Preclinical tolerance evaluation of the addition of a cisplatin-based dry powder for inhalation to the conventional carboplatin-paclitaxel doublet for treatment of non-small cell lung cancer.

Despite the advances in targeted therapies and immunotherapy for non-small cell lung cancer (NSCLC) patients, the intravenous administration of carboplatin (CARB) and paclitaxel (PTX) in well-spaced cycles is widely indicated for the treatment of NSCLC from stage II to stage IV. Our strategy was to add a controlled-release cisplatin-based dry-powder for inhalation (CIS-DPI-ET) to the conventional CARB-PTX-IV doublet, administered during the treatment off-cycles to intensify the therapeutic response while avoiding the impairment of pulmonary, renal and haematological tolerance of these combinations. The co-administration of CIS-DPI-ET (0.5 mg/kg) and CARB-PTX-IV (17-10 mg/kg) the same day showed a higher proportion of neutrophils in BALF (35 ± 7% vs 1.3 ± 0.8%), with earlier regenerative anaemia than with CARB-PTX-IV alone. A first strategy of CARB-PTX-IV dose reduction by 25% also induced neutrophil recruitment, but in a lower proportion than with the first combination (20 ± 6% vs 0.3 ± 0.3%) and avoiding regenerative anaemia. A second strategy of delaying CIS-DPI-ET and CARB-PTX-IV administrations by 24 h avoided both the recruitment of neutrophils in BALF and regenerative anaemia. Moreover, all these groups showed higher cytotoxicity (LDH activity, protein content) with no higher renal toxicities. These two strategies seem interesting to be assessed in terms of antitumor efficacy in mice.

Pulmonary and renal tolerance of cisplatin-based regimens combining intravenous and endotracheal routes for lung cancer treatment in mice.

Despite recent advances, platinum-based chemotherapy (partially composed of cisplatin, CIS) remains the backbone of non-small-cell lung cancer treatment. As CIS presents a cumulative and dose-limiting nephrotoxicity, it is currently administered with an interruption phase of 3-4 weeks between treatment cycles. During these periods, the patient recovers from the treatment side effects but so does the tumour. Our strategy is to increase the treatment frequency by delivering a cisplatin controlled-release dry powder for inhalation (CIS-DPI) formulation during these off-cycles to expose the tumour environment for longer to CIS, increasing its effectiveness. This is promising as long as the pulmonary and renal toxicities remain acceptable. The aim of the present investigation was to evaluate the pulmonary and renal tolerance of CIS-DPI (three times per cycle) and CIS using the intravenous (IV) route (CIS-IV) (one time per cycle) as monotherapies and to optimize their combination in terms of dose and schedule. At the maximum tolerated dose (MTD), combining CIS-DPI and CIS-IV impaired the pulmonary and the renal tolerance. Therefore, pulmonary tolerance was improved when the CIS-IV dose was decreased by 25% (to 1.5 mg/kg) while maintaining the MTD for CIS-DPI. In addition to this dose adjustment, a delay of 24 h between CIS-DPI and CIS-IV administrations limited the acute kidney injury.

Stability study of full-length antibody (anti-TNF alpha) loaded PLGA microspheres.

Antibodies (Abs) require the development of stable formulations and specific delivery strategies given their susceptibility to a variety of physical and chemical degradation pathways. In this study, the encapsulation of an antibody into polylactide-co-glycolide (PLGA) based microspheres was explored to obtain a controlled-release of the incorporated drug. In order to avoid stability issues, a solid-in-oil-in-water (s/o/w) method was preferred. The solid phase was made of anti-TNF alpha monoclonal antibody (MAb) spray-dried microparticles, and the PLGA microspheres were produced using two different polymers (i.e., Resomer(®) RG505 and Resomer(®) RG755S). The stability of the MAb incorporated into the microspheres was investigated under three conditions (5 ± 3°C, 25 ± 2°C/60% RH and 40 ± 2°C/75% RH) for 12 weeks. During this stability study, it was demonstrated that the MAb loaded PLGA microspheres were stable when stored at 5 ± 3°C and that the Resomer(®) RG755S, composed of 75%(w/w) lactic acid as PLGA, was preferred to preserve the stability of the system. Storage at temperatures higher than 5°C led to antibody stability issues such as aggregation, fragmentation and loss of activity. The release profiles were also altered. Physical ageing of the system associated with changes in the glass transition temperature and enthalpy of relaxation was noticed during the storage of the MAb loaded PLGA microspheres.

The cost of surgical training: analysis of operative time for laparoscopic cholecystectomy.

BACKGROUND: Duration of surgery is a main cost factor of surgical training. The purpose of this analysis of operative times for laparoscopic cholecystectomies (LC) was to quantify the extra time and related costs in regards to the surgeons' experience in the operating room (OR). METHODS: All LC performed between January 01, 2005 and December 31, 2008 in 46 hospitals reporting to the database of the Swiss Association for Quality Management in Surgery (AQC) were analyzed (n = 10,010). Four levels of seniority were specified: resident (R), junior consultant (JC), senior consultant (SC), and attending surgeon (AS). The differences in operative time according to seniority were investigated in a multivariable log-linear and median regression analysis controlling for possible confounders. The OR costs were calculated by using a full cost rate in a teaching hospital. RESULTS: A total of 9,208 LC were available for analysis; 802 had to be excluded due to missing data (n = 212) or secondary major operations (n = 590). Twenty-eight percent of the LC were performed by R as teaching operations (n = 2,591). Compared with R, the multivariable analysis of operative time showed a median difference of -2.5 min (-9.0; 4.8) for JC and -18 min (-25; -11) for SC and -28 min (-35; -10) for AS, respectively. The OR minute costs were 17.57, resulting in incremental costs of 492 (159; 615) per operation in case of tutorial assistance. CONCLUSIONS: The proportion of LC performed as tutorial assistance for R remains low. Surgical training in the OR causes relevant case-related extra time and therefore costs.

Sentinel lymph node biopsy is associated with improved survival compared to level I & II axillary lymph node dissection in node negative breast cancer patients.

OBJECTIVE: The few long-term follow-up data for sentinel lymph node (SLN) negative breast cancer patients demonstrate a 5-year disease-free survival of 96-98%. It remains to be elucidated whether the more accurate SLN staging defines a more selective node negative patient group and whether this is associated with better overall and disease-free survival compared with level I & II axillary lymph node dissection (ALND). METHODS: Three-hundred and fifty-five consecutive node negative patients with early stage breast cancer (pT1 and pT2< or =3 cm, pN0/pN(SN)0) were assessed from our prospective database. Patients underwent either ALND (n=178) in 1990-1997 or SLN biopsy (n=177) in 1998-2004. All SLN were examined by step sectioning, stained with H&E and immunohistochemistry. Lymph nodes from ALND specimens were examined by standard H&E only. Neither immunohistochemistry nor step sections were performed in the analysis of ALND specimen. RESULTS: The median follow-up was 49 months in the SLN and 133 months in the ALND group. Patients in the SLN group had a significantly better disease-free (p=0.008) and overall survival (p=0.034). After adjusting for other prognostic factors in Cox proportional hazard regression analysis, SLN procedure was an independent predictor for improved disease-free (HR: 0.28, 95% CI: 0.10-0.73, p=0.009) and overall survival (HR: 0.34, 95% CI: 0.14-0.84, p=0.019). CONCLUSIONS: This is the first prospective analysis providing evidence that early stage breast cancer patients with a negative SLN have an improved disease-free and overall survival compared with node negative ALND patients. This is most likely due to a more accurate axillary staging in the SLN group.

Molecular mechanisms involved in vasoactive intestinal peptide receptor activation and regulation: current knowledge, similarities to and differences from the A family of G-protein-coupled receptors.

An actual paradigm for activation and regulation of the GPCR (G-protein-coupled receptors)/seven-transmembrane helix family of receptors essentially emerges from extensive studies of the largest family of receptors, the GPCR-A/rhodopsin family. The mechanisms regulating the GPCR-B family signal transduction are less precisely understood due in part to the lack of the conserved signatures of the GPCR-A family (E/DRY, NPXXY) and in part to the absence of a reliable receptor modelling, although some studies suggest that both families share similar features. Here, we try to highlight the current knowledge of the activation and the regulation of the VIP (vasoactive intestinal peptide) receptors, namely VPAC (VIP/pituitary adenylate cyclase-activating peptide receptor) 1 and 2. This includes search for amino acids involved in the stabilization of the receptor active conformation and in coupling to G-proteins, signalling pathways activated in response to VIP, agonist-dependent receptor down-regulation, phosphorylation and internalization as well as pharmacological consequences of receptor hetero-dimerization.

Breast cancer sagittal/horizontal plane location influences axillary lymph node involvement.

AIM: To assess the influence of tumour location on axillary lymph node involvement (ALNI) and prognosis in breast cancer by evaluating the significance of the sagittal/horizontal alignment. METHODS: We compared 57 patients with superficially located breast carcinomas up to 3.0 cm with patients having lesions in posterior planes of the breast. Both groups were matched according to age, time of diagnosis, tumour size, grade, hormonal receptor status and tumour site within the frontal plane. Histologic evidence of skin involvement, excluding tumours fulfilling the criteria for pT4b, was defined as inclusion criteria and reference plane for superficial tumour location. RESULTS: Tumours situated in the superficial region of the breast, compared to those located in deeper planes, have an increased risk of ALNI (p=0.023), whereas no difference was observed with reference to disease-specific survival (p=0.203). CONCLUSION: This study shows that ALNI is dependent on sagittal/horizontal as well as frontal tumour location. Clinicians should be aware that tumours lying posteriorly may be at increased risk of occult spread outside axillary lymph nodes.

Scope and significance of non-uniform classification practices in breast cancer with non-inflammatory skin involvement: a clinicopathologic study and an international survey.

BACKGROUND: The study evaluates the scope of non-uniform classification practices concerning breast carcinomas with non-inflammatory skin involvement. PATIENTS AND METHODS: We compared the clinical course of patients with histologically proven non-inflammatory skin involvement: 119 (65.4%) with clinically obvious 'classical' skin changes (Group A) and 63 (34.6%) with no or only discreet changes (Group B). A questionnaire was circulated to pathology departments in 24 countries to assess the practice concerning the placement of skin- involved breast carcinomas in the TNM classification. RESULTS: Patients in Group B showed a significantly better disease specific survival (P=0.0002). Eighty-six respondents (70.5%) of the survey preferred the 'histological view' and classified tumors with only histological proven skin involvement as T 4 b/stage IIIB. The opposing classification principle ('clinical view'), which dictates that T 4 b breast cancer is a clinical diagnosis and the classical signs must be present, was supported by 31 respondents (25.4%). CONCLUSIONS: A large number of breast cancer patients with non-inflammatory skin involvement are only histologically proven and show, compared with cases exhibiting the classical clinical signs, significant differences in clinical course and prognosis. In general, both subsets were aggregated in one T category/stage (T 4 b/IIIB). This results in a considerable distortion of the reported statistical data.

Retroperitoneoscopic living donor nephrectomy: a comparison with the open approach in respect of early postoperative pain management.

INTRODUCTION: We retrospectively compared perioperative donor outcomes and early postoperative pain control after retroperitoneoscopic (RLDN) and standard open (OLDN) living donor nephrectomy. METHODS: One hundred donors included fifty after RLDN (37 women/13 men) and 50 after OLDN (35 women/15 men) were retrospectively analyzed for basic analgesics, for opioid consumption, and for visual analog scale (VAS) to verify the experienced pain. The donors were questioned in the morning and evening of the first through fifth postoperative days. RESULTS: The mean age of both groups was equal. The mean operating time was 149.7 +/- 48.2 minutes (60 to 270) for RLDN and 164.1 +/- 30.3 minutes (60 to 240) for OLDN (P = NS). The mean warm ischemia time was 120 +/- 36 seconds (50 to 240) and 114 +/- 31 seconds (60 to 190) for the RLDN and OLDN groups, respectively (P = NS). The mean evening VAS for RLDN versus OLDN on postoperative days 1 to 5 was: 2.1 versus 2.2 (P = NS), 0.9 versus 1.8 (P = .009), 0.5 versus 1.3 (P = .016), 0.1 versus 0.7 (P = .013), and 0.1 versus 0.7 (P = .013), respectively. In both groups there was a tendency toward a higher VAS score in the morning than in the evening. RLDN donors showed significantly earlier period free of pain (VAS = 0) than those after OLDN. There was a significant difference of being free from any opiate between both groups after surgery. CONCLUSIONS: After RLDN donors experienced less postoperative pain than after OLDN over the early postoperative days. Therefore, postoperative regional anesthesia is not necessary for donors operated by a retroperitoneoscopic approach.

Long-term outcomes of breast cancer patients after endoscopic axillary lymph node dissection: a prospective analysis of 52 patients.

BACKGROUND: Reports on long-term outcomes after endoscopic axillary lymph node dissection (ALND) of breast cancer patients are still lacking in the medical literature. The objective of this prospective study was to assess the oncological and functional outcomes in breast cancer patients after endoscopic ALND. METHODS: Fifty-five breast cancer patients were prospectively enrolled, of whom 52 were available for follow-up with a median of 71.9 months (range 11-96). The following oncological and functional endpoints were evaluated during follow-up at several time points: occurrence of local, axillary and distant metastases, seroma or infection, shoulder mobility (range of motion), numbness, pain, presence of lymphoedema as well as restriction in activities of daily living. RESULTS: In 52 patients endoscopic ALND of level I and II was successfully performed. Two port-site metastases (2/52, 4%) occurred, one of which in a patient with negative axillary lymph nodes. The same patient suffered from the only axillary recurrence (1/52, 2%). Three patients (3/52, 6%) developed lymphoedema. No other functional adverse events (shoulder mobility, pain, numbness, hypertrophic scar) were noticed at the end of the observation period. CONCLUSION: The present investigation with long-term follow-up after endoscopic ALND--the first one in the literature--reveals minor morbidity, good functional and cosmetic results. In contrary to conventional surgery, the endoscopic procedure is associated with the occurrence of port-site metastases, not seen in the open approach. Axillary recurrences do not appear more frequently when compared with results after conventional ALND. In the meantime the less invasive sentinel lymph node (SLN) biopsy is the established standard technique in evaluating the axillary lymph node status.

Changes in bone mineral content after surgical treatment of morbid obesity.

Weight loss reduces bone mass and increases the risk of osteoporosis. This study was undertaken to assess changes of bone metabolism following Roux-en-Y gastric bypass (RYGB) and adjustable silicone gastric banding (ASGB) as compared to nonoperated controls of morbidly obese subjects. Fourteen female and 5 male patients with a mean (+/-SEM) age of 44.3 +/- 1.8 years participated in the 24-month prospective study. Nine patients underwent ASGB, 4 patients RYGB operation, and 6 patients were included in the control group. Bone metabolism was assessed by determination of serum parathyroid hormone (PTH), osteocalcin, urinary deoxypyridinoline, and dual energy x-ray absorptiometry (DXA) before, and 6, 12, and 24 months after intervention. The body mass index (BMI) decreased from 41.0 +/- 1.1 to 34.0 +/- 1.4 kg/m2 in the ASGB group (P = .001), from 42.7 +/- 2.2 to 30.5 +/- 2.2 kg/m2 in the RYGB group (P = .006), and remained unchanged in the control group (from 41.2 +/- 1.2 to 41.4 +/- 1.4 kg/m2) after 24 months. Bone mineral content (BMC) showed no significant change in the ASGB group (from 3,079 +/- 140 to 3,064 +/- 129 g) and in the control group (from 2,945 +/- 130 to 2,940 +/- 111 g), whereas it decreased from 2,968 +/- 111 to 2,621 +/- 139 g in the RYGB group (P = .005). The loss in BMC was accompanied by significant increases in urinary deoxypyridinoline (P < .05) and in serum osteocalcin (P < .01) after RYGB, suggesting both, increased bone resorption and increased bone formation. The authors were aware of the fact that the study groups were small and conclusions need to be regarded as preliminary. However, the RYGB operation resulted in enhanced weight loss and significant net loss of bone mass in comparison to ASGB and obese control subjects. Patients losing large amounts of body weight should be monitored regularly regarding prevention of osteoporosis.

Diagnosis of malignant hyperthermia susceptibility during CABG surgery.

A 55-year-old man presented for coronary artery bypass graft (CABG) surgery. Malignant hyperthermia (MH) was suspected in his family. This case report describes a diagnostic approach to obtain a definite MH diagnosis by performing an in vitro contracture test at the time of CABG surgery in combination with molecular genetic investigations.

Lysine 195 and aspartate 196 in the first extracellular loop of the VPAC1 receptor are essential for high affinity binding of agonists but not of antagonists.

The role in ligand recognition and receptor activation of two adjacent charged residues (lysine 195 and aspartate 196) in the first extracellular loop of the human VPAC(1) receptor was investigated in stably transfected CHO cells expressing the wild type or point mutated receptors.Replacement of lysine 195 by glutamine or of aspartate 196 by asparagine reduced the agonists' ability to stimulate adenylate cyclase activity; VIP behaved like a partial agonist and a partial agonist behaved as an antagonist. The receptor's capacity to recognize agonists was reduced but antagonists' affinity was unaffected. Both results suggesting that the two charged residues are essential for VPAC(1) receptor activation. On the other hand, the double mutant was less severely affected than single mutants suggesting that hydrogen bonds may partially compensate the loss of charged residues. But the inversion of the residues affected receptor recognition and activation more markedly suggesting that the two charged residues do not interact directly.

Peritoneal mesothelioma after environmental asbestos exposure.

Mesothelioma are primary malignant neoplasms of the serous membranes. They usually involve the pleura and rarely the pericardium, the peritoneum and the tunica vaginalis testis. About 90% are associated with exposure to asbestos. The exposure is generally occupational, an environmental inhalation of asbestos and asbestiform fibers in areas in Turkey has been observed and presents a major health problem. This report of a patient from Anatolia with peritoneal mesothelioma after environmental exposure outlines the importance of considering this pathology in the differential diagnosis of a Turkish patient presenting with ascites.

A small sequence in the third intracellular loop of the VPAC(1) receptor is responsible for its efficient coupling to the calcium effector.

The stimulatory effect of vasoactive intestinal peptide (VIP) on the intracellular calcium concentration ([Ca(2+)](i)) has been investigated in Chinese hamster ovary cells stably transfected with the reporter gene aequorin, and expressing human VPAC(1), VPAC(2), chimaeric VPAC(1)/VPAC(2) or mutated receptors. The VIP-induced increase in [Ca(2+)](i) was linearly correlated with receptor density, and was higher in cells expressing VPAC(1) receptors than in cells expressing a similar density of VPAC(2) receptors. The study was performed to establish the receptor sequence responsible for this difference. VPAC(1)/VPAC(2) chimaeric receptors were first used for broad positioning: those receptors having the third intracellular loop (IC3) of the VPAC(1) or the VPAC(2) receptor behaved, in this respect, phenotypically like VPAC(1) and VPAC(2) receptors respectively. Replacement in the VPAC(2) receptor of the sequence comprising residues 315-318 (VGGN) within IC3 by its VPAC(1) receptor counterpart (residues 328-331; IRKS) and the introduction of VGGN instead of IRKS into VPAC(1) was sufficient to mimic VPAC(1) and VPAC(2) receptor characteristics respectively. Thus a small sequence in the IC3 domain of the VPAC(1) receptor is responsible for the efficient agonist-stimulated increase in [Ca(2+)](i).

Two tyrosine residues in the first transmembrane helix of the human vasoactive intestinal peptide receptors play a role in supporting the active conformation.

1: We investigated the human vasoactive intestinal polypeptide (VIP) receptors VPAC(1) and VPAC(2) mutated at conserved tyrosine residues in the first transmembrane helix (VPAC(1) receptor Y146A and Y150A and VPAC(2) receptor Y130A and Y134A). 2: [(125)I]-Acetyl-His(1) [D-Phe(2), K(15), R(16), L(27)]-VIP (1-7)/GRF (8-27) (referred to as [(125)I]-VPAC(1) antagonist) labelled VPAC(1) binding sites, that displayed high and low affinities for VIP (IC(50) values and per cent of high affinity binding sites: wild-type, 1 nM (57+/-9%) and 160 nM; Y146A, 30 nM (40+/-8%) and 800 nM; Y150A, 4 nM (27+/-8%) and 300 nM). [R(16)]-VIP behaved as a "super agonist" at both mutated VPAC(1) receptors and the efficacies of VIP analogues modified in positions 1, 3 and 6 were significantly decreased. 3: VIP was less potent at the Y130A and Y134A mutated VPAC(2) receptors (EC(50) 200 and 400 nM, respectively) than at the wild-type VPAC(2) receptor (EC(50) 7 nM). Furthermore, [hexanoyl-His(1)]-VIP behaved as a "super agonist" at the two mutated VPAC(2) receptors, and VIP analogues modified in positions 1, 3 and 6 were less potent and efficient at the mutated than at wild-type VPAC(2) receptors. However, the Y130A and Y134A mutants could not be studied in binding assays. 4: Our results suggest that the conserved tyrosine residues do not interact directly with the VIP His(1), Asp(3) or Phe(6) residues (that are necessary for receptor activation), but stabilize the correct active receptor conformation.

[Malignant transformation of perianal Buschke-Löwenstein tumor. Extensive abdominoperineal rectum excision and reconstruction with transpelvic myocutaneous rectus abdominis muscle flap]. / Maligne entarteter perianaler Buschke-Löwenstein-Tumor. Ausgedehnte abdominoperineale Rektumamputation und Rekonstruktion mit transpelvinem myokutanem M.-rectus-abdominis-Lappen.

INTRODUCTION: The Buschke Löwenstein tumor (giant condyloma) in its perianal variant is an extremely rare disease caused by human papilloma virus. Although of histologically benign appearance, it infiltrates and destroys the surrounding tissue. There is a high risk of local recurrence and malignant transformation. The treatment of choice is wide surgical resection. CASE: A 56-year-old woman presented with perianal giant condyloma infiltrating the rectum and vagina. The extensive soft tissue defect resulting from wide resection was filled with a transpelvic myocutaneous rectus abdominis flap. Histology showed a squamous cell carcinoma arising in the Buschke Löwenstein tumor with clear resection margins. Therefore, the patient was irradiated locally after uneventful primary wound healing. CONCLUSION: A simultaneous reconstruction of a large pelvinoperineal soft tissue defect with the transpelvic myocutaneous rectus abdominis flap allows primary healing, accelerated rehabilitation, and safe adjuvant radiotherapy without risk of serious radiation damage to the small bowel by preventing it from protruding into the pelvic defect.

Vasoactive intestinal peptide (VIP) stimulates [Ca2+]i and cyclic AMPin CHO cells expressing Galpha16.

The stimulatory effect of vasoactive intestinal peptide (VIP) and analogues on [Ca2+]i has been investigated in chinese hamster ovary (CHO) cells stably transfected with the reporter gene aequorin, and expressing either the human VPAC1or VPAC2 receptor in absence or in presence of the Galpha16. In cells that were not transfected with Galpha16 and expressed a similar density of receptors, the VIP induced [Ca2+]i ncrease was higher in VPAC1 than in VPAC2 receptor expressing cells. In aequorin/Galpha16 cotransfected cells, the VIP-induced response was higher, reaching 70 to 80% of the maximal calcium response, obtained after digitonin treatment, in response to both VPAC1 and VPAC2 receptor stimulation. The results suggest that in hematopoietic cells, which express both VIP receptors and Galpha16, the signalling pathway of VIP could be mediated through both cyclic AMP and [Ca2+]i increase.

Mutational analysis of the human vasoactive intestinal peptide receptor subtype VPAC(2): role of basic residues in the second transmembrane helix.

1. We investigated the role of two conserved basic residues in the second transmembrane helix arginine 172 (R172) and lysine 179 (K179) of the VPAC(2) receptor. 2. Vasoactive intestinal polypeptide (VIP) activated VPAC(2) receptors with an EC(50) value of 7 nM, as compared to 150, 190 and 4000 nM at R172L, R172Q and K179Q-VPAC(2) receptors, respectively. It was inactive at K179I mutated VPAC(2) receptors. These results suggested that both basic residues were probably implicated in receptor recognition and activation. 3. The VPAC(2)-selective VIP analogue, [hexanoyl-His(1)]-VIP (C(6)-VIP), had a higher affinity and efficacy as compared to VIP at the mutated receptors. 4. VIP, Asn(3)-VIP and Gln(3)-VIP activated adenylate cyclase through R172Q receptors with EC(50) values of 190, 2 and 2 nM, respectively, and through R172L receptors with EC(50) values of 150, 12 and 8 nM, respectively. Asn(3)-VIP and Gln(3)-VIP behaved as partial agonists at the wild type receptor, with E(max) values (in per cent of VIP) of 75 and 52%, respectively. In contrast, they were more efficient than VIP (E(max) values of 150 and 150% at the R172Q VPAC(2) receptors, and of 400 and 360% at the R172L receptors, respectively). These results suggested that the receptor's R172 and the ligand's aspartate 3 are brought in close proximity in the active ligand-receptor complex. 5. The K179I and K179Q mutated receptors had a lower affinity than the wild-type receptors for all the agonists tested in this work: we were unable to identify the VIP amino acid(s) that interact with K179.