Jane de Lartigue, PhD

in the late 1980s. Since then, these tyrosine

kinases have also been proven to transmit a variety of signals into the cells with many biological consequences, adding to the interest in the targetability of the JAK pathway.

Dysregulation of the JAK pathway plays a role in the development of numerous tumor types; it is particularly central to the pathophysiology of myelofibrosis (MF) and has long been recognized as a potentially valuable therapeutic target in that malignancy. Ruxolitinib (Jaka ), the first JAK inhibitor to gain FDA approval, has become a centerpiece in the treatment of patients with MF. JAK inhibition has not proved effective in other tumor types, and thus far, no other JAK-targeting therapies have become available.

That picture, however, may be changing. A better understanding of the complexities of JAK signaling in normal and cancerous cells and the design of rational drug combinations are poised to expand the use of JAK inhibitors in anticancer therapy for hematologic malignancies, and possibly solid tumors as well. Ruxolitinib continues to be explored in multiple malignancies, and several promising novel agents are being evaluated in clinical trials (TABLE).

JAKs are now known as the Janus kinases, after the 2-faced Roman god of the same name, in recognition of their most striking feature: the presence of 2 near-identical kinase domains. There are 4 known members of the JAK family: JAK1, JAK2, JAK3, and TYK2, all sharing 7 conserved JAK homology (JH) domains within their protein structures.

At one end of the protein are JH1 and JH2, which comprise the 2 kinase domains. The rst has tyrosine kinase activity, while the second is a pseudokinase domain with no catalytic activity that is thought to negatively regulate the rst.3,4 At the other end, 3 of the JH domains collectively make up a larger FERM domain through which the JAKs are recruited to and bound by the cell-surface receptors that activate JAK signaling. A third domain, SH2, facilitates signal transducer and activator of transcription (STAT) signaling&nbsp;(FIGURE 1 ).

Cytokines and their receptors are among the most significant activators of the JAK pathway. Unlike tyrosine kinase receptors, cytokine receptors have no catalytic activity and are reliant upon the JAKs to perform kinase functions for them. JAKs bind to the part of the cytokine receptor that protrudes into the cell via their FERM domain. When the cytokine receptor is activated, it pairs up with another receptor molecule and this brings 2 JAKs close enough that they are able to phosphorylate and activate one another.

Activated JAKs then phosphorylate other target proteins, including the cytokine receptor to which they are bound, forming a binding platform for proteins containing an SH2 domain, primarily STAT proteins. Once they dock to the cytokine receptor, the STATs themselves are phosphorylated by the JAK proteins and become activated (FIGURE 2).

Activated STATs subsequently disengage from the cytokine receptor, pair up with one another and move into the nucleus, where they attach to specific sequences within the DNA to either activate or, less commonly, repress the transcription of a host of target genes. In this way, the interaction between JAKs and STATs transmits a signal from outside of the cell into the nucleus to coordinate a response to the signal.

Although this at first appears to be a relatively simple signaling cascade, the activity is complicated by the fact that there are 7 STATs. The diversity of possible JAK-STAT pairings, along with the range of different ligands and receptors that can activate JAKs upstream, allows for the multiplicity of biolog- ical effects coordinated by this pathway.

In addition, the pathway is negatively regulated in several ways. One involves protein tyrosine phosphatases (PTPs), which are enzymes that can deactivate JAK by reversing its phosphorylation. The suppressors of cytokine signaling (SOCS) are a family of proteins that are activated by STATs, and turn off the JAK-STAT pathway, thereby forming a negative feedback mechanism. They do this in sev- eral ways: directly inhibiting JAK kinase activity; blocking recruitment of downstream effector pro- teins; or mediating proteasomal degradation of JAKs. Finally, protein inhibitors of activated STATs (PIAS) block the activity of STATs in the nucleus through mechanisms that remain somewhat unclear.

Further adding to the complexity is the significant cross-talk with other signaling pathways, of which we currently have a relatively poor understanding. The best characterized interaction is with the mitogen-activated protein kinase pathway, which crosses paths with JAK-STAT signaling at multiple points.

As a result of its myriad roles in essential cellular processes such as growth, proliferation, survival, inflammation, and immunity, JAK pathway signaling is tightly regulated in normal cells by a delicate balance between activators and inhibitors.

Disruption of this balance and aberrant activation of the JAK pathway contributes to the development and progression of numerous cancer types by promoting many of the hallmark abilities of cancer cells, such as unchecked cell growth and proliferation. Because of its important role in regulating cytokine signaling, dysregulation of the JAK pathway can also contribute to in ammation and immunosuppression in the tumor microenvironment.

The mechanisms by which the JAK pathway becomes dysregulated include mutations in the receptors that sit upstream of JAK, which drive persistent activation of JAK, or in downstream effectors, which would bypass the need for JAK activation. Alternatively, the function of molecules involved in deactivation of the pathway, such as PTPs and SOCS, may be lost.

Fusions, mutations, deletions, and insertions in the JAK1 and JAK3 genes have been noted in intestinal, skin, stomach, and endometrial cancers, among oth- ers, whereas mutations in JAK2 have been identi ed in various hematologic malignancies.

In particular, the JAK pathway is central to the pathogenesis of myeloproliferative neoplasms (MPNs), including MF. These are cancers of the blood stem cells that cause an overproduction of white or red blood cells or platelets. Three mutually exclusive activating mutations in JAK2 are found in approxi- mately 90% of patients with MF. The most commonly observed mutation is JAK2 V617F, which drives ligand-independent activation of the JAK pathway, and is observed in 40% to 60% of patients.

Activation of STATs is also commonly reported in several cancer types, including prostate cancer, nonsmall cell lung cancer (NSCLC), glioblastoma, and thyroid cancer, but is particularly prevalent in breast cancer. Upstream of JAK, increased expression of the cytokine interleukin-6 (IL-6) is observed almost universally across tumor types.

Ruxolitinib, a small-molecule inhibitor of JAK1 and JAK2, received FDA approval in 2011 for the treatment of patients with intermediate- or high-risk MF, including primary MF, or MF that follows polycythemia vera (PV) or essential thrombocythemia.

Approval in this indication was based on data from 2 randomized phase III trials, COMFORT-I and COMFORT-II, which collectively involved more than 500 patients. Ruxolitinib more effectively reduced spleen size and MF symptom burden compared with placebo (COMFORT-I) and best available therapy (BAT; COMFORT-II). These studies also demonstrated an improvement in overall survival (OS) with ruxolitinib treatment. Ruxolitinib was well tolerated; grade 3/4 adverse events (AEs) included thrombocytopenia and anemia, but these were manageable and rarely led to treatment discontinuation.

The survival bene t was con rmed in recently published 5-year follow-up data for both trials, which also showed that long-term treatment with ruxolitinib was safe, with no signi cant cumu- lative toxicity. Long-term data from COMFORT-I also suggested that ruxolitinib improved patient quality of life.12-15 Various clinical trials of ruxolitinib are ongoing, including in other MPNs, hematologic malignancies, and solid tumors.

Although there are no other FDA-approved JAK inhibitors, other drugs have generated positive data in phase III clinical trials. Momelotinib, also a JAK1/2 inhibitor, has been evaluated in the phase III SIMPLIFY trials in patients with MF.

In SIMPLIFY-1, momelotinib was compared with ruxolitinib in patients with JAK inhibitorna&iuml;ve MF. The noninferiority of momelotinib was established in terms of the primary endpoint of a &ge;35% reduction in spleen volume at 24 weeks. However, ruxolitinib was superior to momelotinib in the key secondary endpoint of a &ge;50% reduction in total symptom score (42.2% vs 28.4%, respectively). There was also a higher rate of infections and treatment-emergent peripheral neuropathy with momelotinib.

Conversely, in the SIMPLIFY-2 trial in patients previously treated with ruxolitinib, participants who received momelotinib for 24 weeks did not experience superior outcomes for splenic response (&ge;35% reduction in spleen volume) compared with BAT (6.7% vs 5.8%, respectively). Nevertheless, momelotinib was signi cantly better than BAT at improving disease-related symptoms, de ned as a &ge;50% reduction in total symptom score (26.2% vs 5.9%) and in the transfusion independence rate (43.3% vs 21.2%). Given these mixed results, momelotinib&rsquo;s future is uncertain as researchers attempt to deter- mine the best next steps in its evaluation.

Pacritinib is a small-molecule inhibitor of JAK2 and the cytokine receptor FLT3. This drug has shown significant efficacy in a phase III clinical program in MF. In the PERSIST-1 trial, 327 patients were randomized to receive pacritinib or BAT as rst-line treatment. At 24-weeks, spleen volume was reduced by &ge;35% in 19.1% of patients treated with pacritinib compared with just 5% of those treated with BAT.

In the PERSIST-2 trial, pacritinib (at 2 different dosages400 mg once daily or 200 mg twice daily) was also compared with BAT in 311 patients with MF and thrombocytopenia. The primary endpoint of a &ge;35% reduction in spleen volume at 24 weeks was achieved in a significantly greater number of patients treated with either dosage of pacritinib than in those who received BAT (18.1% vs 2%, respectively). In terms of this primary endpoint, the twice-daily dose was more effective than the once-daily regimen (21.6% vs 14.7%, respectively). Pacritinib was also superior in terms of total symptom score, with 25% of patients demonstrating a &ge;50% reduction compared with 14% for BAT. Additionally, pacritinib reduced transfusion dependence.

Despite this display of substantial ef cacy, higher death rates from intracranial hemorrhage, heart fail- ure, and cardiac arrest in the PERSIST-2 trial prompted the FDA to place a clinical hold on pacritinib in February 2016. However, less than a year later, CTI Bio-Pharma, the company developing the drug, reported that the clinical hold had been lifted after submission of nal data from the PERSIST trials.

A new clinical trial examining the safety of pacritinib in patients with primary MF who have previously received ruxolitinib has been initiated (PAC203; NCT03165734).

The development of several other promising JAK inhibitors has been discontinued due to severe toxicities. AZD1480 caused neurological AEs, and lestaurtinib (CEP-701) resulted in frequent gastrointestinal toxicity. Fedratinib (SAR302503), a selective JAK2 inhibitor that showed significant efficacy in patients with MF in the phase III JAKARTA-1 trial, was put on a clinical hold by the FDA in 2013 after reports of potential cases of Wernicke encephalopathy. Sano subsequently discontinued its development.

However, fedratinib may be making a comeback. The rights for global development have been acquired by Impact Biomedicines. The company announced that the clinical hold was lifted based on a review of addi- tional data from 18 studies in more than 800 patients, including JAKARTA-1 and the follow-up JAKARTA-2 study, which recruited patients who were resistant to all available therapies including ruxolitinib.

In JAKARTA-2, 55% of patients treated with fedratinib experienced a &ge;35% reduction in spleen size. The most common AEs were anemia, nausea, diarrhea, and vomiting, and there were no cases of encephalopathy. No clinical trials of fedratinib are ongoing, but Impact Biomedicines plans to commercialize the drug for patients with MF and PV.21-23

The complexity of the JAK pathway, the pleiotropic nature of its biological effects, and substantial amount of cross-talk between it and other important signaling pathways presents a signi cant challenge to the effectiveness of JAK inhibitors. The development of rational combinations of drugs could help to improve outcomes and these are starting to be studied in clinical trials.

Preclinical study results have suggested synergy between JAK inhibitors and epigenetic modi ers, such as histone deacetylase and DNA methyltransferase inhibitors (DNMTs); agents targeting the Hedgehog (Hh) pathway, which plays a critical role in hematopoietic cell development and stem cell survival; inhibitors of PI3K, a key downstream effector of JAK signaling; and immune checkpoint inhibitors targeting PD-1 and its ligands.

The results of an ongoing phase II trial of ruxoli- tinib plus azacitidine, a DNMT inhibitor, in patients with MPNs, including MF, and myelodysplastic syn- drome (MDS) were recently published. Among the 35 patients evaluable for response, 9 patients (45%) responded after the addition of azacytidine and 64% experienced >50% reduction in palpable splenomeg- aly at 24 weeks.24 Results from an ongoing phase I/ II study of ruxolitinib plus decitabine, also a DNMT inhibitor, in accelerated phase MPN or post-MPN acute myeloid leukemia demonstrated a complete response (CR) or CR with incomplete blood count recovery in 33% of patients.25 A phase I/II study of ruxolitinib in combination with the Hh inhibitor sonidegib in patients with MF is ongoing, but not actively recruiting participants. The regimen has demonstrated a &ge;35% reduction in spleen size at 24 weeks in 44% of patients and was well tolerated.

Meanwhile, the combination of JAK and PI3K inhibition has also been investigated in early-stage clinical trials. The phase Ib HARMONY trial showed promise for the combination of ruxolitinib plus buparlisib, and although studies of that combination are no longer ongoing, ruxolitinib is being evaluated in combinations with TGR-1202 and INCB050465, which are inhibitors of the delta isoform of PI3K. Pre- liminary data from the TGR-1202 study showed that the combination was well tolerated, with a clinical bene t rate of 89%.

Despite initial promise, JAK inhibitors have suffered several high-profile failures in clinical trials in patients with solid tumors. Incyte has halted its development of ruxolitinib in solid malignancies, following lack of efficacy in patients with pancreatic and colon cancers. However, combination therapy also could help to boost ef cacy in this instance and several trials involving other JAK inhibitors are underway.

For example, there is evidence that activation of the JAK pathway could be a contributing factor to EGFR inhibitor resistance in EGFR-mutant NSCLC. A phase I study of ruxolitinib in combination with afatinib (Gilotrif) in patients with acquired resistance to EGFR inhibitors is ongoing, and preliminary data were recently presented. The combination demon- strated promising antitumor activity and was well tolerated.28 Also in this setting, itacitinib, a JAK1 inhibitor, is being evaluated in combination with osimertinib (Tagrisso).

There is also some interest in targeting other com- ponents of the JAK pathway, with the cytokine IL-6 emerging as the most promising target thus far. Mono- clonal antibodies have been developed against IL-6, primarily for the treatment of in ammatory diseases such as rheumatoid arthritis. Siltuximab (Sylvant), an IL-6 inhibitor, is being evaluated in multiple myeloma.

As the principal downstream target of JAK signaling, the STAT proteins also make rational targets, but have proved to be a therapeutic challenge thus far. Several types of STAT inhibitors, predominantly aimed at STAT3 and STAT5, have been developed, including small-molecule inhibitors and, more recently, antisense oligonucleotides. The latter are short stretches of DNA or RNA that can bind to specific stretches of messenger RNA, and prevent its translation into a protein. AZD1950 is one example of this technology that has demonstrated single-agent antitumor activity in patients with treatment-refractory lymphoma or NSCLC.

Wilks AF. Two putative protein-tyrosine kinases identi ed by application of the polymerase chain reaction. Proc Natl Acad Sci U S A. 1989;86(5):1603-1607.

Wilks AF. The JAK kinases: not just another kinase drug discovery target. Semin Cell Dev Biol. 2008;19(4):319-328. doi: 10.1016/j.semcdb.2008.07.020.

Articles

Since the identification of the role of the JAK kinase family&nbsp;in the late 1980s, awareness of this&nbsp;has grown significantly. These tyrosine<br />
kinases have been proven to transmit a variety of signals into the cells with many biological consequences, adding to the interest in the targetability of the JAK pathway. However,&nbsp;better understanding of the complexities of JAK signaling are being evaluated in clinical trials.

Novel Approaches Tackle the JAK Pathway Beyond Ruxolitinib and Myelofibrosis

Chimeric antigen receptor (CAR) T cells have hit the prime time, with 2 FDA approvals already for this class of cellbased therapy. Undoubtedly this will be a game changer for patients with B-cell malignancies who have a small number of treatment options; however, questions regarding the realworld application of CAR T-cell therapies remain. Several challenges may affect the adoption of CAR T-cell therapy, which range from the logistical complexities of therapeutic development to the potentially fatal toxicities of treatment, as Stephan A. Grupp, MD, PhD, indicated yesterday in his keynote address. A number of innovations are already entering the scene, including next-generation &ldquo;armored&rdquo; CARs and the application of state-of-the-art genome-editing technologies. These innovations and more will be discussed during tomorrow&rsquo;s plenary session entitled &ldquo;Genetically Modified Cell Therapy,&rdquo; starting with an explanation of gene-edited CAR T cells by Martin Pule, PhD. It will be several years before the true impact of CAR T cells becomes apparent, but it is evident that long-term success requires unlocking the potential of CAR T-cell therapy beyond B-cell malignancies, particularly in solid tumors, which account for the vast majority of cancer deaths.

CAR T-cell therapy falls under the banner of adoptive cellular therapy (ACT), in which the effector cells of the immune system, predominantly the T cells, are transplanted into a patient. The graft-versus-tumor effect observed in patients undergoing allogeneic hematopoietic stem cell transplantation served as the stimulus for pursuing ACT. T cells present in the transplanted material were seemingly capable of mounting an effective antitumor immune response, but this was limited by the development of graft-versus host-disease (GvHD), wherein the transplanted T cells also attacked healthy tissue, causing severe toxicity.

Genetically modifying host T cells to endow them with tumor-killing specificity originated in the 1980s. Seminal studies demonstrated that replacing the region of the T-cell receptor (TCR) responsible for antigen recognition with the single- chain variable fragment (scFv) of an antibody could couple antibody specificity for a tumorassociated antigen (TAA) with the T-cell activating machinery of the TCR.

This permitted direct activation of T cells by tumor cells that express the target antigen, in a manner that bypasses the need for antigen presentation by the major histocompatibility complex.

CAR T cells are generated by collecting host T cells through leukapheresis, where the CAR is introduced to the T cells through a viral vector. Clinical trials of the resulting CAR T cells, which can then be infused back into the patient, were pioneered by investigators at Memorial Sloan Kettering Cancer Center and the University of Pennsylvania.

The prototypical CAR was composed of an extracellular portion, consisting of the antibody scFv, joined to a hinge or spacer and then a transmembrane domain that spans into the cell and links to an intracellular signaling region.

The latter mediates T-cell activation and was most often derived from the CD3&zeta; chain of the TCR, which harbors 3 immunoreceptor tyrosine- based activation motifs (ITAMs). When the CAR engages its target antigen, the ITAMs become phosphorylated and serve as adaptors for a panel of signaling proteins that coordinate T-cell activation and proliferation.

Subsequent generations have been developed to improve on the efficacy of the first generation. The realization that full activation of T cells required 2 signals led to the incorporation of costimulatory domains in second- and third-generation CARs, with enhanced T-cell activity and persistence and greater antitumor efficacy.

In this rapidly evolving field of immuno-oncolytic therapeutic development, the most clinically advanced CAR T cells are currently those engineered against CD19, a cell surface receptor constitutively expressed in hematologic malignancies. Several pharmaceutical companies raced to be the first to secure regulatory approval, but ultimately the firstto- market advantage went to Novartis&rsquo; CTL019 (tisagenlecleucel; Kymriah).

In August, tisagenlecleucel was approved for the treatment of pediatric and young-adult patients (up to 25 years) with relapsed/refractory B-cell precursor acute lymphoblastic leukemia (ALL). The efficacy and safety of tisagenlecleucel was demonstrated in the phase II ELIANA trial that led to its approval for this indication. A total of 68 pediatric and young-adult patients with CD19-positive B-cell precursor ALL were infused with a single dose of CAR T cells. After a follow-up of 3 months, 63 patients treated with tisagenlecleucel were evaluable for response; 83% achieved complete remission (CR) or incomplete CR within 3 months of infusion with minimal residual disease (MRD)negative status achieved in the bone marrow.

Hot on Novartis&rsquo; heels was Kite Pharma, whose CD19-targeted CAR T-cell therapy, axicabtagene ciloleucel (KTE-C19; axi-cel; Yescarta), received approval as treatment for adults with relapsed or refractory non-Hodgkin lymphoma based on results of the ZUMA-1 trial in October. Axi-cel was administered as a single infusion of modified autologous T cells at a target dose of 2 x 106 CAR-positive T cells/kg.

As of January 2017, 101 patients enrolled in the ZUMA-1 trial were treated with axi-cel. The study completed its primary endpoint of combined objective response rate (ORR) for patients with diffuse large B-cell lymphoma (DLBCL), primary mediastinal B-cell lymphoma (PMBCL), or transformed follicular lymphoma (FL) after 6 months&rsquo; follow-up. The majority of the population enrolled in the ZUMA-1 trial were patients with DLBCL (n = 77); the ORR and CR rates in this population were 82% and 49%, respectively.7 Among the patients with PMBCL/ FL (n = 24), the ORR was 83% and the CR rate was 71%.

Both axicabtagene ciloleucel and tisagenlecleucel are also being evaluated in clinical trials for additional indications (TABLE 1). Results from the ongoing phase I/II ZUMA-3 trial for axicabtagene ciloleucel in adults with relapsed/refractory B-cell ALL were reported at the 2017 ASCO Annual Meeting. Among 8 patients evaluable for efficacy, 6 achieved an MRD-negative CR or a CR with partial or incomplete hematopoietic recovery.

The ongoing phase II JULIET trial is investigating the safety and efficacy of tisagenlecleucel treatment in adult patients with relapsed/refractory DLBCL. Preliminary results from JULIET were presented at the 2017 International Conference on Malignant Lymphoma. Of 85 patients who received a single dose of tisagenlecleucel, 51 were evaluable for tisagenlecleucel response with at least 3 months&rsquo; follow-up. For these patients, the ORR was 59% (95% CI, 44.2%-72.4%); 43% of patients had a CR, and 16% had a partial remission.

Juno Therapeutics&rsquo; CD19-targeted CAR T-cell therapy, JCAR015, was originally slated to secure regulatory approval first. However, the development of JCAR015 was discontinued following 2 clinical holds placed on the phase II ROCKET trial, in which an adverse safety profile revealed an increased incidence of deaths due to cerebral edema, as was discussed yesterday in a presentation by Mark J. Gilbert, MD, of neurotoxicities seen in the ROCKET trial.

Despite the disappointing safety concerns with JCAR015, Juno Therapeutics is continuing clinical development of a substantial CAR T-cell therapy pipeline. The ongoing phase I TRANSCEND study is investigating the safety and efficacy of another pipeline CD19-targeted CAR T-cell therapy, JCAR017, and has enrolled patients with relapsed/refractory B-cell NHL, including DLBCL, PMBCL, FL, and mantle cell lymphoma. Interim data were presented at the 2016 ASH Annual Meeting of evaluable patients following treatment with fludarabine, cyclophosphamide, and JCAR017. The majority of the population (n = 25) had DLBCL. Of 20 patients treated with lower-dose JCAR017, the ORR was 80% and 60% achieved a CR.

Juno&rsquo;s experience highlights a complicating factor of clinical implementation of CAR T-cell therapiesthe potential for severe, sometimes fatal, treatment-related toxicities. Most prominent are cytokine release syndrome (CRS)&mdash;a severe immune reaction resulting from the release of cytokines from the targeted cancer cells and by immune effector cells recruited to the area&mdash;and neurotoxicity, resulting in brain swelling, altered consciousness, and seizures.

In the ELIANA trial, CRS occurred in 78% of patients, with 27% of grade 4 severity; however, there were no CRS-related deaths. Grade 3 neuropsychiatric events occurred in 15% of patients, but there was no incidence of cerebral edema. In the ZUMA-1 trial, adverse events of CRS and neurotoxicity of grade &ge;3 severity occurred in 13% and 28% of patients, respectively; all cases resolved except for a grade 1 memory impairment. The first death from cerebral edema in this trial was recently reported, and although studies will continue a, patient consent forms will now be amended to reflect the risk.

All CAR T-cell therapies developed to date cause these and other toxicities in at least some patients, but there is substantial variation in the severity and type of toxicity. A prospective registry and monitoring study were both proposed to assess the safety of tisagenlecleucel for up to 15 years after initiation to gain a better understanding of the toxicity profile and longterm effects of the approved treatment.

Understanding the reasons for these toxicities is a central focus of ongoing research, in addition to the question of longterm safety relating to the risks of using viral vectors and the persistence of CAR T cells in the body after treatment.

The success of CD19-targeted CAR T cells has prompted exploration of other TAAs. Of particular note is the B-cell maturation antigen (BCMA), a member of the tumor necrosis factor receptor superfamily. Until recently, CAR T-cell therapies had generated mixed results in multiple myeloma, an incurable B-cell malignancy, but BCMA is expressed nearly universally on multiple myeloma cells, making it a promising target for CAR T-cell therapy in this tumor type.

Bluebird Bio and the Chinese company Nanjing Legend Biotech are pursuing the development of rival CAR T-cell therapies for the treatment of relapsed/refractory multiple myeloma. Both therapies created a buzz at this year&rsquo;s ASCO Annual Meeting.

Nanjing Legend Biotech&rsquo;s LCAR-B38M is uniquely engineered, in which the antigen-binding region of the CAR is a non-scFv structure targeting 2 regions of the BCMA antigen. Researchers hope this design will allow for more effective antitumor activity. To address safety concerns, LCAR-B38M is also being administered as 3 smaller infusions rather than a single large infusion.12 Nineteen patients have been treated with LCAR-B38M to date, 7 of whom were monitored for more than 6 months. The ORR was 100%; 95% of patients (n = 18) reached near CR without relapse, including 7 patients who achieved CR and were MRD-negative at a median follow-up of 6 months. CRS was observed in 74% of patients but was mostly mild or manageable. Two patients experienced grade 3 or 4 CRS; the patient with grade 4 CRS recovered fully after treatment.

In Bluebird Bio&rsquo;s phase I trial, bb2121 was administered to 11 patients with relapsed/refractory multiple myeloma in 4 escalating dose cohorts (doses of 5, 15, 45, and 80 x 107 CAR T cells). The ORR in the 6 evaluable patients treated with doses of &ge;15 x 107 CAR T cells was 100%, including stringent CR in 2 patients and MRD-negative responses in 2 patients. There were no reported adverse events of CRS or neurotoxicity of &ge;grade 3, and there were no dose-limiting toxicities.

The unprecedented clinical efficacy of CAR T-cell therapy in patients with hematologic malignancies has not translated into solid tumors to date. However, the safety and efficacy of CAR T-cell therapy for solid tumors, the major cause of cancer-related mortality, have been the focus of a growing number of clinical trials.

Solid tumors are thought to present a number of challenges to the effectiveness of CAR T-cell therapy, including in selecting appropriate target antigens given the substantial heterogeneity even within an individual tumor. CD19 has proven to be a success story in part because of its near uniform expression on B-cell malignancies and minimal presence on normal B cells.

The success of drugs targeting members of the human epidermal growth factor receptor (HER) protein family makes them obvious targets for CAR T-cell therapy in solid tumors. Extracellular glycoproteins such as mucin-1, mucin-16, and mesothelin, as well as other cell surface proteins like fibroblast activation protein and L1 cell adhesion molecule, are also being explored for targeting moieties (TABLE 2).

Positive data have been demonstrated in trials of HER2- targeted CARs in patients with sarcoma. Findings from a small phase I study were presented at the 2017 ASCO meeting. Among 6 patients with refractory or metastatic HER2-positive sarcoma, 1 patient achieved a CR, 2 had stable disease, and 3 had progressive disease. The median overall survival in 2 patients who were still alive was 14.2 months.

CARsgen Therapeutics is developing CAR T cells that target glypican-3 (GPC-3), a cell surface protein that is highly expressed in hepatocellular carcinoma (HCC). Phase I clinical trial data, presented at the 2017 ASCO meeting, showed that it was a safe and feasible treatment for Chinese patients with relapsed/refractory GPC-3positive HCC.

This study has been terminated for reasons not related to safety, efficacy,or feasibility. A phase II trial in this indication is planned, and a phase I/II trial in pediatric solid tumors is ongoing.

The need to expand the impact of CAR T-cell therapy by addressing challenges posed by solid tumors, as well as to improve its safety and efficacy, is driving the development of several innovative strategies for next-generation CAR design. One strategy to overcome TAA heterogeneity in solid tumors is to broaden the specificity of CAR T cells, either by transferring more than 1 population of CAR T cells targeting different antigens or by producing CARs that target multiple TAAs, so-called tandem or bispecific CARs. Several of the latter are currently being evaluated in clinical trials (NCT03097770, NCT03097770, NCT03271515, NCT03271632).

The biopharmaceutical company Endocyte is collaborating with Seattle Children&rsquo;s Research Institute to develop a novel approach that could create a universal CAR T-cell system capable of targeting any TAA. Their CAR T cells are engineered to bind to small adaptor molecules composed of a fluorescein isothiocyanate (FITC) molecule conjugated to a ligand that targets overexpressed receptors on tumor cells. Instead of the CAR T cells binding to a TAA, an anti-FITC receptor on the surface of the CAR T cell will bind to the FITC end of the adaptor molecule, which allows the activation of the targeted TAA on tumor cells by the free ligand component of the adaptor molecule.

Another unique hurdle to CAR T-cell therapy in solid tumors is the tumor microenvironment, which presents physical, chemical, and biological barriers preventing entry to the tumor site. Many solid tumors are surrounded by a dense stroma and abnormal vasculature that impede access. The hypoxic, nutrient-deprived, acidic microenvironment is inhospitable for infiltrating T cells. In addition, both the tumor and the microenvironment exert immunosuppressive effects that hinder T-cell migration to the tumor site, and inhibitory immune checkpoints that switch off infiltrating T cells.

Armored CARs and TRUCKs (T cells redirected for universal cytokine killing) are being explored in clinical trials. These are designed to deliver a payload, such as cytokine interleukin-12 (IL-12), on antigen binding, which can help to make the tumor microenvironment more hospitable to immune cells. In a mouse model of ovarian cancer, IL-12secreting CAR T cells were found to be more effective and improved survival.

TRUCKs expressing catalase to protect T cells from oxidative stress, or heparanase, and to improve T cell penetration through the stroma, are also being explored.

To overcome the ability of tumor cells to suppress the antitumor immune response through exploitation of immune checkpoint pathways, combinatorial trials of CAR T-cell therapy and immune checkpoint inhibitors are underway. CAR T cells that express antibodies against the PD-1 and cytotoxic T-lymphocyte antigen-4 are in early clinical trials.

Efforts to make CAR T-cell therapies safer are also under way, in particular seeking ways to control CAR T-cell activity once the cells have been infused into the patient, to minimize their impact on healthy cells. One such strategy being explored is to introduce a safety switch into the CAR design, such as suicide genes or inducible apoptotic molecules.

Another is the development of so-called inhibitory CARs, designed to provide a T-cell inhibitory signal when engaging an antigen on healthy cells.

A particularly exciting research avenue is the application of state-of-the-art genome-editing technologies to the production of CAR T cells. The CRISPR/Cas9 system could be used to create a coveted &ldquo;off the shelf&rdquo; CAR T-cell therapy from universal donor cells, which would help to overcome some of the complexities associated with the manufacturing of CAR T cells.

The use of donor T cells is limited by the presence of endogenous TCR and human leukocyte antigens on their surface, which can activate an immune response and result in GvHD. Genome editing is being used to knock out the TCR and HLA genes to bypass these problems, in addition to knocking out genes that can dampen the antitumor immune response, such as PD-1.

The first human trial of CRISPR immunotherapy was recently given the green light by the FDA and will evaluate the feasibility and safety of knocking out the PD-1 gene in the T cells of patients with lung cancer (NCT02793856). Another CRISPR study seeking FDA approval will knock out the PD-1 and TCR genes in NY-ESO-1targeted TCR-engineered T cells. CRISPR could also be used to improve the delivery of CAR genes. While viral delivery randomly inserts the CAR into the T-cell DNA, CRISPR could more precisely place it in a specific part of the genome. The potential benefit of this approach has been demonstrated in a preclinical study, in which CRISPR was used to insert a CAR into the T-cell receptor alpha constant locus, which generated a more uniform expression of the CAR and produced CAR T cells that outperformed traditional ones in mice with ALL.

Gross G, Waks T, Eshhar Z. Expression of immunoglobulin-T-cell receptor chimeric molecules as functional receptors with antibody-type specificity. Proc Natl Acad Sci U S A. 1989;86(24):10024-10028.

Feinberg BA, Fillman J, Simoncini J, Nabhan C. CAR-T cells: the next era in immuno-oncology. Am J of Manag Care. 2017;23(spec no 2):SP48-SP52.

Zhang C, Liu J, Zhong JF, Zhang X. Engineering CAR-T cells. Biomark Res. 2017;5:22. doi: 10.1186/s40364-017-0102-y.

Wang Z, Guo Y, Han W. Current status and perspectives of chimeric antigen receptor modified T cells for cancer treatment. Protein Cell. May 2, 2017. doi: 10.1007/s13238-017-0400-z.

Jackson HJ, Rafiq S, Brentjens RJ. Driving CAR T-cells forward. Nat Rev Clin Oncol. 2016;13(6):370-383. doi: 10.1038/nrclinonc.2016.36.

Chimeric antigen receptor T cells have hit the prime time, with 2 FDA approvals already for this class of cellbased therapy. Undoubtedly this will be a game changer for patients with B-cell malignancies who have a small number of treatment options; however, questions regarding the realworld application of CAR T-cell therapies remain.

CAR T-Cell Therapy Hits Prime Time; Challenges Remain

Metastatic disease accounts for the vast majority of cancer-related deaths. Ensuring a definitive diagnosis and the most effective treatment in a timely fashion is essential for extending life expectancy.

Despite a spurt in knowledge regarding treatment-related biomarkers and availability of the latest technologies to probe these in an efficient manner, the uptake of these technologies in community practice has been highly suboptimal. In some cases, this lag can create a perception that community practices are not &ldquo;keeping up&rdquo; relative to academic centers with respect to the use of these latest technologies.

Ralph Boccia, MD, FACP, is medical director of the Center for Cancer and Blood Disorders and clinical associate professor of medicine at Georgetown University, as well as a consulting medical director of the International Oncology Network clinical program and chairman of their medical advisory panel. He has been involved in assessing the clinical value of some of these technologies and agrees that the use of genomics beyond the usual breast and lung cancer assays has had slow uptake in general practice in large part because universal, broadbased understanding of how best to use these tools is lacking.

The Challenge of Accurate Diagnosis in Metastatic Disease

With a clear need for more practice-friendly solutions, the ultimate goal is to integrate accurate diagnosis with comprehensive biomarker information to achieve precision therapy that reflects the heterogeneity of the disease. A San Diego-based company has created a suite of genomic-based tests called bioT3 that aims to provide community oncologists with the tools to achieve these goals, while being conscious of high actionability, low tissue use, cost and turn-around time.

Identification of a single confirmed diagnosis of tumor type and subtype and of the most effective therapeutic options for metastatic disease are critical aspects for ensuring optimal treatment and extending life expectancy. Typically, a diagnosis is made using standard pathologic techniques: examining the morphology of the cancer cells and performing immunohistochemical (IHC) analyses to identify specific cellular markers. However, there is frequent ambiguity in the diagnosis of metastatic cancer; each year in the United States, as many as 90,000 to 130,000 newly diagnosed metastatic patients have unclear diagnoses, about 25% to 40% of which are classified as cancer of unknown primary (CUP).

IHC, which is the current standard of care for tumor classification has been shown to be accurate in only ~2 out of 3 metastatic cases,

consistent with older studies which also point to similar challenges related to achieving an accurate diagnosis.

More recently, a recent study at MD Anderson

shed light on the problem of diagnostic ambiguity. A total of 2714 cancer cases were reexamined by an expert pathologist. In a surprisingly high number of cases (25%), the diagnosis was changed upon the expert review. In recent years, molecular tumor classification has emerged as an important technique in tumor diagnostics that is proving to be effective in identifying tumor type. By providing a more accurate diagnoses, molecular classification can be used to help a physician choose the most effective treatment, particularly in the context of targeted therapy. Conversely, lack of a definitive diagnosis limits potential therapeutic options. F. Anthony Greco, MD, director of the Sarah Cannon Cancer Center in Nashville, Tennessee explained, &ldquo;Understanding where a tumor is coming from is extremely important [because] we treat cancers based on their site of origin. Once you feel confident with what type of cancer the patient has, there are a number of other molecular tests that are now standard.&rdquo;

Recently, there has been debate in the academic community on the taxonomy of cancers,

with some in the industry arguing for the use of sequencing alone to treat patients with unclear diagnosis.

Cohen and Settleman wrote a comprehensive review,

which put the debate into a practical clinical context. The authors (from Genentech and Calico) explain that use of driver mutation alone to treat patients is inconsistent with clinical evidence to-date. Firstly, most clinical trials have inclusion criteria that are cancer specific. Secondly, there are a variety of examples that show that targeted therapies linked to specific biomarkers show different responses depending on the cancer type. For example:

Vemurafenib is effective in patients with melanoma who have BRAF V600E mutations, while clinical study data did not show significant efficacy in colorectal cancer patients with the same mutation.

Overexpression of the human epidermal growth factor receptor 2 (HER2) is indicative of trastuzumab efficacy in breast cancer and gastric cancer, but not in non-small cell lung cancer (NSCLC).

Cetuximab is approved for patients with colorectal cancer who have wild-type KRAS mutational status, while clinical study data show that patients with advanced biliary tract cancer who have the same wildtype KRAS status do not benefit.

Tamoxifen is an established and effective therapy for patients with estrogen receptor-positive (ER+) breast cancer, but is not effective in patients with ER+ melanoma. Finally, without a single confirmed diagnosis, ease of access to and reimbursement of targeted agents can be problematic. Thus, with the ever-expanding list of targeted biomarker-driven agents tied to specific cancer types, accurate diagnosis is now more important than ever before.

Boccia agrees that biomarker information alone is unhelpful and may in fact hinder clinical decision making. He outlined the importance of genomic profiling technologies in this endeavor, &ldquo;When you offer people the profile that may show multiple abnormalities, it leaves them floundering with what to do with that information,&rdquo; he said. Boccia believes that BioT3 offers community practice doctors a valuable service by combining diagnosis and biomarker analysis in a single platform. &ldquo;We treat patients based on the tumor type or cellular context of the tumor rather than the sites of metastases, and simply knowing what biomarkers are present is not always helpful.&rdquo; Citing the example of BRAF mutations, he stressed that in these cases, it is critical to distinguish the primary tumor type.

&ldquo;We can now rely on some of these assays, with bioTheranostics being one of the sources using assays like BioT3, to help us do that, to better predict where tumors are coming from in patients in whom the presenting picture is not clear, so we can better initiate therapy based on that indication and interpretation.&rdquo;

CancerTYPE ID from bioTheranostics in San Diego is the most commonly used gene expression based tumor-classification assay. It identifies the type and subtype of a tumor by performing a realtime RT-PCR assay of 92 genes on formalin-fixed paraffin-embedded tissue biopsies. The resulting 92-gene expression profile can help diagnose 28 tumor types and 50 subtypes.

Boccia discussed the importance of improved diagnostic tools in the metastatic setting. &ldquo;Cancer-Type ID helps us more accurately identify the tumor type in a patient with advanced cancer when not obvious or apparent, or in situations where the presentation is unusual and the diagnosis or primary site is questionable.&rdquo;

Greco and other researchers at the Sarah Cannon Research Institute are among those who have been involved in the clinical evaluation of this assay. &ldquo;It&rsquo;s quite reliable in identifying the tissue of origin or the cancer type in patients with unclear diagnosis,&rdquo; Greco said. Important clinical studies that validated the platform include:

A large, blinded, multi-institutional study across Massachusetts General, Mayo and UCLA, CancerTYPE ID, which demonstrated 87% accuracy in identifying the 28 main tumor types and 82% accuracy for the 50 subtypes, with consistently high accuracy across several clinically relevant subsets (eg, cases with extremely limited tissue based on FNA)

A prospective trial of 252 patients with CUP, in which CancerTYPE ID was able to identify a single diagnosis of tumor type in 98% of patients, showed a 37% improvement in OS relative to historical controls, and identified 60% of patients who had tumors that were more responsive to site-directed therapy

A blinded comparator study of 122 patients with difficult to diagnose, poorly differentiated or undifferentiated tumors, in which CancerTYPE ID showed statistically higher accuracy overall versus IHC, the current standard of care for diagnosis of tumor type

&ldquo;When you put it together with clinical examination and standard pathology and other specialized pathology tests, in the majority of patients we can now tell where the cancer&rsquo;s coming from; then, we can treat according to the type of cancer they have rather than just using this sort of shotgun therapy as we&rsquo;ve been doing for 30 years, so-called empiric chemotherapy,&rdquo; Greco said.

Studies have also demonstrated the clinical utility of CancerTYPE ID. Interim results from a 28-site registry study, involving 134 patients, showed that its use changed the treatment decision in more than half of cases.

Further, a physician-reported clinical utility study

found that CancerTYPE ID identified a primary tumor type not previously considered in one-fourth of all patients. The test has been widely used across the US with more than 22,000 tumor samples analyzed to-date and is covered by Medicare Part B reimbursement with no patient copay, and by an increasing number of commercial insurance companies.

Once an accurate diagnosis has been established, an equally important step is to identify the mutational profile of a tumor. Because genetic alterations may be indicative of response or resistance to certain drugs, understanding the profile of a tumor is important to achieve optimal patient outcomes &ldquo;We now know that there are certain genetic abnormalities that you can treat with drugs that are on the market and FDA-approved, and several others have prominent associated clinical trials,&rdquo; explained Greco.

CancerTREATMENT NGS+ is designed to address 4 critical needs of the community oncologist that were inadequately addressed:

Low tissue use and rejection rates (&ldquo;QNS&rdquo;)

According to BioTheranostics, the company aims to address these needs through specific design aspects of their platform:

Actionability is defined strictly based on either NCCN recommendations for prominent cancer types or major phase II/III clinical trials, which they state will allow the use of minimal tissue while keeping the turnaround time and cost low

Use of a 50-gene Ion Torrent next-generation sequencing panel from Life Technologies, which offers high speed and coverage of the most actionable genes

A selected list of FISH and IHC tests that they believe will fill the key gaps in actionability, while keeping turnaround time, cost, and QNS low

Tumor-specific biomarker panels are offered for cancers with biomarker-driven FDA-approved treatment options, including NSCLC, CRC, breast cancer, and melanoma, while a more comprehensive panel is offered for all other cancers to identify potential driver mutations and clinical trials that a patient may be eligible to enter. Boccia explained that for these 5 cancer types, &ldquo;We have very well-studied, documented and proven uses for some of these tools.&rdquo; He continued, &ldquo;There are relationships where you can associate mutations in certain pathways with certain drug behaviors and responses.&rdquo; Clinicians can use these alterations to guide therapeutic decision-making. According to Boccia, &ldquo;There&rsquo;s no doubt that use of this profiling analysis among academic and community physicians can help us to plan therapies based on what we know about those biomarkers.&rdquo;

Greco pointed out that numerous companies now offer next-generation sequencing of biopsy specimens and, while some companies such as Foundation Medicine use a sequencing approach alone, others like bioTheranostics take a more comprehensive approach by including both genetic mutation and protein expression (IHC). Boccia explained the benefits of this approach, &ldquo;Besides sequencing, CTID provides, additionally, FISH and IHC testing, which further enhances high actionability of results; their information and reports are concise and offer these benefits.&rdquo;

BioTheranostics believes addition of IHC tests, which measure the level of a specific protein&rsquo;s expression, allows examination of new mechanisms relevant in melanoma and NSCLC such as PD-L1,

as well as older biomarkers such as HER2, ER, and PR expression relevant in breast cancer. For patients with NSCLC, they perform ALK rearrangement testing with FISH (vs NGS), because this is consistent with the inclusion criteria of the crizotinib pivotal studies,

By combining NGS with a specific list FISH/IHC tests, the company aims to achieve good turn-around times:

5-7 days for patients with clear diagnosis, which involves testing CancerTREATMENT NGS+ alone

7-10 days for patients with unclear diagnosis, which involves testing CancerTREATMENT NGS+ with CancerTYPE ID

&ldquo;Extraordinarily important to practices is fast turn-around,&rdquo; said Boccia. There is nothing worse than making patients wait long periods of time to get results that allow us to select and tailor the best therapies for patients rather than taking the old shotgun approach.&rdquo;Greco stated that he considers the cost of obtaining these tests by bioTheranostics to be competitive. Boccia agreed and explained why he felt cost to be an important factor, &ldquo;[bioTheranostics] cost is less than some of the other large labs currently offering biomarker analyses in a variety of ways; given rising health care costs in general, and payors sometimes challenging expensive testing, and given there are now new reimbursement models that have cost and risk sharing, cost becomes more and more important.&rdquo;

In the past several decades, we have begun to capitalize on technological advances and a deeper understanding of the molecular heterogeneity of cancer to improve diagnosis and treatment. The creation of more practice-friendly solutions is an important step in ensuring wider use of these state-of-the-art technologies aimed at more optimal patient outcomes.

Fong TH, Govindan R, Morgensztern D. Cancer of unknown primary.

Anderson GG, Weiss LM. Determining tissue of origin for metastatic cancers: meta-analysis and literature review of immunohistochemistry performance.

Weiss LM, Chu P, Schroeder B, et al. Blinded comparator study of immunohistochemical analysis versus a 92-gene cancer classifier in the diagnosis of the primary site in metastatic tumors.

Laouri M, Schroeder B, Chen E, Erlander G, Schnabel C. Diagnostic utility of molecular profiling for cancers of uncertain primary.

Raab SS, Grzybicki DM, Janosky JE, et al. Clinical impact and frequency of anatomic pathology errors in cancer diagnoses.

Middleton LP, Feeley TW, Albright HW, Walters R, Hamilton SH. Secondopinion pathologic review is a patient safety mechanism that helps reduce error and decrease waste.

Hoadley KA, Yau C, Wolf DM, et al. Multiplatform analysis of 12 cancer types reveals molecular classification within and across tissues of origin.

Ross et al., J Clin Oncol 32:5s, 2014 (suppl; abstr 11048)

Cohen, RL and Settleman J. From cancer genomics to precision oncology tissue&rsquo;s still an issue.

bioTheranostics. CancerTYPE ID. http://www.biotheranostics.com/ healthcare-professionals/hcp/ctid/. Accessed August 29, 2014.

Kerr SE, Schnabel C, Sullivan PS, et al. Multisite validation study to determine performance characteristics of a 92-gene molecular cancer classifier.

Hainsworth JD, Rubin MS, Spigel DR, et al. Molecular gene expression profiling to predict the tissue of origin and direct site-specific therapy in patients with carcinoma of unknown primary site: a prospective trial of the Sarah Cannon research institute.

Thomas SP et al. Molecular profiling with the 92-gene assay and decision-impact on cancer treatment: Interim results from a prospective, multidisciplinary study.

Kim B, Schroeder B, Schnabel C, Erlander G, Malin JL. Physician-reported clinical utility of the 92-gene molecular classifier in tumors with uncertain diagnosis following standard clinicopathologic evaluation.

Life Technologies. Ion Torrent&trade;. http://www.lifetechnologies.com/us/ en/home/brands/ion-torrent.html. Accessed August 29, 2014

Topalian SL, Hodi FS, Brahmer JR, et al. Safety, activity, and immune correlates of anti-PD-1 antibody in cancer.

Brahmer JR, Horn L, Gandhi L, et al. Nivolumab (antiPD-1, BMS-936558, ONO-4538) in patients (pts) with advanced non&ndash;small-cell lung cancer. Presented at the ASCO Meeting 2014. Chicago, IL. Abstract 8112.

Shaw AT, Kim DW, Nakagawa K, et al. Crizotinib versus chemotherapy in advanced ALK-positive lung cancer,

Leighl NB, Rekhtman N, Biermann WA, et al. Molecular testing for selection of patients with lung cancer for epidermal growth factor receptor and anaplastic lymphoma kinase tyrosine kinase inhibitors: American society of clinical oncology endorsement of the college of american pathologists/ international society for the study of lung cancer/association of molecular pathologists guideline.

BioT3 Integrates Diagnosis With Actionable Biomarkers in Metastatic Cancer

The 5-year survival rate for patients with metastatic colorectal cancer (CRC) remains dismal.

Liver metastases are a particularly common occurrence, developing in approximately half of all patients with colorectal cancer.

While surgical resection is the current standard of care, only 10% to 15% of patients are eligible at diagnosis.

Several lines of systemic chemotherapy and targeted agents are used in patients with unresectable disease, but these treatments are associated with significant toxicity, and patients eventually become refractory to them.

In an effort to improve outcomes for patients with metastatic CRC, liver-directed regional therapies that exploit the differential blood supply of the tumor and healthy tissue have been developed. Among them, selective internal radiation therapy (SIRT) has been proved safe and effective in a number of clinical settings. Here, we describe the clinical development of SIRT using SIR-Spheres microspheres, and the SIRFLOX trial, the results of which are eagerly anticipated.

External beam radiation therapy (ERT) has shown limited utility against primary tumors and metastases of the liver, as healthy liver tissue is highly sensitive to radiation. SIRT is a form of RT designed to specifically target the tumor with high doses of radiation, while sparing normal liver tissue.

Also known as radioembolization, SIRT capitalizes on the fact that healthy liver tissue derives the majority of its blood supply via the portal vein, while the tumor, which requires oxygenated blood, uses the hepatic artery. Via SIRT, doses of internal radiation up to 40 times higher than conventional RT can be noninvasively targeted to the liver.

FDA-Approved SIR-Spheres: Safety and Efficacy in Metastatic CRC

The radioisotope yttrium-90 is most commonly used with SIRT. A high-energy, beta-emitting isotope, it ensures localized spread of radiation as it penetrates only around 2 mm into the tissue. It is also beneficial because of its medically useful half-life of 64 hours and benign safety profile.

SIR-Spheres (Sirtex Medical Inc) microspheres are resin-based delivery devices for yttrium-90 administration, which are approved by the US Food and Drug Administration for the treatment of colorectal liver metastases. They consist of biocompatible polymer microspheres with an average diameter of 32.5 &mu;mabout the size of 4 red blood cells. They are injected into the hepatic artery via a femoral catheter and travel to and lodge in the tumor vasculature, where the yttrium-90 irradiates the tumor over its half-life of approximately 64 hours, inducing tumor cell death.

SIRT can be performed in the outpatient setting as it has a good safety profile. Postembolization syndrome (severe nausea, vomiting, and abdominal pain that requires ongoing hospitalization) is common with certain liver-directed therapies, such as drugeluting beads and transarterial chemoembolization, but is not a common occurence following after SIRT and typically doesn&rsquo;t require hospitalization when it does.

To reduce the risk of complications, careful patient selection and a meticulous pretreatment assessment are performed. Angiography is used to examine the arterial anatomy; extra-hepatic vessels branching off of the hepatic artery are prophylactically occluded to prevent delivery of microspheres outside the liver. Technetium- 99m-labelled macroaggregated albumin (99mTc- MAA), which closely mimics yttrium-90 distribution, is used to evaluate the degree of arterial shunting from the liver to the lungs and gastrointestinal (GI) tract. Shunting of >20% is a contraindication to therapy, as it can lead to radiation pneumonitis or GI ulceration.

The implantation procedure is performed approximately 1 week after assessment is completed. Dosimetry is based on whole liver infusion; the calculated activity (GBq) of the whole liver is multiplied by the ratio of the target site (the whole liver). Several methods of dosimetry are available, but the body surface area method is the most widely used and recommended.

FDA-approval of SIR-Spheres was granted in 2002 as a result of a trial of 74 patients with nonresectable liver metastases from primary CRC. SIR-Spheres were administered both as first-line therapy and in patients who had received prior chemotherapy. There was a significant improvement in response rate and time to progression.15 Since then, a number of prospective clinical trials have demonstrated the safety and efficacy of SIR-Spheres as first-line, second-line, and salvage therapy.

According to Michael Pishvaian, MD, PhD, assistant professor in the Department of Hematology/ Oncology at the Lombardi Comprehensive Cancer Center in Washington, DC, &ldquo;Traditionally, [Sir- Spheres] have been used primarily for refractory patients who have no other therapeutic options, but there&rsquo;s a growing body of data indicating that using them earlier in the course of therapy may improve therapeutic outcomes, and that combining them with chemotherapy concurrently is safe.&rdquo;

Several ongoing studies are investigating the utility of SIR-Spheres treatment in combination with standard chemotherapy earlier in the course of disease. One such study is the SIRFLOX trial (NCT00724503), an international, multicentered, randomized, controlled study evaluating the combination of SIRSpheres and mFOLFOX-6 chemotherapy (oxaliplatin, leucovorin, and 5-fluorouracil) compared with FOLFOX alone in patients with nonresectable liver metastases from primary CRC who have not yet received chemotherapy for advanced disease. Investigators were also allowed to add bevacizumab (Avastin) to either arm at their discretion. The study completed enrollment (532 patients) in April 2013; results are expected to be available in early 2015.

&ldquo;It is hypothesized that the addition of SIR-Spheres will improve progression-free survival in the SIRSpheres + FOLFOX arm compared with FOLFOX,&rdquo; stated Marwan Fakih, MD, professor of medical oncology and experimental therapeutics and director of GI medical oncology at City of Hope Comprehensive Cancer Center in Duarte, California. According to Fakih, &ldquo;The sample size to achieve an 80% power in confirming this hypothesis is 450 patients, so with over 530 patients enrolled, the study is well powered.&rdquo;

Providing further detail on the 2 treatment arms, Fakih explained, &ldquo;Patients on the SIR-Spheres arm receive SIR-Spheres during the first cycle of FOLFOX (day 4) and the dose of oxaliplatin is attenuated on cycles 1 to 3 of treatment to reduce the risk of severe bone marrow suppression. In the event that an institute elects to use bevacizumab as part of the treatment arms on SIRFLOX, bevacizumab is added starting with cycle 1 on the FOLFOX arm and starting with cycle 4 on the FOLFOX + SIR-Spheres arm.&rdquo;

More than 100 hospitals have participated over the past 6 years, including 21 sites in the United States, and enrollment was completed in April 2013. The primary end point is progression-free survival (PFS), while secondary endpoints include response rate, overall survival, and liver PFS. Fakih stated that the first reports on PFS data are expected in early 2015.

Pishvaian pointed out that Sirtex has already presented favorable interim safety data, and he expects that the final results from SIRFLOX will provide further confirmation of this. Among 122 patients, the safety profile was comparable to or better than standard chemotherapy; the most common adverse events were nausea (59%), vomiting (31%), diarrhea (47%), constipation (22%) and abdominal pain (18%).

Both Fakih and Pishvaian noted that a similar international study, the FOXFIRE trial, is also currently enrolling patients. Data analysis from these 2 front line studies will be pooled to further increase the power of the studies and to definitively determine the overall survival endpoint.

Fakih explained that combining SIR-Spheres with standard chemotherapy is an important strategy that may have the potential to downstage hepatic metastases and increase resectability rates, or even prolong disease control. He believes that if these trials confirm the efficacy of this combination, &ldquo;It will result in a shift in the standard of care and move the application of SIRT from the later lines of treatment to the frontline in liver metastases [that are] dominant [in] CRC.&rdquo; Pishvaian also believes that these studies will provide &ldquo;critical data to see whether this paradigm actually improves survival.&rdquo;

Although experts in the field agree that SIR-Spheres should help to downstage tumors and increase the proportion of patients who are eligible for resection, in a recent special report for Clinical Oncology News, N. Joseph Espat, MD, professor, director of the Cancer Center, and chief of surgical oncology at Boston University, observed that there is still a common misconception that surgical resection is not possible after SIRT.

&ldquo;The data on surgical resection after SIR-Spheres [are] really limited, but at the same time, it does seem promising from the point of view of safety,&rdquo; stated Pishvaian. Espat and Pishvaian both pointed out that there have been a series of anecdotal reports of patients who have successfully undergone surgical resection after SIRT. Pishvaian believes that Sirtex has been gathering these anecdotes together, with a view to publishing them in the near future.

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The 5-year survival rate for patients with metastatic colorectal cancer (CRC) remains dismal. Liver metastases are a particularly common occurrence, developing in approximately half of all patients with colorectal cancer.

Jane de Lartigue, PhD | Authors

Articles

Novel Approaches Tackle the JAK Pathway Beyond Ruxolitinib and Myelofibrosis

CAR T-Cell Therapy Hits Prime Time; Challenges Remain

BioT3 Integrates Diagnosis With Actionable Biomarkers in Metastatic Cancer

SIR-Spheres Microspheres for the Treatment of Liver Mets From CRC: Results of the SIRFLOX Trial Eagerly Anticipated