A phase I study of binimetinib (MEK 162), a MEK inhibitor, plus carboplatin and pemetrexed chemotherapy in non-squamous non-small cell lung cancer.

INTRODUCTION: MEK inhibition is a potential therapeutic strategy in non-small cell lung cancer (NSCLC). This phase I study evaluates the MEK inhibitor binimetinib plus carboplatin and pemetrexed in stage IV non-squamous NSCLC patients (NCT02185690). METHODS: A standard 3 + 3 dose-escalation design was used. Binimetinib 30 mg BID (dose level 1 [DL1]) or 45 mg BID (dose level 2 [DL2]) was given with standard doses of carboplatin and pemetrexed using an intermittent dosing schedule. The primary outcome was determination of the recommended phase II dose (RP2D) and safety of binimetinib. Secondary outcomes included efficacy, pharmacokinetics, and an exploratory analysis of response based on mutation subtype. RESULTS: Thirteen patients (6 DL1, 7 DL2) were enrolled: 7 KRAS, 5 EGFR, and 1 NRAS mutation. The RP2D was binimetinib 30 mg BID. Eight patients (61.5%) had grade 3/4 adverse events, with dose limiting toxicities in 2 patients at DL2. Twelve patients were evaluated for response, with an investigator-assessed objective response rate (ORR) of 50% (95% CI 21.1%-78.9%; ORR 33.3% by independent-review, IR), and disease control rate 83.3% (95% CI 51.6%-97.9%). Median progression free survival (PFS) was 4.5 months (95% CI 2.6 months-NA), with a 6-month and 12-month PFS rate of 38.5% (95% CI 19.3%-76.5%) and 25.6% (95% CI 8.9%-73.6%), respectively. In an exploratory analysis, KRAS/NRAS-mutated patients had an ORR of 62.5% (ORR 37.5% by IR) vs. 25% in KRAS/NRAS wild-type patients. In MAP2K1-mutated patients, the ORR was 42.8%. CONCLUSION: The addition of binimetinib to carboplatin and pemetrexed appears to have manageable toxicity with evidence of activity in advanced non-squamous NSCLC.

Consensus recommendations for optimizing biomarker testing to identify and treat advanced EGFR-mutated non-small-cell lung cancer.

The advent of personalized therapy for non-small-cell lung carcinoma (nsclc) has improved patient outcomes. Selection of appropriate targeted therapy for patients with nsclc now involves testing for multiple biomarkers, including EGFR. EGFR mutation status is required to optimally treat patients with nsclc, and thus timely and accurate biomarker testing is necessary. However, in Canada, there are currently no standardized processes or methods in place to ensure consistent testing implementation. That lack creates challenges in ensuring that all appropriate biomarkers are tested for each patient and that the medical oncologist receives the results for making informed treatment decisions in a timely way. An expert multidisciplinary working group was convened to create consensus recommendations about biomarker testing in advanced nsclc in Canada, with a primary focus on EGFR testing. Recognizing that there are biomarkers beyond EGFR that require timely identification, the expert multidisciplinary working group considered EGFR testing in the broader context of integration into complex lung biomarker testing. Primarily, the panel of experts recommends that all patients with nonsquamous nsclc, regardless of stage, should undergo comprehensive reflex biomarker testing at diagnosis with targeted next-generation sequencing. The panel also considered the EGFR testing algorithm and the challenges associated with the pre-analytic, analytic, and post-analytic elements of testing. Strategies for funding testing by reducing silos of single biomarker testing for EGFR and for optimally implementing the recommendations presented here and educating oncology professionals about them are also discussed.

OCTANE (Ontario-wide Cancer Targeted Nucleic Acid Evaluation): a platform for intraprovincial, national, and international clinical data-sharing.

Cancer is a genetic disease resulting from germline or somatic genetic aberrations. Rapid progress in the field of genomics in recent years is allowing for increased characterization and understanding of the various forms of the disease. The Ontario-wide Cancer Targeted Nucleic Acid Evaluation (octane) clinical trial, open at cancer centres across Ontario, aims to increase access to genomic sequencing of tumours and to facilitate the collection of clinical data related to enrolled patients and their clinical outcomes. The study is designed to assess the clinical utility of next-generation sequencing (ngs) in cancer patient care, including enhancement of treatment options available to patients. A core aim of the study is to encourage collaboration between cancer hospitals within Ontario while also increasing international collaboration in terms of sharing the newly generated data. The single-payer provincial health care system in Ontario provides a unique opportunity to develop a province-wide registry of ngs testing and a repository of genomically characterized, clinically annotated samples. It also provides an important opportunity to use province-wide real-world data to evaluate outcomes and the cost of ngs for patients with advanced cancer. The octane study is attempting to translate knowledge to help deliver precision oncology in a Canadian environment. In this article, we discuss the background to the study and its implementation, current status, and future directions.

Crizotinib inhibition of ROS1-positive tumours in advanced non-small-cell lung cancer: a Canadian perspective.

The ros1 kinase is an oncogenic driver in non-small-cell lung cancer (nsclc). Fusion events involving the ROS1 gene are found in 1%-2% of nsclc patients and lead to deregulation of a tyrosine kinase-mediated multi-use intracellular signalling pathway, which then promotes the growth, proliferation, and progression of tumour cells. ROS1 fusion is a distinct molecular subtype of nsclc, found independently of other recognized driver mutations, and it is predominantly identified in younger patients (<50 years of age), women, never-smokers, and patients with adenocarcinoma histology. Targeted inhibition of the aberrant ros1 kinase with crizotinib is associated with increased progression-free survival (pfs) and improved quality-of-life measures. As the sole approved treatment for ROS1-rearranged nsclc, crizotinib has been demonstrated, through a variety of clinical trials and retrospective analyses, to be a safe, effective, well-tolerated, and appropriate treatment for patients having the ROS1 rearrangement. Canadian physicians endorse current guidelines which recommend that all patients with nonsquamous advanced nsclc, regardless of clinical characteristics, be tested for ROS1 rearrangement. Future integration of multigene testing panels into the standard of care could allow for efficient and cost-effective comprehensive testing of all patients with advanced nsclc. If a ROS1 rearrangement is found, treatment with crizotinib, preferably in the first-line setting, constitutes the standard of care, with other treatment options being investigated, as appropriate, should resistance to crizotinib develop.

A Canadian guideline on the use of next-generation sequencing in oncology.

Rapid advancements in next-generation sequencing (ngs) technology have created an unprecedented opportunity to decipher the molecular profile of tumours to more effectively prevent, diagnose, and treat cancer. Oncologists now have the option to order molecular tests that can guide treatment decisions. However, to date, most oncologists have received limited training in genomics, and they are now faced with the challenge of understanding how such tests and their interpretation align with patient management. Guidance on how to effectively use ngs technology is therefore needed to aid oncologists in applying the results of genomic tests. The Canadian guideline presented here describes best practices and unmet needs related to ngs-based testing for somatic variants in oncology, including clinical application, assay and sample selection, bioinformatics and interpretation of reports performed by laboratories, patient communication, and clinical trials.

Impact of somatic molecular profiling on clinical trial outcomes in rare epithelial gynecologic cancer patients.

OBJECTIVES: Conducting clinical trials in rare malignancies is challenging due to the limited number of patients and differences in biologic behavior. We investigated the feasibility and clinical utility of using genomic profiling for rare gynecologic malignancies. METHODS: Rare epithelial gynecologic cancer patients were analyzed for somatic variants through an institutional molecular profiling program using the Sequenom MassArray platform or the TruSeq Amplicon Cancer Panel on the MiSeq platform. Clinical trial outcomes by RECIST 1.1, and time on treatment were evaluated. RESULTS: From March 2012 to November 2015, 767 gynecologic patients were enrolled and 194 (27%) were classified as rare epithelial malignancies. At least one somatic mutation was identified in 72% of patients, most commonly in TP53 (39%), KRAS (28%) and PIK3CA (27%). A total of 14% of patients were treated on genotype-matched trials. There were no significant differences in overall response rate between genotype-matched versus unmatched trials, nor in median time on treatment between genotype trials and the immediate prior systemic standard treatment. Among 13 evaluable Low Grade Serous ovarian cancer patients treated on genotype-matched trials with MEK inhibitor-based targeted combinations, there were four partial responses. CONCLUSIONS: Somatic molecular profiling is feasible and enables the identification of patients with rare gynecologic cancers who are candidates for genotype-matched clinical trials. Genotype-matched trials, predominantly MEK-based combinations in KRAS and/or NRAS mutant Low Grade Serous ovarian cancer patients, and genotype-unmatched trials, have shown potential clinical activity. Prospective trials with integrated genotyping are warranted to assess the clinical utility of next generation sequencing tests as a standard clinical application in rare malignancies.

The Somatic Curation and Interpretation Across Laboratories (SOCIAL) project-current state of solid-tumour variant interpretation for molecular pathology in Canada.

Background: Practices in somatic variant interpretation and classification vary between Canadian clinical molecular diagnostic laboratories, and understanding of current practices and perspectives is limited. To define gaps and future directions, including consensus guideline development, the Somatic Curation and Interpretation Across Laboratories (social) project examined the present state of somatic variant interpretation in Canadian molecular laboratories, including testing volumes and methods, data sources and evidence criteria, and application of published classification guidelines. Methods: Individuals who perform somatic variant interpretation in Canadian centres were invited to participate in an online survey. Invitees included laboratory directors (certified as Fellows of the Canadian College of Medical Geneticists or the American College of Medical Geneticists), md or md and phd molecular pathologists, and other phd experts, including phd specialists in variant annotation or bioinformatics. Current testing methods, volumes, and platforms in next-generation sequencing, use of variant annotation resources and evidence criteria, and preference for variant classification schemes were evaluated. Results: Responses were received from 37 participants in 8 provinces. A somatic variant classification scheme jointly supported by the Association for Molecular Pathology (amp), the American Society of Clinical Oncology (asco), and the College of American Pathologists (cap) was used by 47% of respondents; an alternative guideline or a combination of published guidelines was used by 35% of respondents. The remaining 18% did not use a published scheme. Only 41% of respondents used a published scheme without alteration. Although all respondents indicated that there is a need for Canadian laboratories to adopt a somatic variant classification guideline, only 38% of respondents felt that it should be mandatory to adopt the amp/asco/cap-endorsed guideline. Conclusions: Data from the social project identified high variability in current practice, yet strong support for standardization of solid-tumour somatic variant interpretation across Canadian institutions. Aligning classification methods will reduce variation in cross-institutional classification and reporting practices, aiding in consistent practice nationwide.

Inter-laboratory proficiency testing scheme for tumour next-generation sequencing in Ontario: a pilot study.

Background: A pilot inter-laboratory proficiency scheme for 5 Ontario clinical laboratories testing tumour samples for the Ontario-wide Cancer Targeted Nucleic Acid Evaluation (octane) study was undertaken to assess proficiency in the identification and reporting of next-generation sequencing (ngs) test results in solid tumour testing from archival formalin-fixed, paraffin-embedded (ffpe) tissue. Methods: One laboratory served as the reference centre and provided samples to 4 participating laboratories. An analyte-based approach was applied: each participating laboratory received 10 ffpe tissue specimens profiled at the reference centre, with tumour site and histology provided. Laboratories performed testing per their standard ngs tumour test protocols. Items returned for assessment included genes and variants that would be typically reported in routine clinical testing and variant call format (vcf) files to allow for assessment of ngs technical quality. Results: Two main aspects were assessed:■ Technical quality and accuracy of identification of exonic variants■ Site-specific reporting practicesTechnical assessment included evaluation of exonic variant identification, quality assessment of the vcf files to evaluate base calling, variant allele frequency, and depth of coverage for all exonic variants. Concordance at 100% was observed from all sites in the technical identification of 98 exonic variants across the 10 cases. Variability between laboratories in the choice of variants considered clinically reportable was significant. Of the 38 variants reported as clinically relevant by at least 1 site, only 3 variants were concordantly reported by all participating centres as clinically relevant. Conclusions: Although excellent technical concordance for ngs tumour profiling was observed across participating institutions, differences in the reporting of clinically relevant variants were observed, highlighting reporting as a gap where consensus on the part of Ontario laboratories is needed.

Evidence-based best practices for EGFR T790M testing in lung cancer in Canada.

Epidermal growth factor receptor (egfr) tyrosine kinase inhibitors (tkis) are recommended as first-line systemic therapy for patients with non-small-cell lung cancer (nsclc) having mutations in the EGFR gene. Resistance to tkis eventually occurs in all nsclc patients treated with such drugs. In patients with resistance to tkis caused by the EGFR T790M mutation, the third-generation tki osimertinib is now the standard of care. For optimal patient management, accurate EGFR T790M testing is required. A multidisciplinary working group of pathologists, laboratory medicine specialists, medical oncologists, a respirologist, and a thoracic radiologist from across Canada was convened to discuss best practices for EGFR T790M mutation testing in Canada. The group made recommendations in the areas of the testing algorithm and the pre-analytic, analytic, and post-analytic aspects of clinical testing for both tissue testing and liquid biopsy circulating tumour dna testing. The recommendations aim to improve EGFR T790M testing in Canada and to thereby improve patient care.

Data resources for the identification and interpretation of actionable mutations by clinicians.

Following initial characterization of the reference human genome, initiatives have evolved worldwide to identify genomic aberrations in cancer with the aim of deriving diagnostic, prognostic and predictive information. However, the functional and clinical relevance of many somatic variants in cancer are presently unknown and there is no consensus definition of 'actionability' for genomic aberrancies. Therefore, while robust detection of a variety of genetic aberrations in clinical specimens remains a technical hurdle, the greater challenge lies in the interpretation of these alterations. Critical evaluation of genomic variation in cancer requires the integration of available clinical and preclinical evidence related to their frequencies, functions and roles as therapeutic targets. Many publicly accessible data resources have compiled such evidence to facilitate the understanding of genomic results and ultimately translating results to clinical action. Information for these data resources is derived from various sources including large population genomic datasets, curation of published literature, and data sharing by the scientific community. Currently, there is no widely accepted guidance to definitively assess and integrate the diagnostic, prognostic and predictive information of somatic variants using these knowledge databases. This review will describe data resources pertinent to the identification and interpretation of actionable genomic aberrations by clinicians, and highlight relevant issues in the clinical application of tumor molecular profiling results.

Genotype-matched treatment for patients with advanced type I epithelial ovarian cancer (EOC).

BACKGROUND: Genomic alterations that activate the MAPK signaling pathway frequently occur in Type I Epithelial Ovarian Cancers (EOCs). We evaluated therapeutic response outcomes in patients with type I EOC treated with genotype-matched therapy on clinical trials enrolled in a prospective molecular profiling program. MATERIAL AND METHODS: Formalin fixed paraffin embedded tumor tissues were prospectively screened for genomic alterations using MALDI-ToF mass-spectrometry platform or targeted sequencing using the Illumina MiSeq TruSeq Amplicon Cancer Panel. Treatment outcomes on genotype-matched trials were retrospectively reviewed using RECIST version 1.1 and Gynecological Cancer Intergroup CA125 related-response criteria RESULTS: 55 patients with type I EOC underwent molecular profiling, 41 (75%) low grade serous (LGS), 9 (16%) clear cell (CC), and 5 (9%) mucinous (MC) histologies. Thirty-five patients (64%) were found to have ≥1 somatic mutations: 23 KRAS, 6 NRAS, 5 PIK3CA, 2 PTEN, 1 BRAF, 1 AKT, 1 TP53, and 1 CTNNB1. Fifteen patients were subsequently enrolled in genotype-matched phase I or II trials, including 14 patients with KRAS/NRAS mutations treated with MEK inhibitor targeted combinations. Among 14 RECIST evaluable patients, there were 7 partial responses (PR), 7 stable disease (SD) and 1 disease progression (PD). CA125 responses were observed in 10/10 evaluable KRAS/NRAS mutant patients treated with MEK inhibitor combinations CONCLUSIONS: Genotyping and targeted sequencing of Type I EOCs frequently identifies actionable mutations. Matched treatment with MEK-based combination therapy in KRAS and/or NRAS mutant type I EOC patients is an active therapeutic strategy.

Multiple effects of childhood deafness on cortical activity in children receiving bilateral cochlear implants simultaneously.

OBJECTIVE: Auditory development is disrupted without normal hearing but might proceed to some extent depending on the type and onset of deafness. We therefore hypothesized that activity in the auditory cortex would be highly variable in children who are deaf. METHODS: To answer this, activity in the deaf brain was evoked by electrical pulses from newly provided bilateral cochlear implants (CIs) in 72 children (n=144 responses). RESULTS: Responses were categorized by visual inspection into 3 main types which were validated by principal component cluster analyses; 49% had a negative amplitude wave similar to that previously reported in pre-term infants, 26% were dominated by a positive peak typical of responses in young normal hearing children and experienced paediatric CI users, 25% were novel multi-peaked responses. No significant demographic differences, including duration and onset of deafness, were found between response types. However, children with severe biallelic mutations of GJB-2 showed predominately negative peak type responses (79%) as compared with their peers without these mutations who had a more equal distribution between cortical response types. CONCLUSION: Cortical development in children who are deaf is heterogeneous but can be better predicted when the genotype is known to be a GJB-2 mutation. SIGNIFICANCE: Remediation of childhood deafness seeks to restore normal development and function of central auditory functions and thus may need to be tailored to account for effects specific to the aetiology of deafness.

The role of molecular microsatellite identity testing to detect sampling errors in prenatal diagnosis.

OBJECTIVE: The objective was to determine the risk of sampling error in amniocentesis and chorionic villus sampling (CVS) in singleton and multiple pregnancies. Data from this and other published studies were used to discuss current practice guidelines for molecular identity testing. METHOD: Clinical and laboratory records of all patients undergoing molecular-based identity testing in our clinical laboratory from July 2002 until March 2008 were reviewed. DNA microsatellite testing was performed to determine zygosity in multiple pregnancies and maternal cell contamination (MCC) in both singleton and multiple pregnancies. RESULTS: MCC was detected in 6/148 (4%) CVS and 1/87 (1%) amniotic fluids from singleton pregnancies. In two of the CVS, only maternal cells were found. In 2/24 (8%) twin pregnancies, the same fetus was tested twice. In a total of 285 pregnancies (235 singleton, 24 twin, 26 with >or= 3 fetuses), without molecular identity testing, four women would have received erroneous results. CONCLUSION: Current guidelines recommend molecular identity testing for MCC in conjunction with molecular diagnostic testing, but not for cytogenetic testing. No published guidelines were found for zygosity testing in multiple pregnancies. We suggest that identity testing be considered for all prenatal testing of multiple pregnancies, especially if CVS is performed.

Functional disomy of Xp: prenatal findings and postnatal outcome.

We report on trisomy of the short arm of the X chromosome (Xp11.2 --> pter) due to a de novo unbalanced X;13 translocation diagnosed prenatally in a female fetus. Amniocentesis was performed at 20-weeks' gestation following ultrasound finding of a Dandy-Walker malformation. The trisomy of Xp11.2 --> pter was confirmed with fluorescence in situ hybridization (FISH), using an X chromosome painting probe and telomeric FISH probes specific for the short arm of chromosome X. The karyotype was defined as 46,XX,der(13)t(X;13)(p11.2;p11.2). Molecular analysis suggested that the extra Xp material was of paternal origin. FISH analysis with an XIST probe showed that the derivative chromosome 13 did not include the XIST locus at the X-inactivation center (XIC). A complex phenotype was seen at birth including macrosomia, facial dysmorphism with preauricular tag, congenital heart defects, and structural brain malformations. Because the derivative chromosome was not subject to X inactivation, functional disomy of Xp11.2 --> pter most likely accounts for the abnormal phenotype in this patient.

Delivery of recombinant product from subcutaneous implants of encapsulated recombinant cells in canines.

Delivering recombinant therapeutic proteins from a universal microencapsulated cell line is an alternate method for gene therapy. It has proved effective in the treatment of several murine models of human genetic diseases. However, in scaling up to large animal models, intraperitoneal Implantations of these microcapsules in canines were associated with excessive Inflammatory response and rapid degradation. We now show that subcutaneous implantation of microencapsulated cells in canines is effective in delivering recombinant product systemically for extended periods, provides a surgically benign site, leads to less inflammatory response, and permits longer-term survival of microcapsules. Allogeneic MDCK cells engineered to secrete human growth hormone (hGH) were microencapsulated in alginate-poly-L-lysine-alginate and implanted subcutaneously. Systemic delivery of hGH was evident within 4 hours and peaked by day 1 after implantation in all dogs. The gradual decline of hGH in the circulation in the first 2 weeks coincided with the development of anti-hGH antibodies by day 11. The high titer persisted for more than 1 month, demonstrating indirectly the persistent delivery of hGH. Microcapsules retrieved from the subcutaneous implant maintained their structure throughout the experiment and were free of host cellular adhesions. The mechanical integrity of the subcutaneously implanted microcapsules also appeared superior to that of the intraperitoneal implant. Hence the subcutaneously implanted microcapsules required minimal surgical intervention and led to a low level of inflammatory response, and the implant survived for at least 1 month, thus demonstrating the feasibility of systemic delivery of recombinant products via subcutaneous implantation in large animals.

The in vivo delivery of heterologous proteins by microencapsulated recombinant cells.

The microencapsulation of recombinant cells is a novel and potentially cost-effective method of heterologous protein delivery. A 'universal' cell line, genetically modified to secrete any desired protein, is immunologically protected from tissue rejection by enclosure in microcapsules. The microcapsule can then be implanted in different recipients to deliver recombinant proteins in vivo.

Suppression of immunological response against a transgene product delivered from microencapsulated cells.

A potential obstacle to successful gene therapy for some patients is the in vivo production of neutralizing antibodies against the recombinant therapeutic product delivered. To mimic this clinical situation, we implanted microencapsulated recombinant cells producing human growth hormone into C57B1/6 mice to provoke antihuman growth hormone antibody production. We then investigated the efficacy of different immunosuppressive treatments to inhibit the development of neutralizing antibodies. The experimental mice were treated with either an immunosuppressive drug (FK506 or cyclophosphamide), a cytokine (interferon-gamma [IFN-gamma] or interleukin-12 [IL-12], or a monoclonal antibody (anti-CD4, anti-gp39, or CTLA4-Ig). Serum human growth hormone and mouse anti-human growth hormone antibody levels were measured by enzyme-linked immunosorbent assay (ELISA) for 4 weeks. There were three patterns of response noted among the seven treatment groups. First, the mice receiving IFN-gamma, IL-12, anti-gp39, or CTLA4-Ig were similar to the untreated controls-no suppression of anti-hGH antibodies and no improvement in delivery of hGH. Next, the mice receiving FK506 or cyclosphosphamide showed > or = 90% suppression of antibodies but also no improvement in product delivery. Last, the mice receiving anti-CD4 showed almost complete antibody suppression over 1 month postimplantation. Furthermore, only anti-CD4 permitted a sustained level of human growth hormone delivery to day 28, in contrast to the controls whose human growth hormone delivery was undetectable by day 14 postimplantation. Hence, the use of anti-CD4 inhibited formation of neutralizing antibodies against a recombinant gene product delivered in vivo, and allowed prolonged delivery of a foreign protein. Its role as adjunct treatment for appropriate patients receiving gene therapy should be examined further.

Non-autologous transplantation with immuno-isolation in large animals--a review.

Transplantation has become a successful method for the management of functional failure of a variety of tissues or organs. However, the majority of clinical transplantations use non-autologous allogeneic donor tissue implanted from one human to another. In order to prevent rejection of the allogeneic tissue, methods to overcome the immune barrier are necessary. Although prevention of organ rejection is currently achieved with pharmacological immune suppression, the undesirable side effects of this method have incited interest in novel methods to overcome the immune barrier. One such novel method of preventing immune reaction is immuno-isolation, in which the non-autologous tissues are physically isolated from the host tissues by placement in devices with perm-selective membranes. The membranes of these devices allow release of the therapeutic product required from the transplanted tissues, as well as diffusion of nutrients and waste necessary for survival of the non-autologous tissues. The membranes also prevent host immune mediators from contacting the non-autologous cells, thus preventing immune rejection. This technology has been tested for efficacy in large animal models, and is currently in the process of clinical trials in humans. This review will discuss the progress made in using immuno-isolation of non-autologous tissues in large animals. Immuno-isolation can be subdivided into two major areas of interest based on whether the non-autologous tissue used in the immuno-isolation device is genetically altered (gene therapy) or not. Studies using non-genetically altered non-autologous cells for immune-isolation have been dominated by the use of pancreatic islet cells for the treatment of diabetes. This work has been tested in large animal models of diabetes, including canine and primate model animals, and human clinical trials are underway. As well, there has also been work on treatment of neurological disorders such as Parkinson's disease or chronic pain using non-autologous immuno-isolated adrenal chromaffin cells or dopaminergic PC12 cells in large animals such as sheep and primates. This work will be reviewed in detail as to the types of disorders, immuno-isolation devices used and the type of large animals involved. Immune-isolation for gene therapy is a more recently developed field of research. In this case, the non-autologous cells used are first genetically altered to secrete a recombinant therapeutic product before placement in the immune-isolation devices. Genetic engineering of the non-autologous cells is beneficial, as it allows the use of a cell type that tolerates well the environment of the immune-isolation device, while still delivering the therapeutic product of interest. This form of gene therapy has been tested in our laboratory for delivery of marker products such as human growth hormone to canines. As several large animal models of human genetic disorders are available, such as canines affected with hemophilia or the lysosomal storage disease mucopolysaccharidosis, testing the efficacy of immuno-isolation for gene therapy in large animal models is an important prelude to human clinical trials. This review will discuss the topics outlined above, as well as some further considerations of the usefulness of large animal models in studying immune-isolation for non-autologous transplantation. Large animals may be more appropriate model organisms than rodents in which to study immune-isolation, as issues such as biocompatibility and immune response in a larger animal can be addressed. As well, large animal studies of immune isolation may provide data that are more relevant than rodent studies to the eventual application to human clinical trials.