14 March 2018

MIT study finds that the probability of clinical trial success is nearly 40% higher than previously thought

By |2018-09-12T21:41:44+00:00March 14, 2018|Biomarkers, Cancer, Drug Development, Haberman Associates, Immunology, Personalized Medicine, Recent News, Strategy and Consulting, Translational Medicine|

NIH Clinical Center

On December 7, 2017 we published an article on this blog entitled ”Improving Candidate Selection: Translating Molecules into Medicines”. This article was based on a December 4, 2017 symposium sponsored by Aptuit entitled “Improving Candidate Selection: Translating Molecules into Medicines”. The focus of the meeting was on improving drug candidate selection in order to improve development success.

Our article stated that “Only about 10% of drug candidates make their way from first-in-humans trials to regulatory approval. The greatest amount of attrition occurs in Phase 2. Approximately half of candidates fail at that stage, mainly due to lack of efficacy.” As we also stated in that article, drug attrition numbers have not changed since our 2009 publications, “Approaches to Reducing Phase II Attrition” and “Overcoming Phase II Attrition Problem”.

However, especially since the year 2000, drug developers have been working with increasingly newer classes of drugs. They attribute continuing high attrition rates to difficulties in working with ever-changing classes of drugs designed to treat complex diseases. Attrition thus continues to be a moving target.

Several more recent estimates of clinical trial success are comparable to those cited by participants in the Aptuit symposium, and in our own 2009 publications. For example, as pointed out by Endpoints News, BIO (the Biotechnology Innovation Organization) in a recent publication analyzing clinical development success rate from 2006 to 2015, determined that the overall likelihood of approval from Phase 1 for all drug candidates was 9.6%, and 11.9% for all indications other than cancer. (The likelihood of approval for oncology candidates was 5.1%; this is about the same as the figure for oncology success cited in our 2009 report.) Meanwhile, AstraZeneca cited a 5% success rate for its own candidates in a January 2018 analysis.

Now comes a January 2018 study by Andrew W Lo, Ph.D. and his colleagues at MIT that concludes that 13.8% of all drug development programs eventually lead to approval. This study was discussed in a February 1, 2018 article in Endpoints News by John Carroll. Dr. Lo is the Director of the MIT Laboratory for Financial Engineering.

As with earlier studies, the success rates depend on the particular indication. For example, infectious disease vaccines have the highest rate of success, 33.4%. Oncology drugs—as in most such studies—have the lowest rate of success—3.4%.

Dr. Lo’s study represents a Big Data approach to determining drug development success rates.The MIT group analyzed a large dataset of over 40,000 entries from nearly 186,000 clinical trials of over 21,000 compounds. To analyze this dataset, the researchers developed automated algorithms designed to trace each drug development path and compute probability of success (POS) statistics in a matter of hours. If generating POS estimates had been done by traditional manual methods, it would have taken months or years.

Despite the intense focus of the biopharmaceutical industry, investors, and the general public on cancer, the POS for oncology drugs has been consistently abysmal for years—as shown by our 2009 report, the 2016 BIO report, and the Lo et al. 2018 MIT study. However, according to the MIT study, although the POS for oncology drugs had the lowest overall approval rate of 3.4% in 2013, it rose to 8.3% in 2015. Both Dr. Lo’s group and John Carroll of Endpoint News attribute this sharp rise to the advent of immuno-oncology drugs.

As we discussed in our February 22, 2018 blog article, “JP Morgan 2018 (JPM18) panel optimistic for new breakthrough immuno-oncology therapies despite a crowded field”, leading researchers in academia and industry believe that because of the strong emergence of immuno-oncology therapies, now is probably the best time for progress in oncology in several decades. This is consistent with the findings of Dr. Lo’s group. However, as we stated in our previous blog article (based on the conclusions of the JPM18 panel), “This historic opportunity would be maximally capitalized if people from academia, industry, regulatory agencies, and nonprofit organizations work together, especially in adopting novel collaborative study design, aimed at bringing the promise of cancer immunotherapies to patients, sooner rather than later.”

Another issue discussed by Dr. Lo and his colleagues in their study is role of biomarkers in the success of clinical trials. The researchers compared POS estimates for trials that stratified patients using biomarkers to those that did not use biomarkers. They found that trials that utilized biomarkers tended to be more successful (by nearly a factor of 2) than those that did not. However, biomarker-stratified trials studied by the MIT group were nearly all in oncology. Therefore, it was not possible for the MIT researchers to obtain valid conclusions on the role of biomarkers for therapeutic areas outside of oncology.

Nevertheless, with the continuing development of oncology biomarkers, coupled with breakthrough R&D results in immuno-oncology, the MIT researchers expect that the rates of approval of cancer drugs will continue to improve.

Conclusions

Dr. Lo’s group intends to provide continuing information on the success rates of clinical trials, beyond this initial study. The goal is to provide greater risk transparency to drug developers, investors, policymakers, physicians, and patients, order to assist them in their decisions.

Moreover, our book-length report, Cancer Immunotherapy: Building on Initial Successes to Improve Clinical Outcomes can help you understand the role of advances in immuno-oncology in the current and expected increases in drug development success in the cancer field.

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to your company, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

22 June 2017

New Report Published By CHI Insight Pharma Reports Highlights Progress in Cancer Immunotherapy

By |2018-09-12T21:30:33+00:00June 22, 2017|Biomarkers, Cancer, Drug Development, Drug Discovery, Haberman Associates, Immunology, Recent News|

CAR T cells attacking a cancer cell. (Source: National Cancer Institute)

On May 3, 2017 Cambridge Healthtech Institute’s (CHI’s) Insight Pharma Reports announced the publication of a new book-length report, Cancer Immunotherapy: Building on Initial Successes to Improve Clinical Outcomes, by Allan B. Haberman, Ph.D.

The new 2017 report includes an updated discussion of approved and clinical stage agents in immuno-oncology. It also addresses the means by which researchers and companies are attempting to build on prior achievements in immuno-oncology to achieve improved outcomes for more patients. This approach is often referred to as “immuno-oncology 2.0.” The American Society of Clinical Oncology (ASCO) named “immunotherapy 2.0” as its “Advance of the Year” for 2017.

As discussed in the report, researchers have found that checkpoint inhibitors such as pembrolizumab (Merck’s Keytruda) and nivolumab (Bristol-Myers Squibb’s Opdivo) produce tumor responses by reactivating TILs (tumor infiltrating lymphocytes). As a result, they have been developing biomarkers that distinguish inflamed (i.e. TIL-containing) tumors—which are susceptible to checkpoint inhibitor therapy—from “cold” tumors, which are not. They have also been working to develop means to render “cold” tumors inflamed, via treatment with various conventional therapies and/or development of novel agents. These studies constitute the major theme of immuno-oncology 2.0.

Meanwhile, cellular immunotherapy has also been advancing, with two chimeric antigen receptor (CAR) T-cell therapies (from Novartis and Kite Pharma) in preregistration with the FDA as of March 2017.

These and other areas of current cancer immunotherapy R&D are discussed in the new report.

The first wave of immuno-oncology 2.0 treatments has begun to achieve regulatory approval:

  • On May 12, 2017, Merck gained FDA approval to market a combination of pembrolizumab with chemotherapy (specifically, carboplatin plus pemetrexed) for first-line treatment of non-small cell lung cancer (NSCLC). This is based on a Phase 2 clinical study that showed that the chemo/pembrolizumab combination resulted in a much higher statistically-significant overall response than chemo alone — 55% vs. 29%. As we discuss in our report, certain types of chemotherapy can induce immune responses that convert “cold” tumors into inflamed tumors, thus making them susceptible to checkpoint inhibitor treatment.
  • On May 23, 2017, the FDA awarded accelerated approval to Merck’s pembrolizumab for the treatment of adult and pediatric patients with unresectable or metastatic solid tumors that exhibit high microsatellite instability (MSI-H) or are mismatch repair deficient (dMMR). This indication includes patients with solid tumors that have progressed following prior treatment, and who have no satisfactory alternative treatment options. It also includes patients with colorectal cancer that has progressed following treatment with chemotherapy. This is the first approval of an anticancer agent based on a tumor’s biomarker, regardless of where the tumor originated in the body. As we discuss in our report, mismatch-repair deficiency results in a large somatic mutation load. This supports a large and diverse population of TILs, which are specific for mutation-associated neoantigens. Treatment with checkpoint inhibitors may reactivate these TILs, resulting in effective antitumor immune responses.

Our report is designed to enable readers to understand current and future developments in immuno-oncology, especially including new developments in immunotherapy 2.0. It is also designed to inform the decisions of leaders in companies and in academic groups that are working in areas that relate to cancer R&D and treatment.

For more information on the report, or to order it, see the CHI Insight Pharma Reports website.

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to your company, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

22 January 2016

Can adoptive cellular immunotherapy successfully treat metastatic gastrointestinal cancers?

By |2018-09-12T21:37:26+00:00January 22, 2016|Biomarkers, Cancer, Drug Development, Drug Discovery, Gene Therapy, Haberman Associates, Immunology, Monoclonal Antibodies, Personalized Medicine, Rare Diseases, Translational Medicine|

Steven Rosenberg

Steven Rosenberg

On September 6, 2014, we published an article on this blog announcing the publication of our book-length report, Cancer Immunotherapy: Immune Checkpoint Inhibitors, Cancer Vaccines, and Adoptive T-cell Therapies, by Cambridge Healthtech Institute (CHI).

In that article, we cited the example of the case of a woman with metastatic cholangiocarcinoma (bile-duct cancer), which typically kills the patient in a matter of months. The patient, Melinda Bachini, was treated via adoptive immunotherapy with autologous tumor-infiltrating T cells (TILs) resulting in survival over a period of several years, with a good quality of life.

Our report includes a full discussion of that case, as of the date of the May 2014 publication of a report in Science by Steven A. Rosenberg, M.D., Ph.D. and his colleagues at the National Cancer Institute (NCI). Ms. Bachini’s story was also covered in a May 2014 New York Times article.

Now comes the publication, in Science on December 2015, of an update from the Rosenberg group on their clinical studies of TIL-based immunotherapy of metastatic gastrointestinal cancers. This article discusses the results of TIL treatment of ten patients with a variety of gastrointestinal cancers, including cancers of the bile duct, the colon or rectum, the esophagus, and the pancreas. The case of Ms. Bachini (“patient number 3737”) was included.

Ms. Bachini, a paramedic and a married mother of six children, and a volunteer with the Cholangiocarcinoma Foundation, was 41 years old when first diagnosed with cancer. She remains alive today—a five-year survivor—at age 46.

The Foundation produced a video, dated March 13, 2015, in which Ms. Bachini gives her “patient perspective”. This video includes her story “from the beginning”—from diagnosis through surgery and chemotherapy, and continuing with adoptive immunotherapy at the NCI under Dr. Rosenberg. Although her tumors continue to shrink and she remains alive, she still is considered to have “Stage 4” (metastatic) cancer. Ms. Bachini is a remarkable woman.

The Cholangiocarcinoma Foundation has also produced an on-demand webinar (dated October 21, 2014) on the adoptive cellular therapy trial in patients with various types of metastatic gastrointestinal cancers, led by Drs. Eric Tran and Steven Rosenberg. Ms. Bachini is also a presenter on that webinar. The December 2015 Science article is an updated version of the results of this trial.

The trial, a Phase 2 clinical study (NCT01174121) remains ongoing, and is recruiting new patients.

The particular focus of Dr. Tran’s and Dr. Rosenberg’s study in TIL treatment of gastrointestinal cancers is whether TILs derived from these tumors include T-cell subpopulations that target specific somatic mutations expressed by the cancers, and whether these subpopulations might be harnessed to successfully treat patients with these cancers. Of the ten patients who were the focus of the December 2015 publication, only Ms. Bachini had a successful treatment. In the case of Ms. Bachini, she received a second infusion of TILs that were enriched for CD4+ T cells that targeted a unique mutation in a protein known as ERBB2IP. It was this second treatment that resulted in the successful knockdown of her tumors, which continues to this day.

Despite the lack of similar successes in the treatment of the other nine patients, the researchers found that TILs from eight of these patients contained CD4+ and/or CD8+ T cells that recognized one to three somatic mutations in the patient’s own tumors. Notably, CD8+ TILs isolated from a colon cancer tumor of one patient (patient number 3995) recognized a mutation in KRAS known as KRAS G12D. This mutation results in an amino acid substitution at position 12 in KRAS, from glycine (G) to aspartic acid (D). KRAS G12D is a driver mutation that is involved in causation of many human cancers.

Although two other patients (numbers 4032 and 4069, with colon and pancreatic cancer, respectively) had tumors that expressed KRAS G12D, the researchers did not detect TILs that recognized the KRAS mutation in these patients. The researchers concluded that KRAS G12D was not immunogenic in these patients. The TILs from patient 3995 were CD8+ T cells that recognized KRAS G12D in the context of the human leukocyte antigen (HLA) allele HLA-C*08:02. [As with all T cells, TILs express T-cell receptors (TCRs) that recognize a specific antigenic peptide bound to a particular major histocompatibility complex (MHC) molecule—this is referred to as “MHC restriction”.] The two patients for whom KRAS G12D was not immunogenic did not express the HLA-C*08:02 allele.

The results seen with KRAS G12D-expressing tumor suggest the possibility of constructing genetically-engineered CD8+ T cells that express a TCR that is reactive with the KRAS mutation in the context of the HLA-C*08:02 allele. The KRAS G12D driver mutation is expressed in about 45% of pancreatic adenocarcinomas, 13% of colorectal cancers, and at lower frequencies in other cancers, and the HLA-C*08:02 allele is expressed by approximately 8% and 11% of white and black people, respectively, in the U.S. Thus, in the U.S. alone, thousands of patients per year with metastatic gastrointestinal cancers would potentially be eligible for immunotherapy with this KRASG12D-reactive T cell.

Although only Ms. Bachini (“patient number 3737”) was a long-term survivor, the researchers were able to treat three other patients with enriched populations of TILs targeting predominantly one mutated tumor antigen. Patient 4069 experienced a transient regression of multiple lung metastases of his pancreatic adenocarcinoma, but patients 4007 and 4032 had no objective response. Whereas 23% of circulating T cells at one month after treatment were adoptively transferred mutation-specific TILs in the case of Ms. Bachini, the other three patients treated with enriched populations of mutation-specific TILs showed no or minimal persistence. The researchers concluded that they will need to develop strategies designed to enhance the potency and persistence of adoptively transferred mutation-specific TILs. Nevertheless, the researchers concluded that nearly all patients with advanced gastrointestinal cancers harbor tumor mutation-specific TILs. This finding may serve as the basis for developing personalized adoptive cellular therapies and/or vaccines that can effectively target common epithelial cancers.

Conclusions

Dr. Rosenberg pioneered the study and development of adoptive cellular immunotherapy, beginning in the 1980s. Most studies with TIL-based adoptive immunotherapy have been in advanced melanoma. Adoptive cellular immunotherapy is the most effective approach to inducing complete durable regressions in patients with metastatic melanoma.

As we discussed in our cancer immunotherapy report, melanoma tumors have many more somatic mutations (about 200 nonsynonymous mutations per tumor) than most types of cancer. This appears to be due to the role of a potent immunogen—ultraviolet light—in the pathogenesis of melanoma. The large number of somatic mutations in melanomas results in the infiltration of these tumors by TILs that target the mutations. As discussed in our report, Dr. Rosenberg and his colleagues cultured TIL cell lines that addressed specific immunodominant mutations in patients’ melanomas. Treatment with these cell lines in several cases resulted in durable complete remissions of the patients’ cancers.

Dr. Rosenberg and his colleagues used the same strategy employed in identification of TIL cell lines that targeted specific mutations in melanomas to carry out the study in gastrointestinal cancers, as discussed in our report. However, the small number of somatic mutations and of endogenous TILs in gastrointestinal cancers and in most other epithelial cancers has made studies in these cancers more difficult than studies in melanoma.

in addition, the susceptibility of melanoma to treatment with checkpoint inhibitors such as the PD-1 blockers pembrolizumab (Merck’s Keytruda) and nivolumab (Bristol-Myers Squibb’s Opdivo) correlates with the large number of somatic mutations in this type of cancer. As we discussed in our December 15, 2014 article on this blog, immune checkpoint inhibitors work by reactivating endogenous tumor-infiltrating T cells (TILs). In the case of melanoma, these endogenous TILs target the numerous somatic mutations found in these cancers, and—as suggested by Dr. Rosenberg’s studies with cultured TIL cell lines—those endogenous TILs that target immunodominant mutations can induce durable compete remissions. As discussed in our December 15, 2014 blog article, the three major types of immuno-oncology treatments—immune checkpoint inhibitors, cancer vaccines, and adoptive T-cell therapies, work via related mechanisms.

In 2015, researchers showed that other types of cancers that have numerous somatic mutations are especially susceptible to checkpoint inhibitor treatment. These include, for example, non-small cell lung cancers (NSCLCs) that have mutational signatures that indicate that the cancers were caused by smoking, and cancers that have mutations in genes involved in DNA repair. (Mutations in genes involved in DNA repair pathways result in the generation of numerous additional mutations.)

Moreover, as discussed in our December 15, 2014 blog article, cancer immunotherapy researchers have been expanding the types of tumors that can be treated with checkpoint inhibitors. Genentech/Roche’s PD-L1 inhibitor that was discussed in that article, MPDL3280A, is now called atezolizumab. The clinical trials of atezolizumab discussed in that article and in our report have continued to progress. In a pivotal Phase 2 study in locally advanced or metastatic urothelial bladder cancer (UBC), atezolizumab shrank tumors in 27 percent of people whose disease had medium and high levels of PD-L1 expression and had worsened after initial treatment with platinum chemotherapy. These responses were found to be durable. According to Genentech, these results may represent the first major treatment advance in advanced UBC in nearly 30 years. Atezolizumab also gave positive results in Phase 2 clinical trials in patients with NSCLC that expresses medium to high levels of PD-L1.

Meanwhile, NewLink Genetics (Ames, IA) has entered Phase 3 clinical trials in pancreatic cancer with its HyperAcute cellular immunotherapy vaccine therapy. A Phase 2 trial of the company’s HyperAcute cellular immunotherapy algenpantucel-L in combination with chemotherapy and chemoradiotherapy in resected pancreatic cancer (clinical trial number NCT00569387) appears to be promising.

Dr. Rosenberg’s studies of TIL therapies of gastrointestinal cancers represent another approach to moving immuno-oncology treatments beyond melanoma, based on mutation-specific targeting. The types of cancers that form the focus of these studies—gastrointestinal epithelial cancers—have proven difficult to treat. Moreover, several of them are among the most common of cancers. The researchers and patients involved in these and other immuno-oncology studies are heroes, and oncologists appear to be making measured progress against cancers that have been until recently considered untreatable.

As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to your company, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

15 December 2014

Immune checkpoint inhibitors work by reactivating tumor-infiltrating T cells (TILs)

By |2018-09-12T21:48:43+00:00December 15, 2014|Biomarkers, Cancer, Drug Development, Immunology, Monoclonal Antibodies, Personalized Medicine|

Cancer Cell

Cancer Cell

The 27 November issue of Nature contains a wealth of new studies on how immune checkpoint inhibitors target various types of cancer, and how researchers and physicians might be able to identify the patients who are most likely to benefit from treatment with these agents.

These studies are described in five papers published in that issue of Nature. This issue also contains a “News & Views” commentary on these articles by Drs. Jedd D. Wolchok and Timothy A. Chan (both at the Memorial Sloan Kettering Cancer Center). This article serves as an introduction to the five research articles.

In addition, Science Magazine published a commentary on these articles, entitled “Multiple boosts for cancer immunotherapy”, by contributing correspondent Mitch Leslie.

Checkpoint inhibitors can be used to treat several types of cancer

One important result of these studies is the expansion of the range of cancers that can be treated via immunotherapy beyond melanoma, kidney cancer, and non-small cell lung cancer (NSCLC). The papers by Powles et al. and Herbst et al. contain results from a Phase 1 clinical trial of Genentech’s monoclonal antibody (MAb) PD-L1 blocker MPDL3280A. Herbst et al. reported that MPDL3280A showed therapeutic responses in patients with NSCLC, melanoma, renal cancer, and head and neck cancer. Powles et al. focused on the effects of this agent in a larger group of patients with metastatic urothelial bladder cancer (UBC). In both reports, researchers documented that a subset of patients experienced durable responses, and that the treatment showed low toxicity.

We discussed earlier presentations of the results of the Phase 1 trial of MPDL3280A in our Insight Pharma Report (IPR), Cancer Immunotherapy: Immune Checkpoint Inhibitors, Cancer Vaccines, and Adoptive T-Cell Therapies. As we discussed in this report, the FDA granted breakthrough therapy designation for MPDL3280A for treatment of UBC. Roche/Genentech has initiated a Phase 2 clinical trial (clinical trial number NCT02108652) of MPDL3280A in UBC. UBC is the ninth most common cancer in the world. Metastatic UBC is associated with a poor prognosis, and has few treatment options. There have been no new treatment advances in nearly 30 years.

Checkpoint inhibitors work by reactivating tumor-infiltrating T cells (TILs)

Perhaps the most important finding of the research published in the November 27th issue of Nature is that checkpoint inhibitors work via reactivating endogenous tumor-infiltrating T cells. (These T cells are often called “TILs”, which is an acronym for “tumor-infiltrating lymphocytes”.)

For example, as described in the Powles et al. report, Genentech’s PD-L1 blocker MPDL3280A was found to be especially effective in treating patients whose tumors contained PD-L1-positive TILs. As we discussed in our IPR report, Genentech researchers found that MPDL3280A not only targets PD-L1 on the surface of tumor cells, but also PD-L1 on the surface of TILs. PD-L1 on activated T cells interacts not only with PD-1, but also with B7 on the surface of antigen presenting cells, sending a negative signal to the T cells. MPDL3280A targets the PD-L1-B7 interaction, thus enabling reactivation of PD-L1-bearing TILs so that they can attack the tumor.

As we also discuss in our report, targeting PD-1, PD-L1, and CTLA-4 may also be important in reversing immunosuppression by regulatory T cells (Tregs), which typically heavily infiltrate tumors. This provides another mechanism by which checkpoint inhibitors can reactivate TILs and thus induce anti-tumor immune responses.

As described in Powles et al, MPDL3280A was engineered with a modification in the Fc domain that eliminates antibody-dependent cellular cytotoxicity (ADCC). Genentech researchers did this because PD-L1 is expressed on activated T cells, and they wanted an anti-PD-L1 MAb agent that would reactivate these T cells, not destroy them via ADCC.

In the studies described by Herbst et al., researchers showed that Genentech’s PD-L1 blocker MPDL3280A gives antitumor response across multiple types of cancer, in tumors that expressed high levels of PD-L1. These responses especially occurred when PD-L1 was expressed by TILs. The studies suggest that MPDL3280A is most effective against tumors in which endogenous TILs are suppressed by PD-L1, and are reactivated via anti-PD-L1 MAb targeting.

In the Tumeh et al. study, the researchers found that patients responding to treatment with Merck’s MAb PD-1 blocker pembrolizumab (Keytruda) showed proliferation of intratumoral CD8+ T cells that correlated with reduction in tumor size. Pretreatment tumor samples taken from responding patients showed higher numbers of CD8, PD-1, and PD-L1 expressing cells at the invasive tumor margin and within tumors, with a close proximity between PD-1 and PD-L1, and a clonal TCR repertoire.

Based on this information, the researchers developed a predictive model based on CD8 expression at the invasive tumor margin. They validated this model in an independent 15-patient cohort. The researchers concluded that tumor regression due to treatment with the PD-1 blocker pembrolizumab requires preexisting CD8+ T cells whose activity has been blocked by PD-1/PD-L1 adaptive resistance. This study, like those of Powles et al. and Herbst et al., thus indicate that checkpoint inhibitors work against cancer by reactivating TILs. The Tumeh et al. study also indicates that CD8 expression at the invasive tumor margin is a predictive biomarker for sensitivity of patient tumors to treatment with anti-PD-1 checkpoint inhibitors.

The Powles, Herbst, and Tumeh reports all involved studies in human patients. However, the other two papers—Yadav et al. and Gubin et al. involve studies in mouse tumor models.

In the study of Yadav et al., the researchers used their mouse model to develop a method for discovering immunogenic mutant peptides in cancer cells that can serve as targets for T cells. They sequenced the exomes of two mouse cancer cell lines, and looked for differences with the corresponding normal mouse exomes. They also identified which of the neoantigens that they identified via exome sequencing could bind to histocompatibility complex class I (MHCI) proteins, and thus could be presented to T cells. They then modeled the MHC1/peptide complexes, and used these models to predict which of these neoantigens were likely to be immunogenic.

These methods identified only a few candidate neoantigens. Vaccination of tumor-bearing mice with these neoantigens resulted in therapeutically active T-cell responses. In addition, the researchers developed methods for monitoring the antitumor T cell response to peptide vaccination.

In the study of Gubin et al., the researchers used similar genomic and bioinformatic approaches to those of Yadav et al., and identified two neoantigens that were targeted by T cells following therapy with anti-PD-1 and/or anti-CTLA-4 antibodies. [Human CTLA-4 is the target of the checkpoint blockade inhibitor ipilimumab (Medarex/ Bristol-Myers Squibb’s Yervoy).] As with PD-1 and PD-L1 blockers, we discussed this agent in our IPR report. T cells specific for these neoantigens (in the context of MHCI proteins expressed by the mice) were present in the tumors. These T cells were reactivated by anti-PD-1 and/or anti-CTLA-4 antibodies, enabling the mice to reject the tumors.

As in the study of Yadav et al., the Gubin et al. researchers performed experiments in which they vaccinated tumor-bearing mice with peptides that incorporated the mutant epitopes. This vaccination induced specific tumor rejection that was comparable to treatment with checkpoint blockade inhibitors. As in the case of Yadav et al, the Gubin et al. researchers concluded that specific mutant antigens were targets of checkpoint inhibitor therapy in their mouse models, and that the mutant antigens could also be used to develop personalized cancer vaccines.

Since the studies of Yadav et al. and Gubin et al. were carried out using mouse tumor models, the results are not directly applicable to cancer in human patients. However, the studies suggest that immune checkpoint inhibitors work by reactivating endogenous TILs, and that anti tumor TILs work by attacking specific neoantigens on the tumors.

As we discussed in our IPR report, Dr. Steven Rosenberg (National Cancer Institute, Bethesda, MD) identified specific antigens that were the targets of TILs, both in metastatic melanoma and in metastatic cholangiocarcinoma (a type of epithelial bile duct cancer). However, these target antigens were from human cancers, and they were targets of TILs that has been isolated from patient tumors, cultured and expanded ex vivo, and used in adoptive cellular immunotherapy.

Moreover, the antigens were targets of TIL therapies that resulted in a durable compete remission in the case of the melanoma patient, and long-term tumor regression in the case of the metastatic cholangiocarcinoma patient. The metastatic cholangiocarcinoma case was highlighted in our September 16, 2014 Biopharmconsortium Blog article.

The Yadav et al. paper referenced the Rosenberg group’s work. However, this paper stated that “few mutant epitopes have been described because their discovery required the laborious screening of patient tumour-infiltrating lymphocytes for their ability to recognize antigen libraries constructed following tumour exome sequencing.”

The methods of Yadav et al. (and of Gubin et al.) are thus designed to simplify and accelerate the discovery of immunogenic mutant peptides. They carried out their studies in mouse models, which helped these researchers to develop methods that could potentially discover greater numbers of neoantigens more efficiently. However, it remains to be seen to what extent they can apply their methods to human patients.

Unifying the field of immuno-oncology

As can be seen, for example, from the title of our IPR report, the three major approaches to immuno-oncology in 2014/2015 are development of immune checkpoint inhibitors, of cancer vaccines, and of adoptive T-cell therapies.

In the immuno-oncology papers published in the 27 November issue of Nature, researchers show that checkpoint inhibitors work via reactivating of endogenous TILs. They also (in mouse tumor models) identified neoantigens that are targets of these reactivated TILs, and designed peptide vaccines that were as effective as checkpoint inhibitor therapy in the mouse models. In principle, one can isolate TILs that are reactive to particular neoantigens in the mouse tumors, culture and expand them ex vivo, and infuse them back into the mice to target their tumors. Thus the studies in the 27 November issue of Nature serve as a template for the unification of the immuno-oncology field as it now exists.

However, it will be necessary to apply the methodologies developed by Yadav et al. and Gubin et al. to human patients. And at least so far, peptide vaccines have not been very successful in treating patients, as compared to TIL therapy (in the subset of patients in whom TIL therapy can be done). It is thus possible that once these methods of neoantigen identification are applied to human patients, it will be found that targeting the neoantigens with ex vivo-expanded TILs will be more successful than therapy with peptide vaccines. However, whether this is true awaits the application of the new methodologies to neoantigen identification in human tumors.


As the producers of this blog, and as consultants to the biotechnology and pharmaceutical industry, Haberman Associates would like to hear from you. If you are in a biotech or pharmaceutical company, and would like a 15-20-minute, no-obligation telephone discussion of issues raised by this or other blog articles, or of other issues that are important to your company, please contact us by phone or e-mail. We also welcome your comments on this or any other article on this blog.

31 October 2013

Chemokine receptors and the HIV-1 entry inhibitor maraviroc

By |2018-05-05T16:59:58+00:00October 31, 2013|Biomarkers, Chemistry, Drug Development, Drug Discovery, Infectious Disease, Monoclonal Antibodies, Strategy and Consulting|

Maraviroc

Maraviroc

In April 2012, Informa’s Scrip Insights published our book-length report, “Advances in the Discovery of Protein-Protein Interaction Modulators.” We also published a brief introduction to this report, highlighting the strategic importance of protein-protein interaction (PPI) modulators for the pharmaceutical industry, on the Biopharmconsortium Blog.

The report included a discussion on discovery and development of inhibitors of chemokine receptors. Chemokine receptors are members of the G-protein coupled receptor (GPCR) superfamily. GPCRs are seven-transmembrane (7TM) domain receptors (i.e. integral membrane proteins that have seven membrane-spanning domains). Compounds that target GPCRs represent the largest class of drugs produced by the pharmaceutical industry. However, in the vast majority of cases, these compounds target GPCRs that bind to natural small-molecule ligands.

Chemokine receptors, however, bind to small proteins, the chemokines. These proteins constitute a class of small cytokines that guide the migration of immune cells via chemotaxis. Chemokine receptors are thus a class of GPCRs that function by forming PPIs. Direct targeting of interactions between chemokines and their receptors (unlike targeting the interactions between small-molecule GPCR ligands and their receptors) thus involves all the difficulties of targeting other types of PPIs.

However, GPCRs–including chemokine receptors–appear to be especially susceptible to targeting via allosteric modulators. Allosteric sites lie outside the binding site for the protein’s natural ligand. However, modulators that bind to allosteric sites change the conformation of the protein in such a way that it affects the activity of the ligand binding site. (Direct GPCR modulators that bind to the same site as the GPCR’s natural ligands are known as orthosteric modulators.) In the case of chemokine receptors, researchers can in some cases discover small-molecule allosteric modulators that activate or inhibit binding of the receptor to its natural ligands. Discovery of such allosteric activators is much easier than discovery of direct PPI modulators.

Chemokines bind to sites that are located in the extracellular domains of their receptors. Allosteric sites on chemokine receptors, however, are typically located in transmembrane domains that are distinct from the chemokine binding sites. Small-molecule allosteric modulators that bind to these sites were discovered via fairly standard medicinal chemistry and high-throughput screening, sometimes augmented with structure-based drug design. This is in contrast to attempts to discover small molecule agents that directly inhibit binding of a chemokine to its receptor, which has so far been extremely challenging.

Our report describes several allosteric chemokine receptor modulators that are in clinical development, as well as the two agents that have reached the market. One of the marketed agents, plerixafor (AMD3100) (Genzyme’s Mozobil), is an inhibitor of the chemokine receptor CXCR4. It is used in combination with granulocyte colony-stimulating factor (G-CSF) to mobilize hematopoietic stem cells to the peripheral blood for autologous transplantation in patients with non-Hodgkin lymphoma and multiple myeloma. The other agent, which is the focus of this blog post, is maraviroc (Pfizer’s Selzentry/Celsentri).

Maraviroc is a human immunodeficiency virus-1 (HIV-1) entry inhibitor. This compound is an antagonist of the CCR5 chemokine receptor. CCR5 is specific for the chemokines RANTES (Regulated on Activation, Normal T Expressed and Secreted) and macrophage inflammatory protein (MIP) 1α and 1β.  In addition to being bound and activated by these chemokines, CCR5 is a coreceptor (together with CD4) for entry of the most common strain of HIV-1 into T cells. Thus maraviroc acts as an HIV entry inhibitor; this is the drug’s approved indication in the U.S. and in Europe. Maraviroc was discovered via a combination of high-throughput screening and optimization via standard medicinal chemistry.

New structural biology studies of the CCR5-maraviroc complex

Now comes a report in the 20 September 2013 issue of Science on the structure of the CCR5-maraviroc complex. This report was authored by a mainly Chinese group led by Beili Wu, Ph.D. (Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai); researchers at the University of California at San Diego and the Scripps Research Institute, San Diego were also included in this collaboration. A companion Perspective in the same issue of Science was authored by P. J. Klasse, M.D., Ph.D. (Weill Cornell Medical College, Cornell University, New York, NY).

As described in the Perspective, the outer surface of the HIV-1 virus displays numerous envelope protein (Env) trimers, each including the outer gp120 subunit anchored in the viral membrane by gp41. When gp120 binds to the cell-surface receptor CD4, this enables interaction with a specific chemokine receptor, either CCR5 or CXCR4. Interaction with both CD4 and the chemokine receptor triggers complex sets of changes in the Env complex, eventually resulting in the fusion of the viral membrane and the cell membrane, and the entry of the virus particle into the host cell.

HIV-1 gp120 makes contact with CCR5 at several points. The interactions between CCR5 and the variable region of gp120 called V3 are especially important for the tropism of an HIV-1 strain, i.e., whether the virus is specific for CCR5 (the “R5 phenotype”) or CXCR4 (the “X4 phenotype”). In the case of R5-tropic viruses, the tip of the V3 region interacts with the second extracellular loop (ECL2) of CCR5, while the base of V3 interacts with the amino-terminal segment of CCR5. Modeling of the interactions between the V3 domain of gp120 of either R5 or X4-tropic viruses with CCR5 or CXCR4 explains coreceptor use, in terms of forming strong bonds or–conversely–weak bonds and steric hindrance.

Monogram Biosciences (South San Francisco, CA) has developed and markets the Trofile assay. This is a molecular assay designed to identify the R5, X4, or mixed tropism of a patient’s HIV strain. If a patient’s strain is R5-tropic, then treatment with maraviroc is appropriate. However, a patient’s HIV-1 strain may undergo a tropism switch, or may mutate in other ways to become resistant to maraviroc.

Dr Wu and her colleagues determined the high-resolution crystal structure of the complex between maraviroc and a solubilized engineered form of CCR5. This included determining the CCR5 binding pocket for maraviroc, which was determined both by Wu et al’s X-ray crystallography, and by site-directed mutagenesis (i.e., to determine amino acid residues that are critical for maraviroc binding) that had been published earlier by other researchers.

The structural studies of Dr. Wu and her colleagues show that the maraviroc-binding site is different from the recognition sites for gp120 and for chemokines, as expected for an allosteric inhibitor. The X-ray structure shows that maraviroc binding prevents the helix movements that are necessary for binding of g120 to induce the complex sequence of changes that result in fusion between the viral and cellular membranes. (These helix movements are also necessary for induction of signal transduction by binding of chemokines to CCR5.)

Structural studies of CXCR4 and its inhibitor binding sites

In addition to their structural studies of the CCR5-maraviroc complex, Dr. Wu and her colleagues also published structural studies of CXCR4 complexed with small-molecule and cyclic peptide inhibitors in Science in 2010. These inhibitors are IT1t, a drug-like orally-available isothiourea developed by Novartis, and CVX15, a 16-residue cyclic peptide that had been previously characterized as an HIV-inhibiting agent. IT1t and CVX15 bind to overlapping sites in CXCR4. Other researchers have found evidence that the binding site for plerixafor also overlaps with the IT1t binding site.

As discussed in Wu et al’s 2013 paper, CCR5 and CXCR4 have similar, but non-identical structures. The binding site for IT1t in CXCR4 is closer to the extracellular surface than is the maraviroc binding site in CCR5, which is deep within the CCR5 molecule. The entrance to the CXCR4 ligand-binding pocket is partially covered by CXC4’s N terminus and ECL2, but the CCR5 ligand-binding pocket is more open.

Mechanisms of CXCR4 and CCR5 inhibition, and implications for discovery of improved HIV entry inhibitors

The chemokine that specifically interacts with the CXCR4 receptor is known as CXCL12 or stromal cell-derived factor 1 (SDF-1). Researchers have proposed a hypothesis for how CXCL12 interacts with CXCR4; this hypothesis appears to be applicable to the interaction between other chemokines and their receptors as well. This hypothesis is know as the “two-step model” or the “two-site model” of chemokine-receptor activation. Under the two-site model, the core domain of a chemokine binds to a site on its receptor (known as the “chemokine recognition site 1” or “site 1”) defined by the receptor’s N-terminus and its ECLs. In the second step, the flexible N-terminus of the chemokine interacts with a second site (known as “chemokine recognition site 2” or “site 2” or the “activation domain”) deeper within the receptor, in transmembrane domains. This result in activation of the chemokine receptor and intracellular signaling.

Under the two-site model, CXCR4 inhibitors (e.g., IT1t, CVX15, and  plerixafor), which bind to sites within the ECLs of CXCR4, are competitive inhibitors of binding of the core domain of CXCL12 to CXCR4 (i.e.., step 1 of chemokine/receptor interaction). They are thus orthosteric inhibitors of CXCR4. (This is contrary to the earlier assignment of plerixafor as an allosteric inhibitor of CXCR4.)  The CCR5 ligand maraviroc, however, binds within a site within the transmembrane domains of CCR5, which overlaps with the activation domain of CCR5. Dr. Wu and her colleagues propose two alternative hypotheses: 1. Maraviroc may inhibit CCR5 activation by chemokines by blocking the second step of chemokine/chemokine receptor interaction, i.e., receptor activation. 2. Maraviroc may stabilize CCR5 in an inactive conformation. It is also possible that maraviroc inhibition of CCR5 may work via both mechanisms.

Dr. Wu and her colleagues further hypothesize that the interaction of  HIV-1 gp120 with CCR5 (or CXCR4) may operate via similar mechanisms to the interaction of chemokines with their receptors. As we discussed earlier in this article, the base (or the stem region) of the gp120 V3 domain interacts with the amino-terminal segment of CCR5. The tip (or crown) of the V3 domain interacts with the ECL2 of CCR5, and–according to Dr. Wu and her colleagues–also with amino acid residues inside the ligand binding pocket; i.e., the activation site of CCR5. The HIV gp120 V3 domain may thus activate CCR5 via a similar mechanism to the two-step  model utilized by chemokines.

Based on their structural biology studies, Dr. Wu and her colleagues have been building models of the CCR5-R5-V3 and CXC4-X4-V3 complexes, and are also planning to determine additional structures needed to fully understand the mechanisms of HIV-1 tropism. The researchers will utilize their studies in the discovery of improved, second-generation HIV entry inhibitors for both R5-tropic and X4-tropic strains of HIV-1.

The bigger picture

The 17 October 2013 issue of Nature contains a Supplement entitled “Chemistry Masterclass”. In that Supplement is an Outlook review entitled “Structure-led design”, by Nature Publishing Group Senior Editor Monica Hoyos Flight, Ph.D. The subject of this article is structure-based drug design of modulators of GPCRs.
This review outlines progress in determining GPCR structures, and in using this information for discovery of orthosteric and allosteric modulators of GPCRs.

According to the article, the number of solved GPCR structures has been increasing since 2008, largely due to the efforts of the Scripps GPCR Network, which was established in that year. Dr. Wu started her research on CXCR4 and CCR5 as a postdoctoral researcher in the laboratory of Raymond C. Stevens, Ph.D. at Scripps in 2007, and continues to be a member of the network. The network is a collaboration that involves over a dozen academic and industrial labs. Its goal has been to characterize at least 15 GPCRs by 2015; it has already solved 13.

Interestingly, among the solved GPCR structures are those for the corticotropin-releasing hormone receptor and the glucagon receptor. Both have peptide ligands, and thus work by forming PPIs.

One company mentioned in the article, Heptares Therapeutics (Welwyn Garden City, UK), specializes in discovering new medicines that targeting previously undruggable or challenging GPCRs. In addition to discovering small-molecule drugs, Heptares, working with monoclonal antibody (MAb) leaders such as MorphoSys and MedImmune, is working to discover MAbs that act as modulators of GPCRs. Among Heptares’ targets are several GPCRs with peptide ligands.

Meanwhile, Kyowa Hakko Kirin Co., Ltd. has developed the MAb drug mogamulizumab (trade name Poteligeo), which is approved in Japan for treatment of relapsed or refractory adult T-cell leukemia/lymphoma. Mogamulizumab targets CC chemokine receptor 4 (CCR4).

Thus, aided in part by structural biology, the discovery of novel drugs that target GPCRs–including those with protein or peptide targets such as chemokine receptors–continues to make progress.


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