High-throughput protein precipitation method with 96-well plate for determination of doxepin and nordoxepin in human plasma using LC-MS/MS.

The aim of this study was to establish a high-throughput and sensitive LC-MS/MS method for the determination of doxepin and its major active metabolite nordoxepin in human plasma. It has been designed for bioequivalence study for formulations containing 25 mg of doxepin. Doxepin and nordoxepin were extracted from human plasma samples by protein precipitation with acetonitrile by using protein precipitation 96-well plates. The analyte was separated using a Phenomenex Kinetex Biphenyl column (100 × 2.1 mm, 2.6 µm) using isocratic elution with a mobile phase of 20 mM ammonium formate (30%) and acetonitrile:methanol 3:7 v:v (70%) at a flow rate of 0.5 mL/min and an injection volume of 10 µL. The detection was performed using a triple quadrupole mass spectrometer by multiple reaction monitoring mode to monitor the precursor-to-product ion transitions of m/z 280.4 → 107.0 and 283.4 → 235.0 for doxepin and doxepin-D3, respectively, and 266.3 → 106.9 and 269.3 → 235.0 for nordoxepin and nordoxepin-D3, respectively, in positive electrospray ionization mode. The total run time was 3.5 min. The method was validated over a concentration range of 50-10,000 pg/mL using a Triple Quad 4500 MS System (Sciex) for both analytes. The developed and validated method can be successfully used to study the bioequivalence/pharmacokinetics of doxepin and nordoxepin.

Protein precipitation method for determination of clobazam and N-desmethylclobazam in human plasma by LC-MS/MS.

A protein precipitation method for the determination of clobazam (CLB) and its major active metabolite N-desmethylclobazam (N-CLB) in human plasma by liquid chromatography tandem mass spectrometry (LC-MS/MS) was established. CLB and N-CLB were extracted from human plasma samples by protein precipitation with methanol. Analyte separation was done using a Phenomenex Kinetex™ Biphenyl (50 × 2.1 mm, 1.7 µm) column using isocratic elution with a mobile phase of 5 mm ammonium formate with 0.01% ammonium hydroxide (40%) and methanol (60%) at a flow rate of 0.4 mL/min and an injection volume of 10 µL. The detection was performed on a triple quadrupole mass spectrometer in multiple reaction monitoring mode to monitor the precursor-to-product ion transitions of m/z 301.1 → 259.0, 306.0 → 263.9 for CLB and CLB-D5 and 287.0 → 245.0, 292.0 → 250.0 for N-CLB and N-CLB-D5 in positive electrospray ionization mode, respectively. The method was validated over a concentration range of 2.0-750 ng/mL for CLB and 0.7-200 ng/mL for N-CLB on SCIEX Triple Quad 4500 MS System. Total run time was 5 min. This method has been designed for bioequivalence study for formulations containing 20 mg of CLB.

Feasibility study of biweekly capecitabine, oxaliplatin, and irinotecan in patients with untreated advanced gastric cancer.

BACKGROUND: Capecitabine in combination with oxaliplatin and irinotecan (COI regimen) is active and well tolerated in metastatic colorectal cancer. Since there is no internationally adopted standard regimen, we have conducted a pilot study of COI in untreated advanced gastric cancer. METHODS: Patients received irinotecan, 180 mg/m2 infused over 90 min on day 1, followed by oxaliplatin, 85 mg/m2 in a 3-hr infusion on day 2, and capecitabine, 1000 mg/m2/day orally twice daily from days 2 to 6 of a biweekly schedule. Treatment was continued up to 8 cycles or until progression of disease occurred. Response (RECIST criteria) was assessed after the first three cycles and was to be confirmed at least 4 weeks following the first response. RESULTS: A total of 12 patients (5 men and 7 women) with a median age of 54 years (range, 42-65) was prospectively enrolled. Most of the patients (83%) had metastatic disease. Three complete responses, four partial responses and two disease stabilizations occurred in the intention-to-treat cohort, with an overall response rate of 58% (95% confidence interval, 28-85%). Median time to progressive disease and overall survival were 6.4 and 12 months, respectively. A total of 68 cycles was administered, with a median of 6 cycles per patient (range, 1-8). Grade 3 neutropenia occurred in two patients. The most common non-hematologic grade 3 toxicities were nausea (3 patients) and diarrhea 12 patients). CONCLUSIONS: These preliminary findings suggest that biweekly COI is a feasible and promising triplet for the first-line treatment of advanced gastric cancer. A large multi-institutional phase II study of the combination has already been planned in this setting.

Clinical update on palonosetron in the management of chemotherapy-induced nausea and vomiting.

The need to control chemotherapy-induced nausea and vomiting is continuously stimulating research to find better options for the optimal antiemetic care. Palonosetron is different from conventional serotonin receptor antagonists not only by the fact of having a longer half-life but also by higher binding affinity for serotonin receptors. It is the first agent in the class which is approved for preventing both delayed and acute emesis induced by moderately emetogenic chemotherapy. Recent studies using palonosetron-based antiemetic regimens, as well as in the clinical setting of multiple-day chemotherapy, have been reported. Palonosetron plus dexamethasone given as a pre-treatment infusion was effective for preventing acute and delayed emesis after moderately emetogenic chemotherapy. Palonosetron in combination with dexamethasone and aprepitant was highly effective in preventing emesis in the days following administration of moderately emetogenic chemotherapy. Treatment was well tolerated, with no unexpected adverse events. Multiple-day dosing of palonosetron plus dexamethasone was safe and effective for prevention of emesis induced by 5-day cisplatin-based chemotherapy. There was no evidence of cumulative toxicity when palonosetron was given three times over 5 days. Further evidence from ongoing clinical trials with palonosetron with or without dexamethasone will be available soon. Palonosetron represents an useful addition to the therapeutic armamentarium for the management of chemotherapy-induced nausea and vomiting. Further studies are needed to assess the effectiveness of palonosetron in combination with dexamethasone compared with that of older serotonin receptor antagonists combined with dexamethasone. However, palonosetron may offer advantages of convenience over the short-acting older antagonists due to its ability to be given as a single intravenous dose prior to chemotherapy.

Safety and activity of sorafenib in different histotypes of advanced renal cell carcinoma.

BACKGROUND: The aim of our study was to evaluate the efficacy and safety in unresectable or advanced renal carcinoma treated with sorafenib, in a situation closely similar to the everyday medical practice. PATIENTS AND METHODS: One hundred and thirty-six patients have been treated with 400 mg b.i.d. of sorafenib administered orally until disease progression or unacceptable toxicity. They were either previously untreated or relapsed after one or more previous treatments with systemic therapy. Most of them had clear cell renal carcinoma (RCC), but other histological types such as papillary, chromophobe, Bellini ducts, sarcomatoid and mixed forms were also represented. RESULTS: Overall disease control of 70.6% was achieved with 7.9% of partial remissions. Response was observed in the majority of patients with RCC, but also in some patients with non-clear cell RCC. Safety was acceptable, with the most common adverse events consisting of hand-foot skin reaction, cutaneous rash, diarrhoea, fatigue and hypertension. CONCLUSIONS: The results confirm previous ones reported in the literature concerning the efficacy and the safety of sorafenib as second-line treatment in patients with RCC. In addition, they disclose the hypothesis that sorafenib could be effective also in patients who underwent multiple previous treatments and in those with histology different from clear cells.

Renal cell cancer and sorafenib: skin toxicity and treatment outcome.

We describe the case of a young man with refractory renal cell carcinoma who achieved an objective response in a metastatic lesion after biotherapy with the multikinase inhibitor sorafenib and also developed a severe skin reaction. The patient had been previously treated with various combinations of immunochemotherapy without any clinical benefit. We performed a brief review of the literature where similar cases were documented with the use of various anti-EGFR agents. The hypothesis of the correlation of skin toxicity with disease response is not new, but in the absence of any strong evidence remains controversial.

Feasibility of sequential therapy with FOLFIRI followed by docetaxel/cisplatin in patients with radically resected gastric adenocarcinoma. A randomized phase III trial.

OBJECTIVE: Combination therapies of fluorouracil (FU) with irinotecan (CPT-11) and docetaxel plus cisplatin have been proven to be active in metastatic gastric cancer. In this paper, we present the results of a phase III trial in which these two combinations given sequentially were compared to mitomycin C (MMC) monochemotherapy in an adjuvant setting. METHODS: 169 patients with radically resected gastric cancer were randomized to receive CPT-11 (180 mg/m2 day 1), leucovorin (100 mg/m2 days 1-2), FU (400-600 mg/m2 days 1-2, q 14; for four cycles; FOLFIRI regimen), followed by docetaxel (85 mg/m2 day 1), cisplatin (75 mg/m2 day 1, q 21; for three cycles; arm A), or MMC (8 mg/m2 days 1-2 as 2-hour infusion, q 42; for four cycles; arm B). All patients had histologically confirmed gastric carcinoma with nodal positivity or pT3/4. A total of 166 patients (85 in arm A and 81 in arm B) were treated. Adjuvant treatment was completed in 76% of the patients in arm A and in 70% of the patients in arm B. The main grade 3/4 side effects recorded were neutropenia in 35%, with only 1 febrile patient, and diarrhea in 11% in arm A, and thrombocytopenia in 10% and neutropenia in 7% in arm B. The FOLFIRI regimen and docetaxel/cisplatin given in sequence was well tolerated and feasible in adjuvant setting. This sequence treatment currently represents the experimental arm of an ongoing multicenter trial.

Irinotecan, fluorouracil and folinic acid (FOLFIRI) as effective treatment combination for patients with advanced gastric cancer in poor clinical condition.

AIMS AND BACKGROUND: Irinotecan (CPT-11) has been tested as a single agent in several studies, and response rates of 18-23% have been reported in first-line gastric cancer therapy. In the present study we report the safety and efficacy results combining CPT-11 with 5-fluorouracil (5-FU) and folinic acid (FA). PATIENTS AND METHODS: Thirty consecutive patients with metastatic gastric cancer, considered in poor clinical condition, were treated with CPT-11 and 5-FU/FA according to the FOLFIRI regimen. All enrolled cases were evaluable for toxicity and drug activity. RESULTS: The main grade 3-4 toxicity (according to the NCI-CTC criteria) was neutropenia (16%, grade 4 in 1 patient); non-hematological grade 3 toxicity consisted mainly in vomiting and diarrhea reported in 1 patient. No treatment-related serious adverse events were observed. Response was obtained in 12 patients (40%), stable disease in 2 patients (7%), while progression was documented in 16 patients (53%). CONCLUSIONS: These results are very promising, and suggest that the combination of CPT-11 plus 5-FU/FA is active and well tolerated and can be considered as useful treatment in patients with metastatic gastric cancer in poor clinical condition.

Trastuzumab cardiac toxicity: a problem we put our heart into.

Trastuzumab has gained a central role in the treatment of HER2-positive breast cancer, revolutionizing care in this setting. However, since the beginning of its use there have been concerns about its cardiac safety. Trastuzumab can give rise to cardiac toxicity, in some cases leading to discontinuation of treatment. Growing evidence shows that trastuzumab toxicity can be detected early or averted by means of new chemotherapy regimens using different anthracycline formulations or avoiding anthracyclines altogether. Moreover, patients in whom cardiac toxicity develops can be treated effectively and full cardiac function can be restored thanks to progress in drug management. Knowing the characteristics and proper management of trastuzumab toxicity is of paramount importance to grant patients in this setting the best available treatment.

HER2-Positive Neuroendocrine Breast Cancer: Case Report and Review of Literature.

BACKGROUND: Neuroendocrine carcinoma is an uncommon histology for breast cancer. CASE REPORT: Our patient underwent right quadrantectomy for a neuroendocrine carcinoma in 1984 and had a bone relapse 30 years later. After thorough pathological and immunohistochemical analysis the diagnosis was confirmed and HER2 amplification was observed. Here we discuss the management, rationale and results of HER2-targeted therapy in advanced neuroendocrine breast carcinoma.

Body mass index and clinical benefit of fulvestrant in postmenopausal women with advanced breast cancer.

PURPOSE: Obesity is a known risk factor for breast cancer and has been linked to increased risk of recurrence and death in breast cancer patients. Little is known about the predictive value of obesity. As endocrine therapy is widely used for breast cancer treatment worldwide, we aimed at correlating baseline body mass index (BMI) with clinical benefit derived from fulvestrant in postmenopausal women with advanced breast cancer. METHODS: We analyzed consecutive patients treated with fulvestrant in our center between January 2009 and March 2015. Patients were categorized as normal (BMI 18.5-24.9 kg/m2), overweight (BMI 25-29 kg/m2) and obese (BMI >30 kg/m2). The antitumor activity of fulvestrant was evaluated in terms of the clinical benefit rate (CBR). RESULTS: Seventy-five consecutive patients matched the eligibility criteria for analysis. Fulvestrant was administered as first-line therapy in 4 (5%) cases, as second line in 27 (36%) and as third line and beyond in 44 (59%) cases. According to BMI, 44 (59%) patients were classified as normal weight, 19 (25%) as overweight, and 12 (16%) as obese. No difference in estrogen receptor expression was found in relation to BMI. CBR was 53% overall, but rose to 70.5% in normal-weight patients and dropped to 31.6% and 25% in overweight and obese patients, respectively (p<0.001). CONCLUSIONS: Increased BMI has a negative influence on treatment outcome. Even with the limitation of the relatively small sample size, it appears that patients of normal weight are 2.5-fold more likely to benefit from fulvestrant as overweight and obese patients.

Circulating biomarkers in advanced colorectal cancer patients randomly assigned to three bevacizumab-based regimens.

The need to identify biomarkers for bevacizumab-based treatment in advanced colorectal cancer is imperative. The aim of this study was to investigate the prognostic role of circulating VEGF, PDGF, SDF-1, osteopontin and CEA in patients randomly assigned to three bevacizumab-based regimens. Plasma samples from 50 patients treated at a single Institution were analysed using the multiplex assay BioPlex™ 2200 (Bio-Rad Laboratories, Inc, Berkeley, CA, USA) at baseline, before first three cycles and subsequently every three cycles until disease progression. Prognostic analyses of baseline values were performed using multivariable Cox models, including disease extension >10 cm or ≤10 cm (measured as the sum of the diameters for all target lesions) as adjustment factor. The association between progression-free and overall survival and biomarkers modulation during treatment was studied using multivariable Cox models, which included summary statistics synthesizing during-treatment modulation together with disease extension. The biomarkers significantly associated with disease extension were baseline CEA (p = 0.012) and SDF-1 (p = 0.030). High values of VEGF and SDF-1 tended to be associated with worse prognosis, especially in terms of overall survival. The negative prognostic trend was more marked for baseline CEA as compared to other biomarkers; increasing values during treatment was significantly related to worse prognosis independently of disease extension (p = 0.007 and 0.016 for progression-free and overall survival, respectively). VEGF is related to bevacizumab pharmacodynamics and is associated to other angiogenic cytokines; some of the proposed biomarkers such as SDF-1 and CEA should be further validated for prognosis assessment and monitoring of bevacizumab-based treatment of advanced colorectal cancer.

Chronomodulated capecitabine and adjuvant radiation in intermediate-risk to high-risk rectal cancer: a phase II study.

OBJECTIVES: The aim of this study was to evaluate the feasibility and tolerability of capecitabine administration according to a specific time schedule, combined with adjuvant radiation therapy, in intermediate-risk to high-risk rectal cancer patients treated with an upfront surgery. The primary endpoint was the rate of grade 3 to 4 diarrhea during chemoradiation (CRT). MATERIALS AND METHODS: Stage II and III rectal cancer patients received, after total mesorectal excision, 2 cycles of XELOX regimen (oxaliplatin 130 mg/m(2) on day 1; capecitabine 1000 mg/m(2) bid on day 1 to 14, q21), followed by capecitabine (800 mg/m(2) bid daily; 20% dose at 12:00 AM and 80% dose at 12:00 PM) administered continuously during pelvic radiation (total 50.4 Gy in 28 fractions, 1.8 Gy daily dose between 2:00 and 4:00 PM). Four additional cycles of XELOX were administered after chemoradiotherapy. RESULTS: Fifty-one radically resected rectal cancer patients were enrolled. All, but one, cases were evaluated for safety of CRT. We reported a grade 3 and 4 diarrhea rate of 14% (7 of 50 patients), whereas no grade 3 and 4 leukopenia was observed. Grade 1 and 2 proctitis was observed in 26 (52%) cases, whereas grade 1 and 2 cystitis in 5 (10%) patients. Only 2 cases of grade 3 proctitis and cystitis were reported, respectively. The CRT phase was feasible and was completed by 43 (84%) patients. Three patients developed actinic enteritis 60 days after the end of the radiotherapy program. CONCLUSIONS: Capecitabine timetable administration combined with adjuvant radiation therapy of rectal cancer is well tolerated and feasible. Further investigation of this chronomodulated schedule in terms of efficacy is warranted in neoadjuvant setting.

Role of the antiangiogenic agent bevacizumab in the treatment of elderly patients with metastatic colorectal cancer.

Although major progress has been achieved in the treatment of metastatic colorectal cancer with the employment of biological antiangiogenic agents, several questions remain open for discussion regarding the use of this therapy in elderly patients with metastatic colorectal cancer. In Western countries, the total number of elderly patients with colorectal cancer is expected to increase in the future. As adverse physical or socioeconomic conditions are more common in the elderly, an assessment of the patient's suitability for this therapy should be performed before a treatment decision is made. Most patients in clinical trials of the antiangiogenic drug bevacizumab were aged <65 years and thus the efficacy and tolerability of this agent in older patients has been less well explored. However, this article shows that older and younger patients with metastatic colorectal cancer appeared to derive similar survival benefit from bevacizumab treatment. Elderly patients were also found to have significant prolongation of median progression-free survival with the addition of bevacizumab to their treatment, with a similar magnitude of improvement in this outcome being observed in younger and older patients. It should be emphasized that the patients included in the studies discussed in this article were eligible for clinical trials and therefore may not be representative of a more general elderly population. Careful selection of patients and monitoring of treatment effects are required to optimize use of the antiangiogenic agent bevacizumab in older patients.

Dose finding study of erlotinib combined to capecitabine and irinotecan in pretreated advanced colorectal cancer patients.

PURPOSE: This study evaluated the maximum tolerated dose (MTD) and the dose limiting toxicity (DLT) of erlotinib when combined to irinotecan and capecitabine in pre-treated metastatic colorectal cancer patients. METHODS: Five dose level combinations with irinotecan (from 180 to 240 mg/m(2), day 1, q21), capecitabine (1,500-2,000 mg/m(2) per day, days 2-15, q21) and erlotinib (50-150 mg per day, continuously) were planned. Patients were enrolled in cohorts of three, and evaluated for first cycle acute toxicity. RESULTS: Twenty-one patients were treated. In the first cohort, no DLT was reported, in the second: one DLT (G4 neutropenic fever associated with G3 cutaneous rash and mucositis); in the third dose level: two DLT (G3 diarrhea and G4 neutropenic fever). To confirm these results, other six patients were additionally included and no DLT was observed. CONCLUSIONS: The results documented that erlotinib at the dose of 100 mg per day, irinotecan 180 mg/m(2) and capecitabine 1,500 mg/m(2) per day for 14 days has an acceptable safety profile and appears suitable for further phase II studies.