EVIDENCE THAT THE <scp>L</scp>-ASPARAGINASE OF GUINEA PIG SERUM IS RESPONSIBLE FOR ITS ANTILYMPHOMA EFFECTS

Broome, J. D.

doi:10.1084/jem.118.1.121

Cited by 154 publications

(49 citation statements)

References 14 publications

(9 reference statements)

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“…1B presents a schematic of the neutral drift screen as applied to the chemotherapeutic enzyme L-Asparaginase II (EcAII, EC 3.5.1.1). EcAII has been a cornerstone component of chemotherapeutic protocols for the treatment of ALL for over 40 y (30)(31)(32)(33). In ALL, lymphoblasts lack or express low levels of L-asparagine synthetase (AS) (34) and therefore require the uptake of L-Asn from serum for cell proliferation (6).…”

mentioning

confidence: 99%

Therapeutic enzyme deimmunization by combinatorial T-cell epitope removal using neutral drift

Cantor¹,

Yoo²,

Dixit

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

108

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A number of heterologous enzymes have been investigated for cancer treatment and other therapeutic applications; however, immunogenicity issues have limited their clinical utility. Here, a new approach has been created for heterologous enzyme deimmunization whereby combinatorial saturation mutagenesis is coupled with a screening strategy that capitalizes on the evolutionary biology concept of neutral drift, and combined with iterative computational prediction of T-cell epitopes to achieve extensive reengineering of a protein sequence for reduced MHC-II binding propensity without affecting catalytic and pharmacological properties. Escherichia coli L-asparaginase II (EcAII), the only nonhuman enzyme approved for repeated administration, is critical in treatment of childhood acute lymphoblastic leukemia (ALL), but elicits adverse antibody responses in a significant fraction of patients. The neutral drift screening of combinatorial saturation mutagenesis libraries at a total of 12 positions was used to isolate an EcAII variant containing eight amino acid substitutions within computationally predicted T-cell epitopes-of which four were nonconservative-while still exhibiting k cat ∕K M ¼ 10 6 M −1 s −1 for L-Asn hydrolysis. Further, immunization of HLA-transgenic mice expressing the ALL-associated DRB1*0401 allele with the engineered variant resulted in significantly reduced T-cell responses and a 10-fold reduction in anti-EcAII IgG titers relative to the existing therapeutic. This significant reduction in the immunogenicity of EcAII may be clinically relevant for ALL treatment and illustrates the potential of employing neutral drift screens to achieve large jumps in sequence space as may be required for the deimmunization of heterologous proteins. directed evolution | protein engineering

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mentioning

confidence: 99%

Therapeutic enzyme deimmunization by combinatorial T-cell epitope removal using neutral drift

Cantor¹,

Yoo²,

Dixit

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

108

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“…Surprisingly this value is virtually the same in both sensitive and resistant forms of 6C3HED. In resistant tumors the level falls even lower during the next 20 hr, to 17.7 nmoles/g, a change which has no detectable effect on the growth rate of palpable tumors and produces no microscopic evidence of damage (4). By contrast, sensitive tumors with similar asparagine levels show marked cytotoxic effects at 6 hr, and at 20 hr, by which time the free asparagine level has risen to 58 nmoles/g, they are substantially necrotic (1).…”

Section: Asparagine Levels Of Normal Mouse Tissues and Of 6c3hed Lympmentioning

confidence: 92%

“…Earlier work has established that aspamginase-sensitive cells of Lymphoma 6C3HED are L-asparagine dependent in tissue culture, while resistant variants are asparaglne independent (4). Asparagine is a constituent of most if not all proteins, and when deprived of the amino acid, protein synthesis in sensitive cells is inhibited, as has been demonstrated by Sobin and Kidd, and by others (11,12).…”

mentioning

confidence: 85%

“…The identification of L-asparaginase as the constituent of guinea pig serum responsible for its tumor-inhibitory properties is now generally accepted (1)(2)(3)(4)(5). A number of other asparaginases having high substrate avidity have recently been found to ~nh~bit tumors, notably, as first shown by Wriston and his coworkers, enzymes from Esckerickia coli and other bacteria (6--8).…”

mentioning

confidence: 99%

“…The subline, as others used earlier, is completely resistant and has maintained the character continuously. In the absence of asparaginase treatment both the sensitive line and resistant sublines have the same growth rate in vivo (4).…”

mentioning

confidence: 99%

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Studies on the Mechanism of Tumor Inhibition by L-Asparaginase

Broome

1968

The Journal of Experimental Medicine

Self Cite

193

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L-asparaginases of agouti serum and Escherichia coli cause a profound lowering in the level of free asparagine in the blood of treated mice and also in the tissues. During treatment, normal tissues and resistant 6C3HED lymphomas survive unharmed with intracellular asparagine levels which are critically low for sensitive lymphomas. An explanation for this contrast between the two types of lymphoma is provided by the finding that resistant cells have not only a higher asparagine synthetic capacity than sensitive cells but appear able to utilize endogenous asparagine preferentially for protein synthesis. Cell-free extracts of resistant cells contain an asparaginase synthetase, but this is not found in preparations from sensitive cells.

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Treatment of lymphosarcoma in the dog with L-asparaginase

Old

Boyse

Campbell

et al. 1967

Cancer

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Three dogs with advanced lymphosarcoma were treated with L‐asparaginase from E. coli. All three responded to therapy by marked regression of lymph nodes and dramatic improvement in general condition without evidence of toxicity. Two dogs returned temporarily to normal health and were clinically free of disease; the other showed a marked partial remission. One of the three dogs has remained in good health for 50 days after treatment was discontinued although the disease now shows signs of recurrence. Early relapse in the other two cases may be attributed to insufficient dosage of L‐asparaginase.

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EVIDENCE THAT THE L-ASPARAGINASE OF GUINEA PIG SERUM IS RESPONSIBLE FOR ITS ANTILYMPHOMA EFFECTS

Cited by 154 publications

References 14 publications

Therapeutic enzyme deimmunization by combinatorial T-cell epitope removal using neutral drift

Therapeutic enzyme deimmunization by combinatorial T-cell epitope removal using neutral drift

Studies on the Mechanism of Tumor Inhibition by L-Asparaginase

Treatment of lymphosarcoma in the dog with L-asparaginase

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