Best practices for the management of local-regional recurrent chordoma: a position paper by the Chordoma Global Consensus Group.

Chordomas are rare, malignant bone tumors of the skull-base and axial skeleton. Until recently, there was no consensus among experts regarding appropriate clinical management of chordoma, resulting in inconsistent care and suboptimal outcomes for many patients. To address this shortcoming, the European Society of Medical Oncology (ESMO) and the Chordoma Foundation, the global chordoma patient advocacy group, convened a multi-disciplinary group of chordoma specialists to define by consensus evidence-based best practices for the optimal approach to chordoma. In January 2015, the first recommendations of this group were published, covering the management of primary and metastatic chordomas. Additional evidence and further discussion were needed to develop recommendations about the management of local-regional failures. Thus, ESMO and CF convened a second consensus group meeting in November 2015 to address the treatment of locally relapsed chordoma. This meeting involved over 60 specialists from Europe, the United States and Japan with expertise in treatment of patients with chordoma. The consensus achieved during that meeting is the subject of the present publication and complements the recommendations of the first position paper.

Mechanism of action of trabectedin in desmoplastic small round cell tumor cells.

BACKGROUND: Desmoplastic small round cell tumor (DSRCT) is a rare and highly aggressive disease, that can be described as a member of the family of small round blue cell tumors. The molecular diagnostic marker is the t(11;22)(p13;q12) translocation, which creates an aberrant transcription factor, EWS-WT1, that underlies the oncogenesis of DSRCT. Current treatments are not very effective so new active drugs are needed. Trabectedin, now used as a single agent for the treatment of soft tissue sarcoma, was reported to be active in some pre-treated DSRCT patients. Using JN-DSRCT-1, a cell line derived from DSRCT expressing the EWS-WT1 fusion protein, we investigated the ability of trabectedin to modify the function of the chimeric protein, as in other sarcomas expressing fusion proteins. After detailed characterization of the EWS-WT1 transcripts structure, we investigated the mode of action of trabectedin, looking at the expression and function of the oncogenic chimera. METHODS: We characterized JN-DSRCT-1 cells using cellular approaches (FISH, Clonogenicity assay) and molecular approaches (Sanger sequencing, ChIP, GEP). RESULTS: JN-DSRCT-1 cells were sensitive to trabectedin at nanomolar concentrations. The cell line expresses different variants of EWS-WT1, some already identified in patients. EWS-WT1 mRNA expression was affected by trabectedin and chimeric protein binding on its target gene promoters was reduced. Expression profiling indicated that trabectedin affects the expression of genes involved in cell proliferation and apoptosis. CONCLUSIONS: The JN-DSRCT-1 cell line, in vitro, is sensitive to trabectedin: after drug exposure, EWS-WT1 chimera expression decreases as well as binding on its target promoters. Probably the heterogeneity of chimera transcripts is an obstacle to precisely defining the molecular mode of action of drugs, calling for further cellular models of DSRCT, possibly growing in vivo too, to mimic the biological complexity of this disease.

Circulating miR-378 in plasma: a reliable, haemolysis-independent biomarker for colorectal cancer.

BACKGROUND: Plasma circulating tumour-specific microRNAs (miRNAs) are promising biomarkers of tumour presence and recurrence, especially for diseases whose best chance of successful treatment requires early diagnosis and timely surgery of an already malignant but not yet invasive tumour, such as colorectal cancer (CRC). METHODS: Expression levels of miRNAs previously found to be differently expressed in tumour vs normal colon tissues were investigated by quantitative real-time PCR in plasma from CRC patients and from healthy donors and confirmed in independent case control series. The validated miRNAs were also measured after surgery. Analyses were repeated on the subsets of haemolysis-free samples. RESULTS: We identified four miRNAs differently expressed between the compared groups, two (miR-21 and miR-378) of which were validated. miR-378 expression decreased in non-relapsed patients 4-6 months after surgery and miR-378 ability to discriminate CRC patients from healthy individuals was not influenced by haemolysis levels of plasma samples. CONCLUSION: The miRNA analysis on plasma samples represents a useful non-invasive tool to assess CRC presence as well as tumour-free status at follow-up. Plasma levels of miR-378 could be used to discriminate CRC patients from healthy individuals, irrespective of the level of haemoglobin of plasma samples.

Adaptive immune contexture at the tumour site and downmodulation of circulating myeloid-derived suppressor cells in the response of solitary fibrous tumour patients to anti-angiogenic therapy.

BACKGROUND: Host immunity is emerging as a key player in the prognosis and response to treatment of cancer patients. However, the impact of the immune system and its modulation by therapies are unknown in rare soft tissue sarcomas such as solitary fibrous tumours (SFTs), whose management in the advanced forms includes anti-angiogenic therapy. Here, we studied the in situ and systemic immune status of advanced SFT patients and the effects of sunitinib malate (SM) in association with the clinical efficacy. METHODS: Immune contexture of SFTs was assessed by immunohistochemistry in lesions from untreated or SM-treated patients. Frequency of circulating myeloid-derived suppressor cells (MDSCs), regulatory T cells (Tregs) and T-cell functions was assessed ex vivo in SFT patients prior and during anti-angiogenic therapy. Patients with long-term tumour control were included to correlate immune profiles and clinical responses. RESULTS: Anti-angiogenic naïve SFT lesions were heavily infiltrated by CD163(+)CD14(+)CD68(-) and CD163(+)CD14(-)CD68(-) myeloid cells but devoid of T cells. Conversely, post-SM tumours acquired a new subset of CD68(+)CD14(+) myeloid cells and displayed traits of an on-going adaptive immunity, strongly enriched in activated CD8(+) and CD4(+) T cells. These changes at the tumour site paralleled the alleviation of systemic immunosuppression and the drop in the frequency of circulating monocytic MDSCs (mMDSCs) and granulocytic MDSCs (gMDSCs). Rebound in the number of mMDSCs, but not of gMDSCs occurred at disease progression, and a reduced percentages of mMDSCs, comparable to those found in healthy donors (HDs), endured only in the SM-responsive patients. CONCLUSIONS: The immune contexture of SFT patients is heavily involved in anti-angiogenic therapy and it could be exploited to achieve more durable disease control through immune-based combination strategies.

Treatment-related outcome of oropharyngeal cancer patients differentiated by HPV dictated risk profile: a tertiary cancer centre series analysis.

BACKGROUND: To date, no treatment modality has been identified as more effective for oropharyngeal cancer (OPC), and no predictive factors are known to guide treatment decision for this disease. This retrospective study evaluates the differential effects of diverse treatment options for OPC according to patient risk profiles. PATIENTS AND METHODS: We considered two series of locally advanced squamous cell OPC patients treated with either surgery followed by radiotherapy (surgical series) or chemoradiation (CRT) with/without induction docetaxel, cisplatin and 5-fluorouracil (TPF) chemotherapy (CRT series). Smoking habits, tumor p16 expression/human papillomavirus (HPV) status and T and N stage were analyzed to stratify the patients according to Ang's risk profile (low, intermediate and high risk). Overall survival (OS) and disease-free survival were calculated with the Kaplan-Meier method. RESULTS: Globally, 171 patients were considered, 56 in surgical and 115 in CRT series. Patients were stratified in low- (20% of surgical and CRT groups), intermediate- (23% and 41%) and high-risk (57% and 39%) groups. In the surgical series, 5-year OS was 54.5%, 46.9% and 40.0% in low, intermediate and high Ang's risk profiles, respectively, whereas in the CRT series those were 100%, 78.9% and 46.7%, respectively. In the multivariable analyses, adjusting for inhomogeneity between the treatment group, the CRT effect was significantly higher in the low- and intermediate-risk groups (P-value for the interaction treatment risk group = 0.034 in the OS analysis). CONCLUSIONS: In this retrospective analysis, low- and intermediate-risk OPC patients had a better survival when treated with CRT compared with open surgery followed by radiation therapy. These data suggest that different treatment approaches might be essential in determining outcome results.

Phase II study on lapatinib in advanced EGFR-positive chordoma.

BACKGROUND: To report on a prospective, investigator-driven, phase II study on lapatinib in epidermal growth factor receptor (EGFR)-positive advanced chordoma patients. PATIENTS AND METHODS: From December 2009 to January 2012, 18 advanced progressing chordoma patients entered this study (median age: 61 years; disease extent: metastatic 72% and locally advanced 28%). Epidermal growth factor receptor (EGFR) expression and activation were evaluated by immunohistochemistry and/or phospho-arrays, real-time polimerase chain reaction, fluorescence immunostaining. Fluorescence in situ hybridization analysis was also carried out. Patients received lapatinib 1500 mg/day (mean dose intensity = 1282 mg/day), until progression or toxicity. The primary study end point was response rate (RR) as per Choi criteria. Secondary end points were RR by Response Evaluation Criteria in Solid Tumor (RECIST), overall survival, progression-free survival (PFS) and clinical benefit rate (CBR; RECIST complete response + partial response (PR) + stable disease (SD) ≥ 6 months). RESULTS: All patients were evaluable for response. Six (33.3%) patients had PR and 7 (38.9%) SD, as their best Choi responses, corresponding to RECIST SD in all cases. Median PFS by Choi was 6 [interquartile (IQ) range 3-8] months. Median PFS by RECIST was 8 (IQ range 4-12) months, with a 22% CBR. CONCLUSIONS: This phase II study showed a modest antitumor activity of lapatinib in chordoma. The clinical exploitation of EGFR targeting in chordoma needs to be further investigated, both clinically and preclinically. Clinical trial Registration No: EU Clinical Trials Register trial no. 2009-014456-29.

Gemcitabine in advanced angiosarcoma: a retrospective case series analysis from the Italian Rare Cancer Network.

BACKGROUND: Angiosarcoma is a highly aggressive soft tissue sarcoma. Responses to anthracyclines plus/minus ifosfamide, and taxanes alone or in combination with gemcitabine are well documented. Very few data are available on gemcitabine as a single agent. PATIENTS AND METHODS: We retrospectively reviewed all cases of advanced progressive angiosarcoma treated with gemcitabine as a single agent (1000 mg/m(2) i.v. every week for 3 weeks every 4 weeks), at Istituto Nazionale Tumori and within the Italian Rare Cancers Network from January 2008 to November 2010. RESULTS: Twenty-five patients [mean age: 52 years; radiation therapy (RT)-related: 8] received gemcitabine. Best tumor response by RECIST was as follows: complete response = 2, partial response = 14, stable disease = 2, progressive disease = 7 cases, for an overall response rate (PR + CR) of 68%. Six of eight post-RT angiosarcomas responded to treatment. Median overall survival (OS) was 17 months. Median progression-free survival (PFS) was 7 months (range 1-40 months). One patient with a locally advanced thyroid angiosarcoma became resectable after 5 months of gemcitabine, with <10% residual viable tumor cells seen on surgical specimen. Overall, gemcitabine was well tolerated. CONCLUSIONS: Gemcitabine is active in both RT- and non-RT-related angiosarcoma, with dimensional and possibly long-lasting responses. A formal phase II study on gemcitabine as a single agent is warranted.

Sunitinib malate in solitary fibrous tumor (SFT).

BACKGROUND: To report on sunitinib activity in a retrospective series of 35 solitary fibrous tumor (SFT) treated at a single institution. PATIENTS AND METHODS: From April 2008, 35 patients with progressive advanced SFT (male/female: 20/15; mean age: 58 years; meningeal/extrameningeal: 6/29; locally advanced/metastatic: 15/20; prior chemotherapy: 25) were treated, on an individual use basis, with continuous-dosing sunitinib 37.5 mg/day. Platelet-derived growth factor receptor beta (PDGFRB) and vascular endothelial growth factor receptor 2 (VEGFR2) status were assessed by immunohistochemistry and, in a subgroup of patients, by real time PCR. RESULTS: Thirty-one patients were assessable for response by RECIST (one early death; three early interruptions). Best responses were 2 partial response (PR), 16 stable disease, 13 progressive disease. A <30% decrease in size was observed in three patients. Fourteen of 29 patients assessable by Choi criteria had a PR. Median progression-free survival by RECIST was 6 months (range 1-22). In two of six patients, resistance to sunitinib was overcome by increasing sunitinib to 50 mg/day. PDGFRB and/or VEGFR2 were positive in all cases and not predictive of response; a less aggressive morphology corresponded to an increased response rate (53% PR by Choi in the malignant SFT, 20% PR in the pleomorphic/dedifferentiated SFT). CONCLUSIONS: Sunitinib is active in SFT. Response can be long-lasting.

Phase II clinical trial of neoadjuvant trabectedin in patients with advanced localized myxoid liposarcoma.

BACKGROUND: To evaluate neoadjuvant trabectedin (1.5 mg/m(2) 24-h i.v. infusion every 3 weeks; three to six cycles) in patients with locally advanced myoxid liposarcoma (ML) previously untreated with chemotherapy or radiation. PATIENTS AND METHODS: Primary efficacy end point was pathological complete response (pCR) or tumoral regression rate. Objective response according to RECIST (v.1.0) was a secondary end point. RESULTS: Three of 23 assessable patients had pCR [13%; 95% confidence interval (CI), 3% to 34%]. Furthermore, very good and moderate histological responses were observed in another 2 and 10 patients, respectively. Histological decrement in the cellular and vascular tumor component and maturation of tumor cells to lipoblasts were observed in both myoxid and myoxid/round cell variants. Seven patients had partial response according to RECIST (objective response rate of 24%; 95% CI, 10% to 44%). No disease progression was reported. Neoadjuvant trabectedin was usually well tolerated, with a safety profile similar to that described in patients with soft tissue sarcoma or other tumor types. CONCLUSION: Trabectedin 1.5 mg/m(2) given as a 24-h i.v. infusion every 3 weeks is a therapeutic option in the neoadjuvant setting of ML.

Role of EGFR family receptors in proliferation of squamous carcinoma cells induced by wound healing fluids of head and neck cancer patients.

BACKGROUND: Mounting evidence suggests that recurrence of resected head and neck squamous cell carcinomas (HNSCCs) is due to the outgrowth of unrecognized residual tumor cells as well as to the premalignant and/or precursor-field epithelial cells. We studied the impact of processes triggered by HNSCC surgery in stimulating both residual tumor cells [demonstrated to overexpress epidermal growth factor receptor (EGFR)], and premalignant cells surrounding the resected lesion. PATIENTS AND METHODS: EGFR expression/activation by immunohistochemistry/biochemistry and gene status by FISH were investigated in 23 primary HNSCCs and surrounding tissues. The ability to induce cell proliferation of wound healing drainages collected from 12 relapsed and 11 not relapsed patients was evaluated by a colorimetric assay in squamous cell carcinoma cell lines A431 (carrying EGFR amplification) and CAL27 (carrying three EGFR copies) in the presence/absence of EGFR therapeutic inhibitors. RESULTS: Primary tumors showed intermediate/high EGFR expression (91%), EGFR phosphorylation and EGFR-positive FISH (35%). Normal, metaplastic and dysplastic epithelium surrounding the resected tumor displayed EGFR overexpression. EGFR activation and gene amplification were observed in normal and dysplastic epithelium, respectively. Each tested wound healing drainage induced the cells to proliferate and the proliferation was significantly higher in relapsed compared with not relapsed HNSCC patients (P = 0.02 and P = 0.03). Anti-EGFR treatments inhibited the drainage-induced proliferation, with the highest inhibitory efficiency by cetuximab on A431 cells, while CAL27 cell growth was more efficiently inhibited by tyrosine kinase inhibitors. CONCLUSIONS: Surgery could favor the proliferation of cells showing EGFR overexpression/activation/amplification such as residual tumor cells and/or precursor-field epithelial cells already present after surgery. Treatment with anti-EGFR reagents inhibits wound-induced stimulation, according to the EGFR family status.

Sunitinib in advanced alveolar soft part sarcoma: evidence of a direct antitumor effect.

BACKGROUND: The purpose of this study was to confirm sunitinib activity in alveolar soft part sarcoma (ASPS) and to report on new insights into the molecular bases thereof. PATIENTS AND METHODS: From July 2007, nine patients with progressive metastatic ASPS received sunitinib 37.5 mg/day, within a named use program. Cryopreserved material was available for five naive patients, among whom three received sunitinib. Immunofluorescence (IF)/confocal microscopy, biochemical, and molecular/cytogenetic analyses were carried out, complemented by antiproliferative and activation assays in a short-term culture derived from one case. RESULTS: All patients were eligible for response. Best RECIST response was partial response in five cases, stable disease in three, and progression in one. The median progression-free survival was 17 months. Positron emission tomography results were consistent. Two cases of interval progressions were recorded. Antiproliferative assays and biochemistry on short-term culture showed that sunitinib is able to markedly impair ASPS cells growth and switch-off PDGFRB. IF/confocal microscopy demonstrated coexpression and physical association between PDGFRB/vascular endothelial growth factor receptor 2 (VEGFR2) and RET/VEGFR2 in ASPS cells, which was validated by biochemistry. PDGFRB, RET, and MET ligand-dependent activation was confirmed. CONCLUSIONS: We confirm the clinical efficacy of sunitinib in ASPS, mediated by PDGFRB, VEGFR2, and RET, which are all expressed in tumor cells. A direct antitumor effect was shown in a short-term cell culture.

An expanded peripheral T cell population to a cytotoxic T lymphocyte (CTL)-defined, melanocyte-specific antigen in metastatic melanoma patients impacts on generation of peptide-specific CTLs but does not overcome tumor escape from immune surveillance in metastatic lesions.

It is not known if immune response to T cell-defined human histocompatibility leukocyte antigen (HLA) class I-restricted melanoma antigens leads to an expanded peripheral pool of T cells in all patients, affects cytotoxic T lymphocyte (CTL) generation, and correlates with anti-tumor response in metastatic lesions. To this end, a limiting dilution analysis technique was developed that allowed us to evaluate the same frequency of peptide-specific T cells as by staining T cells with HLA-peptide tetrameric complexes. In four out of nine patients, Melan-A/Mart-1(27-35)-specific CTL precursors (CTLp) were >/=1/2,000 peripheral blood lymphocytes and found mostly or only in the CD45RO(+) memory T cell subset. In the remaining five patients, a low (<1/40,000) peptide-specific CTLp frequency was measured, and the precursors were only in the CD45RA(+) naive T cell subset. Evaluation of CTL effector frequency after bulk culture indicated that peptide-specific CTLs could be activated in all patients by using professional antigen-presenting cells as dendritic cells, but CTLp frequency determined the kinetics of generation of specificity and the final number of effectors as evaluated by both limiting dilution analysis and staining with HLA-A*0201-Melan-A/Mart-1 tetrameric complexes. Immunohistochemical analysis of 26 neoplastic lesions from the nine patients indicated absence of tumor regression in most instances, even in patients with an expanded peripheral T cell pool to Melan-A/Mart-1 and whose neoplastic lesions contained a high frequency of tetramer-positive Melan-A/Mart-1-specific T cells. Furthermore, frequent lack of a "brisk" or "nonbrisk" CD3(+)CD8(+) T cell infiltrate or reduced/absent Melan-A/Mart-1 expression in several lesions and lack of HLA class I antigens were found in some instances. Thus, expansion of peripheral immune repertoire to Melan-A/Mart-1 takes place in some metastatic patients and leads to enhanced CTL induction after antigen-presenting cell-mediated selection, but, in most metastatic lesions, it does not overcome tumor escape from immune surveillance.

Oncogenic and ligand-dependent activation of KIT/PDGFRA in surgical samples of imatinib-treated gastrointestinal stromal tumours (GISTs).

As the range of receptor tyrosine kinase (RTK) inhibitors widens, a detailed understanding of the activating mechanisms of KIT/platelet-derived growth factor receptor (PDGFR)A and the related downstream pathways involved in the development and maintenance of GISTs is becoming increasingly important. We analysed areas with different histological response ratios in surgical specimens taken from imatinib-treated and untreated GIST patients in order to investigate KIT and PDGFRA expression/activation, the presence of their cognate ligands and the activation of downstream signalling, by means of biochemistry, immunohistochemistry and flow cytometry. All of the cases showed KIT and PDGFRA co-expression. In addition to the oncogenic activation of mutated receptors, activation of wild-type KIT and wild-type PDGFRA, sustained by heterodimerization and an autocrine-paracrine loop, was demonstrated by the presence of their specific ligands, stem cell factor (SCF) and PDGFA. To confirm RTK activation further, all of the samples (including those with the highest regression ratios) were investigated for downstream effectors, and all proved to have activated downstream signalling. The results show that after the mutated receptors are switched off, heterologous wild-type receptors become important in imatinib-treated GISTs as a means of maintaining signalling activation. Taken together, our findings suggest that drugs targeting wild-type receptors should be tested in imatinib-treated GIST patients.

PI3KCA/PTEN deregulation contributes to impaired responses to cetuximab in metastatic colorectal cancer patients.

BACKGROUND: It has been reported that KRAS mutations (and to a lesser extent KRAS mutations with the BRAF V600E mutation) negatively affect response to anti-epidermal growth factor receptor (EGFR) mAbs in metastatic colorectal cancer (mCRC) patients, while the biological impact of the EGFR pathway represented by PI3K/PTEN/AKT on anti-EGFR treatment is still not clear. PATIENTS AND METHODS: We analysed formalin-fixed samples from a cohort of 32 mCRC patients treated with cetuximab by means of EGFR immunohistochemistry, EGFR and PTEN FISH analysis, and KRAS, BRAF, PI3KCA, and PTEN genomic sequencing. RESULTS: Ten (31%) of 32 patients showed a partial response to cetuximab and 22 (69%) did not [nonresponder (NR)]. EGFR immunophenotype and FISH-based gene status did not predict an anti-EGFR mAb response, whereas KRAS mutations (24%) and PI3K pathway activation, by means of PI3KCA mutations (13%) or PTEN mutation (10%)/loss (13%), were significantly restricted to, respectively, 41% and 37% of NRs. CONCLUSION: These findings suggested that KRAS mutations and PI3KCA/PTEN deregulation significantly correlate with resistance to cetuximab. In line with this, patients carrying KRAS mutations or with activated PI3K profiles can benefit from targeted treatments only by switching off molecules belonging to the downstream signalling of activated EGFR, such as mammalian target of rapamycin.

Trabectedin in myxoid liposarcomas (MLS): a long-term analysis of a single-institution series.

BACKGROUND: Trabectedin has been approved in Europe as second-line therapy for advanced soft tissue sarcomas. A previous analysis showed that myxoid liposarcomas (MLS) are particularly sensitive to the drug. We report on the long-term efficacy of trabectedin in a subgroup of that series. METHODS: Since September 2002, 32 advanced pretreated MLS patients received trabectedin at our center. Data were reviewed focusing on their long-term outcome. RESULTS: Trabectedin was given as a 24-h continuous infusion every 21 days. A total of 376 and a median of 12 courses per patient (range 2-26; interquartiles range (IQR) 8-15) were delivered. Response rate per RECIST was 50% [95% confidence interval (CI) 32% to 68%], median progression-free survival (PFS) was 17 months (95% CI 13.5-30.1) and median overall survival is still not reached. In 10 patients, therapy was stopped in the absence of any evident disease, mostly after complete surgery of residual lesions. In these 10 patients, at a median follow-up of 25 months, PFS was 28.1 months (95% CI 25.6-36.4) from treatment start. DISCUSSION: These data indicate that the high response rate of MLS to trabectedin translates into prolonged PFS. Surgery of residual metastatic disease is already used quite extensively in metastatic MLS. Trabectedin may give further significance to this kind of surgery.

Response to imatinib plus sirolimus in advanced chordoma.

BACKGROUND: Imatinib (IM) is active in advanced chordoma. The evidence of upstream and/or downstream mammalian target of rapamycin (mTOR) pathway activation prompted us to combine an mTOR inhibitor, sirolimus, to IM in IM-resistant advanced chordoma. PATIENTS AND METHODS: Since July 2007, 10 progressive advanced chordoma patients with secondary resistance to IM, and biochemical and/or immunohistochemical evidence of upstream and/or downstream mTOR effector activation, started IM (400 mg/day) plus sirolimus (2 mg/day) on a named basis. RESULTS: The mean treatment duration was 9 months. Of nine patients assessable for response, at 3 months, we had one RECIST partial response (PR), seven stable disease (SD) and one progressive disease (PD). According to Choi criteria applied even to magnetic resonance imaging, we had seven PR (> or =10% decrease in size in four cases), one SD and one PD. Seven patients had a positron emission tomography response. The clinical benefit [RECIST complete response + PR + SD > or =6 months] was 89%. Pretreatment mTOR effectors analysis carried out in nine cases was positive in all patients (AKT activation in six patients, S6Sp6 expression/activation in seven). Post-treatment biopsy in one responsive patient confirmed S6 switch off. CONCLUSION: In addition to PDGFRB, mTOR pathway can be activated in chordomas and the combination of IM plus rapalogs may be effective in IM-resistant chordomas.

Quantitative and qualitative characterization of plasma DNA identifies primary and recurrent colorectal cancer.

Because plasma DNA may be a useful tool for cancer detection, we screened primary tumors and related multiple plasma samples at the time of surgery and during the follow-up period for plasma DNA level as well as for K-Ras mutations and p16INK4a promoter hypermethylation in colorectal cancer patients. At the time of surgery, DNA levels were higher in tumor patients than in healthy donors, and K-Ras and p16INK4a alterations were detected in 7 and 11 cancers respectively, and in all related plasma samples. During the follow-up, plasma DNA levels decrease progressively but rapidly increased when a relapse occurred, whereas K-Ras and p16INK4a alterations were detected only in relapsed patients. Therefore, combined quantitative and qualitative analyses of plasma DNA confirm the presence of colorectal cancer, define disease-free status and indicate the presence of relapse.

Functional analyses and molecular modeling of two c-Kit mutations responsible for imatinib secondary resistance in GIST patients.

Imatinib-acquired resistance related to the presence of secondary point mutations has become a frequent event in gastrointestinal stromal tumors. Here, transient transfection experiments with plasmids carrying two different KIT-acquired point mutations were performed along with immunoprecipitation of total protein extracts, derived from imatinib-treated and untreated cells. The molecular mechanics/Poisson Boltzmann surface area computational techniques were applied to study the interactions of the wild-type and mutated receptors with imatinib at the molecular level. Biochemical analyses showed KIT phosphorylation in cells transfected with vectors carrying the specific mutant genes. Imatinib treatment demonstrated that T670I was insensitive to the drug at all the applied concentrations, whereas V654A was inhibited by 6 microM of imatinib. The modeling of the mutated receptors revealed that both substitutions affect imatinib-binding site, but to a different extent: T670I substantially modifies the binding pocket, whereas V654A induces only relatively confined structural changes. We demonstrated that T670I and V654A cause indeed imatinib-acquired resistance and that the former is more resistant to imatinib than the latter. The application of molecular simulations allowed us to quantify the interactions between the mutated receptors and imatinib, and to propose a molecular rationale for this type of drug resistance.

Gene p53 mutations are restricted to poorly differentiated and undifferentiated carcinomas of the thyroid gland.

The p53 gene was analyzed in tumor specimens obtained from 52 patients with various types of carcinoma of the thyroid gland by a combined molecular and immunocytochemical approach. The histologic types included 37 well-differentiated papillary and follicular carcinomas, 8 poorly differentiated, and 7 undifferentiated carcinomas. The p53 gene was shown to be unaffected in all differentiated tumors by single-strand conformation polymorphism analysis. However, in two out of eight (25%) of poorly differentiated carcinomas and five out of seven (71%) undifferentiated carcinomas, p53 mutations were identified and subsequently characterized by DNA sequencing. One undifferentiated carcinoma displayed two areas with varying degrees of differentiation. The comparative analysis of the p53 gene, in both the more and the less differentiated area of this tumor, clearly showed that the p53 mutation was confined to the latter component of the tumor specimen. These results indicate that mutations of the p53 gene are associated with the most aggressive histologic types of thyroid tumors, such as the undifferentiated carcinoma and, to a certain extent, the poorly differentiated carcinoma, and that the alterations of this gene represent a late genetic event in human thyroid carcinogenesis.

Patient-derived solitary fibrous tumour xenografts predict high sensitivity to doxorubicin/dacarbazine combination confirmed in the clinic and highlight the potential effectiveness of trabectedin or eribulin against this tumour.

BACKGROUND: Preclinical models that mimic pathological and molecular features of solitary fibrous tumour (SFT) represent an important tool to select effective regimes and novel compounds to be tested in the clinic. This study was aimed at developing two preclinical models of SFT, assessing their predictive value in the clinic and selecting potential novel effective treatments. MATERIAL AND METHODS: Two dedifferentiated-SFT (D-SFT) models obtained from patients' biopsies were grown in immunodeficient mice. The antitumour activity on these models of doxorubicin, dacarbazine (DTIC), ifosfamide (monotherapy or combination), trabectedin and eribulin was tested. Twelve SFT patients were treated with doxorubicin and DTIC. Response by RECIST, progression-free survival and overall survival were retrospectively evaluated, distinguishing malignant-SFT (M-SFT) and D-SFT. RESULTS: Two D-SFT patient-derived xenografts (PDXs) that represent the first available preclinical in vivo models of SFT were developed and characterised. Doxorubicin/DTIC, DTIC/ifosfamide, doxorubicin/ifosfamide combinations consistently induced better antitumour activity than the single-agents. Particularly, doxorubicin/DTIC combination caused a max tumour volume inhibition >80% in both models. Doxorubicin/DTIC combo showed activity also in the case-series. Best RECIST responses were: 6 responses (M-SFT = 2 of 7, D-SFT = 4 of 5), 1 stable disease, 5 progressions, with a 6-month median progression-free survival (M-SFT = 6, D-SFT = 10 months). The PDXs were very sensitive to trabectedin and eribulin. CONCLUSION: Doxorubicin plus DTIC combination was effective in our two D-SFT mice models and appeared to be active also in the clinic, especially in high-grade D-SFT patients. Among additional drugs tested in the PDXs, trabectedin and eribulin were highly effective, providing a rational to test these drugs in D-SFT patients.