Residual ctDNA after treatment predicts early relapse in patients with early-stage non-small cell lung cancer.

BACKGROUND: Identification of residual disease in patients with localized non-small cell lung cancer (NSCLC) following treatment with curative intent holds promise to identify patients at risk of relapse. New methods can detect circulating tumour DNA (ctDNA) in plasma to fractional concentrations as low as a few parts per million, and clinical evidence is required to inform their use. PATIENTS AND METHODS: We analyzed 363 serial plasma samples from 88 patients with early-stage NSCLC (48.9%/28.4%/22.7% at stage I/II/III), predominantly adenocarcinomas (62.5%), treated with curative intent by surgery (n = 61), surgery and adjuvant chemotherapy/radiotherapy (n = 8), or chemoradiotherapy (n = 19). Tumour exome sequencing identified somatic mutations and plasma was analyzed using patient-specific RaDaR™ assays with up to 48 amplicons targeting tumour-specific variants unique to each patient. RESULTS: ctDNA was detected before treatment in 24%, 77% and 87% of patients with stage I, II and III disease, respectively, and in 26% of all longitudinal samples. The median tumour fraction detected was 0.042%, with 63% of samples <0.1% and 36% of samples <0.01%. ctDNA detection had clinical specificity >98.5% and preceded clinical detection of recurrence of the primary tumour by a median of 212.5 days. ctDNA was detected after treatment in 18/28 (64.3%) of patients who had clinical recurrence of their primary tumour. Detection within the landmark timepoint 2 weeks to 4 months after treatment end occurred in 17% of patients, and was associated with shorter recurrence-free survival [hazard ratio (HR): 14.8, P <0.00001] and overall survival (HR: 5.48, P <0.0003). ctDNA was detected 1-3 days after surgery in 25% of patients yet was not associated with disease recurrence. Detection before treatment was associated with shorter overall survival and recurrence-free survival (HR: 2.97 and 3.14, P values 0.01 and 0.003, respectively). CONCLUSIONS: ctDNA detection after initial treatment of patients with early-stage NSCLC using sensitive patient-specific assays has potential to identify patients who may benefit from further therapeutic intervention.

Whom are we treating with adaptive servo-ventilation? A clinical post hoc analysis.

BACKGROUND: Recent evidence has shown that adaptive servo-ventilation (ASV) is contraindicated in patients with predominant central sleep apnea (CSA) and reduced left ventricular ejection fraction (LVEF ≤45%). The objective of this study was to assess the clinical usage of ASV in patients at the time-point of the release of a safety warning by type of SDB, breathing pattern and LVEF. METHODS: Patients of a cardiac and a respirology sleep center, both in Germany, who received ASV therapy were contacted between May and October 2015. Retrospective analyses included diagnostic polysomnography, polysomnography with continuous positive airway pressure prior to ASV initiation and echocardiography. Treatment emergent CSA was diagnosed after an appropriate treatment period on CPAP. RESULTS: 285 patients receiving ASV therapy (91 in the cardiac and 194 in the respirology setting) underwent diagnostic polysomnography. 233 (82%) patients had severe SDB, 94 (33%) predominant CSA, and 185 (65%) periodic breathing. 20% (n = 52) of patients had an LVEF of ≤45%. The most common indications for ASV were CSA in heart failure (41%) in the cardiac setting and treatment emergent CSA (80%) diagnosed after an appropriate period on CPAP in the respirology setting. The proportion of patients in whom ASV was contraindicated (CSA and LVEF ≤45%) was 16% in the cardiac setting and 9% in the respirology setting. CONCLUSION: Clinical usage of ASV changed for a small subgroup of patients after release of the SERVE-HF results. Nevertheless, ASV treatment should be monitored and evaluated with diligence in the reminder indications.

Novel therapeutic approach to improve hematopoiesis in low risk MDS by targeting MDSCs with the Fc-engineered CD33 antibody BI 836858.

We recently reported that the accumulation of myeloid-derived suppressor cells (MDSC), defined as CD33+HLA-DR-Lin-, has a direct role in the pathogenesis of myelodysplastic syndrome (MDS). In particular, CD33 is strongly expressed in MDSC isolated from patients with MDS where it has an important role in MDSC-mediated hematopoietic suppressive function through its activation by S100A9. Therefore, we tested whether blocking this interaction with a fully human, Fc-engineered monoclonal antibody against CD33 (BI 836858) suppresses CD33-mediated signal transduction and improves the bone marrow microenvironment in MDS. We observed that BI 836858 can reduce MDSC by antibody-dependent cellular cytotoxicity, which correlated with increases in granule mobilization and cell death. BI 836858 can also block CD33 downstream signaling preventing immune-suppressive cytokine secretion, which correlates with a significant increase in the formation of CFU-GM and BFU-E colonies. Activation of the CD33 pathway can cause reactive oxygen species (ROS)-induced genomic instability but BI 836858 reduced both ROS and the levels of double strand breaks and adducts (measured by comet assay and γH2AX). This work provides the ground for the development of a novel group of therapies for MDS aimed at MDSC and their disease-promoting properties with the goal of improving hematopoiesis in patients.

Isoform expression of CD44 adhesion molecules, Bcl-2, p53 and Ki-67 proteins in lung cancer.

CD44, belongs to the cell adhesion molecule family and is expressed on cell surfaces in several isoforms which are generated by alternative splicing of messenger RNA. These splice variants have been shown in several cancer cell types and are thought to be involved in tumor progression. The aim of the current study was to evaluate the expression of selected CD44 variants on lung cancer cells of various histology and to compare these with other markers of tumor spread. Surgical samples of primary lung carcinoma of various histology were subjected to alkaline phosphatase-anti-alkaline phosphatase complex immunohistochemistry using a panel of monoclonal antibodies: anti-CD44 v5, v6, v7/8, v10, anti-Ki-67, anti-Bcl-2 and anti-p53. Positive cells were scored in a semiquantitative way. The patients were subdivided into groups with and without metastases, as found during surgery. All CD44 variants tested could be demonstrated on lung cancer cells, but the incidence of particular isoforms varied, depending on lung cancer histology. In general, CD44 expression was highest in squamous cell tumors and lowest in anaplastic small cell carcinomas. Squamous cell cancers had high expression of v5 and v6 variants, while in anaplastic large cell and small cell carcinomas v10 was abundant. When Ki-67, Bcl-2 and p53 protein expression was compared to the incidence of CD44 variants, coincidence was found for v10 only. Most of the cases positive for v10 were also Ki-67 positive (p = 0.0146). In 12 cases with metastases, tumor cells had high v6 and Ki-67 expression, but these data were not significant compared to cases without metastases. Overall, these data suggest that v5 and v6 variants are of significance in squamous cell lung carcinoma, presumably in the promotion of metastasis, while in anaplastic small cell or large cell cancers only v10 expression seems to correlate with proteins associated with tumor growth and progression.

Safety and biodistribution of 99mTechnetium-labeled anti-CD44v6 monoclonal antibody BIWA 1 in head and neck cancer patients.

The CD44 protein family consists of isoforms, encoded by standard exons and up to nine alternatively spliced variant exons (v2-v10), which are expressed in a tissue-specific way. Expression of v6-containing variants (CD44v6) has been related to aggressive behavior of various tumor types and was shown to be particularly high in squamous cell carcinoma (SCC). Therefore, CD44v6 might be a suitable target for radioimmunoscintigraphy (RIS) and therapy. The present study evaluates the novel high-affinity murine anti-CD44v6 monoclonal antibody (MAb) BIWA 1 for its safety and targeting potential in patients with SCC of the head and neck (HNSCC). Twelve HNSCC patients, who had planned to undergo resection of the primary tumor and neck dissection, were included. Preoperatively, 2, 12, or 52 mg of 99nTc-labeled MAb BIWA 1 was administered. RIS results obtained 21 h after injection were compared with palpation, computed tomography, and magnetic resonance imaging, with histopathology as the gold standard. Moreover, biodistribution of BIWA 1 was evaluated by radioactivity measurement in blood and bone marrow and in biopsies from the surgical specimen obtained 40 h after injection. The distribution of BIWA 1 in tumor biopsies was analyzed by immunohistochemistry. BIWA 1 integrity in the blood was assessed by high-performance liquid chromatography and related to soluble CD44v6 levels in serum samples. No drug-related adverse events were observed. Human antimouse antibody responses were observed in 11 patients. The diagnostic efficacy of RIS appeared to be comparable for the three BIWA 1 dose levels and for the four diagnostic methods. Besides activity uptake in tumor tissue, minimal accumulation of activity was observed in mouth, lungs, spleen, kidney, bone marrow, and scrotal area. Analysis of tissue biopsies revealed high uptake in tumors, with a mean value of 14.2+/-8.4% of the injected dose/kg tumor tissue and a mean tumor:blood ratio of 2.0+/-1.4 at 40 h after injection. Differences among the three dose groups were not statistically significant, although a trend toward lower uptake in the highest dose group was noted. Distribution of BIWA 1 throughout the tumor was heterogeneous for all dose groups, which might be related to the high affinity of the MAb. The mean biological half-life in blood (34.5+/-6.1 h) was not dose dependent. Extensive complex formation of BIWA 1 was observed in the 2-mg group, most probably with soluble CD44v6 present in the blood, and complex formation relatively diminished upon increase of the MAb dose. BIWA 1 is a promising MAb for targeting HNSCC because it can be safely administered to HNSCC patients, while it shows high and selective tumor uptake. However, BIWA 1 is immunogenic, and therefore a chimerized or humanized derivative of BIWA 1 with intermediate affinity will be used in future clinical trials.

Evaluation of soluble CD44v6 as a potential serum marker for head and neck squamous cell carcinoma.

In recent years, the measurement of soluble CD44 levels in the circulation of patients with malignant diseases has been introduced as a new and simple diagnostic tool for the detection of human cancer. The high CD44v6 expression in head and neck squamous cell carcinoma (HNSCC) would enable the use of soluble CD44v6 proteins present in the circulation of HNSCC patients as a marker of disease. In the present study, we determined CD44v6 plasma levels using a domain-specific ELISA in healthy volunteers, non-cancer patients, and HNSCC patients before and after surgical removal of the tumor. A difference between the CD44v6 plasma levels of HNSCC patients and controls could not be observed. Moreover, surgical removal of the tumor did not result in a reduction of the CD44v6 plasma level in the HNSCC patients. In addition, the spectrum of soluble v6-containing CD44 proteins present in the plasma of HNSCC patients and controls was determined by immunoprecipitation experiments, but again, tumor-related isoforms could not be distinguished in patient samples. Additional experiments to unravel the biological source of these circulating proteins indicated surprisingly that the v6-containing proteins present in the circulation of healthy individuals are only released in part, if at all, by activated lymphocytes or other nucleated blood cells. Most circulating CD44v6 proteins seem to be derived from the normal epithelial cell compartments, including breast cells, colon cells, and squamous cells. Taken together, these data do not support the use of soluble CD44v6 as a tumor marker in HNSCC or any other tumor type that has developed from tissues producing soluble isoforms.

Plant-derived acetophenones with antiasthmatic and anti-inflammatory properties: inhibitory effects on chemotaxis, right angle light scatter and actin polymerization of polymorphonuclear granulocytes.

3-Methoxy- and 2-hydroxy-3-methyl substituted 4-hydroxyacetophenones and some of the corresponding 4-O-glucosides revealed inhibitory activity on LTB4-, fMLP- and PAF-induced chemotaxis of polymorphonuclear granulocytes (PMN). Blocking of PMN myeloperoxidase (MPO), which is required for the intracellular activation of acetophenones, caused a loss of activity. By in vitro activation of apocynin (1) with human MPO and H2O2 and subsequent chromatographic purification an apocynin-free fraction was obtained, which contained the major metabolite. The fraction showed strong anti-chemotactic activity, which was largely preserved despite of MPO-blockade. The influences on right angle light scatter and actin polymerization in PMN demonstrate the inhibitory effects of acetophenones on the cytoskeletal rearrangement after stimulation with fMLP. Acetosyringenin (2) was shown to reduce mastoparan-induced PMN migration. The inhibition of receptor independent motility of PMN suggests that the active metabolites of acetophenones may interfere with the post receptor signalling.

Expression of CD44 isoforms on isolated bone marrow plasma cells and peripheral CD19+ B cells of patients with multiple myeloma and healthy individuals.

The expression of certain isoforms of CD44 was shown to correlate with aggressiveness and metastatic potential of various tumour types. We analysed the expression of the adhesion molecule CD44 and its variant domains (v6, v7, v7/8, v10) on isolated bone marrow (BM) plasma cells and peripheral blood (PBL) CD19+ B cells of 21 patients with MM and 15 healthy donors. B cells and plasma cells were isolated by immunomagnetic sorting and analysed by two-colour flow cytometry. The expression of CD44 isoforms was significantly higher on PBL B cells of patients with MM than in healthy controls. The elevated expression of CD44 isoforms (v6, v7/8, v10) on PBL B cells correlated with reduced overall survival in MM. CD44 isoforms were more strongly expressed on "larger", activated B cells. Furthermore, CD44 isoforms were found to be simultaneously expressed with CD38hi and CD56 on both, B lymphocytes and plasma cells of patients with MM. The determination of CD44 isoforms on circulating B cells may be helpful in defining prognostically unfavourable subgroups in MM.

Polycation-based DNA complexes for tumor-targeted gene delivery in vivo.

BACKGROUND: Efficient and target-specific in vivo gene delivery is a major challenge in gene therapy. Compared to cell culture application, in vivo gene delivery faces a variety of additional obstacles such as anatomical size constraints, interactions with biological fluids and extracellular matrix, and binding to a broad variety of non-target cell types. METHODS: Polycation-based vectors, including adenovirus-enhanced transferrinfection (AVET) and transferrin-polyethylenimine (Tf-PEI), were tested for gene delivery into subcutaneously growing tumors after local and systemic application. DNA biodistribution and reporter gene expression was measured in the major organs and in the tumor. RESULTS: Gene transfer after intratumoral application was 10-100 fold more efficient with Tf-PEI/DNA or AVET complexes in comparison to naked DNA. Targeted gene delivery into subcutaneously growing tumors after systemic application was achieved using electroneutral AVET complexes and sterically stabilized PEGylated Tf-PEI/DNA complexes, whereas application of positively charged polycation/DNA complexes resulted in predominant gene expression in the lungs and was associated by considerable toxicity. CONCLUSION: For systemic application, the physical and colloidal parameters of the transfection complexes, such as particle size, stability, and surface charge, determine DNA biodistribution, toxicity, and transfection efficacy. By controlling these parameters, DNA biodistribution and gene expression can be targeted to different organs.

Poor diagnostic value of colonic CD44v6 expression and serum concentrations of its soluble form in the differentiation of ulcerative colitis from Crohn's disease.

BACKGROUND: Increased expression of CD44v6 on colonic crypt epithelial cells in ulcerative colitis has been suggested as a diagnostic tool to distinguish ulcerative colitis from colonic Crohn's disease. AIMS: To investigate colonic CD44v6 expression and serum concentrations of soluble CD44v6 (sCD44v6) in patients with ulcerative colitis and Crohn's disease. METHODS: Colonic biopsy samples were obtained from 16 patients with ulcerative colitis, 13 with ileocolonic Crohn's disease, and 10 undergoing polypectomy. Serum samples were obtained from 15 patients with active ulcerative colitis, 20 with active Crohn's disease, and 20 healthy donors. Colonic CD44v6 expression was evaluated immunohistochemically by monoclonal antibody 2F10 and the higher affinity monoclonal antibody VFF18. Serum sCD44v6 concentrations were measured by ELISA. RESULTS: 2F10 stained colonic epithelium of inflamed ulcerative colitis and Crohn's disease samples in 80% and 40% of cases, respectively, and VFF18 in 95% and 87%, respectively. Both monoclonal antibodies displayed a sensitivity and specificity of 60% and 87% to differentiate ulcerative colitis from colonic Crohn's disease. Serum concentrations of sCD44v6 were lower in patients with ulcerative colitis (median 153 ng/ml; interquartile range (IQR) 122-211) compared with Crohn's disease (219; IQR 180-243) and healthy donors (221; IQR 197-241 (p = 0.002)). Its sensitivity and specificity to discriminate ulcerative colitis from Crohn's disease was 75% and 71%, respectively. CONCLUSION: Colonic CD44v6 and serum sCD44v6 concentrations do not facilitate reliable differential diagnosis between ulcerative colitis and Crohn's disease.

Comparison of CD44 expression in early colorectal carcinomas of the de novo and ex adenoma types.

Small colorectal carcinomas without morphological evidence of origin from an adenoma have been called "de novo" carcinomas. As changes in the expression of the adhesion molecule CD44 and its variants have been described along the adenoma-carcinoma sequence in colorectal carcinoma, we compared patterns of CD44 expression in early de novo and ex-adenoma colorectal carcinomas by staining specimens from a group of early (pT1) colorectal carcinomas by immunohistochemistry for CD44 (standard and variant forms v3, v5, v6, v7, v7/8, v10). We evaluated carcinoma, adenoma (ex-adenoma cases), transitional mucosal areas and apparently nonneoplastic mucosa peripheral to the lesions (when present). A marked increase was seen in numbers and intensity of standard and variant forms of CD44 in carcinomatous areas compared with nonneoplastic mucosa in both groups, with no significant difference between the groups. However, adenoma areas of the ex-adenoma cases and the transitional mucosa of the de novo carcinomas had nearly identical staining patterns. Together with data from other molecular studies, this may be interpreted as evidence for an adenoma-type precursor lesion in so-called de novo colorectal carcinomas.

TGFbeta signaling is necessary for carcinoma cell invasiveness and metastasis.

BACKGROUND: Invasive growth of epithelial tumor cells, a major cause of death from cancer in humans, involves loss of epithelial polarity and dedifferentiation. Transforming growth factor beta (TGFbeta) is regarded as a major tumor suppressor during early tumor development because it inhibits cell-cycle progression and tumor growth. Many dedifferentiated, late-stage tumors are resistant to growth inhibition by TGFbeta, however, and even secrete TGFbeta. In line with this, TGFbeta is involved in angiogenesis, wound healing and epithelial-mesenchymal transition (EMT) during development. Ha-Ras-transformed mammary epithelial cells (EpRas) undergo TGFbeta-induced EMT maintained via a TGFbeta autocrine loop. Thus, we have analyzed whether signal transduction by the TGFbeta receptor (TGFbetaR) is required for local tumor cell invasion and metastasis. RESULTS: A dominant-negative type II TGFbetaR (TGFbetaRII-dn) was expressed using retroviral vectors in EpRas cells and highly metastatic mesenchymal mouse colon carcinoma cells (CT26). In both cell types, TGFbetaRII-dn blocked TGFbetaR signaling and heavily delayed tumor formation. In EpRas cells, TGFbetaRII-dn prevented EMT. In the dedifferentiated mesenchymal CT26 cells, TGFbetaRII-dn caused mesenchymal-to-epithelial transition and inhibited their in vitro invasiveness in several assays. In addition, TGFbetaRII-dn completely abolished metastasis formation by CT26 cells. Furthermore, several human carcinoma lines lost in vitro invasiveness when treated with neutralizing TGFbeta antibodies or soluble receptor variants. Finally, human colon carcinoma cells (hnPCC) expressing a mutated, non-functional TGFbetaRII were non-invasive in vitro, a defect restored by re-expressing wild-type TGFbetaRII. CONCLUSIONS: Cell-autonomous TGFbeta signaling is required for both induction and maintenance of in vitro invasiveness and metastasis during late-stage tumorigenesis. TGFbetaRII therefore represents a potential target for therapeutical intervention in human tumorigenesis.

CD44 and variants in melanocytic skin neoplasms.

Expression of cell surface molecules that mediate cell-matrix and cell-cell interactions largely contributes to the ability of melanoma cells to migrate and spread beyond the primary site of the tumor. CD44, the principal cell-surface receptor for hyaluronate, and its numerous splice variants have been reported to play a crucial role in invasion and the metastatic process of different human neoplasms, including primary malignant melanoma (PMM). The aim of this study was to clarify which isoforms of CD44 (standard CD44 and CD44 variants) are distributed in PMM with a vertical tumor thickness of >1.4 mm. Staining of CD44 standard (CD44s) and splice variants was further examined for diagnostic and prognostic relevance in a panel of melanocytic skin lesions. Ten cases of PMM with Breslow >1.4 mm were analysed by immunohistochemistry using monoclonal antibodies specific for CD44s and the splice variants v3, v5, v6, v7, v7-8, and v10. In addition, using anti-CD44s, v5, and v6 antibodies, 55 melanocytic lesions, including dermal nevi (n=12), Clark nevi (dysplastic nevi) (CN; n=11), melanoma in situ (Mis; n=8), PMM (n=18), and cutaneous metastasis of malignant melanoma (cMMM; n=6) were assessed. Staining intensities were scored visually and evaluated by means of a staining index. In ten cases of PMM with a Breslow index >1.4 mm positive staining was ascertained for CD44s, v5 and for v6 in three cases. No staining was found for v3, v7, v7-8, and v10. Examination of CD44s, v5, and v6 in 55 melanocytic skin lesions revealed a high index for CD44s in all specimens and a weak staining of v5 in Mis; dermal nevi and CN did not stain for v5. However, in PMM and cMMM we found v5 to be strongly positive. The isoform v6 showed a variable index only in PMM, but without connection to established prognostic criteria. We conclude that CD44s and splice variants can not be regarded as indicators for tumor progression in malignant melanomas. However, v5 may potentially serve as a diagnostic marker for melanocytic skin lesions.

Expression of CD44 splice variant v10 in Hodgkin's disease is associated with aggressive behaviour and high risk of relapse.

Expression of CD44 isoforms has been shown to correlate with the progression and prognosis of some malignant tumours. The aim of this study was to investigate the expression of CD44 standard (CD44s) and CD44 splice variants (CD44v) v5, v6, and v10 in lymph node specimens from patients with nodular sclerosing Hodgkin's disease (NSHD), with or without initial bone marrow involvement and with or without relapse. Specimens were studied by immunohistochemistry to determine CD44s and CD44v in Hodgkin- and Reed-Sternberg (HRS) cells. For validation of the immunohistochemical of detection of CD44v10 in paraffin-embedded samples, selected cases were analysed in parallel immunohistochemically using fresh frozen material and by reverse transcription-polymerase chain reaction (RT-PCR). There was high expression of CD44 isoforms containing the variant exon v10 selectively in HRS cells of patients with relapse within 2-3 years or with initial bone marrow involvement. In patients without relapse, however, no or only very few HRS cells were positive. These differences were statistically highly significant (p < or = 0.001), whereas evaluation of CD44s, CD44v5, and v6 expression revealed no marked differences. It is concluded that evaluation of CD44v10 expression could serve as a new prognostic marker in NSHD. These results are considered to be of sufficient importance to initiate a large multi-institutional study for confirmation; furthermore, they might suggest causal involvement of CD44v10 in the progression of NSHD.

Expression and prognostic value of the CD44 splicing variants v5 and v6 in gastric cancer.

In the present study, the expression and prognostic role of the CD44 splicing variants v5 and v6 were immunohistochemically investigated in 418 curatively resected gastric carcinomas. CD44v5 was expressed in 65.3 per cent (n = 273) and CD44v6 in 77.0 per cent (n = 322) of the tumours. Whereas the expression of CD44v5 was correlated with advanced pT categories, with lymph node involvement, and with the presence of blood and lymphatic vessel invasion, such a correlation could not be found for the variant v6. As shown by univariate analysis, patients with CD44v5-positive tumours had a significantly shorter overall survival than patients with CD44v5-negative tumours (P = 0.049). In contrast, expression of CD44v6 had no impact on prognosis (P = 0.574). In a multivariate analysis including the prognostic parameters pT category and pN category, as well as blood and lymphatic vessel invasion, the prognostic impact of CD44v5 expression could not, however, be maintained. Although in the present study the expression of CD44v5 was correlated with a more aggressive tumour type, these data suggest that neither CD44v5 nor CD44v6 can predict survival in patients with gastric cancer, nor is their expression a suitable tool for identifying subgroups of patients who may be at higher risk.

Spontaneous malignant craniopharyngioma in an albino rat.

Craniopharyngioma is a rare neoplasm in the rat, and few cases have been described. These lesions are thought to originate from squamous cell remnants of Rathke's pouch, an evagination of primitive stomatodeum. This neoplasm is usually locally invasive, and neither cranial nor extracranial metastases have been described. A spontaneously occurring malignant, metastasizing craniopharyngioma arising from the neurohypophysis was detected in a 2-year-old male albino rat. The infiltrative growth was observed in the wall of the vessels of the circle of Willis, in the perivascular space of Virchow and Robin, in the submeningeal space near the hypothalamus, through the fissura chorioidea, in the medulla oblongata, and along the optic nerve into the periocular region. Metastases were detected in the thalamus and hippocampus. The diagnosis was made on the basis of microscopic, immunocytochemical and ultrastructural findings.

Characterization of a high-affinity monoclonal antibody specific for CD44v6 as candidate for immunotherapy of squamous cell carcinomas.

Variant isoforms of CD44, a family of cell-surface glycoproteins generated by alternative splicing and post-translational modifications, are expressed in a variety of human tumors and play important roles in tumor progression and metastasis formation. The murine monoclonal IgG1 antibody VFF18, specific for an epitope encoded by human CD44 variant exon 6, binds with high affinity to the recombinant protein (Kd = 1.7 x 10(-10) M) as well as to tumor cell lines in vitro, and is suitable for immunohistochemical analysis of human tumors. Screening of more than 500 tumor samples of different histogenesis showed that VFF18 most strongly and uniformly reacts with squamous cell carcinomas (SCC). Detailed analysis of 185 SCC (head and neck, lung, skin) confirmed reactivity of the antibody with 99% of the samples, with intense and homogeneous staining of the tumor cells in the majority of cases, whereas reactivity of VFF18 with normal tissues is limited to certain epithelia and activated lymphocytes. When radiolabelled VFF18 was administered to nude mice bearing human epidermoid carcinoma (A-431) xenograft, fast and selective tumor uptake of the radioimmunoconjugate with a maximum of 18% of the injected dose per gram of tissue was observed. Taken together, these data suggest that mAb VFF18 is a promising targeting vehicle for radioimmunotherapy of squamous cell carcinomas in humans.

Expression of CD44 isoforms in human skin cancer.

In animal models, isoforms of CD44 (CD44v) containing sequences encoded by one or several of ten different exons (v1-v10) contribute to tumour metastasis. In certain human cancers, CD44v6 expression is associated with poor prognosis. This paper examines CD44v expression in skin carcinogenesis and skin cancer metastasis. CD44v expression was studied in basal cell carcinoma (BCC), squamous cell carcinoma (SCC), primary malignant melanoma (PMM), metastases of MM (MMM), benign melanocytic naevi (BMN) and normal skin (NS) by immunohistochemistry and reverse transcript polymerase chain reaction (RT-PCR). BCC, SCC and NS expressed several CD44v, including v6, albeit in different distributions and intensities. PMM, MMM and BMN expressed isoforms containing v7/8 and v10, but failed to express epitopes encoded by v5 or v6. Thus, different CD44 isoforms are found in human skin cancers and are modulated during carcinogenesis. However, we did not observe a correlation of CD44v6 expression with metastatic potential.