between FLT3 mutation and OS in gliomas.

Singleton birth at term: an old alarm or a new debate?

In 2004, Human Reproduction published a debate series focusing on the rising tide of multiple pregnancy associated with IVF. The premise of the primary report in that debate was that by considering IVF outcomes differently-by focusing on healthy singleton birth at term rather than clinical pregnancy, the standard currency at that time-the necessary shift toward reduced numbers of embryos transferred might be accelerated. The choice of end-point in that debate-Birth Emphasizing a Successful Singleton at Term (BESST)-was not an effort to 'dumb down' the complex equation linking risks and benefits. That balance is a dynamic and various mix of issues that clinicians discuss with patients on a daily basis. And BESST was certainly not proposed as a new primary outcome for application to other treatment modalities in reproductive medicine, such as ovulation induction. It was simply a responsible and brave call for change in the accelerating and competitive world of IVF.

An integrated approach to male-factor subfertility: bridging the gap between fertility specialists trained in urology and gynaecology.

Subfertile men and women are usually cared for by different clinicians, namely urologists and gynaecologists. While these doctors share each other's goals, they may not always appreciate the content or implications of their opposite number's clinical decisions; to some degree they may practice in "silos." We address this problem by reviewing the effectiveness of medical treatments for male factor subfertility in the context of female factors. The effectiveness of treatments for couples with male factor subfertility, other than IVF with ICSI, appears modest. However, data from randomized controlled trials suggest benefits from some treatments: clomiphene and tamoxifen for the male (common odds ratio for pregnancy [COR] 2.42; 95% CI 1.47 to 3.94), antioxidants (COR 4.18; 95% CI 2.65 to 6.59) and surgical management of a clinical varicocele (COR 2.39; 95% CI 1.56 to 3.66). Nevertheless, close attention to female age and the duration of subfertility help to avoid lost opportunity through delays in treatment when IVF with ICSI is indicated. Making treatment decisions squarely in the context of the couple's overall prognosis is key for optimal outcomes. Future trials of male fertility treatments should focus on pregnancy as the primary outcome, rather than less important surrogates such as sperm quality.

Les hommes et les femmes hypofertiles obtiennent habituellement leurs soins auprès de cliniciens distincts, soit des urologues et des gynécologues, respectivement. Bien que ces professionnels de la santé aient des objectifs communs, il est possible qu'ils ne comprennent pas toujours le contenu ou les implications des décisions cliniques de leurs homologues; on pourrait même en venir à affirmer qu'ils agissent de façon cloisonnée. Nous traitons de ce problème en analysant l'efficacité des traitements médicaux visant l'hypofertilité masculine dans le contexte des facteurs féminins. À l'exception de l'utilisation concomitante de la FIV et de l'IICS, l'efficacité des traitements offerts aux couples qui font face à une hypofertilité attribuable à des causes imputables à l'homme semble modeste. Toutefois, des données issues d'essais comparatifs randomisés semblent indiquer que certains traitements offrent des avantages : clomiphène et tamoxifène administrés à l'homme (rapport de cotes commun pour ce qui est de la grossesse [RCC], 2,42; IC à 95 %, 1,47 - 3,94), antioxydants (RCC, 4,18; IC à 95 %, 2,65 - 6,59) et prise en charge chirurgicale d'une varicocèle clinique (RCC, 2,39; IC à 95 %, 1,56 - 3,66). Quoi qu'il en soit, le fait de bien porter attention à l'âge de la femme et à la durée de l'hypofertilité aide à éviter les occasions manquées en raison de délais dans la mise en œuvre du traitement, dans les cas où l'utilisation concomitante de la FIV et de l'IICS s'avère indiquée. Le fait de prendre des décisions en s'assurant de tenir absolument compte du pronostic global du couple est d'une importance capitale pour l'obtention d'issues optimales. L'obtention d'une grossesse (et non des critères auxiliaires moins importants, tels que la qualité des spermatozoïdes) devrait constituer le critère d'évaluation principal des futurs essais visant les traitements contre l'infertilité masculine.

Validation and Implementation of a Somatic-Only Tumor Exome for Routine Clinical Application.

Next-generation sequencing-based genomic testing is standard of care for tumor workflows. However, its application across different institutions continues to be challenging given the diversity of needs and resource availability among different institutions globally. Moreover, the use of a variety of different panels, including those from a few individual genes to those involving hundreds of genes, results in a relatively skewed distribution of care for patients. It is imperative to obtain a higher level of standardization without having to be restricted to specific kits or requiring repeated validations, which are generally expensive. We show the validation and clinical implementation of the DH-CancerSeq assay, a tumor-only whole-exome-based sequencing assay with integrated informatics, while providing similar input requirements, sensitivity, and specificity to a previously validated targeted gene panel and maintaining similar turnaround times for patient care.

Chondrolipoma of the Breast: A Myofibroblastoma Variant or a Distinct Lesion?

The entity commonly referred to as chondrolipoma is a rare and enigmatic breast lesion with unclear histogenesis and a complete lack of molecular characterization. It is uncertain whether it represents a hamartoma, choristoma, or a distinct neoplasm, including possibly a variant of mammary-type myofibroblastoma. We report two additional chondrolipomatous lesions of the breast. The lesions had varying histologic and immunohistochemical features similar to myofibroblastoma, including the loss of retinoblastoma (Rb) protein expression in one lesion. Molecular analysis by chromosomal microarray analysis performed on a second lesion did not demonstrate a loss of 13q14 or 16q typical of myofibroblastoma. Our findings further support the concept that at least a subset of breast lesions that historically have been classified as chondrolipoma are related to myofibroblastoma. However, the lack of myofibroblastoma-specific molecular alterations in one lesion suggests chondrolipomas may also have varying origins.

A Droplet Digital Polymerase Chain Reaction-Based Tool to Aid in Melanoma Diagnosis.

CONTEXT.­: Detecting copy number variations (CNVs) at certain loci can aid in the diagnosis of histologically ambiguous melanocytic neoplasms. Droplet digital polymerase chain reaction (ddPCR) is a rapid, automated, and inexpensive method for CNV detection in other cancers, but not yet melanoma. OBJECTIVE.­: To evaluate the performance of a 4-gene ddPCR panel that simultaneously tests for ras responsive binding element protein 1 (RREB1) gain; cyclin-dependent kinase inhibitor 2A (CDKN2A) loss; MYC proto-oncogene, bHLH transcription factor (MYC) gain; and MYB proto-oncogene, transcription factor (MYB) loss in melanocytic neoplasms. DESIGN.­: One hundred sixty-four formalin-fixed, paraffin-embedded skin samples were used to develop the assay, of which 65 were used to evaluate its performance. Chromosomal microarray analysis (CMA) data were used as the gold standard. RESULTS.­: ddPCR demonstrated high concordance with CMA in detecting RREB1 gain (sensitivity, 86.7%; specificity, 88.9%), CDKN2A loss (sensitivity, 80%; specificity, 100%), MYC gain (sensitivity, 70%; specificity, 100%), and MYB loss (sensitivity, 71.4%; specificity, 100%). When one CNV was required to designate the test as positive, the 4-gene ddPCR panel distinguished nevi from melanomas with a sensitivity of 78.4% and a specificity of 71.4%. For reference, CMA had a sensitivity of 86.2% and a specificity of 78.6%. Our data also revealed interesting relationships with histology, namely (1) a positive correlation between RREB1 ddPCR copy number and degree of tumor progression; (2) a statistically significant correlation between MYC gain and nodular growth; and (3) a statistically significant correlation between MYB loss and a sheetlike pattern of growth. CONCLUSIONS.­: With further validation, ddPCR may aid both in our understanding of melanomagenesis and in the diagnosis of challenging melanocytic neoplasms.

A pilot study assessing art therapy as a mental health intervention for subfertile women.

BACKGROUND Subfertility is a common but hidden source of anxiety, depressive symptoms and hopelessness. Counselling reduces this emotional burden and may even enhance the likelihood of pregnancy. Art therapy may be a useful intervention, because it facilitates the expression of feelings, both visually and verbally, and may ease emotional distress. METHODS Weekly 2-h art therapy group courses were held for a total of 21 subfertile women. The impact of subfertile women's support systems and barriers to coping were all explored. The effectiveness of art therapy was assessed using Beck Hopelessness, Depression and Anxiety Inventories, administered before and after participation, as well as a qualitative exit questionnaire. RESULTS The mean age of participants was 35.7 (SD 2.1) years and duration of infertility was 64 (12.0) months. Mean Beck Hopelessness Scale fell from 6.1 (3.8) to 3.5 (3.1, P = 0.01) after therapy. Beck Depression Inventory-II Score fell from 19.8 (11.0) to 12.5 (10.2, P = 0.01) and Beck Anxiety Inventory Score changed from 12.4 (8.4) to 8.4 (5.2, P = 0.3). Based on the exit questionnaire, women felt the course was insightful, powerful and enjoyable. CONCLUSIONS Art therapy is an inexpensive, non-pharmacological intervention, which was associated with decreased levels of hopelessness and depressed mood in subfertile women. It also provides insight into the meaning and emotional implications of subfertility for patients and caregivers. This pilot study highlights the need for further research in this field.

Using evidence from randomized trials in fertility practice.

Randomized controlled trials (RCTs) are central to the understanding of treatment effectiveness and diagnostic test utility. If they are to be relied upon in clinical practice, data from trials should have three main attributes: validity (be free from bias); clinical relevance (be based on patients similar to your own, reporting outcomes that matter to them); and importance (demonstrate effect sizes that are large enough to justify the costs and risks entailed). With these principles in mind, this brief article reviews key questions to pose while deciding whether new evidence from RCTs should influence subfertility patient care.

Cross-border reproductive care in North America: a pilot study testing a prospective data collection program for in vitro fertilization clinics in Canada and the United States.

OBJECTIVE: To develop and test a nonidentifying prospective data collection system for cross-border reproductive care (CBRC) in Canada and the United States (U.S.). DESIGN: Survey and cross-sectional study. SETTING: Fertility clinics. PATIENT(S): Women traveling to and from Canada and the U.S. for reproductive care. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Patients' home country, reason for crossing borders, and type of care received. RESULT(S): Of 32 Canadian and 440 U.S. clinics contacted, seven and 46, respectively, responded to the initial questionnaire. Three out of seven Canadian and 44 out of 46 U.S. clinics reported providing CBRC. Seventy five percent agreed that nonidentifying data on country of origin and reason for travel should be collected. However, only one of seven Canadian and none of 46 U.S. clinics that expressed initial interest actually collected data, despite multiple communications. CONCLUSION(S): Although CBRC is a major component of assisted reproductive technology in North America (3%-10% of IVF cycles are provided to out-of-country patients in Canada and the U.S.), clinicians are not motivated to collect the simplest of data regarding CBRC patients. Despite this, reliable data are needed to help better understand the reasons for and impact of CBRC.

Are health care providers adequately educating couples for embryo disposition decisions?

OBJECTIVE: To determine the effectiveness of education provided by health care professionals during and after IVF treatment in preparing couples for surplus embryo disposition decisions. DESIGN: Cross-sectional survey. SETTING: University-affiliated fertility center. PATIENT(S): Couples with embryos cryopreserved for more than 2 years. INTERVENTION(S): Self-administered questionnaire. MAIN OUTCOME MEASURE(S): A Likert scale was used to rate the response to questions about patients' preparedness to make decisions regarding their surplus embryos. RESULT(S): The survey response rate was 70% (131 of 187). Education provided by health care professionals before initiating treatment met the needs of the majority of participants (n = 86). After treatment, the education received was not adequate to assist couples in making embryo disposition decisions. Of the 127 respondents who provided feedback on their intention for their cryopreserved embryos, 37% (n = 47) had no intention of using cryopreserved embryos for their own reproduction, 24% (n = 30) intended to use embryos for procreation, and the remaining 39% (n = 50) remained undecided regarding their future use of their embryos. Participants with more than 3 years of infertility (n = 49) were most likely to feel conflicted about the decision after treatment. CONCLUSION(S): The education received after treatment was considered inadequate. Couples with a long duration of infertility and those conflicted about final embryo disposition may be appropriate targets for further intervention. More written information and/or counseling services after treatment may help patients make informed and timely decisions regarding their surplus embryos.

Randomized clinical trials: the meeting place of medical practice and clinical research.

Randomized clinical trials (RCTs) have been used to assess interventions affecting health since biblical times. They provide the most valid means of measuring the true effects of intervention compared with no treatment or placebo. Although they can also be used to assess the value of diagnostic tests, this article focuses on randomized trials in the context of treatment. Key elements of an RCT include: the explicit definition of a clinically relevant question; the identification of an appropriate sample of patients; a clearly defined and reproducible intervention; an appropriate intervention and comparator; clinically relevant, measurable outcomes; and appropriate tools for measurement and analysis. It is also essential to establish that the question posed is ethical, the methods of study are valid, and that follow-up is complete, with an "intent to treat" analysis. Results are best presented using both proportions and absolute numbers. By providing clinical decision-makers with numbers needed to treat or harm, decisions may be better informed and easier to understand, than if proportions alone are used.

The diagnosis and management of ovarian hyperstimulation syndrome: No. 268, November 2011.

OBJECTIVE: To review the clinical aspects of ovarian hyperstimulation syndrome and provide recommendations on its diagnosis and clinical management. OUTCOMES: These guidelines will assist in the early recognition and management of ovarian hyperstimulation. Early recognition and prompt systematic supportive care will help avert poor outcomes. EVIDENCE: Medline, Embase, and the Cochrane database were searched for relevant articles, using the key words "ovarian hyperstimulation syndrome" and "gonadotropins," and guidelines created by other professional societies were reviewed. VALUES: The quality of evidence was rated using the criteria described in the Report of the Canadian Task Force on Preventive Health Care. Recommendations for practice were ranked according to the method described in that report (Table 1).

Art therapy as a healing tool for sub-fertile women.

Although fertility is fundamental to spiritual health, it is often taken for granted. When a desired pregnancy fails to occur, stress and grief frequently follow. Visual expression of feelings through "art therapy" has proved a powerful healing tool for women brave enough to give it a try at the McMaster University Fertility Clinic. The objective and subjective findings of this ongoing project suggest that through simple visual self-expression, stress, anxiety and hopelessness may be reduced. This form of art therapy also provides a joyful social experience of sharing with other women, who are dealing with these issues.

Cross-border fertility services in North America: a survey of Canadian and American providers.

OBJECTIVE: To identify the scope and volume of cross-border fertility services in Canada and the U.S. and to evaluate the three-way communication between patients and their service providers in 2008. DESIGN: Mail and on-line surveys of cross-border fertility care activity were sent to 34 Canadian and 392 American fertility clinics and clinicians. MAIN OUTCOME MEASURE(S): Clinician and patient experience with assisted reproductive technologies. RESULT(S): The most commonly reported cross-border treatment sought by Canadians was anonymous donor-oocyte in vitro fertilization (IVF; 363 out of 452, 80%). For patients entering Canada to receive fertility treatment, the largest demand was for IVF (106 out of 146, 73%). The majority of out-of-country patients received by U.S. clinics sought standard IVF (927 out of 1,809, 51%), most of these coming from Europe (25%) and Latin America (39%). The largest proportion of patients leaving the U.S. to receive IVF (41%) or donor-egg IVF (52%) traveled to India/Asia. Concurrence was seen between Canadian and U.S. clinics' ratings of key data that should be provided along with returning patients. Experience of earlier patients with individual centers and perceived safety and effectiveness of care are the key factors in choice of destination. CONCLUSION(S): Anonymous donor-oocyte IVF is the main assisted reproductive technology sought by Canadians traveling to the U.S. India and Asia are the main destinations for U.S. women leaving the country for their fertility care. Three-way communication between patients and sending and receiving clinics is an important element of safe and effective care.

Dual positive and negative regulation of wingless signaling by adenomatous polyposis coli.

The evolutionarily conserved Wnt/Wingless signal transduction pathway directs cell proliferation, cell fate, and cell death during development in metazoans and is inappropriately activated in several types of cancer. The majority of colorectal carcinomas contain truncating mutations in the adenomatous polyposis coli (APC) tumor suppressor, a negative regulator of Wnt/Wingless signaling. Here, we demonstrate that Drosophila Apc homologs also have an activating role in both physiological and ectopic Wingless signaling. The Apc amino terminus is important for its activating function, whereas the beta-catenin binding sites are dispensable. Apc likely promotes Wingless transduction through down-regulation of Axin, a negative regulator of Wingless signaling. Given the evolutionary conservation of APC in Wnt signal transduction, an activating role may also be present in vertebrates with relevance to development and cancer.