Andres Chiesa-Vottero

The Role of Proteasome Inhibition With Bortezomib in the Treatment of Antibody-Mediated Rejection After Kidney-Only or Kidney-Combined Organ Transplantation

Background. We report our initial experience in using the proteasome inhibitor, bortezomib, to treat established antibody-mediated rejection (AMR) in 20 patients. Methods. There were 16 kidney-only and 4 kidney-combined organ recipients with de novo donor-specific antibody (DSA) and histologic evidence of AMR with peritubular capillaries C4d deposition. AMR was diagnosed 19.8 months (range 1-71 months) posttransplant. Patients received intravenous corticosteroids followed by a 2-week cycle on days 1-4-8-11 of plasmapheresis and 1.3 mg/m2 bortezomib; then 0.5 mg/kg intravenous immunoglobulin four times. Results. De novo class I DSA was detected in 11 (55%) and class II DSA in 18 (90%) recipients. The absolute mean difference between peak-nadir dominant DSA was 68,171 molecules of equivalent soluble fluorochrome (P<0.0001), representing 55%±22%. Only two patients (10%) had undetectable DSA after treatment. Patient survival is 100%, and graft survival is 85% with a mean follow-up of 9.8 months (range 2-20 months). The treatment was generally well tolerated but caused fatigue, gastrointestinal complaints, fluid retention, and thrombocytopenia in a number of patients. The last follow-up estimated glomerular filtration rate was 41.9±16.8 mL/min (range 20.6-72.2 mL/min). However, only 25% returned to their baseline renal function before AMR, and many have proteinuria with urine protein/creatinine more than 0.5 in 41% and more than 1.0 in 18%. Conclusions. The bortezomib-containing regimen demonstrated activity in AMR but seems to be most effective before the onset of significant renal dysfunction (serum creatinine <3 mg/dL) or proteinuria (<1 g/day). The best use of bortezomib to treat AMR should be evaluated in controlled trials using dosing strategies that include longer courses or retreatment schedules.

Immunohistochemical overexpression of p16 and p53 in uterine serous carcinoma and ovarian high-grade serous carcinoma.

Summary: The immunohistochemical expression pattern of p16 in biopsy samples has been useful as part of a panel to distinguish adenocarcinomas arising from the endometrium from those arising from the endocervix. However, no information is available on the expression of p16 in uterine serous carcinoma (USC) or ovarian high-grade serous carcinoma that could be used for diagnostic purposes. Here, we retrospectively analyzed the immunohistochemical expression of p16 in 11 cases of USC (5 pure and 6 mixed with endometrioid adenocarcinoma) and 10 cases of ovarian high-grade serous carcinoma and compared p16 expression with that of p53 in the same samples. p16 was strongly expressed by 100% of tumor cells in all 11 uterine specimens and in 5 of the 10 ovarian specimens; of the other 5 ovarian specimens, 4 showed strong positivity in 20% to 80% of tumor cells, and 1 case showed only weak expression. Positivity for p53 was strong and diffuse (100% of tumor cells) in 5 uterine tumors and in 3 ovarian tumors. p53 expression in 6 of the uterine specimens and 7 of the ovarian specimens was present in fewer tumor cells, of weak intensity, or both. We also performed human papilloma virus (HPV) DNA in situ hybridization in 4 uterine pure serous carcinomas; all 4 were negative. The negative results were confirmed by reverse transcriptase in situ polymerase chain reaction. We conclude that p16, owing to its diffuse expression in USC, should not be interpreted as indicating cervical origin or HPV-induced carcinogenesis; however, p16 may be a better marker (albeit unspecific) than p53 for identifying USC. The overexpression of p16 in USC is unrelated to HPV. Further studies are necessary to determine whether p16 expression is useful in the differential diagnosis of ovarian high-grade serous carcinoma.

Scleroderma renal crisis

OBJECTIVES To discuss the pathophysiology, risk factors, clinical manifestations, diagnosis, treatment, prevention, and outcomes of scleroderma renal crisis (SRC), a serious yet potentially treatable complication of scleroderma (systemic sclerosis). METHODS A PubMed search for articles published up until April 2014 was conducted using the following keywords: scleroderma, systemic sclerosis, scleroderma renal crisis, renal, treatment, and prognosis. Literature was carefully reviewed, and different risk factors, treatment options, prognostic factors, and survival data were assessed. RESULTS SRC occurs in about 10% of all patients with scleroderma. It is characterized by malignant hypertension and progressive renal failure. Around 10% of SRC cases may present with normal blood pressure, termed normotensive renal crisis. The etiopathogenesis is presumed to be a series of insults to the kidneys resulting in endothelial injury, intimal proliferation, and narrowing of renal arterioles leading to decreased blood flow, hyperplasia of the juxtaglomerular apparatus, hyperreninemia, and accelerated hypertension. Risk factors include rapid skin thickening, use of certain medications such corticosteroids or cyclosporine, new-onset microangiopathic hemolytic anemia and/or thrombocytopenia, cardiac complications (pericardial effusion, congestive heart failure, and/or arrhythmias), large joint contractures, and presence of anti-RNA polymerase III antibody. Since the 1970s, with the advent of angiotensin-converting enzyme (ACE) inhibitors, mortality associated with SRC decreased from 76% to <10%. Some patients may progress to end-stage renal disease and need dialysis. Renal transplantation has improved survival, though SRC may recur in transplanted kidneys. CONCLUSIONS More than 60 years after its initial description, SRC still remains an important cause of morbidity and mortality in scleroderma. Since the advent of ACE inhibitors, the prognosis of SRC has improved substantially. Prompt diagnosis and treatment may help prevent adverse outcomes and improve survival.

Comparison of Proliferation Indices in Glioblastoma Multiforme by Whole Tissue Section vs Tissue Microarray

Tissue microarrays (TMAs) reduce the amount of tissue analyzed with the assumption that protein and gene expression patterns are homogeneous throughout tumors. Many tumor types, including glioblastoma multiforme (GBM), are heterogeneous in many regards, including cell proliferation. We retrospectively compared Ki-67 labeling indices (LIs) determined by whole tissue section (WTS) vs TMA in a series of 50 GBMs from 45 patients. A paired t test indicated that the difference between average LIs obtained from a TMA vs a WTS was not significant (P = .51). There was no correlation between TMA and WTS (r = 0.042; P = .77), indicating that the methods yielded very different results in individual tumors. The Ki-67 LI did not always correlate with the tissue section in an individual tumor; however, when evaluating a large number of tumors on a TMA, the LI range and mean LI were roughly comparable with the LI range and mean LI determined from the WTS.

Primary cervical cancer screening and triage using an mRNA human papillomavirus assay and visual inspection

Objective Mexican Cervical Cancer Screening Study II (MECCS II) seeks to develop a highly sensitive and highly specific screening program able to be adapted to all socioeconomic levels in Mexico. The objectives of MECCS II are (1) to compare the sensitivity and specificity for cervical intraepithelial neoplasia (CIN) 3 or cancer of self-collected vaginal specimens tested for high-risk types of the human papillomavirus (HR-HPV) by APTIMA with those tested for HR-HPV by Hybrid Capture 2 (HC2); and (2) determine the efficacy of cryotherapy in the treatment of HR-HPV–positive and acetic acid–aided visual inspection (VIA)–positive and -negative women after VIA triage. Methods The study was conducted in rural Mexico. Women aged 30 to 50 years, nonpregnant, with no history of hysterectomy or pelvic irradiation and varied histories of screening, participated. A direct endocervical sample was tested for cytology, HC2, and APTIMA assay (AHPV). Subjects positive on any test were recalled for triage VIA, biopsies, and immediate cryotherapy. Tests were compared using McNemar test. Results Two thousand forty-nine patients have complete results. Mean age of the patients was 39.2 years; 7.7% presented with ≥atypical squamous cells of uncertain significance (ASCUS), 1.8% ≥low-grade squamous intraepithelial neoplasia, and 0.5% ≥high-grade squamous intraepithelial neoplasia. Two percent of patients had ≥CIN2, and 0.78% had ≥CIN3 (including 2 with invasive disease). The sensitivity of ThinPrep (>ASCUS), HC2, and AHPV for >CIN3 for direct endocervical collection was 87.5%, 100%, and 100%, respectively. The specificity of ThinPrep (>ASCUS), HC2, and AHPV for >CIN3 was 94.1%, 92.2%, and 93.5%, respectively. Specificities of HC2 and AHPV differed significantly. The overall percentage of agreement among HPV assays (HC2 vs APTIMA) is 97%. Four hundred sixty-nine women returned for VIA. Two hundred ninety-one women were treated with cryotherapy. Conclusions The specificity of the APTIMA assay along with high sensitivity is an advantage for primary screening. Follow-up evaluation will be important to determine the true impact of potential undertreatment in the screening algorithm. Self-sampling applications are explored.

Platelets in Early Antibody-Mediated Rejection of Renal Transplants

Antibody-mediated rejection is a major complication in renal transplantation. The pathologic manifestations of acute antibody-mediated rejection that has progressed to functional impairment of a renal transplant have been defined in clinical biopsy specimens. However, the initial stages of the process are difficult to resolve with the unavoidable variables of clinical studies. We devised a model of renal transplantation to elucidate the initial stages of humoral rejection. Kidneys were orthotopically allografted to immunodeficient mice. After perioperative inflammation subsided, donor-specific alloantibodies were passively transferred to the recipient. Within 1 hour after a single transfer of antibodies, C4d was deposited diffusely on capillaries, and von Willebrand factor released from endothelial cells coated intravascular platelet aggregates. Platelet-transported inflammatory mediators platelet factor 4 and serotonin accumulated in the graft at 100- to 1000-fold higher concentrations compared with other platelet-transported chemokines. Activated platelets that expressed P-selectin attached to vascular endothelium and macrophages. These intragraft inflammatory changes were accompanied by evidence of acute endothelial injury. Repeated transfers of alloantibodies over 1 week sustained high levels of platelet factor 4 and serotonin. Platelet depletion decreased platelet mediators and altered the accumulation of macrophages. These data indicate that platelets augment early inflammation in response to donor-specific antibodies and that platelet-derived mediators may be markers of evolving alloantibody responses.

Diagnosis of BK Viral Nephropathy in the Renal Allograft Biopsy: Role of Fluorescence in Situ Hybridization

Early recognition of BK viral nephropathy is essential for successful management. Our aim in this study was to evaluate a novel fluorescence in situ hybridization (FISH) assay for detection of BK virus in renal transplant biopsies in the context of standard detection methods. Renal allograft biopsies (n = 108) were analyzed via H&E, immunohistochemistry (IHC) for simian virus 40, and FISH for BK virus. BK virus was detected in 16 (14.8%) cases by H&E, 13 (12%) cases by IHC, 18 (16.6%) cases by FISH, and 19 (17.6%) cases by real-time PCR; 24 of 108 showed a discrepancy in ≥1 testing modalities. Comparison of H&E, IHC, and FISH showed no statistical difference in detection of BK virus. However, performing comparisons between the different tissue-based assays in the context of plasma or urine real-time PCR results showed significant improvement in detection of BK by FISH over H&E (P = 0.02) but not IHC (P = 0.07). This novel FISH-based approach for BK virus identification in renal allograft biopsy tissue mirrored real-time PCR results and showed superior performance to detection of inclusions by H&E. Therefore, use of FISH for BK virus detection in the setting of renal allograft biopsy is a useful and sensitive detection method and could be adopted in any laboratory that currently performs FISH analysis.

Association of specific PTEN/10q haplotypes with endometrial cancer phenotypes in African-American and European American women

OBJECTIVE Endometrial carcinoma (EC), the most common gynecologic malignancy in the United States, affects European American (EA) women more frequently than African-American (AA) women. Yet, AA women are more likely to die from EC. Proposed etiologies for this racial disparity, such as socioeconomic status, aggressive, non-endometrioid tumor histology, and comorbid conditions, do not account for the entire disparity experienced by AA women, suggesting an unexplored genetic component. Germline mutations in PTEN cause Cowden syndrome (CS), which increases lifetime risk of endometrial cancer. In addition, somatic PTEN silencing is one of the most common initiating events in sporadic EC. Therefore, we hypothesized that specific PTEN haplotypes in the AA population may directly predispose AA women to unfavorable tumor characteristics when diagnosed with EC. METHODS We conducted a case-control association study of germline variations in and around the PTEN/10q region between 53 EA and 51 AA EC cases and ethnic controls. RESULTS Eighteen tag SNPs with minor allele frequency ≥0.1, were genotyped and used to reconstruct haplotypes. Forty-eight ancestry informative markers were genotyped control for population stratification. Two haplotypes were overrepresented in AA, and there was a trend towards tumors with higher stage and grade in patients with these haplotypes. One haplotype was overrepresented in the EA population with a trend towards more endometrioid tumors. CONCLUSIONS We show that specific PTEN/10q haplotypes are significantly different between EA and AA individuals (p≤0.02), and specific haplotypes may increase the risk of unfavorable tumor phenotypes in AA women diagnosed with EC.

Mexican Cervical Cancer Screening Study II: 6-month and 2-year follow-up of HR-HPV women treated with cryotherapy in a low-resource setting.

Objective To determine the efficacy and tolerance of cryotherapy in a visual inspection with acetic acid (VIA) triage protocol after primary human papillomavirus (HPV) screening in a low-resource setting. Materials and Methods This continuous series conducted over 2 years enrolled nonpregnant, high-risk HPV (HR-HPV)–positive women between the ages of 30 and 50 years, who resided in the state of Michoacán, Mexico, and had a history of no Pap smear screening or knowledge of Pap smear results within the last 3 years. These women were initially enrolled in the Mexican Cervical Cancer Screening Study II (MECCS II) trial and were treated with cryotherapy after VIA triage. They subsequently followed up at 6 months and 2 years for repeat VIA, colposcopy, and biopsy. Results A total of 291 women were treated with cryotherapy, of whom 226 (78%) followed up at 6 months. Of these 226 women, 153 (68%) were HR-HPV–negative; there were no findings of cervical intraepithelial neoplasia grade 2 (CIN2) or worse. The remaining 73 women (32%) were HR-HPV–positive; of these women, 2 had CIN2 and 3 had CIN3. Only 137 women followed up at 2 years. Of these 137 women, 116 were HR-HPV–negative and 21 were HR-HPV–positive. Of the 21 women positive for HR-HPV, 9 had negative biopsy results, 11 had CIN1, and 1 had no biopsy. The clearance rate of HR-HPV was 83% (95% confidence interval: 0.78–0.87). There were no biopsy findings of CIN2 or worse at 2 years. Before cryotherapy, of the 226 women, 15 (6.6%) were positive for endocervical curettage (ECC) and 5 (2.2%) were referred for surgical management. Of these 15 ECC-positive women, 10 (67%) followed up at 6 months and it was shown that no patient was ECC positive at that time point. Moreover, of the 15 ECC-positive women, 11 (73%) followed up at 2 years and it was shown that no patient was ECC positive at that time point. In our study, VIA had a false-positive rate of 5%. Conclusions Cryotherapy was an effective, acceptable, and well-tolerated means of treating cervical dysplasia in a low-resource setting.

Ovarian Tissue Cryopreservation for Benign Gynecologic Disease: A Case of Ovarian Torsion and Review of the Literature

Ovarian tissue cryopreservation (OTC) is an emerging method for fertility preservation. Although OTC has been previously proposed for benign indications, to our knowledge this is the first report highlighting the use of OTC for the indication of ovarian torsion. A 36-year-old nulligravid woman with a history of recurrent ovarian torsion presented with an acute episode of ovarian torsion confirmed by ultrasound. She requested a laparoscopic oophorectomy because her previous oophoropexy had failed, and in light of this was counseled to undergo concurrent OTC. On laparoscopy, 10 strips of ovarian cortex were obtained. A portion of this tissue was sent for pathological analysis, which revealed a primordial follicle density of 167 follicles/mm(3), a primary follicle density of 38 follicles/mm(3), and minimal ischemic damage. Although the clinical application of OTC continues to evolve and requires further investigation, the possibility of expanding the indications for benign gynecologic conditions is promising.