Targeting Breast Cancer Using 177 Lu-Labeled Trastuzumab and Trastuzumab Fragment : First-in-Human Clinical Experience.

PURPOSE: A monoclonal antibody, trastuzumab, is used for immunotherapy for HER2-expressing breast cancers. Large-sized antibodies demonstrate hepatobiliary clearance and slower pharmacokinetics. A trastuzumab fragment (Fab; 45 kDa) has been generated for theranostic use. PATIENTS AND METHODS: Fab was generated by papain digestion. Trastuzumab and Fab have been radiolabelled with 177 Lu after being conjugated with a bifunctional chelating. The affinity and target specificity were studied in vitro. The first-in-human study was performed. RESULTS: The bifunctional chelating agent conjugation of 1-2 molecules with trastuzumab and Fab was detected at the molar ratio 1:10 in bicarbonate buffer (0.5 M, pH 8) at 37°-40°C. However, 2-3 molecules of bifunctional chelating agent were conjugated when DMSO in PBS (0.1 M, pH 7) was used as a conjugation buffer at a molar ratio of 1:10. The radiolabelling yield of DOTA-conjugated Fab and trastuzumab at pH 5, 45°C to 50°C, with incubation time 2.5-3 hours was 80% and 41.67%, respectively. However, with DOTAGA-conjugated trastuzumab and Fab, the maximum radiolabelling yield at pH 5.5, 37°C, and at 2.5-3 hours was 80.83% and 83%, respectively. The calculated K d of DOTAGA Fab and trastuzumab with HER2-positive SKBR3 cells was 6.85 ± 0.24 × 10 -8 M and 1.71 ± 0.10 × 10 -8 M, respectively. DOTAGA-Fab and trastuzumab showed better radiolabelling yield at mild reaction conditions.177 Lu-DOTAGA-Fab demonstrated higher lesion uptake and lower liver retention as compared with 177 Lu-DOTAGA-trastuzumab. However, 177 Lu-DOTAGA-Fab as compared with 177 Lu-DOTAGA-trastuzumab showed a relatively early washout (5 days) from the lesion. CONCLUSIONS: 177 Lu-DOTAGA-Fab and trastuzumab are suitable for targeting the HER2 receptors.

Bio-evaluation of poly(lactic-co-glycolic) acid nanoparticles loaded with radiolabelled rifampicin.

AIMS: The poly(lactic-co-glycolic) acid (PLGA) nanoparticles of tubercular drugs have been demonstrated to have a sustained release profile over 7 days. There is no information on the location or mode of release of these nanoparticles in living systems. Therefore, we have planned the study to explore the pharmacokinetics and biodistribution of PLGA rifampicin nanoparticles in healthy human volunteers. We aim to study the distribution pattern of PLGA-loaded nano-formulation of radiolabelled rifampicin in humans. METHODS: Rifampicin was labelled with 99m Tc by indirect method and nanoparticles were prepared by a single emulsion evaporation method. To investigate the pharmacokinetics and biodistribution of nanoparticles, a single dose of 450 mg of rifampicin was administered orally to healthy human volunteers divided into two different groups. RESULTS: Following a single oral dosage of the rifampicin nanoformulation, the pharmacokinetic (PK) parameters were significantly different between the nanoparticle and conventional groups: area under the concentration-time curve (AUC = 113.8 vs. 58.6; P < .001), mean residence time (MRT = 16.2 vs. 5.8; P < .01) and elimination rate constant (Ke = 0.04 vs. 0.10; P < .05). Also, Single-photon emission computed tomography/computed tomography (SPECT/CT) images revealed biodistribution of nanoparticles in the distal portions of the intestine, which is consistent with our dosimetry analysis. CONCLUSIONS: Significant difference in PK parameters and biodistribution of nanoparticles in spleen and lymph nodes with maximum deposition were observed in the large intestine. The nanoparticle distribution pattern may be advantageous for the treatment of intestinal or lymph node tuberculosis (TB) and has the potential to result in a lower dose of rifampicin nanoformulation for the treatment of pulmonary TB.

Improving outcomes for high-risk DLBCL: a pilot study looking at the role of fractionated cyclophosphamide with RCHOP chemo-immunotherapy (SCUBA-1 trial).

The outcomes for patients with high-risk DLBCL are suboptimal, especially in Low-middle income countries in comparison to published data from the western world. Most newer therapies aimed at improving outcomes are either unavailable or out of reach for the majority of patients in low-middle income countries. Cyclophosphamide is an easily available and accessible drug that forms the backbone for therapy for DLBCL. We conducted a single-center, open-label randomized pilot study comparing standard RCHOP to RCHOP with fractionated cyclophosphamide (RfCHOP) in patients with newly diagnosed, high-risk DLBCL. Fifty-five patients were randomized- 28 to RfCHOP and 27 to the RCHOP arm. RfCHOP was associated with a higher complete response rate than RCHOP at the end of 6 cycles of therapy (81.2% vs. 59.3%; p-0.062). Grade III/IV adverse events were comparable in both arms with the use of prophylactic GCSF in the RfCHOP arm. At a median follow-up of 22 months, the Median EFS and OS was not reached in either arm. RfCHOP may represent a therapeutic option for patients with newly-diagnosed, high-risk DLBCL, especially in Low-middle income countries. Larger studies are required to confirm these findings.

Development 68Ga trastuzumab Fab and bioevaluation by PET imaging in HER2/neu expressing breast cancer patients.

INTRODUCTION: Receptors on breast cancer cells play a crucial role in the management of patients. Trastuzumab is a widely used drug for the treatment of HER2/neu expressing tumors. ImmunoPET with trastuzumab is not feasible due to slow pharmacokinetics. Fragment of antigen-binding (Fab) radiolabeled with positron emitters can be used for immunoPET. METHODS: Fab has been generated by papain digestion and conjugated with the bifunctional chelating agent NOTA. The SDS-PAGE and MALDI-TOF were used to see the integrity of Fab and conjugated Fab. In-vitro stability and target specificity for HER2/neu receptors were performed in plasma and receptor binding with bio-layer interferometry (BLI) techniques. Radiolabeling was standardized with 68GaCl3 and PET imaging was performed in seven patients showing 18F fluorodeoxyglucose (18F-FDG) uptake and correlated with HER2/neu expression by immunohistochemistry. RESULTS: Fab production was optimized at molar ratio 23:1 of trastuzumab and papain at 37 °C with a constant stirrer at 850 rpm for 22-24 h, at pH 8. Conjugation with NOTA was standardized at molar ratio 1:25 of trastuzumab Fab and NOTA. Molecular mass of trastuzumab Fab-NOTA was found approximately 46.3 kDa (~1/3 of intact antibody). Trastuzumab Fab-NOTA showed radiolabelling efficiency of 48-70% with incubation time 15 min at 37-40 °C and pH 4.5-5.0. BLI demonstrated the affinity of trastuzumab, trastuzumab Fab and trastuzumab Fab-NOTA towards HER2/neu receptor with KD of <1pM, ~0.5nM and ~20nM, respectively. All immunohistochemistry proven patients showed uptake in primary breast lesion and lymph nodes. CONCLUSION: Trastuzumab Fab-NOTA is suitable for radiolabelling with 68Ga and ImmunoPET imaging of HER2/neu receptor.

Prospective observational study evaluating acute and delayed treatment related toxicities of prophylactic extended field volumetric modulated arc therapy with concurrent cisplatin in cervical cancer patients with pelvic lymph node metastasis.

PURPOSE: To evaluate the treatment related acute and delayed toxicities of extended field Volumetric modulated arc therapy (VMAT) with concurrent chemotherapy in patients of locally advanced cervical cancer with pelvic lymph nodes. MATERIAL AND METHODS: From 2014 to 2016, 15 patients of locally advanced cervical cancer with Fluoro-deoxyglucose positron emission tomography (FDG-PET) positive pelvic lymph nodes were treated with extended field Simultaneous integrated boost (SIB)-VMAT 45 Gy/55 Gy/25#/5weeks and concurrent cisplatin. Acute toxicities were documented according to common terminology criteria for adverse events version 4 (CTCAE v.4). Dose volume parameters and patient characteristics were analyzed for association with toxicities. RESULTS: Median age of patients at diagnosis was 48 years. 40% (6 patients) were stage IIB & 60% (9 patients) were stage IIIB. Median number of involved pelvic lymph nodes was 2 (range, 1-4), commonest location was external iliac lymph node region (86%). Median number of concurrent chemotherapy cycles received was five. Treatment was well tolerated and there were no grade ≥ 3 acute toxicities. Commonest acute toxicities observed were vomiting (≥grade2 -13.3%) followed by & nausea (grade ≥ 2 in 6%) and were associated with volume of bowel bag receiving 45 Gy. Constitutional symptoms (≥grade 2) were observed in 6% patients and had no dosimetric associations. At a median follow up of 43 months, delayed ≥ grade1, 2, 3 toxicity were observed in 80%, 0%, and 0% respectively with diarrhea being the commonest. CONCLUSION: Prophylactic para aortic extended field VMAT with concurrent chemotherapy for locally advanced cervical cancer is well tolerated with acceptable acute toxicity profile. Significant grade 3 acute/delayed toxicities were not observed in this cohort of patients.

Molecular Imaging Targeting Corticotropin-releasing Hormone Receptor for Corticotropinoma: A Changing Paradigm.

BACKGROUND: Corticotrophin-releasing hormone (CRH) is the major regulator of adrenocorticotrophic hormone (ACTH) secretion from the anterior pituitary and acts via CRH-1 receptors (CRH-1R). Corticotropinoma though autonomous, still retain their responsiveness to CRH and hence, we hypothesize that in vivo detection of CRH-1 receptors on pituitary adenoma using Gallium-68 (68Ga)-tagged CRH can indicate the functionality of adenoma, and combining it with positron emission tomography-computed tomography (PET-CT) can provide requisite anatomical information. METHODS: Subjects with ACTH-dependent Cushing's syndrome (CS) (n = 27, 24 with Cushing's disease [CD], 3 with ectopic CS [ECS]) underwent 68Ga CRH PET-CT. Two nuclear medicine physicians read these images for adenoma delineation and superimposed them on magnetic resonance imaging (MRI) sella. The information provided was used for intraoperative navigation and compared with operative and histopathological findings. FINDINGS: 68Ga CRH PET-CT correctly delineated corticotropinoma in all the 24 cases of CD, including the 10 cases with adenoma size < 6mm (4 cases were negative on MRI). Corticotropinoma location on 68Ga CRH PET fusion images with MRI were concordant with operative findings and were further confirmed on histopathology. There was no tracer uptake in the pituitary in 2 patients with ECS, while, in another, the diffuse uptake in pituitary suggested ectopic CRH production. CONCLUSION: 68Ga CRH PET-CT represents a novel, noninvasive molecular imaging, targeting CRH receptors that not only delineate corticotropinoma and provides the surgeon with valuable information for intraoperative tumor navigation, but also helps in differentiating a pituitary from an extra-pituitary source of ACTH-dependent CS. FUNDING: None.

Efficacy of Bacopa Monnieri (Brahmi) and Donepezil in Alzheimer's Disease and Mild Cognitive Impairment: A Randomized Double-Blind Parallel Phase 2b Study.

OBJECTIVES: Alzheimer's disease (AD) is the most common cause of dementia worldwide in the older population. There is no disease-modifying therapy available for AD. The current standard of care drug therapy for AD is cholinesterase inhibitors, including donepezil. Bacopa monnieri or brahmi is used in traditional Indian medicine for memory loss. We conducted a phase 2b randomized controlled trial (RCT) to find out the efficacy of brahmi and donepezil in AD and mild cognitive impairment (MCI). PATIENTS AND METHODS: The study was planned as a 52 week, randomized, double-blind, parallel-group, phase-2 single-center clinical trial comparing the efficacy and safety of Bacopa monnieri (brahmi) 300 mg OD and donepezil 10 mg OD for 12 months in 48 patients with AD and MCI-AD including cognitive and quality of life outcomes. The primary outcome was differences in the change from baseline of the neuropsychological tests [Alzheimer's disease assessment scale-cognitive subscale (ADAS-Cog) and postgraduate institute (PGI) memory scale] at 12 months between the intervention group (brahmi) and active comparison group (donepezil). RESULTS: The study was terminated after 3 years and 9 months, after recruiting 34 patients, because of slow recruitment and a high dropout rate. Intention to treat analysis after adjusting for baseline confounders showed no difference in the rate of change in ADAS-Cog score from baseline at any time point, including the last follow-up. There was no difference in the rate of change in PGI Memory scale (PGIMS) at 3, 6, and 9 months. In the last follow-up, there was a significant difference in the change in total PGIMS score between brahmi and donepezil, while there was no difference in individual scores of the PGI memory scale. CONCLUSION: This phase-2 RCT on the efficacy of brahmi vs. donepezil showed no significant difference between them after 1 year of treatment. Larger phase-3 trials, preferably multicentric, are required to find the superiority of brahmi over donepezil.

Positron emission tomography/computed tomography guided percutaneous biopsies of Ga-68 avid lesions using an automated robotic arm.

PURPOSE: The purpose of this prospective study was to evaluate the feasibility of positron emission tomography/computed tomography (PET/CT)-guided biopsy of Ga-68 avid lesions using an automated robotic arm and determine the diagnostic yield of this technique. MATERIAL AND METHODS: Patients who underwent Ga-68 labelled tracers imaging followed by PET/CT-guided biopsies of tracer-avid lesions were prospectively included. Biopsies were performed using a dedicated automated-robotic-arm assisted PET/CT-guided biopsy device on the same-day of diagnostic PET/CT-imaging. The tissue samples were retrieved after confirming the position of needle-tip in the target lesion. Procedure-related complications and radiation exposure of the interventionist were recorded. Histopathological reports were reviewed for diagnostic yield. RESULTS: A total of 25 patients (19 men, six women) with a mean age of 50.8±17.3 (SD) years (range: 17-83 years) were included. The biopsies were performed after PET/CT using Ga-68 DOTANOC (n=16) or Ga-68 PSMA (n=8) and Ga-68 chemokine-analogue (n=1). The biopsy samples were obtained from the liver (n=9), bone (n=8), lymph-nodes (n=3), lung (n=1), pancreas (n=1), anterior mediastinal lesion (n=1), peritoneal-deposit (n=1) and thigh-lesion (n=1). No immediate or delayed procedure-related complications were documented in any patient. PET/CT-guided molecular sampling was technically successful in all the patients. Histopathology revealed malignancies in all the biopsied specimens without the need for repeat sampling or further invasive-diagnostic workup, with a diagnostic yield of 100%. The estimated absorbed-radiation dose was 566.7µSv/year for the interventionist. CONCLUSION: PET/CT-guided molecular biopsy using Ga-68 labelled radiotracers is feasible and can be performed safely and accurately with a high-diagnostic yield. It is helpful in accurately staging the disease when tracer-avid isolated distant lesion evident on imaging and highly practical in patients with previous inconclusive sampling.

Prevalence and Characteristics of Sensory Processing Abnormalities and its Correlation with FDG-PET Findings in Children with Autism.

OBJECTIVE: To study the prevalence and characteristics of Sensory processing abnormalities (SPAs) in children with autism and to study if there is any correlation between sensory processing abnormalities with FDG-PET findings in children with severe autism. METHODS: One hundred children, aged 3-12 y, diagnosed as Autistic spectrum disorder; ASD (DSM-V) and 100 age and sex matched controls were studied. SPAs were detected using Short sensory profile (SSP) questionnaire. Children with progressive neurological diseases, active epilepsy and structural brain abnormalities were excluded. On Childhood Autism rating scale, 30 children had severe and 70 had mild-moderate autism. The pattern of sensory processing abnormalities in children with severe ASD was compared with mild-moderate ASD. FDG-PET scan was done in children with severe autism and correlated with SPAs. RESULTS: All children with severe autism had sensory processing abnormalities as compared to only 40% children with mild-moderate autism. Underresponsiveness/seeking-sensation was affected in all children with severe ASD and 82% had movement sensitivity. In children with mild-moderate ASD, 45% had auditory filtering, 30% had movement sensitivity and 27% had underresponsiveness/seeking-sensation. FDG-PET was abnormal in 17% of children with severe autism. Diffuse cerebral/ temporal lobe hypometabolism, increased bilateral frontal lobe uptake and moderate reduction in parietal lobe (Lt > Rt) was observed. CONCLUSIONS: All patients with severe autism had SPAs. However, they did not correlate with FDG-PET findings.

68Ga-Glutathione radio-complex as a new diagnostic probe for targeting colon cancer in rats.

OBJECTIVE: Glutathione (GSH) plays an important role in a horde of cellular events that include cell proliferation and apoptosis.The present study describes the radio-synthesis and characterization of gallium-68 (68Ga)-labelled glutathione for its application in radionuclide imaging of cancer. SUBJECT AND METHODS: The radio-synthesis of radio-complex 68Ga-GSH was performed by the direct labeling method. The developed radio-complex was subjected to quality control tests. Colon tumors were developed in healthy male Sprague Dawley (S.D) rats by giving subcutaneous injections of dimethylhydrazine (DMH) in order to monitor the uptake of 68Ga-GSH radio-complex. RESULTS: Gallium-68-labelled glutathione was synthesized with a labeling efficiency of 73.5%±1%. Percentage plasma protein binding and log Po/w values for the radio-complex were found to be 20%-30% and -0.223±0.12, respectively. A significantly higher percentage specific uptake value of newly developed 68Ga-GSH complex was observed in colon tumor in comparison to soft tissue at 90 minutes post administration thereby exhibiting specificity for cancerous cells, which was also witnessed significantly increased overtime from the ratio of colon tumor uptake to normal colon uptake (P≤0.05). CONCLUSION: Therefore, 68Ga-labelled glutathione can further be exploited for radionuclide imaging and assessment of tumor drug resistance in patients.

Two Cases of 188Re Microspheres for Inoperable Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is one of the most lethal malignancies. Often, patients present with inoperable and advanced disease. Selective internal radionuclide therapy offers prolonged survival and improved quality of life by delivering high radiation dose to the tumor with minimal complications. We report 2 inoperable cases of HCC treated with therapeutic dose of indigenously developed Re microspheres delivered to the hepatic lesions by transarterial catheterization. Follow-up CT revealed necrosis within the lesion, suggesting response to selective internal radionuclide therapy. Re microspheres may be a potential treatment option for inoperable HCC with or without portal vein thrombosis.

Radio-pathological Correlation of 18F-FDG PET in Characterizing Gallbladder Wall Thickening.

AIM: Thick-walled gallbladder is difficult to characterize on conventional imaging. 18F-FDG PET was used to differentiate benign and malignant wall thickness and compared with histopathology. METHODS: Thirty patients with gallbladder (GB) wall thickening (focal > 4 mm and diffuse > 7 mm), underwents uspected on  ultrasound, or CT scan, and underwent 18F-FDG PET. Histopathology of the specimen was compared with imaging findings. RESULTS: The mean age was 48.22 ± 31.33 years with a M:F 1:4 ratio. Twenty patients had diffuse and 10 had focal thickening. On 18F-FDG PET, lesion was benign in 12, malignant in 13, and indeterminate in 5. Histopathology was malignancy in 12; benign in 18-chronic cholecystitis in 11, xanthogranulomatous in 4, IgG4 related in 2, and polyp in 1. The mean GB wall thickness was 7.79 ± 3.59 mm (10.34 malignant and 6.10 in benign, p = 0.001). At a cutoff of 8.5 mm, the sensitivity and specificity of detecting malignancy was 94% and 67%. The mean SUV uptake was 7.46 (benign 4.51, malignant 14.26, p = 0.0102). At a cutoff of 5.95, the sensitivity and specificity of detecting malignancy was 92% and 79%. For 18F-FDG PET, overall sensitivity was 91%, specificity 79%, PPV 77%, NPV 92%, and diagnostic accuracy was 84%. CONCLUSION: 18F-FDG PET is a reliable method of differentiation between benign and malignant thickening of the gallbladder particularly when wall thickness and SUV value is taken into account.

18F-fluorodeoxyglucose positron emission tomography-based evaluation of systemic and vascular inflammation and assessment of the effect of systemic treatment on inflammation in patients with moderate-to-severe psoriasis: A randomized placebo-controlled pilot study.

BACKGROUND: Psoriasis is a systemic inflammatory disorder associated with an increased risk of cardiovascular disease. OBJECTIVE: To evaluate the utility of [[18]F]-fluorodeoxyglucose positron emission tomography/computed tomography in identifying vascular and systemic inflammation in psoriasis patients with moderate-to-severe disease and to analyze its usefulness in assessing the effect of systemic treatment. METHODS: This was a randomized, double-blind pilot study conducted in a tertiary care center. Baseline standardized uptake value score was estimated by18F-fluorodeoxyglucose positron emission tomography/computed tomography in patients with moderate-to-severe psoriasis and compared with historical controls. Patients were then randomized using computer-generated randomization list into methotrexate or placebo (with or without pioglitazone) groups.18F-fluorodeoxyglucose positron emission tomography/computed tomography was repeated at 12 weeks and composite standardized uptake value score determined. The correlation between Psoriasis Activity and Severity Index and SUVmax was assessed. RESULTS: A total of 16 patients were randomized to different treatment groups. Significant increase in mean SUVmax was observed in the ascending aorta in psoriasis patients as compared to historical controls (2.03 ± 0.53 vs 1.51 ± 0.36, P < 0.03). There was no difference in composite standardized uptake value score after 12 weeks of treatment in any of the treatment groups (P = 0.82), although an improvement in Psoriasis Activity and Severity Index score in the methotrexate arm was observed. No correlation was found between mean SUVmax and Psoriasis Activity and Severity Index scores in various aortic segments (r = 0.3-0.7). LIMITATIONS: Small sample size, short follow-up, historical controls, exclusion of patients with comorbid conditions and lack of surrogate markers of systemic inflammation. CONCLUSION: 18F-fluorodeoxyglucose positron emission tomography imaging showed higher vascular inflammation in ascending aorta of psoriasis patients as compared to historical controls. Systemic treatment with methotrexate and pioglitazone did not influence the vascular inflammation in the short term.

Endoscopic Management of Vesicoureteral Reflux and Long-term Follow-up.

OBJECTIVE: To report our experience with endoscopic management of vesicoureteral reflux (VUR) by injection of a tissue bulking substance - Dextranomer/ hyaluronic acid co-polymer at vesicoureteric junction. DESIGN: Retrospective analyses of case records. SETTING: Pediatric Surgery department in a tertiary care government Institute. PARTICIPANTS: 500 children (767 renal units) consecutively referred to the out-patient department with vesicoureteral reflux noted on micturating cysto-urethrogram (MCU) over a period of 13 years (2004-2016). INTERVENTION: Preoperative VUR grading and renal scars on radionuclide scans were documented. Dextranomer hyaluronic acid copolymer was injected through a cystoscope at the vesicoureteral junction as a day care procedure under short anesthesia. Patients were followed (average duration 27.3 mo) with clinical assessment, periodic urine cultures and renal scans. MAIN OUTCOME MEASURE: Cessation of VUR and symptomatic relief / clinical success postoperatively at 3 months. RESULTS: Complete symptomatic relief was obtained in 482 (96.4%) patients. In 681 units where MCU was available, 614 (90%) units showed resolution of VUR. CONCLUSIONS: Endoscopic injection of tissue bulking substances at vesicoureteric junction to stop VUR seems to be an effective intervention.