Long-Term Follow-Up of Anti-CD19 Chimeric Antigen Receptor T-Cell Therapy

Abstract

PURPOSE

Anti-CD19 chimeric antigen receptors (CARs) are artificial fusion proteins that cause CD19-specific T-cell activation. Durability of remissions and incidence of long-term adverse events are critical factors determining the utility of anti-CD19 CAR T-cell therapy, but long-term follow-up of patients treated with anti-CD19 CAR T cells is limited. This work provides the longest follow-up of patients in remission after anti-CD19 CAR T-cell therapy.

METHODS

Between 2009 and 2015, we administered 46 CAR T-cell treatments to 43 patients (ClinicalTrials.gov identifier: NCT00924326). Patients had relapsed B-cell malignancies of the following types: diffuse large B-cell lymphoma or primary mediastinal B-cell lymphoma (DLBCL/PMBCL; n = 28), low-grade B-cell lymphoma (n = 8), or chronic lymphocytic leukemia (CLL; n = 7). This report focuses on long-term outcomes of these patients. The CAR used was FMC63-28Z; axicabtagene ciloleucel uses the same CAR. Cyclophosphamide plus fludarabine conditioning chemotherapy was administered before CAR T cells.

RESULTS

The percentages of CAR T-cell treatments resulting in a > 3-year duration of response (DOR) were 51% (95% CI, 35% to 67%) for all evaluable treatments, 48% (95% CI, 28% to 69%) for DLBCL/PMBCL, 63% (95% CI, 25% to 92%) for low-grade lymphoma, and 50% (95% CI, 16% to 84%) for CLL. The median event-free survival of all 45 evaluable treatments was 55 months. Long-term adverse effects were rare, except for B-cell depletion and hypogammaglobulinemia. Median peak blood CAR-positive cell levels were higher among patients with a DOR of > 3 years (98/µL; range, 9-1,217/µL) than among patients with a DOR of < 3 years (18/µL; range, 0-308/μL, P = .0051).

CONCLUSION

Complete remissions of a variety of B-cell malignancies lasting ≥ 3 years occurred after 51% of evaluable anti-CD19 CAR T-cell treatments. Remissions of up to 9 years are ongoing. Late adverse events were rare.

INTRODUCTION

Chimeric antigen receptors (CARs) are artificial proteins containing antigen recognition domains and T-cell signaling domains.^1-6 CAR T cells targeting the B-cell antigen CD19 are used to treat relapsed B-cell malignancies.^7-15 Patients with chemotherapy-refractory diffuse large B-cell lymphoma (DLBCL) have poor prognoses, with an objective response rate of 26% and a median overall survival of 6.3 months when treated with standard salvage regimens.¹⁶ The two US Food and Drug Administration (FDA)–approved anti-CD19 CAR products, axicabtagene ciloleucel and tisagenle-cleucel, have complete remission (CR) rates of 40%-54% against DLBCL.^17,18 Mantle cell lymphoma, follicular lymphoma, and chronic lymphocytic leukemia (CLL) have also been successfully treated with anti-CD19 CAR T cells.^7,8,19-22

CONTEXT

• Key Objective

• Chimeric antigen receptor (CAR) T cells targeting CD19 can cause complete remissions in relapsed B-cell lymphoma, but the long-term durability of these remissions is unclear. Here, we examined the long-term outcomes and adverse effects in patients with lymphoma receiving anti-CD19 CAR T cells.

• Knowledge Generated

• Our data demonstrate that anti-CD19 CAR T-cell treatment often leads to highly durable remissions in patients with B-cell lymphoma that are ongoing up to 113 months after CAR T-cell therapy. These are the longest reported responses to anti-CD19 CAR T-cell treatment. Response and durability of response were associated with higher peak CAR T-cell levels. Long-term adverse events were rare.

• Relevance

• These highly durable responses raise the possibility that anti-CD19 CAR T cells may be curative for B-cell lymphoma.

A critical factor for any lymphoma therapy is durability of response. The long-term durability of response after anti-CD19 CAR T-cell therapy for lymphoma is still being determined. In a recent report of patients who received axicabtagene ciloleucel, 36% of patients were in ongoing responses at last follow-up, with a median follow-up time of 27.1 months.²³

The most commonly reported toxicities during long-term follow-up after anti-CD19 CAR T-cell therapy are decreased blood B-cell counts and hypogamma‐globulinemia.^10,23-27 Late-onset infections and secondary malignancies appear to be uncommon, but the duration of follow-up has been limited in published studies.^17,23,24,28

We conducted, to our knowledge, the first clinical trial of anti-CD19 CAR T cells to show responses against B-cell malignancies¹²; therefore, we have patients with the longest follow-up after anti-CD19 CAR T-cell therapy. Here, we present the long-term results of patients treated with anti-CD19 CAR T cells at the National Cancer Institute (NCI) between 2009 and 2015. Many patients have ongoing CRs > 5 years after treatment, and long-term adverse events have been rare.

METHODS

Trial Design

This phase I, single-arm clinical trial (Clinical Trials.gov identifier: NCT00924326; CONSORT diagram in Data Supplement) evaluated an anti-CD19 CAR designated FMC63-28Z (Data Supplement).² Initial results of this trial have been reported; this report focuses on long-term outcomes.^7-9 All patients gave informed consent. The research was approved by the NCI Institutional Review Board and allowed by the US FDA. After CAR T-cell infusion, patients were removed from the clini-cal trial when progression of lymphoma or initiation of new lymphoma therapy occurred, so reported response durations are only a result of the CAR T-cell protocol therapy. After removal from the CAR T-cell trial, patients were evaluated on a long-term gene therapy follow-up protocol (ClinicalTrials.gov identifier: NCT00923026).

Lymphoma and CLL response assessment was performed using standard approaches.^29,30 Lymphoma and CLL staging was performed using standard methods.^31,32 Duration of response (DOR) was defined as the time from first response (stable disease [SD], partial remission [PR], or CR) until lymphoma progression or initiation of any antimalignancy therapy. For event-free survival (EFS), events were defined as death, lymphoma progression, or initiation of a new lymphoma therapy. When development of a second malignancy, loss to follow-up, or treatment with allogeneic stem-cell transplantation occurred, DOR ended, and patients were censored on EFS plots. Statistics are described in the figure legends and the Data Supplement.

Clinical Protocol

Protocol treatment evolved over the course of this clinical trial. The full protocol is in the Data Supplement. The trial can be divided into three cohorts based on different chemotherapy conditioning regimens (Fig 1A) and CAR T-cell production methods (Data Supplement). Cohort 1 had a 24-day CAR T-cell production process. Patients in cohort 1 received a conditioning regimen of high-dose cyclophosphamide plus fludarabine. These patients received high-dose intravenous interleukin-2 (IL-2) after CAR T-cell infusion.⁷ Cohort 2 had a shortened CAR T-cell production process that lasted 10 days. These patients received a conditioning regimen of high-dose cyclophosphamide plus fludarabine; exogenous IL-2 was not adminstered.⁸ Cohort 3 was similar to cohort 2 except the conditioning regimen was fludarabine 30 mg/m² daily for 3 days plus lower cyclophosphamide doses of 300 mg/m² (n = 18) or 500 mg/m² (n = 4) on the same days as fludarabine.⁹

FIG 1.

Trial design and responses. (A) Schematics of clinical treatment protocols are shown. Patients in cohort 1 received cyclophosphamide (Cy) 60 mg/kg on days −7 and −6, fludarabine (Flu) 25 mg/m² on days −5 to day −1, and chimeric antigen receptors (CAR) T-cell infusion on day 0. High-dose intravenous interleukin-2 (IL-2) administration started on day 0. IL-2 administration continued until either patient toxicity required cessation or 6 days of IL-2 administration were completed. Patients in cohort 2 received Cy doses of 30 or 60 mg/kg on days −7 and −6, Flu 25 mg/m² on days −5 to −1, and CAR T-cell infusion on day 0. IL-2 was not administered. Patients in cohort 3 received daily doses of Cy 300 or 500 mg/m² + Flu 30 mg/m² on days −5 to −3. CAR T-cell infusion was on day 0 with no IL-2 administration. (B) Best responses of each patient and durations of responses are shown. Each bar represents one patient. Three patients were retreated with CAR T-cells; patient 1 (follicular lymphoma), patient 3 (CLL), and patient 4 (splenic marginal zone lymphoma) were each treated twice. These retreatments are shown. Patients 2 and 11 were not evaluable for response because of early death and are not shown. Patient 18 was omitted because this patient was not evaluable for response due to patient noncompliance. Patients who were no longer evaluable due to development of a second malignancy, loss to follow-up, or treatment with allogeneic stem-cell transplantation while in ongoing response are indicated on the figure. Ongoing responses are indicated by black arrows. CAR+, Chimeric antigen receptor positive; CLL, chronic lymphocytic leukemia; DLBCL, diffuse large B-cell lymphoma; PMBCL, primary mediastinal B-cell lymphoma.

Quantification of CAR-Positive T Cells, Lymphocytes, and Immunoglobulins

Blood CAR-positive (CAR⁺) cells were quantified by quantitative real-time polymerase chain reaction (PCR). PCR techniques are detailed in the Data Supplement and have been described previously.^8,9 Blood B-cell counts were determined by flow cytometry for CD19+ lymphocytes. T-cell and natural killer (NK) cell counts were also determined by flow cytometry. Serum immunoglobulins were determined by standard clinical immunoturbidimetric assay.

RESULTS

Patient Characteristics

Forty-three patients were treated. Patient characteristics are listed in Table 1. Twenty-eight patients had DLBCL or primary mediastinal B-cell lymphoma (PMBCL). Eight patients had low-grade B-cell lymphomas (follicular lymphoma, n = 5; splenic marginal zone lymphoma, n = 1; mantle cell lymphoma, n = 1; and unspecified low-grade lymphoma, n = 1). Seven patients had CLL (Table 1 and Data Supplement). Of the 28 patients with DLBCL/PMBCL, 19 had chemotherapy-refractory lymphoma, and six additional patients with DLBCL/PMBCL had lymphoma that had relapsed after autologous stem-cell transplantation as the last treatment before protocol enrollment. Of the patients with DLBCL/PMBCL, 16 had a high second-line age-adjusted International Prognostic Index (sAAIPI), 10 had an intermediate sAAIPI, and two had a low sAAIPI (Data Supplement). sAAIPI was calculated as described previously.³³ Individual patient CAR T-cell doses are provided in the Data Supplement. Double-hit DLBCL (rearrangement in C-myc plus either B-cell lymphoma [Bcl]-2 or Bcl-6) was present in three of 15 evaluable patients with DLBCL/PMBCL (Data Supplement). The median number of prior lines of treatment of all patients was four (range, one to 12 lines; Table 1).

TABLE 1.

Baseline Characteristics According to Malignancy Type

DORs

Forty-three patients were treated, and three patients received two CAR T-cell treatments, so 46 total CAR T-cell treatments were performed. Outcomes of each treatment are summarized by malignancy type in Figure 1B. Twenty-five (58%; 95% CI, 42% to 73%) of 43 total evaluable treatments resulted in a best response of CR, whereas 10 (23%; 95% CI, 12% to 39%) resulted in a best response of PR (Fig 1B, Table 2, and Data Supplement). The objective remission rate (CR+PR) was 81% (95% CI, 67% to 92%). Four responses initially reported as PRs eventually converted to CRs after longer follow-up (patients 1, 4, 7, and 21).^7,8 No patient whose best response was PR or SD had a durable response. Nineteen (76%) of 25 CRs were still ongoing at the time of last follow-up. The DORs among these ongoing CRs ranged from 43 to 113 months at last follow-up, except for patient 41 who was lost to follow-up while in CR with a DOR of 14 months (Fig 1B and Data Supplement). Responses are summarized by large-cell lymphoma subtype in the Data Supplement. Six (24%) of 25 CRs ended as a result of relapse. Overall, the percentage of evaluable CAR T-cell treatments that resulted in a DOR of > 3 years was 51% (21 of 41 evaluable treatments; 95% CI, 35% to 67%). The percentages of treatments resulting in a DOR of > 3 years were 48% (95% CI, 28% to 69%) for DLBCL/PMBCL, 63% (95% CI, 25% to 92%) for low-grade lymphoma, and 50% (95% CI, 16% to 84%) for CLL (Data Supplement). A variety of demographic and baseline clinical characteristics were not associated with the incidence of a DOR > 3 years (Data Supplement). The median time for best response to be achieved was 67 days (range, 25-2,370 days).

TABLE 2.

Responses by Lymphoma Type

EFS and Overall Survival

For all patients, the median follow-up time was 42 months (range, 1-123 months). The median EFS for all 45 evaluable treatments was 55 months (Fig 2A). Patient 18 was the only patient not evaluable for EFS because of noncompliance with all response assessments. The median overall survival (OS) for patients enrolled on the study was not reached (Fig 2B). The median EFS for all patients with DLBCL/PMBCL was 15 months, the median EFS for patients with low-grade lymphoma was 55 months, and the median EFS for patients with CLL was 40.5 months. The median OS was not reached for any of the malignancy types. There was no statistically significant difference in either EFS or OS when any two of the three B-cell malignancy types were compared (Figs 2C and 2D). We assessed EFS in the three patient cohorts defined in Figure 1A. The median EFS was 12 months for cohort 1, 66 months for cohort 2, and 20 months for cohort 3 (Fig 2E). The EFS was not statistically different when any two of the three cohorts were compared. The median OS was not reached for any of the cohorts (Data Supplement). The median EFS for the 25 patients who achieved CR was not reached (Fig 2F). Nineteen of 22 patients treated on cohort 3 had DLBCL/PMBCL. The median EFS of these 19 patients in cohort 3 with DLBCL/PMBCL was 15 months, and nine of the 19 patients had ongoing lymphoma remissions at their last follow-up (Data Supplement).

FIG 2.

Anti-CD19 chimeric antigen receptor (CAR) T cells generated long-term complete remissions (CRs). (A) Event-free survival (EFS) curves of all 45 evaluable CAR T-cell treatments. For the three patients who received two CAR T-cell treatments, both treatments are included separately on this plot. The median EFS was 55 months. (B) Overall survival (OS) of all 42 evaluable patients. For patients who received second CAR T-cell treatments, the OS from the time of the second CAR T-cell infusion is shown. Median OS was not reached. (C) EFS curves of each CAR T-cell treatment divided by malignancy type. The median EFS for patients with diffuse large B-cell lymphoma (DLBCL) or primary mediastinal B-cell lymphoma (PMBCL) was 15 months. The median EFS for patients with low-grade lymphoma was 55 months, and the median EFS for patients with chronic lymphocytic leukemia (CLL) was 40.5 months. No statistically significant differences were found when the EFS curves of each malignancy type were compared. For patients 1, 3, and 4 who received two CAR T-cell treatments, both the first and second treatments are included. (D) OS curves divided by malignancy type are shown. The median OS was not reached for any of the malignancy types. For patients who received two CAR T-cell treatments, OS was calculated starting from the time of the second treatment. No statistically significant differences were found when the OS curves of each malignancy type were compared. (E) EFS curves of each treatment cohort are shown. (continued on following page)(Continued). The median EFS of cohorts 1, 2, and 3 were 12, 66, and 20 months, respectively. For patients who received retreatments, both treatments are included on the plot. No statistically significant differences were found when the EFS curves of each cohort were compared. (F) The plot shows EFS of treatments that resulted in CRs. Patient 3 received two treatments that both resulted in CR, and both treatments are included on this graph. The median EFS was not reached. All plots were prepared using the Kaplan-Meier method, and all curve comparisons were made using the log-rank test. Patients who were no longer evaluable for response due to diagnosis of non–skin cancer second malignancies or loss to follow-up were censored and are indicated with a red mark. Red marks are also present for patients in ongoing remission at the time of final data analysis. Patient 18 was not included in the analysis for EFS or OS due to patient noncompliance with follow-up.

Long-Term Adverse Events

Seven of the 43 patients developed a second malignancy after protocol treatment. Patient 3 had a history of resected prostate cancer before protocol enrollment and developed recurrent prostate cancer 82 months after CAR T-cell infusion. The following patients developed new solid tumors after CAR T-cell infusions: patient 8, laryngeal cancer; patient 19, prostate cancer and GI stromal tumor; patient 35, localized melanoma; and patient 42, hepatocellular carcinoma. Patient 14 developed myelodysplastic syndrome (MDS) 39 months after CAR T-cell infusion, and patient 23 developed MDS 20 months after cell infusion. The presence of the CAR gene was assessed using quantitative PCR in the bone marrow of patients 14 and 23 at the time of MDS diagnosis. In patient 14, 0.04% of bone marrow mononuclear cells contained the CAR gene. In patient 23, the CAR gene was below the PCR limit of detection in bone marrow mononuclear cells. Hypothyroidism in patient 15 was the only probable autoimmune diagnosis among the patients.

We previously reported infections occurring acutely after CAR T-cell infusion in these patients.^7-9 Because anti-CD19 CAR T-cell therapy causes B-cell depletion and decreased immunoglobulin levels, we assessed infections occurring ≥ 6 months after CAR T-cell infusion. Four patients required hospital admission for infections. Patient 1 had disseminated herpes zoster 6 months after his second CAR T-cell infusion. Patient 9 had pneumonia 3 years after cell infusion. Patient 33 had Citrobacter bacteremia 6 months after cell infusion. Patient 35 was hospitalized 1 year after CAR T-cell infusion for influenza, 2 years after infusion for pneumonia, and 3 years after infusion for pneumonia. Of note, patients routinely received prophylactic medications for herpes zoster and Pneumocystis pneumonia until the blood CD4+ T-cell count recovered to 200/μL. Testing for replication-competent retroviruses (RCRs) was performed after CAR T-cell infusion; no RCRs were detected (Data Supplement).

DOR Was Associated With High Peak Blood CAR⁺ Cell Levels

Peak blood levels of CAR⁺ cells were higher among patients who obtained best responses of CR than among patients who obtained best responses of PR, SD, or progressive disease (PD; Fig 3A). Peak CAR⁺ cell levels occurred between day 6 and day 17 after CAR T-cell infusion for all treatments except four treatments with peak CAR⁺ cell levels between day 26 and day 55 after infusion. To determine the impact of persistence on response, we also measured blood CAR⁺ cell levels 28-56 days after CAR T-cell infusion by quantitative PCR. CAR⁺ cell levels 28-56 days after CAR T-cell infusion were not statistically different between patients who achieved a best response of CR versus PR, SD, or PD (Fig 3B). Patients with DORs of > 3 years had higher peak blood CAR⁺ cell levels when compared with patients with DORs of < 3 years (Fig 3C). The CAR⁺ cell level at days 28-56 did not differ between patients who had a DOR of > 3 years versus < 3 years (Fig 3D). The CAR T-cell production process was substantially different for cohort 1 compared with cohorts 2 and 3 (Data Supplement). We compared peak blood CAR⁺ cell levels of patients on each cohort (Data Supplement). Patients on cohort 1 had significantly lower peak CAR⁺ cell numbers compared with patients on cohort 2 or 3.

FIG 3.

Association of higher peak chimeric antigen receptor–positive (CAR⁺) cell levels with anti-lymphoma responses. (A) The peak numbers of blood CAR⁺ cells for CAR T-cell treatments that resulted in best responses of complete remission (CR) are compared with peak numbers of blood CAR⁺ cells for CAR T-cell treatments that resulted in responses of partial remission (PR), stable disease (SD), or progressive disease (PD; n = 41 of 46 total administered treatments). Two treatments were not evaluable due to lack of cell samples for polymerase chain reaction (PCR) analysis, and three treatments were not evaluable due to lack of response assessment data. (B) The levels of blood CAR⁺ cells at 28-56 days after CAR T-cell infusion are compared for treatments resulting in best responses of CR versus PR, SD, or PD (n = 38 of 46 total treatments). Three patients were not evaluable due to lack of response assessment data, and five patients were not evaluable due to lack of samples for PCR. (C) The peak levels of blood CAR⁺ cells are compared for CAR T-cell treatments that resulted in durations of response (DORs) > 3 years versus DORs < 3 years (n = 39 of 46 total treatments). Five treatments lacked lymphoma response follow-up for this analysis, and two lacked PCR data due to lack of blood samples. (D) The levels of blood CAR⁺ cells at 28-56 days after CAR T-cell infusion are compared for treatments resulting in DORs > 3 years versus DORs < 3 years (n = 36 of 46 total treatments). Five treatments lacked lymphoma response follow-up for this analysis, and five lacked PCR data due to lack of blood samples. All CAR⁺ cell values were determined using quantitative PCR with an assay specific for the CAR. Data are presented as CAR⁺ cells per microliter, and all values are rounded to the nearest whole number. Median values are indicated by the black horizontal bars. Each dot indicates an individual patient. All statistical comparisons were done using two-tailed Mann-Whitney U tests. Statistically significant P values are shown on the graph. P < .05 was considered statistically significant. NS, not statistically significant.

B-Cell and Immunoglobulin Recovery

Twenty-four CAR T-cell treatments generated a CR and had blood samples available for assessment of blood CD19+ B-cell levels. Recovery to a normal absolute number of B cells never occurred after nine treatments (38%) after a median follow-up time of 51 months (range, 8-82 months). After 15 (63%) of 24 evaluable CAR T-cell infusions, absolute blood B-cell numbers recovered to normal in a median time of 12 months (range, 2-59 months; Fig 4A). Ten (67%) of 15 patients with B-cell recovery to normal levels remain in evaluable, ongoing CR, with a median time from achievement of a normal B-cell level to most recent follow-up of 50 months (range, 37-112 months).

FIG 4.

Time to recovery of B cells and immunoglobulins after chimeric antigen receptor (CAR) T-cell infusion. (A) The number of blood CD19⁺ cells was determined by flow cytometry before conditioning chemotherapy and at the indicated time points after CAR T-cell infusion (normal range, 61-321 cells/µL). (B) Serum immunoglobulin (Ig) G levels (normal range, 700-1,600 mg/dL) were determined before treatment and at the indicated time points after CAR T-cell infusion. (C) Serum IgA levels (normal range, 70-400 mg/dL) were determined before treatment and at the indicated time points after CAR T-cell infusion. (D) Serum IgM levels (normal range, 40-230 mg/dL) were determined before treatment and at the indicated time points after CAR T-cell infusion. Median values are indicated by the black horizontal bars; the blue boxes include the 25th to 75th percentiles; each dot indicates an individual patient; and the whiskers extend from the minimum to the maximum values. The dashed line on each plot indicates the lower limit of normal for each measured parameter. A total of 24 treatments are presented because there were 24 treatments that resulted in complete remission and had blood samples available for long-term immunoglobulin and B-cell monitoring. The treatments were all for unique patients, except both the first and second treatments for patient 3 are included.

Levels of serum immunoglobulin (Ig) G (Fig 4B), IgA (Fig 4C), and IgM (Fig 4D) were quantified over time and are provided for all evaluable patients who achieved a CR. Six (25%) of 24 evaluable patients reached normal levels of all three immunoglobulins during the study period. All six of these patients also reached normal levels of B cells during the study period. The remaining 18 evaluable patients (75%) had a long-standing abnormality in at least one Ig type. IgA levels did not recover to normal levels in 15 (63%) of 24 patients at last follow-up. Eight (33%) of 24 patients had persistently low IgM levels, and five (21%) of 24 patients had persistently low IgG levels. Intravenous Ig (IVIg) was administered when serum IgG levels decreased to < 400-500 mg/dL. Because IVIgs are made up primarily of IgG,³⁴ administration of IVIgs accounts for a portion of the levels of serum IgG in Figure 4B. Of the 24 patients evaluable for long-term Ig levels, 19 (79%) of 24 received IVIgs at some point after cell infusion; at the most recent follow-up visits, four (17%) of 24 patients continued to receive IVIg.

Patients had reduced T-cell and NK-cell counts after administration of the conditioning chemotherapy regimens. NK cells recovered rapidly. T-cell recovery was prolonged in many patients and incomplete in some patients, especially for CD4+ T cells (Data Supplement). Delayed CD4+ T-cell recovery has been previously reported for patients receiving chemotherapy without CAR T cells.³⁵

DISCUSSION

Our results provide the longest follow-up of patients with B-cell lymphoma treated with anti-CD19 CAR T cells. We demonstrated durable ongoing remissions lasting up to 113 months for follicular lymphoma, 99 months for CLL, and 97 months for DLBCL/PMBCL. Long-term adverse effects from anti-CD19 CAR T-cell therapy were rare.

Higher peak blood CAR⁺ cell levels after CAR T-cell infusions have been associated with response in a variety of CAR T-cell studies.^{9,18,20,36-39} The degree of CAR T-cell persistence needed for durable lymphoma remissions is unclear. We showed that peak peripheral-blood CAR T-cell levels are associated with response but that blood CAR T-cell levels 28-56 days after infusion were not associated with response (Fig 3). Because functionally effective anti-CD19 CAR T cells eliminate CD19+ B cells, recovery of normal CD19+ B cells demonstrates a lack of persisting functionally effective anti-CD19 CAR T cells.²⁸ Ten patients from this study had normal blood B-cell levels and ongoing CRs at last follow-up. These data strongly suggest that permanent persistence of CAR T cells is not required for maintenance of remission in lymphoma. Persistent CAR T cells have been associated with maintenance of remission in acute lymphoblastic leukemia and CLL.^20,39,40 It is possible that different malignancy types or different CAR constructs may have varying requirements for CAR T-cell persistence to maintain remissions.

Cohort 1 had a T-cell production process that lasted 24 days, whereas cohorts 2 and 3 had shorter T-cell production times of 10 days and 6-10 days, respectively (Data Supplement). Peak blood CAR⁺ cell levels were lower for CAR T cells produced on cohort 1 compared with cohorts 2 and 3, which suggests lower proliferative capacity of the cells produced on cohort 1 (Data Supplement).

Long-term adverse effects were rare in this study; five of 43 patients developed a nonhematologic malignancy, one of which was recurrence of prostate cancer that preceded CAR T-cell treatment, and two of 43 patients developed MDS. These rates of second malignancies and MDS are not higher than expected in the patients treated on this trial, who all had a history of substantial chemotherapy exposure before protocol enrollment.^41-43 Another previous concern for gene therapy approaches has been occurrence of RCR. Despite several years of follow-up, we never detected RCR. This is in line with other studies that documented an absence of replication-competent γ-retroviruses and lentiviruses after T-cell gene therapy studies.^44,45

We observed prolonged low B-cell levels and hypogammaglobulinemia in many patients (Fig 4), but the rate of severe infections was low. B-cell depletion and hypogammaglobulinemia are both expected after anti-CD19 CAR T-cell therapy; however, the observation that some patients have low blood B-cell levels and low serum Ig levels for up to 5 years after anti-CD19 CAR T-cell infusion has not been described. We also observed that blood B-cell levels and Ig levels usually improved over several years. Serum levels of IgA were less likely to recover than serum levels of IgG or IgM after anti-CD19 CAR T-cell therapy. It has long been noted that IgG and IgM levels increase more rapidly during child development than IgA.^46,47 It is possible that the slow recovery of IgA in our patients reflects this process of B-cell development.

The main strength of this report is that it provides the longest follow-up of patients with lymphoma treated with anti-CD19 CAR T cells. The durability of lymphoma responses and rarity of significant long-term adverse effects reported here, along with prior reports of high shorter-term response rates after anti-CD19 CAR T-cell therapy, argue strongly that anti-CD19 CAR T-cell therapy should be the preferred treatment of relapsed DLBCL.^8,9,13,17,18 The role of allogeneic stem-cell transplantation for refractory DLBCL is being re-evaluated in the context of CAR T-cell therapies.⁴⁸ Our results with anti-CD19 CAR T cells support the idea that allogeneic stem-cell transplantation should be reserved for patients who have persisting lymphoma after CAR T-cell therapy. Our results and results from other studies suggest that anti-CD19 CAR T cells will play an increasing role in treatment of follicular lymphoma, mantle cell lymphoma, and CLL.^11,19,21,22 The durable remissions reported here raise the possibility, but do not prove, that anti-CD19 CAR T cells are curative for CLL and some types of B-cell lymphoma.

DATA SHARING STATEMENT

A data sharing statement provided by the authors is available in the Supplement tab of this article at DOI https://doi.org/10.1200/JCO.20.01467.

AUTHOR CONTRIBUTIONS

Conception and design: Richard M. Sherry, Steven A. Rosenberg, James N. Kochenderfer

Financial support: James N. Kochenderfer

Collection and assembly of data: Kathryn M. Cappell, Richard M. Sherry, James C. Yang, Stephanie L. Goff, Danielle A. Vanasse, Lori McIntyre, James N. Kochenderfer

Data analysis and interpretation: Kathryn M. Cappell, Richard M. Sherry, James C. Yang, Stephanie L. Goff, Steven A. Rosenberg, James N. Kochenderfer

Manuscript writing: All authors

Final approval of manuscript: All authors

Accountable for all aspects of the work: All authors

AUTHORS' DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST

Long-Term Follow-Up of Anti-CD19 Chimeric Antigen Receptor T-Cell Therapy

The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated unless otherwise noted. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO's conflict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/jco/authors/author-center.

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James C. Yang

Research Funding: Kite, a Gilead Company (Inst)

Patents, Royalties, Other Intellectual Property: Royalties on inventions may go to my institution (National Cancer Institute), which may pass a portion (anonymously) to me

Steven A. Rosenberg

Research Funding: Kite Pharma (Inst), Iovance Biotherapeutics (Inst), ZIOPHARM Oncology (Inst)

Patents, Royalties, Other Intellectual Property: Patents held by National Institutes of Health

James N. Kochenderfer

Research Funding: Kite, a Gilead Company (Inst), Celgene (Inst)

Patents, Royalties, Other Intellectual Property: I have received royalties from Kite, a Gilead Company, as an inventor on patents describing new chimeric antigen receptors

Travel, Accommodations, Expenses: Kite, a Gilead Company (Inst)

No other potential conflicts of interest were reported.